benzofurans and efaroxan

Prolonged stimulation of insulin secretion by depolarization and Ca2+ influx regularly leads to a reversible state of decreased secretory responsiveness to nutrient and nonnutrient stimuli. This state is termed "desensitization." The onset of desensitization may occur within 1 h of exposure to depolarizing stimuli. Desensitization by exposure to sulfonylureas, imidazolines, or quinine produces a marked cross-desensitization against other ATP-sensitive K+ channel (KATP channel)-blocking secretagogues. However, desensitized beta-cells do not necessarily show changes in KATP channel activity or Ca2+ handling. Care has to be taken to distinguish desensitization-induced changes in signaling from effects due to the persisting presence of secretagogues. The desensitization by depolarizing secretagogues is mostly accompanied by a reduced content of immunoreactive insulin and a marked reduction of secretory granules in the beta-cells. In vitro recovery from a desensitization by the imidazoline efaroxan was nearly complete after 4 h. At this time point the depletion of the granule content was partially reversed. Apparently, recovery from desensitization affects the whole lifespan of a granule from biogenesis to exocytosis. There is, however, no direct relation between the beta-cell granule content and the secretory responsiveness. Even though a prolonged exposure of isolated islets to depolarizing secretagogues is often associated with the occurrence of ultrastructural damage to beta-cells, we could not find a cogent link between depolarization and Ca2+ influx and apoptotic or necrotic beta-cell death.

Topics: Animals; Benzofurans; Calcium; Humans; Idazoxan; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Membrane Potentials; Potassium Channels

Topics: Agmatine; Animals; Benzofurans; Down-Regulation; Imidazoles; Imidazoline Receptors; Insulin; Insulin Secretion; Islets of Langerhans; Receptors, Drug

We have tested, in a prospective randomized, double-blind, placebo-controlled, crossover, 12-week study, the effects of 2 mg efaroxan, a potent alpha-2 antagonist, given three times per day to 14 patients with progressive supranuclear palsy. Efaroxan did not induce any significant change on any motor assessment criteria. The present data do not confirm the assumption that the blockade of alpha-2 receptors might be a useful pharmacologic strategy to improve patients with progressive supranuclear palsy.

Topics: Adrenergic alpha-Antagonists; Adult; Aged; Benzofurans; Cross-Over Studies; Dose-Response Relationship, Drug; Double-Blind Method; Drug Administration Schedule; Female; Humans; Imidazoles; Male; Middle Aged; Neurologic Examination; Prospective Studies; Supranuclear Palsy, Progressive; Treatment Outcome

Agmatine (AG), idazoxan (IDZ), and efaroxan (EFR) are imidazoline receptor ligands with beneficial effects in central nervous system disorders. The present study aimed to evaluate the interaction between AG, IDZ, and EFR with an opiate, tramadol (TR), in a conditioned place preference (CPP) paradigm. In the experiment, we used five groups with 8 adult male Wistar rats each. During the condition session, on days 2, 4, 6, and 8, the rats received the drugs (saline, or TR, or IDZ and TR, or EFR and TR, or AG and TR) and were placed in their least preferred compartment. On days 1, 3, 5, and 7, the rats received saline in the preferred compartment. In the preconditioning, the preferred compartment was determined. In the postconditioning, the preference for one of the compartments was reevaluated. TR increased the time spent in the non-preferred compartment. AG decreased time spent in the TR-paired compartment. EFR, more than IDZ, reduced the time spent in the TR-paired compartment, but without statistical significance. AG reversed the TR-induced CPP, while EFR and IDZ only decreased the time spent in the TR-paired compartment, without statistical significance.

Topics: Agmatine; Analgesics, Opioid; Animals; Benzofurans; Conditioning, Psychological; Drug Interactions; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Rats; Rats, Wistar; Time Factors; Tramadol

Based on the finding that a central antihypertensive agent with high affinity for I1-type imidazoline receptors - rilmenidine, shows cytotoxic effects on cultured cancer cell lines, it has been suggested that imidazoline receptors agonists might have a therapeutic potential in the cancer therapy. Nevertheless, potential rilmenidine side effects caused by activation of α-adrenoceptors, or other associated receptors and enzymes, might hinder its therapeutic benefits. Considering that human α-adrenoceptors belong to the rhodopsin-like class A of G-protein-coupled receptors (GPCRs) it is reasonable to assume that imidazolines might have the affinity for other receptors from the same class. Therefore, to investigate possible off-target effects of imidazoline ligands we have prepared a reverse docking protocol on class A GPCRs, using imidazoline ligands and their decoys. To verify our in silico results, three ligands with high scores and three ligands with low scores were tested for antagonistic activity on α

Topics: Animals; Area Under Curve; Benzofurans; CHO Cells; Cricetulus; Humans; Idazoxan; Imidazoles; Imidazolines; Ligands; Molecular Docking Simulation; Receptors, Adrenergic, alpha-2; Receptors, G-Protein-Coupled; Reproducibility of Results

Stimulation of α2-adrenoceptor/I1-imidazoline receptors in the rostral ventrolateral medulla decreases the blood pressure via sympathoinhibition. However, alteration of receptor responses in genetically hypertensive rats remains unclear. We examined cardiovascular responses of α2-adrenoceptor/I1-imidazoline receptor agonist and antagonists microinjected into the rostral ventrolateral medulla of conscious spontaneously hypertensive rats and normotensive Wistar Kyoto rats. Injection of 2-nmol clonidine-an α2-adrenoceptor/I1-imidazoline receptor agonist-unilaterally into the rostral ventrolateral medulla decreased the blood pressure, heart rate, and renal sympathetic nerve activity; the responses were significantly enhanced in spontaneously hypertensive rats than in Wistar Kyoto rats. Co-injection of 2-nmol 2-methoxyidazoxan (a selective α2-adrenoceptor antagonist) or 2-nmol efaroxan (an I1-receptor antagonist) with 2 nmol of clonidine attenuated the hypotensive and bradycardic effects of clonidine-only injection. Injection of 2-methoxyidazoxan alone increased the blood pressure and heart rate in spontaneously hypertensive rats, but not in Wistar Kyoto rats. These results suggest enhanced responsiveness of α2-adrenoceptor/I1-imidazoline receptors in the rostral ventrolateral medulla of spontaneously hypertensive rats.

Topics: Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Blood Pressure Determination; Clonidine; Consciousness; Heart Rate; Hypertension; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Medulla Oblongata; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Receptors, Adrenergic, alpha-2; Sympathetic Nervous System

Locomotor sensitization to repeated ethanol (EtOH) administration is proposed to play a role in early and recurring steps of addiction. The present study was designed to examine the effect of agmatine on EtOH-induced locomotor sensitization in mice.. Mice received daily single intraperitoneal injection of EtOH (2.5 g/kg, 20 v/v) for 7 consecutive days. Following a 3-day EtOH-free phase, the mice were challenged with EtOH on day 11 with a single injection of EtOH. Agmatine (10 to 40 μg/mouse), endogenous agmatine enhancers (l-arginine [80 μg/mouse], arcaine [50 μg/mouse], aminoguanidine [25 μg/mouse]), and imidazoline receptor agonist/antagonists were injected (intracerebroventricular [i.c.v.]) either daily before the injection of EtOH during the 7-day development phase or on days 8, 9, and 10 (EtOH-free phase). The horizontal locomotor activity was determined on days 1, 3, 5, 7, and 11.. Agmatine (20 to 40 μg/mouse) administration for 7 days (development phase) significantly attenuated the locomotor sensitization response of EtOH challenge on day 11. Further, the agmatine administered only during EtOH-free period (days 8, 9, and 10) also inhibited the enhanced locomotor activity on the 11th day to EtOH challenge as compared to control mice indicating blockade of expression of sensitization. Daily treatment (i.c.v.) with endogenous agmatine enhancers like l-arginine (80 μg/mouse) or arcaine (50 μg/mouse) and aminoguanidine (25 μg/mouse) restrained the development as well as expression of sensitization to EtOH. Imidazoline I. Inhibition of EtOH sensitization by agmatine is mediated through imidazoline receptors and project agmatine and imidazoline agents in the pharmacotherapy of alcohol addiction.

Topics: Agmatine; Animals; Arginine; Benzofurans; Biguanides; Central Nervous System Sensitization; Dose-Response Relationship, Drug; Drug Interactions; Ethanol; Guanidines; Idazoxan; Imidazoles; Imidazoline Receptors; Infusions, Intraventricular; Male; Mice; Microinjections; Motor Activity

Long-term use of opioid analgesics is limited by tolerance development and undesirable adverse effects. Paradoxically, spinal administration of ultra-low-dose (ULD) G-protein-coupled receptor antagonists attenuates analgesic tolerance. Here, we determined whether systemic ULD α2-adrenergic receptor (AR) antagonists attenuate the development of morphine tolerance, whether these effects extend to the cannabinoid (CB1) receptor system, and if behavioral effects are reflected in changes in opioid-induced spinal gliosis. Male rats were treated daily with morphine (5 mg/kg) alone or in combination with ULD α2-AR (atipamezole or efaroxan; 17 ng/kg) or CB1 (rimonabant; 5 ng/kg) antagonists; control groups received ULD injections only. Thermal tail flick latencies were assessed across 7 days, before and 30 min after the injection. On day 8, spinal cords were isolated, and changes in spinal gliosis were assessed through fluorescent immunohistochemistry. Both ULD α2-AR antagonists attenuated morphine tolerance, whereas the ULD CB1 antagonist did not. In contrast, both ULD atipamezole and ULD rimonabant attenuated morphine-induced microglial reactivity and astrogliosis in deep and superficial spinal dorsal horn. So, although paradoxical effects of ULD antagonists are common to several G-protein-coupled receptor systems, these may not involve similar mechanisms. Spinal glia alone may not be the main mechanism through which tolerance is modulated.

Topics: Adrenergic alpha-2 Receptor Antagonists; Analgesics; Analgesics, Opioid; Animals; Benzofurans; Cannabinoid Receptor Antagonists; Cannabinoids; Dose-Response Relationship, Drug; Drug Tolerance; Gliosis; Imidazoles; Injections, Spinal; Male; Morphine; Neuroglia; Norepinephrine; Pain Measurement; Rats; Rats, Sprague-Dawley; Receptor, Cannabinoid, CB1; Rimonabant; Spine

The dorsal diencephalic conduction system connects limbic forebrain structures to monaminergic mesencephalic nuclei via a distinct relay station, the habenular complexes. Both habenular nuclei, the lateral as well as the medial nucleus, are considered to play a prominent role in mental disorders like major depression. Herein, we investigate the effect of the polyamine agmatine on the electrical activity of neurons within the medial habenula in rat. We present evidence that agmatine strongly decreases spontaneous action potential firing of medial habenular neurons by activating I1-type imidazoline receptors. Additionally, we compare the expression patterns of agmatinase, an enzyme capable of inactivating agmatine, in rat and human habenula. In the medial habenula of both species, agmatinase is similarly distributed and observed in neurons and, in particular, in distinct neuropil areas. The putative relevance of these findings in the context of depression is discussed. It is concluded that increased activity of the agmatinergic system in the medial habenula may strengthen midbrain dopaminergic activity. Consequently, the habenular-interpeduncular axis may be dysregulated in patients with major depression.

Topics: Action Potentials; Agmatine; Animals; Benzofurans; Depression; Female; Habenula; Humans; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Middle Aged; Neurons; Rats, Wistar; Ureohydrolases

The purpose of this work was a comparative analysis of the brain adrenergic and imidazoline receptors activation effects in laboratory rats with normal and elevated activity of the sympathetic nervous system. The experiments were carried out on male rats of Wistar and Wistar-Kyoto lines and spontaneously hypertensive (SHR) line. In freely moving rats the mean arterial pressure, heart beat-to-beat interval and cardiochronotropic component of baroreceptor reflex were recorded and analyzed at rest and emotional stress, caused by a conditioned signal. Agonist of imidazoline and adrenergic receptor clonidine, a primary agonist of imidazoline receptor moxonidine, selective alpha 2-adrenoreceptor blocker yohimbine and blocker of imidazoline and alpha - adrenergic receptor efaroksan were used for neurochemical analysis of adrenergic and imidazoline systems role in mechanisms of regulation of blood circulation. Our data have shown, that if adrenergic systems of the brain provide its participation in the long-term regulation of blood pressure, stimulation of imidazoline receptors activate the central link of baroreceptor reflex arc at the level of the medulla oblongata and also strengthening the processes of its suprabulbar modulation, caused by emotional stress.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Baroreflex; Benzofurans; Blood Pressure; Brain; Clonidine; Heart Rate; Imidazoles; Imidazoline Receptors; Male; Rats; Rats, Inbred SHR; Rats, Wistar; Sympathetic Nervous System; Yohimbine

Hyperglycemia induces damage of vascular endothelial cells leading to diabetic complications. We investigated the effects of insulinotropic compounds and elevated glucose on endothelial cells in the absence or presence of vascular endothelial growth factor (VEGF).. Human umbilical vein endothelial cells (HUVECs) were treated with glibenclamide, repaglinide and insulinotropic imidazolines at high glucose concentration in the presence or absence of VEGF and viability, proliferation and nitric oxide production were measured. Hyperglycemia inhibited pro-survival effects of VEGF on endothelial cells. Glibenclamide and repaglinide decreased HUVEC viability at elevated glucose concentration in the absence but not in the presence of VEGF, without affecting HUVEC proliferation. Repaglinide also had some positive influence on HUVEC function elevating NO production in the presence of VEGF. Imidazolines showed different activities on endothelial cell survival. Efaroxan diminished HUVEC viability at elevated glucose concentration in the presence, however not in the absence of VEGF, while RX871024 decreased HUVEC survival regardless of the presence of VEGF.. Our data demonstrate an important interplay between the actual insulinotropic compounds, VEGF and ambient glucose concentration affecting the survival of the vascular endothelial cells. Consequently, this interplay needs to be taken into consideration when designing novel oral antidiabetic compounds.

Topics: Benzofurans; Carbamates; Cell Proliferation; Cell Survival; Cells, Cultured; Dimethyl Sulfoxide; Glucose; Glyburide; Human Umbilical Vein Endothelial Cells; Humans; Hypoglycemic Agents; Imidazoles; Indoles; Nitric Oxide; Piperidines; Vascular Endothelial Growth Factor A

Centrally acting antihypertensive action of moxonidine is a result of activation of Imidazoline-1 receptor (I1R) in the rostral ventrolateral medulla (RVLM). Hypertension shows an increase in reactive oxygen species (ROS) in the RVLM. The present objective was to determine the phosphoinositide-3 kinase (PI3K) signaling pathway involved in the effect of moxonidine on ROS generation in the RVLM of spontaneously hypertensive rat (SHR).. Wistar-Kyoto rats and SHR received intracisternal infusion (2 weeks) of tested agents which were subjected to subsequent experiments. In-situ ROS in the RVLM was evaluated by the oxidative fluorescence dye. Western blot and PCR analysis were performed to detect the expression levels of PI3K signaling pathway. Lentivirus was injected bilaterally into the RVLM for silencing PI3K signaling.. ROS production in the RVLM was dose-dependently reduced in SHRs treated with infusion of moxonidine (20 nmol/day), which was prevented by the I1R antagonist efaroxan but not by the α2-adrenoceptor antagonist yohimbine. Moxonidine pretreatment significantly blunted cardiovascular sensitivity to injection of tempol (5 nmol) or angiotensin II (10 pmol) into the RVLM in SHR. Expression levels of PI3K/Akt, nuclear factor kappa-B (NFκB), NADPHase (NOX4), and angiotensin type I receptor (AT1R) in the RVLM were markedly decreased in SHR treated with moxonidine. Infection of lentivirus containing PI3K shRNA in the RVLM effectively prevented effects of moxonidine on cardiovascular activity and expression levels of Akt, NFκB, NOX4, and AT1R.. The centrally antihypertensive drug moxonidine decreases ROS production in the RVLM through inactivation of the PI3K/Akt signaling pathway in hypertension.

Topics: Animals; Antihypertensive Agents; Benzofurans; Disease Models, Animal; Hypertension; Imidazoles; Male; Medulla Oblongata; Phosphatidylinositol 3-Kinases; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Reactive Oxygen Species; Signal Transduction

Synthesis and hypotensive properties of centrally acting imidazoline agents: 1-[(imidazolidin-2-yl)imino]-1H-indazole (Marsanidine) and 7-chloro-1-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-1H-indazole (TCS-80) were tested in rats. We have recently synthesized two novel Marsanidine analogues which decrease blood pressure and heart rate in rats: 1-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-1H-indole (TCS-54), and 7-chloro-1-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-1H-indole (TCS-213). Among all these analogues, compound TCS-80 exhibits the highest affinity to I1-imidazoline receptors and the lowest α2/I1 selectivity ratio. The observed cardiovascular effects of the compounds might be mediated through α2-adrenergic and I1-imidazoline receptors and subsequent decrease of the symphathetic nerve activity. The present studies were performed to determine whether α2-adrenergic and/or I1-imidazoline receptors are involved in the decrease of blood pressure and heart rate induced by Marsanidine, TCS-54, TCS-80, and TCS-213 in rats.. Anesthetized rats were infused iv with the tested compounds and selective α2-adrenoceptor antagonist, RX821002, or nonselective α2-adrenergic/I1-imidazoline receptor antagonist, Efaroxan. The mean arterial blood pressure and heart rate were monitored directly and continuously throughout the experiment.. Efaroxan inhibited the hypotensive effect of TCS-80 stronger than RX821002. The degree of inhibition of the hypotensive effect of the remaining compounds was similar for both antagonists. The presence of Efaroxan and RX821002 diminished the heart rate decrease induced by all compounds administration, though the influence on the maximal chronotropic effect was attenuated significantly in the TCS-80 and TCS-213 treated animals only.. Our results indicate that hypotensive and negative chronotropic activities of all tested compounds are mediated by both the α2-adrenergic and I1-imidazoline receptors. Moreover, the circulatory effect of TCS-80 might be mediated to relatively higher degree by the I1-imidazoline receptors than by the α2-adrenergic ones.

Topics: Animals; Benzofurans; Blood Pressure; Heart Rate; Hypotension; Idazoxan; Imidazoles; Imidazolidines; Imidazoline Receptors; Imidazolines; Indazoles; Male; Rats; Receptors, Adrenergic, alpha-2

This in vivo study assessed the potential of the imidazoline receptor (IR) ligands moxonidine (selective I1-IR), BU224 (selective I2-IR) and LSL61122 (mixed I1/I2-IR) to dampen excitotoxic signalling induced by kainic acid (KA; 45 mg/kg) in the mouse brain (hippocampus and cerebral cortex). KA triggered a strong behavioural syndrome (seizures; maximal at 60-90 minutes) and sustained stimulation (at 72 hours with otherwise normal mouse behaviour) of pro-apoptotic c-Jun-N-terminal kinases (JNK) and calpain with increased cleavage of p35 into neurotoxic p25 (cyclin-dependent kinase 5 [Cdk5] activators) in mouse hippocampus. Pretreatment (five days) with LSL61122 (10 mg/kg), but not moxonidine (1 mg/kg) or BU224 (20 mg/kg), attenuated the KA-induced behavioural syndrome, and all three IR ligands inhibited JNK and calpain activation, as well as p35/p25 cleavage after KA in the hippocampus (effects also observed after acute IR drug treatments). Efaroxan (I1-IR, 10 mg/kg) and idazoxan (I2-IR, 10 mg/kg), postulated IR antagonists, did not antagonise the effects of moxonidine and LSL61122 on KA targets (these IR ligands showed agonistic properties inhibiting pro-apoptotic JNK). Brain subcellular preparations revealed reduced synaptosomal postsynaptic density-95 protein contents (a mediator of JNK activation) and indicated increased p35/Cdk5 complexes (with pro-survival functions) after treatment with moxonidine, BU224 and LSL61122. These results showed that I1- and I2-IR ligands (moxonidine and BU224), and especially the mixed I1/I2-IR ligand LSL61122, are partly neuroprotective against KA-induced excitotoxic signalling. These findings suggest a therapeutic potential of IR drugs in disorders associated with glutamate-mediated neurodegeneration.

Topics: Animals; Benzofurans; Cerebral Cortex; Cyclin-Dependent Kinase 5; Glutamic Acid; Hippocampus; Idazoxan; Imidazoles; Imidazoline Receptors; Kainic Acid; Ligands; Male; Mice; Neurodegenerative Diseases; Neuroprotective Agents; Styrenes; Time Factors

Experimental studies in preclinical mouse models of breast cancer have shown that chronic restraint stress can enhance disease progression by increasing catecholamine levels and subsequent signaling of β-adrenergic receptors. Catecholamines also signal α-adrenergic receptors, and greater α-adrenergic signaling has been shown to promote breast cancer in vitro and in vivo. However, antagonism of α-adrenergic receptors can result in elevated catecholamine levels, which may increase β-adrenergic signaling, because pre-synaptic α2-adrenergic receptors mediate an autoinhibition of sympathetic transmission. Given these findings, we examined the effect of α-adrenergic blockade on breast cancer progression under non-stress and stress conditions (chronic restraint) in an orthotopic mouse model with MDA-MB-231HM cells. Chronic restraint increased primary tumor growth and metastasis to distant tissues as expected, and non-selective α-adrenergic blockade by phentolamine significantly inhibited those effects. However, under non-stress conditions, phentolamine increased primary tumor size and distant metastasis. Sympatho-neural gene expression for catecholamine biosynthesis enzymes was elevated by phentolamine under non-stress conditions, and the non-selective β-blocker propranolol inhibited the effect of phentolamine on breast cancer progression. Selective α2-adrenergic blockade by efaroxan also increased primary tumor size and distant metastasis under non-stress conditions, but selective α1-adrenergic blockade by prazosin did not. These results are consistent with the hypothesis that α2-adrenergic signaling can act through an autoreceptor mechanism to inhibit sympathetic catecholamine release and, thus, modulate established effects of β-adrenergic signaling on tumor progression-relevant biology.

Topics: Adrenergic alpha-1 Receptor Antagonists; Adrenergic alpha-2 Receptor Antagonists; Animals; Benzofurans; Cell Proliferation; Disease Progression; Female; Imidazoles; Mammary Neoplasms, Experimental; Mice; Neoplasm Metastasis; Prazosin; Restraint, Physical; Signal Transduction; Stress, Physiological; Stress, Psychological

Various imidazoline receptor (IR) proteins have been proposed to mediate the effects of selective I1- and I2-IR drugs. However, the association of these IR-binding proteins with classic I1- and I2-radioligand binding sites remains somewhat controversial. In this study, three IR antibodies (anti-NISCH and anti-nischarin for I1-IRs; and anti-IRBP for I1/I2-IRs) were used to immunodetect, characterize and compare IR protein patterns in brain (mouse and human; total homogenate, subcellular fractionation, grey and white matter) and some cell systems (neurones, astrocytes, human platelets). Various immunoreactive IRs (specific molecular weight bands coincidently detected with the different antibodies) were related to I1-IR (167 kDa, 105/115 kDa and 85 kDa proteins) or I2-IR (66 kDa, 45 kDa and 30 kDa proteins) types. The biochemical characterization of cortical 167 kDa protein, localized in the membrane/cytosol but not in the nucleus, indicated that this I1-IR also forms part of higher order nischarin-related complexes. The contents of I1-IR (167 kDa, 105/115 kDa, and 85 kDa) proteins in mouse brain cortex were upregulated by treatment with I1-drugs (moxonidine, efaroxan) but not with I2-drugs (BU-224, LSL 61122). Conversely, the contents of I2-IR (66 kDa, 45 kDa and 30 kDa) proteins in mouse brain cortex were modulated by treatment with I2-drugs (decreases after BU-224 and LSL 61122, and increases after idazoxan) but not with I1-drugs (with the exception of moxonidine). These findings further indicate that brain immunoreactive IR proteins exist in multiple forms that can be grouped in the already known I1- and I2-IR types, which are expressed both in neurones and astrocytes.

Topics: Adult; Animals; Antibodies; Benzofurans; Binding Sites; Brain; Cell Membrane; Cytosol; Female; Humans; Idazoxan; Imidazoles; Imidazoline Receptors; Imidazolines; Male; Mice; Styrenes; Up-Regulation

Evidence suggests that intracellular Ca(2+) levels and contractility of cardiomyocytes can be modulated by targeting receptors other than already identified adrenergic or non-adrenergic sarcolemmal receptors. This study uncovers the presence in myocardial cells of adrenergic α2 (α2-AR) and imidazoline I1 (I1R) receptors. In isolated left ventricular myocytes generating stationary spontaneous Ca(2+) transients in the absence of triggered action potentials, the prototypic agonist of both receptors agmatine can activate corresponding signaling cascades with opposing outcomes on nitric oxide (NO) synthesis and intracellular Ca(2+) handling. Specifically, activation of α2-AR signaling through PI3 kinase and Akt/protein kinase B stimulates NO production and abolishes Ca(2+) transients, while targeting of I1R signaling via phosphatidylcholine-specific phospholipase C (PC-PLC) and protein kinase C (PKC) suppresses NO synthesis and elevates averaged intracellular Ca(2+). We identified that endothelial NO synthase (eNOS) is a major effector for both signaling cascades. According to the established eNOS transitions between active (Akt-dependent) and inactive (PKC-dependent) conformations, we suggest that balance between α2-AR and I1R signaling pathways sets eNOS activity, which by defining operational states of myocellular sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) can adjust Ca(2+) re-uptake and thereby cardiac inotropy. These results indicate that the conventional catalog of cardiomyocyte sarcolemmal receptors should be expanded by the α2-AR and I1R populations, unveiling previously unrecognized targets for endogenous ligands as well as for existing and potential pharmacological agents in cardiovascular medicine.

Topics: Adrenergic alpha-2 Receptor Agonists; Agmatine; Animals; Benzofurans; Calcium Signaling; Cells, Cultured; Imidazoles; Imidazoline Receptors; Myocytes, Cardiac; Nitric Oxide; Nitric Oxide Synthase Type III; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, Adrenergic, alpha-2

CR4056 is a novel imidazoline-2 (I2 ) ligand exhibiting potent analgesic activity in animal models of pain. In this study, we investigated the effects of CR4056 in a well-established model of postoperative pain where rats develop hyperalgesia in the injured hind paw.. By measuring paw withdrawal threshold to mechanical pressure, we studied the pharmacology of CR4056, potential sex differences in pain perception and response to treatment, and the pharmacodynamic interaction of CR4056 with morphine.. Oral CR4056 and subcutaneous morphine dose-dependently reversed the hyperalgesic response. Analgesic effects of CR4056 were completely suppressed by the non-selective imidazoline I2 /α2 -adrenoceptor antagonist idazoxan, were partially reduced (~30%; P < 0.05) by the selective α2 -adrenoceptor antagonist yohimbine, but were not influenced by the non-selective I1 /α2 -adrenoceptor antagonist efaroxan or by the μ opioid receptor antagonist naloxone. We found no differences in responses to CR4056 or morphine between male and female rats. However, females had a lower pain threshold than males, and needed lower doses of drugs to reach a significant analgesia. When CR4056 and morphine were combined, their median effective doses were lower than expected for additive effects, both in males and in females. Isobolographic analysis confirmed a synergism between CR4056 and morphine.. CR4056 is a novel pharmacological agent under development for postoperative pain both as stand-alone treatment and in association with morphine. CR4056 has successfully completed Phase I studies for tolerability and pharmacokinetics in healthy volunteers, and is currently entering the first proof-of-concept study in patients.

Topics: Adrenergic alpha-2 Receptor Antagonists; Analgesics; Analgesics, Opioid; Animals; Benzofurans; Binding Sites; Drug Synergism; Female; Hyperalgesia; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Morphine; Naloxone; Narcotic Antagonists; Pain, Postoperative; Quinazolines; Rats, Sprague-Dawley; Yohimbine

Although bupropion has been widely used in the treatment of depression, the precise mechanism of its therapeutic actions is not fully understood. The present study investigated the role of agmatine in an antidepressant like effect of bupropion in mouse forced swim test. The antidepressant like effect of bupropion was potentiated by pretreatment with agmatine (10-20mg/kg, ip) and by the drugs known to increase endogenous agmatine levels in brain viz., l-arginine (40 μg/mouse, icv), an agmatine biosynthetic precursor, ornithine decarboxylase inhibitor, dl-α-difluoromethyl ornithine hydrochloride, DFMO (12.5 μg/mouse, icv), diamine oxidase inhibitor, aminoguanidine (6.5 μg/mouse, icv) and agmatinase inhibitor, arcaine (50 μg/mouse, icv) as well as imidazoline I1 receptor agonists, moxonidine (0.25mg/kg, ip) and clonidine (0.015 mg/kg, ip) and imidazoline I2 receptor agonist, 2-(2-benzofuranyl)-2-imidazoline hydrochloride, 2-BFI (5mg/kg, ip). Conversely, prior administration of I1 receptor antagonist, efaroxan (1mg/kg, ip) and I2 receptor antagonist, idazoxan (0.25mg/kg, ip) blocked the antidepressant like effect of bupropion and its synergistic combination with agmatine. These results demonstrate involvement of agmatine in the antidepressant like effect of bupropion and suggest agmatine and imidazoline receptors as a potential therapeutic target for the treatment of depressive disorders.

Topics: Agmatine; Animals; Antidepressive Agents; Arginine; Benzofurans; Biguanides; Bupropion; Clonidine; Dose-Response Relationship, Drug; Drug Interactions; Eflornithine; Guanidines; Idazoxan; Imidazoles; Immobility Response, Tonic; Injections, Intraventricular; Male; Mice; Motor Activity

Enhancement of renal sympathetic nerve activity during renal ischemia and norepinephrine overflow from the kidney after reperfusion play important roles in the development of ischemic acute kidney injury. Recently, we have found that moxonidine, an α2/imidazoline Ι1-receptor agonist, has preventive effects on ischemic acute kidney injury by suppressing the excitation of renal sympathetic nervous system after reperfusion. In the present study, to clarify the renoprotective mechanisms of moxonidine (360 nmol/kg, i.v.) against ischemic acute kidney injury, we investigated the effect of intravenous (i.v.) and intracerebroventricular (i.c.v.) injection of efaroxan, an α2/Ι1 receptor antagonist, on the moxonidine-exhibited actions. Ischemic acute kidney injury was induced by clamping the left renal artery and vein for 45 min followed by reperfusion, 2 weeks after contralateral nephrectomy. The suppressive effect of moxonidine on enhanced renal sympathetic nerve activity during renal ischemia was not observed in the rat treated with either i.v. (360 nmol/kg) or i.c.v. (36 nmol/kg) of efaroxan. Furthermore, i.v. injection of efaroxan eliminated the preventive effect of moxonidine on ischemia/reperfusion-induced kidney injury and norepinephrine overflow, and i.c.v. injection of efaroxan did not completely inhibit the moxonidine's effects. These results indicate that moxonidine prevents the ischemic kidney injury by sympathoinhibitory effect probably via α2/Ι1 receptors in central nervous system and by suppressing the norepinephrine overflow through α2/Ι1 receptors on sympathetic nerve endings.

Topics: Acute Kidney Injury; Animals; Benzofurans; Cytoprotection; Imidazoles; Imidazoline Receptors; Kidney; Male; Norepinephrine; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sympathetic Nervous System; Veins

The potentiation of oxycodone antinociception by BMS182874 (endothelin-A (ET(A)) receptor antagonist) and agmatine (imidazoline receptor/α(2)-adrenoceptor agonist) is well-documented. It is also known that imidazoline receptors but not α(2)-adrenoceptors are involved in potentiation of oxycodone antinociception by agmatine and BMS182874 in mice. However, the involvement of specific imidazoline receptor subtypes (I(1), I(2), or both) in this interaction is not clearly understood. The present study was conducted to determine the involvement of imidazoline I(1) and I(2) receptors in agmatine- and BMS182874-induced potentiation of oxycodone antinociception in mice. Antinociceptive (tail flick and hot-plate) latencies were determined in male Swiss Webster mice treated with oxycodone, agmatine, BMS182874, and combined administration of oxycodone with agmatine or BMS182874. Efaroxan (imidazoline I(1) receptor antagonist) and BU224 (imidazoline I(2) receptor antagonist) were used to determine the involvement of I(1) and I(2) imidazoline receptors, respectively. Oxycodone produced significant antinociceptive response in mice which was not affected by efaroxan but was blocked by BU224. Agmatine-induced potentiation of oxycodone antinociception was blocked by BU224 but not by efaroxan. Similarly, BMS182874-induced potentiation of oxycodone antinociception was blocked by BU224 but not by efaroxan. This is the first report demonstrating that BMS182874- or agmatine-induced enhancement of oxycodone antinociception is blocked by BU224 but not by efaroxan. We conclude that imidazoline I(2) receptors but not imidazoline I(1) receptors are involved in BMS182874- and agmatine-induced potentiation of oxycodone antinociception in mice.

Topics: Agmatine; Analgesics, Opioid; Animals; Benzofurans; Dansyl Compounds; Drug Synergism; Imidazoles; Imidazoline Receptors; Male; Mice; Oxycodone; Pain Measurement

The role of imidazoline receptors in the regulation of vascular function remains unclear. In this study, we evaluated the effect of agmatine, an imidazoline receptor agonist, on systolic blood pressure (SBP) in spontaneously hypertensive rats (SHRs) and investigated the expressions of imidazoline receptors by Western blot. The isometric tension of aortic rings isolated from male SHRs was also estimated. Agmatine decreased SBP in a dose-dependent manner in SHRs but not in the normal group [Wistar-Kyoto (WKY) rats]. This reduction in SBP in SHRs was abolished by BU224, a selective antagonist of imidazoline I(2) -receptors. Higher expression of imidazoline receptors in SHR was observed. Moreover, agmatine-induced relaxation in isolated aortic rings precontracted with phenylephrine or KCl. This relaxation was also abolished by BU224 but was not modified by efaroxan, an imidazoline I(1) -receptor antagonist. Agmatine-induced relaxation was also attenuated by PNU 37883, a selective blocker of vascular ATP-sensitive potassium (K(ATP) ) channels. Additionally, vasodilatation by agmatine was reduced by an inhibitor of protein kinase A (PKA). We suggest that agmatine can lower blood pressure in SHRs through activation of the peripheral imidazoline I(2) -receptor, which is expressed more highly in SHRs.

Topics: Agmatine; Animals; Antihypertensive Agents; Aorta, Thoracic; Benzofurans; Blood Pressure; Blotting, Western; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelium, Vascular; Humans; Hypertension; Imidazoles; Imidazoline Receptors; KATP Channels; Male; Potassium Channel Blockers; Protein Kinase Inhibitors; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Up-Regulation; Vasodilation; Vasodilator Agents

The group of imidazoline-1 receptors (I(1)-IR) agonists encompasses drugs are currently used in treatment of high blood pressure and hyperglycemia. The I(1)-IR protein structures have not been determined yet, but Nischarin protein that binds numerous imidazoline ligands inducing initiation of various cell-signaling cascades, including apoptosis, is identified as strong I(1)-IR candidate. In this study we examined apoptotic activity of rilmenidine (potent I(1)-IR agonist), moxonidine (moderate I(1)-IR agonist), and efaroxan (I(1)-IR partial agonist) on cancer cell line (K562) expressing Nischarin. The Nischarine domains mapping was performed by use of the Informational Spectrum Method (ISM). The 3D-Quantitative Structure-Activity Relationship (3D-QSAR) and virtual docking studies of 29 I(1)-IR ligands (agonists, partial agonists, and antagonists) were carried out on I(1)-IR receptors binding affinities. The 3D-QSAR study defined 3D-pharmacophore models for I(1)-IR agonistic and I(1)-IR antagonistic activity and created regression model for prediction of I(1)-IR activity of novel compounds. The 3D-QSAR models were applied for design and evaluation of novel I(1)-IR agonists and I(1)-IR antagonists. The most promising I(1)-IR ligands with enhanced activities than parent compounds were proposed for synthesis. The results of 3D-QSAR, ISM, and virtual docking studies were in perfect agreement and allowed precise definition of binding mode of I(1)-IR agonists (Arg 758, Arg 866, Val 981, and Glu 1057) and significantly different binding modes of I(1)-IR antagonists or partial I(1)-IR agonists. The performed theoretical study provides reliable system for evaluation of I(1)-IR agonistic and I(1)-IR antagonistic activity of novel I(1)-IR ligands, as drug candidates with anticancer activities.

Topics: Antineoplastic Agents; Apoptosis; Benzofurans; Cell Proliferation; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Humans; Imidazoles; Imidazoline Receptors; K562 Cells; Ligands; Models, Molecular; Molecular Docking Simulation; Oxazoles; Quantitative Structure-Activity Relationship; Rilmenidine; Tumor Cells, Cultured

Ultra-low doses of alpha-2 (α2)-adrenoceptor antagonists augment spinal morphine antinociception and inhibit tolerance, but the role of receptor specificity in these actions is unknown. We used the stereo-isomers of the α2 adrenoceptor antagonist, efaroxan to evaluate the effect of receptor specificity on the induction of spinal morphine tolerance and hyperalgesia. Tail flick and paw pressure tests were first used to evaluate high dose efaroxan (12.6 μg) and its stereo-isomers on clonidine analgesia in intrathecally catheterized rats. Ultra-low doses of individual isomers (1.3 ng) were then co-administered with morphine (15 μg) to determine their effects on acute antinociceptive tolerance and hyperalgesia induced by low dose spinal morphine (0.05 ng). Results demonstrate that high dose (+) efaroxan antagonized clonidine-induced antinociception, while (-) efaroxan had minimal effect. In addition, an ultra-low dose of (+) efaroxan (1.3 ng), substantially lower than required for receptor blockade, inhibited the development of acute morphine tolerance, while (-) efaroxan was less effective. Racemic (±) efaroxan effects were similar to those of (+) efaroxan. Furthermore, low dose morphine (0.05 ng) produced sustained hyperalgesia in the tail flick test and this was blocked by co-injection of (+) but not (-) efaroxan (1.3 ng). Given the isomer-specific efaroxan effects and their different receptor potencies, we suggest that inhibition of opioid tolerance by ultra-low dose efaroxan involves a specific interaction with spinal α2-adrenoceptors in this model. Likewise, inhibitory effects of adrenoceptor antagonists on morphine tolerance may be due to blockade of opioid-induced hyperalgesia.

Topics: Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-Antagonists; Analgesics; Animals; Benzofurans; Clonidine; Drug Tolerance; Hot Temperature; Hyperalgesia; Imidazoles; Injections, Spinal; Male; Morphine; Pain; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-2; Touch

Agmatine, an endogenous ligand for imidazoline receptor, has been shown to prevent opioid dependence, but not much is known about the mechanisms of the effect of agmatine. In the present study, we investigated the function of I1 imidazoline receptor and its candidate protein imidazoline receptor antisera-selected (IRAS)/Nischarin in morphine dependence as well as in the effect of agmatine inhibiting morphine dependence by pharmacological and molecular approaches. Results showed that inhibition of IRAS or Nischarin did not change the development of morphine dependence in vitro and in vivo under the basal condition. Agmatine could reduce the cyclic 3', 5' adenosine monophosphate (cAMP) overshoot at the concentration of 0.01-10 µM in the primary cultured rat hippocampal neurons and attenuated the withdrawal signals and the elevation of FosB and ΔFosB at the dose of 5 mg/kg in the morphine-dependent mice. The effect of agmatine was inhibited by efaroxan (I1 imidazoline receptor non-specific antagonist) and the RNA interference against IRAS or Nischarin. These findings indicate that I1 imidazoline receptor or IRAS/Nischarin mediates the effect of agmatine on morphine dependence and provide evidence that I1 imidazoline receptor may be a new target for treating morphine dependence.

Topics: Adenosine Monophosphate; Adrenergic alpha-Antagonists; Agmatine; Animals; Benzofurans; Cells, Cultured; Drug Interactions; Gene Knockdown Techniques; Hippocampus; Imidazoles; Imidazoline Receptors; Immune Sera; Intracellular Signaling Peptides and Proteins; Male; Mice; Mice, Inbred BALB C; Morphine Dependence; Neurons; Proto-Oncogene Proteins c-fos; RNA, Small Interfering; Substance Withdrawal Syndrome

Allantoin is known as the agonist of imidazoline receptor, especially the I₂ subtype. Effect of allantoin on imidazoline I₁ receptor (I₁R) relating to reduction of blood pressure and its merit in steatosis are still obscure. Also, farnesoid X receptor (FXR) plays an important role in lipid homeostasis related to I₁R activation. Thus, we administered allantoin into high fat diet (HFD)-fed mice showing hypertriglyceridemia and hypercholesterolemia. Allantoin significantly improved hyperlipidemia in HFD mice after 4 weeks of administration. Pretreatment with efaroxan, at a dose sufficient to inhibit I₁R activation, attenuated the action of allantoin. In addition, in cultured HepG2 cells, allantoin increased the expression of farnesoid X receptor (FXR). The allantoin-induced FXR expression was blocked by efaroxan. Similar changes were observed in the expressions of FXR-targeted genes. Otherwise, allantoin also lowered systolic blood pressure (SBP) in HFD mice that can be blocked by efaroxan. Taken together, allantoin has an ability to activate I₁R for improvement of metabolic disorders.

Topics: Allantoin; Animals; Anticholesteremic Agents; Antihypertensive Agents; Benzofurans; Diet, High-Fat; Fatty Liver; Hep G2 Cells; Humans; Hypercholesterolemia; Hypertriglyceridemia; Hypolipidemic Agents; Imidazoles; Imidazoline Receptors; Liver; Male; Mice; Mice, Inbred C57BL; Receptors, Cytoplasmic and Nuclear; Receptors, LDL; Up-Regulation

Imidazoline I1-receptors (I1R) are known to regulate blood pressure and rilmenidine, an agonist, is widely used as antihypertensive agent in clinic. However, the role of I1R in feeding behavior is still unclear. In the present study, we used the agonist of I1R to investigate the effect on hyperphagia in streptozotocin (STZ)-induced diabetic mice. Rilmenidine decreased the food intake of STZ-diabetic mice in a dose-dependent manner. The reduction of food intake was abolished by pretreatment with efaroxan at the dose sufficient to block I1R. Intracerebroventricular (icv) administration of rilmenidine into STZ-diabetic mice also significantly reduced hyperphagia, which was reversed by icv administration of efaroxan. In addition, similar results were observed in STZ-diabetic mice, which received chronic treatment with rilmenidine 3 times daily (t.i.d.) for 7 days. Moreover, the hypothalamic neuropeptide Y (NPY) level was reduced by rilmenidine that was also reversed by pretreatment with efaroxan. In conclusion, the obtained results suggest that rilmenidine can decrease food intake in STZ-diabetic mice through an activation of I1R to lower hypothalamic NPY level.

Topics: Animals; Benzofurans; Cerebrum; Diabetes Mellitus, Type 1; Disease Models, Animal; Eating; Female; Humans; Hyperphagia; Hypothalamus; Imidazoles; Imidazoline Receptors; Male; Mice; Mice, Inbred BALB C; Neuropeptide Y; Oxazoles; Rilmenidine; Streptozocin

Inspired by recent speculation about the potential utility of α(2A)-antagonism in the treatment of type 2 diabetes, the study examined the contribution of α(2)-antagonism vs other mechanisms to the antihyperglycaemic activity of the imidazoline (±)-efaroxan.. Effects of the racemate and its pure enantiomers on isolated pancreatic islets and beta cells in vitro, as well as on hyperglycaemia in vivo, were investigated in a comparative manner in mice.. In isolated perifused islets, the two enantiomers of efaroxan were equally potent in counteracting inhibition of insulin release by the ATP-dependent K(+) (K(ATP)) channel-opener diazoxide but (+)-efaroxan, the presumptive carrier of α(2)-antagonistic activity, was by far superior in counteracting inhibition of insulin release by the α(2)-agonist UK14,304. In vivo, (+)-efaroxan improved oral glucose tolerance at 100-fold lower doses than (-)-efaroxan and, in parallel with observations made in vitro, was more effective in counteracting UK14,304-induced than diazoxide-induced hyperglycaemia. The antihyperglycaemic activity of much higher doses of (-)-efaroxan was associated with an opposing pattern (i.e. with stronger counteraction of diazoxide-induced than UK14,304-induced hyperglycaemia), which implicates a different mechanism of action.. The antihyperglycaemic potency of (±)-efaroxan in mice is almost entirely due to α(2)-antagonism, but high doses can also lower blood glucose via another mechanism. Our findings call for reappraisal of the possible clinical utility of α(2A)-antagonistic compounds in recently identified subpopulations of patients in which a congenitally higher level of α(2A)-adrenergic activation contributes to the development and pathophysiology of type 2 diabetes.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Brimonidine Tartrate; Calcium; Cells, Cultured; Diabetes Mellitus, Type 2; Diazoxide; Female; Hyperglycemia; Imidazoles; Insulin; Insulin-Secreting Cells; KATP Channels; Male; Membrane Potentials; Mice; Mice, Inbred C57BL; Patch-Clamp Techniques; Quinoxalines; Yohimbine

TVP1022, the S-enantiomer of rasagiline (Azilect®) (N-propargyl-1R-aminoindan), exerts cyto/cardio-protective effects in a variety of experimental cardiac and neuronal models. Previous studies have demonstrated that the protective activity of TVP1022 and other propargyl derivatives involve the activation of p42/44 mitogen-activated protein kinase (MAPK) signaling pathway. In the current study, we further investigated the molecular mechanism of action and signaling pathways of TVP1022 which may account for the cyto/cardio-protective efficacy of the drug. Using specific receptor binding and enzyme assays, we demonstrated that the imidazoline 1 and 2 binding sites (I(1) & I(2)) are potential targets for TVP1022 (IC(50) =9.5E-08 M and IC(50) =1.4E-07 M, respectively). Western blotting analysis showed that TVP1022 (1-20 µM) dose-dependently increased the immunoreactivity of phosphorylated p42 and p44 MAPK in rat pheochromocytoma PC12 cells and in neonatal rat ventricular myocytes (NRVM). This effect of TVP1022 was significantly attenuated by efaroxan, a selective I(1) imidazoline receptor antagonist. In addition, the cytoprotective effect of TVP1022 demonstrated in NRVM against serum deprivation-induced toxicity was markedly inhibited by efaroxan, thus suggesting the importance of I(1)imidazoline receptor in mediating the cardioprotective activity of the drug. Our findings suggest that the I(1)imidazoline receptor represents a novel site of action for the cyto/cardio-protective efficacy of TVP1022.

Topics: Animals; Benzofurans; Binding Sites; Blotting, Western; Cardiotonic Agents; Dose-Response Relationship, Drug; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Indans; Inhibitory Concentration 50; Mitogen-Activated Protein Kinase 1; Myocytes, Cardiac; PC12 Cells; Rats; Signal Transduction

ATP-sensitive potassium channels (K(ATP) channels) in beta cells are a major target for insulinotropic drugs. Here, we studied the effects of selected stimulatory and inhibitory pharmacological agents in islets lacking K(ATP) channels.. We compared insulin secretion (IS) and cytosolic calcium ([Ca(2+)](c)) changes in islets isolated from control mice and mice lacking sulphonylurea receptor1 (SUR1), and thus K(ATP) channels in their beta cells (Sur1KO).. While similarly increasing [Ca(2+)](c) and IS in controls, agents binding to site A (tolbutamide) or site B (meglitinide) of SUR1 were ineffective in Sur1KO islets. Of two non-selective blockers of potassium channels, quinine was inactive, whereas tetraethylammonium was more active in Sur1KO compared with control islets. Phentolamine, efaroxan and alinidine, three imidazolines binding to K(IR)6.2 (pore of K(ATP) channels), stimulated control islets, but only phentolamine retained weaker stimulatory effects on [Ca(2+)](c) and IS in Sur1KO islets. Neither K(ATP) channel opener (diazoxide, pinacidil) inhibited Sur1KO islets. Calcium channel blockers (nimodipine, verapamil) or diphenylhydantoin decreased [Ca(2+)](c) and IS in both types of islets, verapamil and diphenylhydantoin being more efficient in Sur1KO islets. Activation of alpha(2)-adrenoceptors or dopamine receptors strongly inhibited IS while partially (clonidine > dopamine) lowering [Ca(2+)](c) (control > Sur1KO islets).. Those drugs retaining effects on IS in islets lacking K(ATP) channels, also affected [Ca(2+)](c), indicating actions on other ionic channels. The greater effects of some inhibitors in Sur1KO than in control islets might be relevant to medical treatment of congenital hyperinsulinism caused by inactivating mutations of K(ATP) channels.

Topics: Animals; Benzofurans; Calcium; Cytosol; Diazoxide; Female; Imidazoles; Imidazolines; Insulin; Insulin Secretion; Insulin-Secreting Cells; KATP Channels; Mice; Mice, Knockout; Phentolamine; Pinacidil; Potassium Channels; Tolbutamide

The excitation of renal sympathetic nervous system plays an important role in the development of ischemic acute kidney injury in rats. Recently, we found that agmatine, an adrenaline alpha(2)/imidazoline I(1)-receptor agonist, has preventive effects on ischemic acute kidney injury by suppressing the enhanced renal sympathetic nerve activity during renal ischemia and by decreasing the renal venous norepinephrine overflow after reperfusion. In the present study, we investigated preventive mechanisms of agmatine against ischemic acute kidney injury in rats. Ischemic acute kidney injury was induced by clamping the left renal artery and vein for 45 min followed by reperfusion, 2 weeks after the contralateral nephrectomy. Pretreatment with efaroxan (30 mumol/kg, i.v.), an alpha(2)/I(1)-receptor antagonist, abolished the suppressive effects of agmatine on the enhanced renal sympathetic nerve activity during renal ischemia and on the elevated norepinephrine overflow after reperfusion, and eliminated the preventing effects of agmatine on the ischemia/reperfusion-induced renal dysfunction and histological damage. On the other hand, pretreatment with yohimbine (6 mumol/kg, i.v.), an alpha(2)-receptor antagonist, eliminated the preventing effects of agmatine on the ischemia/reperfusion-induced renal injury and norepinephrine overflow, without affecting the lowering effect of agmatine on renal sympathetic nerve activity. These results indicate that agmatine prevents the ischemic renal injury by sympathoinhibitory effect probably via I(1) receptors in central nervous system and by suppressing the norepinephrine overflow through alpha(2) or I(1) receptors on sympathetic nerve endings.

Topics: Adrenergic alpha-2 Receptor Antagonists; Agmatine; Animals; Benzofurans; Imidazoles; Imidazoline Receptors; Kidney Diseases; Male; Norepinephrine; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sympathetic Nervous System; Veins; Yohimbine

The purpose of this study is to assess the change of uveoscleral outflow induced by moxonidine and to investigate whether the increase of uveoscleral outflow induced by moxonidine is mediated by alpha1, alpha2, or I1 receptors.. 0.05% moxonidine was topically and unilaterally administered in rabbit eyes with or without pretreatment of prazosin, yohimbine, efaroxan, or AGN 192403, as indicated. We injected fluorescein isothiocyanate-bovine serum albumin (FITC-BSA) into the anterior chamber and observed the fluorescence intensity of the uveoscleral outflow. Finally, the volume of uveoscleral outflow was calculated based on the fluorescence intensities captured.. A bilateral increase of fluorescence intensity was observed along the uveoscleral outflow pathway following moxonidine administration, especially in the ciliary body and supraciliochoroidal space. Pretreatment with prazosin further enhanced the bilateral increase of fluorescence intensity at between 2 and 4 hours after moxonidine administration. The response of moxonidine was antagonized by either yohimbine, an alpha2 receptor antagonist, or efaroxan, an I1/alpha2 receptor antagonist. The antagonizing effect of yohimbine was more potent than that of efaroxan. The moxonidne-induced response was not antagonized by AGN 192403, an I1 receptor antagonist. The bilateral volumes of aqueous humor within the uveoscleral pathway increased significantly induced by moxonidine (p < 0.01 versus control). The increased bilateral volumes of uveoscleral outflow were 0.381 +/- 0.073 and 0.376 +/- 0.095 mu l/min, respectively.. These results suggest that topical, unilateral administration of moxonidine causes a bilateral increase of aqueous humor via the uveoscleral outflow pathway. The moxonidine-induced increase of uveoscleral outflow is mediated by alpha2 adrenergic receptors, not by I1 imidazoline receptors.

Topics: Administration, Topical; Adrenergic alpha-Antagonists; Animals; Aqueous Humor; Benzofurans; Bridged Bicyclo Compounds; Drug Synergism; Fluorescein-5-isothiocyanate; Heptanes; Imidazoles; Imidazoline Receptors; Prazosin; Rabbits; Receptors, Adrenergic, alpha-2; Sclera; Serum Albumin, Bovine; Uvea; Yohimbine

To determine the effects of the centrally antihypertensive drug moxonidine injected into the rostral ventrolateral medulla (RVLM) on baroreflex function in spontaneously hypertensive rats (SHR).. Baroreflex sensitivity control of renal sympathetic nerve activity (RSNA) and barosensitivity of the RVLM presympathetic neurons were determined following application of different doses of moxonidine within the RVLM.. Three doses (0.05, 0.5, and 5 nmol in 50 nL) of moxonidine injected bilaterally into the RVLM dose-dependently reduced the baseline blood pressure (BP) and RSNA in SHR. At the highest dose (5 nmol) of moxonidine injection, the maximum gain (1.24%+/-0.04%/mmHg) of baroreflex control of RSNA was significantly decreased. However, the lower doses (0.05 and 0.5 nmol) of moxonidine injection into the RVLM significantly enhanced the baroreflex gain (2.34%+/-0.08% and 2.01%+/-0.07%/mmHg). The moxonidine-induced enhancement in baroreflex function was completely prevented by the imidazoline receptor antagonist efaroxan but not by the alpha(2)-adrenoceptor antagonist yohimbine. A total of 48 presympathetic neurons were recorded extracellularly in the RVLM of SHR. Iontophoresis of applied moxonidine (30-60 nA) dose-dependently decreased the discharge of RVLM presympathetic neurons but also significantly increased the barosensitivity of RVLM presympathetic neurons.. These data demonstrate that a low dose of moxonidine within the RVLM has a beneficial effect on improving the baroreflex function in SHR via an imidazoline receptor-dependent mechanism.

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Baroreflex; Benzofurans; Blood Pressure; Dose-Response Relationship, Drug; Imidazoles; Imidazoline Receptors; Iontophoresis; Kidney; Medulla Oblongata; Microinjections; Neurons; Rats; Rats, Inbred SHR; Sympathetic Nervous System; Synaptic Potentials; Yohimbine

To explore the therapeutic potential of imidazoline I(1) receptor ligands in motor dysfunction related to the basal ganglia, rigidity was induced in mice by intraperitoneal administration of reserpine. The imidazoline I(1) receptor agonists moxonidine and tizanidine reduced rigidity in a dose-dependent manner. Although rigidity was reduced by efaroxan (an imidazoline I(1) receptor and alpha(2)-adrenoceptor antagonist) and idazoxan (an imidazoline I(1) and I(2) receptor and alpha(2)-adrenoceptor antagonist), SKF86466 and yohimbine, both of which are alpha(2)-adrenoceptor antagonists with no affinity for imidazoline receptors, also suppressed rigidity, suggesting that activation rather than blockade of imidazoline I(1) receptors contributes to reduction of reserpine-induced muscle rigidity.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antiparkinson Agents; Benzazepines; Benzofurans; Clonidine; Disease Models, Animal; Dose-Response Relationship, Drug; Electromyography; Idazoxan; Imidazoles; Imidazoline Receptors; Injections, Intraperitoneal; Ligands; Male; Mice; Muscle Rigidity; Muscle, Skeletal; Parkinsonian Disorders; Reserpine; Time Factors; Yohimbine

An insufficient number of insulin-producing beta-cells is a major cause of defective control of blood glucose in both type 1 and type 2 diabetes. The aim of this study was to clarify whether the insulinotropic imidazolines can affect the survival of highly proliferating insulin-secreting cells, here exemplified by the MIN6 cell line. Our data demonstrate that RX871024, but not efaroxan, triggered MIN6 cell death and potentiated death induced by a combination of the pro-inflammatory cytokines interleukin-1beta, interferon- gamma and tumor necrosis factor-alpha. These effects did not involve changes in nitric oxide production but correlated with stimulation of c-jun N-terminal kinase (JNK) activity and activation of caspases-1, -3, -8 and -9. Our results suggest that the imidazoline RX871024 causes death of highly proliferating insulin-secreting cells, putatively via augmentation of JNK activity, a finding that may impact on the possibility of using compounds of similar activity in the treatment of diabetes.

Topics: Animals; Benzofurans; Caspases; Cell Death; Cell Line; Cell Proliferation; Cytokines; Enzyme Activation; Humans; Imidazoles; Indoles; Insulin-Secreting Cells; JNK Mitogen-Activated Protein Kinases; Mice; Nitric Oxide

A partial recovery of locomotor functions has been shown in spinal cord-transected (Tx) cats after regular treadmill training and repeated administration of clonidine, an alpha(2)-adrenoreceptor agonist. However, clonidine has generally failed to show prolocomotor effects in other models (e.g., rat or mudpuppy in vitro-isolated spinal cord preparations). The reasons for this discrepancy remain unclear, but they may suggest condition- or species-specific effects induced by clonidine. This study is aimed at examining both the acute (at 6 or 41 days post-Tx) and chronic effects of repeated (once a week for one month) clonidine administration (0.25-5.0 mg/kg i.p.) on hindlimb movement generation in Tx mice (thoracic segment9/10). Locomotor-like (LM) and nonlocomotor movements (NLM) were assessed both in open-field and treadmill conditions. The results show that clonidine consistently failed, in both conditions, to induce LM and NLM at all time points even though control experiments revealed hindlimb movements steadily induced by 8-hydroxy-2-(di-N-propylamino)-tetralin (8-OH-DPAT), a serotonin receptor agonist. In turn, clonidine acutely suppressed (I(1)-imidazoline receptor-mediated) the frequency of spontaneously occurring LM and NLM but apparently increased spinal excitability over time, because the frequency of spontaneous LM and NLM was significantly greater in clonidine-treated (before an injection) than vehicle-treated animals after repeated administration for a few weeks. The results clearly show that clonidine can not acutely induce hindlimb movements in untrained and otherwise nonstimulated (e.g., no tail or perineal pinching) Tx mice, although repeated administration may progressively facilitate the expression of spontaneous hindlimb movements.

Topics: Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Clonidine; Hindlimb; Imidazoles; Imidazoline Receptors; Male; Mice; Mice, Inbred Strains; Movement; Receptors, Adrenergic, alpha-2; Spinal Cord; Spinal Cord Injuries; Yohimbine

The present study examined the antinociceptive effect of diphenyl diselenide (PhSe)2, given orally (p.o.), in the hot-plate test in mice. The administration of diphenyl diselenide (10-100 mg/kg, p.o.) caused a significant inhibition of thermal nociception induced by hot-plate test in mice. Pretreatment of animals by intraperitoneal route (i.p.) with caffeine (10 mg/kg; a non-specific adenosine receptor antagonist) and PSB1115 (1 mg/kg; an adenosine A(2B) receptor antagonist), but not DPCPX (2 mg/kg; an adenosine A(1) receptor antagonist) and SCH5826 (3 mg/kg; an adenosine A(2A) receptor antagonist) significantly blockaded the antinociceptive effect caused by diphenyl diselenide (10 mg/kg, p.o.) in the hot-plate test. Moreover, the pretreatment of animals with efaroxan (1 mg/kg, i.p.; a mixed I(1) imidazoline/alpha(2)-adrenoceptor antagonist) and idazoxan (3 mg/kg, i.p.; a mixed I(2) imidazoline/alpha(2)-adrenoceptor antagonist) did not significantly reverse the antinociception caused by oral administration of diphenyl diselenide (10 mg/kg, p.o.) in the hot-plate test. These results indicate that diphenyl diselenide produced antinociception in a thermal model of pain in mice and its effect was prevented by caffeine and by a selective adenosine A(2B) receptor, but not by imidazoline receptor antagonists in mice.

Topics: Adrenergic alpha-Antagonists; Animals; Behavior, Animal; Benzene Derivatives; Benzofurans; Caffeine; Disease Models, Animal; Dose-Response Relationship, Drug; Hyperalgesia; Imidazoles; Injections, Intraperitoneal; Male; Mice; Neuroprotective Agents; Organoselenium Compounds; Pain Measurement; Phosphodiesterase Inhibitors; Pyrimidines; Triazoles; Xanthines

The imidazoline-type alpha2-adrenoceptor antagonists (+/-)-efaroxan and phentolamine increase insulin secretion and reduce blood glucose levels. It is not known whether they act by antagonizing pancreatic beta-cell alpha2-adrenoceptors or by alpha2-adrenoceptor-independent mechanisms. Many imidazolines inhibit the pancreatic beta-cell KATP channel, which is the molecular target of sulphonylurea drugs used in the treatment of type II diabetes. To investigate the mechanisms of action of (+/-)-efaroxan and phentolamine, alpha2A-adrenoceptor knockout (alpha2A-KO) mice were used.. Effects of (+/-)-efaroxan, 5 mg kg(-1), and phentolamine, 1 mg kg(-1), on blood glucose and insulin levels were compared with those of the non-imidazoline alpha2-adrenoceptor antagonist [8aR,12aS,13aS]-5,8,8a,9,10,11,12,12a,13,13a-decahydro-3-methoxy-12-(ethylsulphonyl)-6H-isoquino[2,1-g][1,6]naphthyridine (RS79948-197), 1 mg kg(-1), and the sulphonylurea glibenclamide, in alpha2A-KO and control (wild type (WT)) mice.. In fed WT mice, (+/-)-efaroxan, phentolamine and RS79948-197 reduced blood glucose and increased insulin levels. Fasting abolished these effects. In fed alpha2A-KO mice, (+/-)-efaroxan, phentolamine and RS79948-197 did not alter blood glucose or insulin levels, and in fasted alpha2A-KO mice, blood glucose levels were increased. Glibenclamide, at a dose only moderately efficacious in WT mice (5 mg kg(-1)), caused severe hyperinsulinaemia and hypoglycaemia in alpha2A-KO mice. This was mimicked in WT mice by co-administration of RS79948-197 with glibenclamide.. These results suggest that (+/-)-efaroxan and phentolamine increase insulin secretion by inhibition of beta-cell alpha2A-adrenoceptors, and demonstrate a critical role for alpha2A-adrenoceptors in limiting sulphonylurea-induced hyperinsulinaemia and hypoglycaemia.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Adrenergic beta-Antagonists; Animals; Atropine Derivatives; Benzofurans; Blood Glucose; Drug Synergism; Fasting; Glyburide; Hypoglycemic Agents; Imidazoles; Insulin; Insulin Secretion; Isoquinolines; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscarinic Antagonists; Naphthyridines; Phentolamine; Propranolol; Receptors, Adrenergic, alpha-2

The aim of the study was to investigate the possible role of MK-801, an NMDA antagonist, in analgesia induced by rilmenidine, an imidazoline (I(1)) agonist, in mice in the formalin test.. 25 microl of formalin 2.5% was injected into the dorsal surface of the right hind paw of the mouse. Pain response was scored after formalin injection for a period of 50 min. A weighted average of nociceptive score, ranging from 0 to 3, was calculated. The mean +/-SEM of scores between 0-5 and 15-40 min after formalin injection was presented.. The study showed that rilmenidine (1.25, 2.5 and 5 mg/kg, i.p.) produced analgesia dose-dependently (p<0.001) in formalin test. In addition, the results demonstrated that efaroxan (0.1 and 1 mg/kg, i.p.) could reduce the antinociceptive effect of rilmenidine (2.5 mg/kg, i.p.) (p<0.01) in animals, however, yohimbine (0.1 and 0.2 mg/kg, i.p.) could not block the analgesia induced by rilmenidine (2.5 mg/kg, i.p.) (p>0.05). On the other hand, MK-801 (0.05 mg/kg, i.p.) reduced the pain related behaviors in mice (p>0.05). Moreover, our findings demonstrated that MK-801 (0.01 mg/kg, i.p.) could potentiate the analgesic effect of rilmenidine (1.25 mg/kg, i.p.) significantly (p<0.01).. The present study suggests that imidazoline (I(1)) receptors play an important role in mediating the antinociception induced by rilmenidine in formalin test. Furthermore, it may be concluded that there is an interaction between NMDA receptors and imidazoline (I(1)) binding sites.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Analgesics; Animals; Benzofurans; Binding Sites; Binding, Competitive; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Excitatory Amino Acid Antagonists; Imidazoles; Imidazoline Receptors; Male; Mice; Nervous System; Neurons, Afferent; Nociceptors; Oxazoles; Pain; Pain Measurement; Receptors, Drug; Receptors, N-Methyl-D-Aspartate; Rilmenidine; Yohimbine

The quality control of chiral drugs requires the determination of their enantiomeric purity. Nowadays, circular dichroism (CD) spectroscopy is gaining increasing importance in pharmaceutical analysis because of the commercially available CD detector in liquid chromatography. The separation of the two enantiomers of a basic drug (efaroxan) was achieved by high performance liquid chromatography using an amylose-derivated column with both UV and CD detections. A baseline-resolved separation (resolution: 5) was obtained after optimization of the mobile phase composition with hexane-ethanol-diethylamine (90:10:0.05; v/v/v). The use of a commercial low-pass electronic noise filter of the CD signal has improved the signal-to-noise ratio by a factor twelve and allowed the quantitation of each enantiomer in the 1.25-300 microg ml(-1) concentration range. The CD linear calibration curve, expressed in terms of stereoisomer height ratio versus concentration ratio, was plotted over the 0.4-6% range. A correlation coefficient greater than 0.999 was obtained by least-squares regression and the limit of detection for the distomer/eutomer ratio was estimated at 0.14%. Although the method validation showed good repeatability on the retention times (RSD < 0.9%), on the peak height ratios (RSD < 8.7%) of each enantiomer only up to 99.2% enantiomeric purity was achieved.

Topics: Adrenergic alpha-Antagonists; Benzofurans; Chromatography, High Pressure Liquid; Circular Dichroism; Imidazoles; Sensitivity and Specificity; Stereoisomerism

The known HPLC method using an achiral C8 silica sorbent and a circular dichroism (CD) detector for the determination of efaroxan enantiomeric excess has been validated. After optimization of the mobile phase, the enantiomers were detected at 278 nm offering maximum ellipticity between two optically active forms. The calibration curve of the anisotropy factor (g) versus the enantiomeric excess was linear with a correlation coefficient (r2) of 0.9985. The accuracy of the method was assessed by comparing the enantiomeric excess obtained by measuring the g factor (C8 column, CD and UV detections) with those determined by enantioselective HPLC (Chiralpak AD-H column, UV detection). Statistical tests (level of confidence of 95%) were assessed to compare the two orthogonal methods. The straight line gave a correlation coefficient of 0.9995, an intercept not significantly different from zero (0.0549) and a slope of 1.026. The precision evaluated on retention time (RSD<0.6%), g factor (RSD<8.3%) and CD peak area (RSD<7.5%) was suitable both in term of intra- and inter-day precisions. The proposed method has the advantages of being fast and precise without using expensive chiral column. Non-enantioselective HPLC-CD was suitable for the simultaneous determination of the optical and chemical purity of efaroxan.

Topics: Benzofurans; Calibration; Chromatography, High Pressure Liquid; Circular Dichroism; Imidazoles; Reproducibility of Results; Sensitivity and Specificity; Spectrophotometry, Ultraviolet; Stereoisomerism

This study investigated the involvement of the imidazoline receptors in the antidepressant-like effect of agmatine in the forced swimming test. The antidepressant-like effects of agmatine (10 mg/kg, i.p.) in the forced swimming test was blocked by pretreatment of mice with efaroxan (1 mg/kg, i.p., an imidazoline I1/alpha2-adrenoceptor antagonist), idazoxan (0.06 mg/kg, i.p., an imidazoline I2/alpha2-adrenoceptor antagonist) and antazoline (5 mg/kg, i.p., a ligand with high affinity for the I2 receptor). A subeffective dose of agmatine (0.001 mg/kg, i.p.) produced a synergistic antidepressant-like effect with clonidine (0.06 mg/kg, i.p, an imidazoline I1/alpha2-adrenoceptor agonist), moxonidine (0.5 mg/kg, i.p., an imidazoline I1/alpha2-adrenoceptor agonist), antazoline (1 mg/kg, i.p.) and MK-801 (0.001 mg/kg, i.p., a non-competitive NMDA receptor antagonist), but not with efaroxan (1 mg/kg, i.p.) and idazoxan (0.06 mg/kg, i.p.). Pretreatment of mice with yohimbine (1 mg/kg, i.p., an alpha2-adrenoceptor antagonist) blocked the synergistic antidepressant-like effect of agmatine (0.001 mg/kg, i.p.) with clonidine (0.06 mg/kg, i.p). A subeffective dose of MK-801 (0.001 mg/kg, i.p.) produced a synergistic antidepressant-like effect with antazoline (5 mg/kg, i.p.), but not with efaroxan (1 mg/kg, i.p.) or idazoxan (0.06 mg/kg, i.p.). In conclusion, this study suggests that the anti-immobility effect of agmatine in the forced swimming test is dependent on its interaction with imidazoline I1 and I2 receptors.

Topics: Agmatine; Animals; Antazoline; Antidepressive Agents; Benzofurans; Clonidine; Female; Imidazoles; Imidazoline Receptors; Male; Mice; Receptors, Adrenergic, alpha-2; Receptors, Drug; Receptors, N-Methyl-D-Aspartate; Swimming

A rapid method for the enantiomeric purity determination of efaroxan has been developed by capillary electrophoresis (CE) using a dual cyclodextrin (CD) system. The influence of the nature and the concentration of CDs on separation parameters has been studied. High resolution (R(s)=7) and peak efficiency (104,000-121,000 theoretical plates) values were obtained for efaroxan enantiomers by adding two cyclodextrins, one neutral (7.5 mM DM-beta-CD) and the other negatively charged (3 mM CM-beta-CD) to the running buffer composed of 100 mM phosphoric acid-triethanolamine (pH 3). These resolution and peak efficiencies values allowed the quantitation of the (S)-enantiomer of efaroxan at very low enantiomeric excess even if the minor component migrates after the major one. This method was fully validated for the enantiomeric impurity determination of the (S)-form of efaroxan at the 0.05% level. Calibration curve, expressed by the peak areas ratio versus the enantiomeric purity was linear over the 0.05-1% enantiomeric impurity range (r2=0.9996). Limits of detection (LOD) and quantification (LOQ), expressed in term of (S)-enantiomer impurity were 0.02% and 0.05%, respectively. The accuracy of the method at 0.12%, 0.50% and 0.80% enantiomeric impurity levels for the (S)-form were determined. Recoveries were in 94-102% range for each quality control sample and were determined with good precision (intra-day R.S.D.=3.54%, inter-day R.S.D.=5.33%).

Topics: Anions; Benzofurans; Buffers; Cyclodextrins; Electrophoresis, Capillary; Imidazoles; Molecular Structure; Sensitivity and Specificity; Stereoisomerism

Moxonidine is a second-generation centrally acting antihypertensive drug that has a high affinity for I(1)-imidazoline receptors (I(1)R). The caudal ventrolateral medulla (CVLM), an important region involved in cardiovascular activity, contains binding sites for centrally acting drugs. Our study aimed to determine the effects of moxonidine injected into the CVLM on cardiovascular activity in anesthetized rats. Unilateral microinjection of moxonidine (0.4 and 4 nmol) into the CVLM dose-dependently increased blood pressure (BP) by 8+/-2 and 18+/-2 mmHg and renal sympathetic nerve activity (RSNA) by 19+/-3 and 48+/-5% without modifying heart rate. Microinjection of the I(1)R/alpha(2)-adrenoceptor antagonist efaroxan (4 nmol) into the CVLM produced significant decreases in baseline BP and RSNA, but also completely abolished the increases in BP (2+/-1 versus 18+/-2 mmHg, P<0.01) and RSNA (3+/-2 versus 45+/-10%, P<0.01) evoked by subsequent injection of moxonidine (4 nmol). However, prior injection of yohimbine (500 pmol), a selective antagonist of alpha(2)-adrenoceptors, into the CVLM had no significant (P>0.05) effect on the moxonidine-induced increase in BP (18+/-2 versus 17+/-3 mmHg) and RSNA (45+/-10 versus 42+/-7%). The current data suggest that moxonidine injection into the CVLM has an excitatory effect on cardiovascular activity, which is mediated by an I(1)R dependent mechanism.

Topics: Adrenergic alpha-Antagonists; Anesthesia; Animals; Benzofurans; Blood Pressure; Dose-Response Relationship, Drug; Drug Interactions; Heart Rate; Imidazoles; Imidazoline Receptors; Male; Medulla Oblongata; Rats; Rats, Sprague-Dawley; Sympathetic Nervous System; Sympatholytics; Yohimbine

The imidazoline compound RX871024 reduces IL-1beta-induced NO production thereby protecting against IL-1beta-induced beta-cell apoptosis. The aim of this study was to evaluate whether imidazolines RX871024 and efaroxan protect beta-cells against death in the presence of a combination of the cytokines IL-1beta, IFNgamma, and TNFalpha. To address this issue, experiments involving different methods for detection of cell death, different concentrations of the cytokines, and a variety of conditions of preparation and culturing of ob/ob mouse islets and beta-cells have been carried out. Thoroughly performed experiments have not been able to demonstrate a protective effect of RX871024 and efaroxan on beta-cell death induced by the combination of cytokines. However, the inhibitory effect of RX871024 on NO production in ob/ob mouse islets and beta-cells was still observed in the presence of all three cytokines and correlated with the decrease in p38 MAPK phosphorylation. Conversely, efaroxan did not affect cytokine-induced NO production. Our data indicate that a combination of pro-inflammatory cytokines IL-1beta, IFNgamma, and TNFalpha, conditions modelling those that take place in type 1 diabetes, induces pancreatic beta-cell death that does not directly correlate with NO production and cannot be counteracted with imidazoline compounds.

Topics: Animals; Benzofurans; Cell Death; Cells, Cultured; Cytokines; Imidazoles; Indoles; Insulin; Insulin Secretion; Insulin-Secreting Cells; Interferon-gamma; Interleukin-1; Mice; Mice, Obese; Nitric Oxide; Tumor Necrosis Factor-alpha

The monomeric G-protein Rhes has been described to be present in pancreatic beta-cells, and a putative role in the control of insulin release has been proposed. Here, we show that treatment of beta-cells with the imidazoline insulin secretagogue efaroxan resulted in a concentration- and time-dependent increase in the expression of Rhes, which peaked after 4h of efaroxan exposure; thereafter, Rhes mRNA levels decreased. Marked stereoselectivity was displayed, with (-)-efaroxan (the selectively insulinotropic enantiomer) being much more effective than (+)-efaroxan at raising Rhes transcript levels. The mechanism by which Rhes gene expression is activated in beta-cells appears to require the influx of extracellular calcium and de novo protein synthesis, and is not directly associated with the release of insulin. The present results confirm our earlier proposal that Rhes is an imidazoline-regulated transcript in pancreatic beta-cells. Studies to understand the role of Rhes as a regulator of beta-cell function are, thus, warranted.

Topics: Adrenergic alpha-Antagonists; Benzofurans; Calcium; Calcium Channel Blockers; Calcium Channels; Dihydropyridines; Glyburide; GTP-Binding Proteins; Humans; Hypoglycemic Agents; Imidazoles; Imidazolines; Insulin-Secreting Cells; RNA, Messenger; Transcription, Genetic

Some studies, suggesting the involvement of I(2)-imidazoline binding sites (I(2)-IBS) in morphine analgesia modulation, prompted us to examine on mice antinociceptive assays the effect produced by 1 (phenyzoline), that in view of its high I(2)-IBS affinity and high I(2)-IBS selectivity with regard to I(1)-IBS, alpha(2)-adrenoreceptors and mu-opioid receptors might be considered the first interesting I(2)-IBS ligand. The study was also applied to its ortho phenyl derivative 2 (diphenyzoline), designed and prepared in order to produce a possible modification of the biological profile of 1. Diphenyzoline (2) retains a significant I(2)-IBS selectivity with regard to I(1)-IBS, alpha(2)-adrenoreceptors and mu-opioid receptors. Moreover, by the functional assays 1 and 2 proved inactive at all alpha(2)-adrenoreceptors subtypes up to 10(-3) M. As expected, phenyzoline and diphenyzoline, which are structurally related, highlighted an interesting "positive" or "negative", respectively, morphine analgesia modulatory effect. In fact, 1 (s.c. 10 mg/kg) enhanced morphine analgesia (60% and 40% in mouse tail-flick and mouse hot-plate, respectively), while 2 (s.c. 10 mg/kg) decreased it (-41% and -20%, respectively). The ability to decrease morphine analgesia had never been observed before in I(2)-IBS ligands. These effects were not affected by i.p. treatment of animals with yohimbine (a selective alpha(2)-adrenoreceptor antagonist, 0.625 mg/kg) or efaroxan (an I(1)-IBS/alpha(2)-adrenoreceptor antagonist, 1.0 mg/kg). In contrast, they were completely reversed by i.p. treatment of animals with idazoxan (an I(2)-IBS/alpha(2)-adrenoreceptor antagonist, 2 mg/kg). Moreover, compound 2, in mouse tail-flick test, was able to potentiate by 23% the naloxone-induced decrease of morphine analgesia. Therefore, the results of this study indicate the crucial involvement of I(2)-IBS in the morphine analgesia modulatory effects of 1 and 2.

Topics: Adrenergic alpha-Antagonists; Analgesics, Opioid; Animals; Benzofurans; CHO Cells; Cricetinae; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Mice; Models, Molecular; Morphine; Naloxone; Narcotic Antagonists; Pain Measurement; Radioligand Assay; Reaction Time; Receptors, Adrenergic, alpha-2; Receptors, Drug; Receptors, Opioid, mu; Yohimbine

Interleukin (IL)-1beta-treated rat islets of Langerhans were exposed in vitro either to the imidazoline compound, Efaroxan, or to the selective inducible nitric oxide synthase (iNOS) inhibitor, 1400W, in a medium containing a high concentration of glucose (16.7 mmol/L). Our data have evidenced the following: (i) addition of Efaroxan to islet cultures inhibited IL-1beta activation of ICE (cysteine protease IL-1beta converting enzyme) while addition of 1400W did not; (ii) Efaroxan completely inhibited IL-1beta-induced suppression of insulin secretion and induction of iNOS mRNA transcripts, and, in addition, counteracted islet beta-cell protein profile alterations, Bax-cytochrome c translocation, caspase activation, and apoptosis; (iii) 1400W inhibited IL-1beta induction of iNOS, but failed to completely counteract the other cytotoxic effects; (iv) the two compounds, moreover, exerted different effects on manganese superoxide dismutase (MnSOD), in fact, while Efaroxan inhibited the early stimulatory effect of IL-1beta on MnSOD, 1400W did not. Thus, Efaroxan completely protected islet beta cells from damage caused by IL-1beta-induced toxicity, while compound 1400W only inhibited NO radical production without altering the cytokine's cytotoxicity. Our observations have evidenced that suppression of ICE activation is required to counteract IL-1beta-mediated islet beta cell toxicity, and that IL-1beta-induced apoptosis is NO-independent and involves the cytochrome c-mitochondrial pathway.

Topics: Aconitate Hydratase; Adrenergic alpha-Antagonists; Amidines; Animals; bcl-2-Associated X Protein; Benzofurans; Benzylamines; Caspases; Cell Death; Cells, Cultured; Cytochromes c; Drug Interactions; Enzyme Inhibitors; Gene Expression Profiling; Imidazoles; Insulin; Interleukin-1; Islets of Langerhans; Mitochondria; Nitrates; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Wistar; RNA, Messenger; Superoxide Dismutase

We have previously shown that acute intravenous injections of moxonidine and clonidine increase plasma atrial natriuretic peptide (ANP), a vasodilator, diuretic and natriuretic hormone. We hypothesized that moxonidine stimulates the release of ANP, which would act on its renal receptors to cause diuresis and natriuresis, and these effects may be altered in hypertension. Moxonidine (0, 10, 50, 100 or 150 microg in 300 microl saline) and clonidine (0, 1, 5 or 10 microg in 300 microl saline) injected intravenously in conscious normally hydrated normotensive Sprague-Dawley rats (SD, approximately 200 g) and 12-14-week-old Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) dose-dependently stimulated diuresis, natriuresis, kaliuresis and cGMP excretion, with these effects being more pronounced during the first hour post-injection. The actions of 5 microg clonidine and 50 microg moxonidine were inhibited by yohimbine, an alpha2-adrenoceptor antagonist, and efaroxan, an imidazoline I1-receptor antagonist. Moxonidine (100 microg) stimulated (P<0.01) diuresis in SHR (0.21+/-0.04 vs 1.16+/-0.06 ml h(-1) 100 g(-1)), SD (0.42+/-0.06 vs 1.56+/-0.19 ml h(-1) 100 g(-1)) and WKY (0.12+/-0.04 vs 1.44+/-0.21 ml h(-1) 100 g(-1)). Moxonidine-stimulated urine output was lower in SHR than in SD and WKY. Moxonidine-stimulated sodium and potassium excretions were lower in SHR than in SD, but not WKY, demonstrating an influence of strain but not of pressure. Pretreatment with the natriuretic peptide antagonist anantin (5 or 10 microg) resulted in dose-dependent inhibition of moxonidine-stimulated urinary actions. Anantin (10 microg) inhibited (P<0.01) urine output to 0.38+/-0.06, 0.12+/-0.01, and 0.16+/-0.04 ml h(-1) 100 g(-1) in SD, WKY, and SHR, respectively. Moxonidine increased (P<0.01) plasma ANP in SD (417+/-58 vs 1021+/-112 pg ml(-1)) and WKY (309+/-59 vs 1433+/-187 pg ml(-1)), and in SHR (853+/-96 vs 1879+/-229 pg ml(-1)). These results demonstrate that natriuretic peptides mediate the urinary actions of moxonidine through natriuretic peptide receptors.

Topics: Animals; Antihypertensive Agents; Benzofurans; Clonidine; Cyclic GMP; Diuresis; Dose-Response Relationship, Drug; Female; Imidazoles; Imidazoline Receptors; Natriuresis; Natriuretic Peptides; Peptides, Cyclic; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-2; Receptors, Drug; Yohimbine

Bath-applied monoamines-dopamine (DA), serotonin (5-HT), and noradrenaline (NE)-strongly suppress the perforant path (PP) input to CA1 hippocampal region with very little effect on the Schaffer collaterals (SC) input. The effect of DA action on PP field excitatory postsynaptic potential (fEPSP) has been characterized in detail, but relatively little is known about the NE and 5-HT effects. Here we show that the maximal inhibition of the PP fEPSP by NE is approximately 55%, whereas 5-HT inhibition is weaker ( approximately 35%). The half-maximal inhibitory concentration of both 5-HT and NE is approximately 1 muM. Neither NE nor 5-HT affected paired-pulse facilitation, suggesting that the effect is not presynaptic. This is in contrast to DA, which does have a presynaptic effect. The NE effect was blocked by alpha2 antagonists, whereas the alpha1 antagonist corynanthine and beta-antagonist propranolol were ineffective. The effect of 5-HT was mimicked by the agonist, 5-carboxamidotryptamine maleate (5-CT), and not affected by adrenergic and dopaminergic antagonists. To determine the 5-HT receptors involved, we tested a number of 5-HT antagonists, but none produced a complete suppression of the 5-HT effect. Of these, only the 5-HT7 and 5-HT2 antagonists produced weak but significant inhibition of 5-HT effect. We conclude that NE inhibits the PP fEPSP through postsynaptic action on alpha2-adrenoceptors and that 5-HT7, 5-HT2, and some other receptor may be involved in 5-HT action in PP.

Topics: Adrenergic alpha-Antagonists; Adrenergic beta-Antagonists; Animals; Animals, Newborn; Benzofurans; Dopamine; Electric Stimulation; Excitatory Postsynaptic Potentials; Hippocampus; Imidazoles; In Vitro Techniques; Neural Inhibition; Neurons; Norepinephrine; Patch-Clamp Techniques; Perforant Pathway; Propranolol; Rats; Rats, Long-Evans; Serotonin; Serotonin Agents; Yohimbine

Our previous study showed that rilmenidine, a selective I(1)-imidazoline receptor agonist, enhanced the phosphorylation of mitogen-activated protein kinase (MAPK)(p42/44), via the phosphatidylcholine-specific phospholipase C pathway in the pheochromocytoma cell line (PC12). In the present study, we tested the hypothesis that enhancement of MAPK phosphorylation in the rostral ventrolateral medulla (RVLM) contributes to the hypotensive response elicited by I(1)-receptor activation in vivo. Systemic rilmenidine (600 microg/kg i.v.) elicited hypotension and bradycardia along with significant elevation in MAPK(p42/44), detected by immunohistochemistry, in RVLM neurons. To obtain conclusive evidence that the latter response was I(1)-receptor-mediated, similar hypotensive responses were elicited by intracisternal (i.c.) rilmenidine (25 microg/rat) or the highly selective alpha(2)-agonist alpha-methylnorepinephrine (4 microg/rat). An increase in RVLM MAPK(p42/44) occurred only after rilmenidine. Furthermore, pretreatment with efaroxan (0.15 microg/rat i.c.), a selective I(1)-imidazoline receptor antagonist, or with PD98059 (2'-amino-3'-methoxyflavone) (5 microg/rat i.c.), a selective extracellular signal-regulated kinase 1/2 inhibitor, significantly attenuated the hypotensive response and the elevation in RVLM MAPK(p42/44) elicited by i.c. rilmenidine. The findings suggest that MAPK phosphorylation in the RVLM contributes to the hypotensive response induced by I(1)-receptor activation and presents in vivo evidence that distinguishes the neuronal responses triggered by the I(1)-receptor from those triggered by the alpha(2)-adrenergic receptor.

Topics: Animals; Benzofurans; Blood Pressure; Heart Rate; Imidazoles; Imidazoline Receptors; Male; Medulla Oblongata; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Oxazoles; Phosphorylation; Rats; Rats, Sprague-Dawley; Receptors, Drug; Rilmenidine; Signal Transduction

Clonidine (a mixed alpha2-adrenoceptor and imidazoline I1 receptor agonist)-evoked hypotension was associated with dissimilar reductions in c-jun gene expression in the rostral ventrolateral medulla (RVLM) and the nucleus tractus solitarius (NTS) in normotensive rats. In the present study, we investigated the relative contribution of the alpha2-adrenoceptor vs. the imidazoline I1 receptor to the reduction in c-jun gene expression in these two brainstem areas. In conscious spontaneously hypertensive rats (SHRs), equihypotensive doses of three centrally acting hypotensive drugs with different selectivity for the two receptors were administered intracisternally (4 microl) to limit their actions to the brain. As a control, a similar hypotensive response was elicited by i.v. hydralazine. Clonidine (0.5 microg), or alpha-methylnorepinephrine (alpha-MNE, 4 microg), a highly selective alpha2-adrenoceptor agonist, similarly reduced c-jun mRNA expression in the NTS and rostral ventrolateral medulla. In contrast, a similar hypotensive response (-37+/-3.5 mm Hg) caused by the selective imidazoline I1 receptor agonist rilmenidine (25 microg) was associated with reduction in c-jun mRNA expression in the rostral ventrolateral medulla, but not in the NTS. Further, intra-rostral ventrolateral medulla rilmenidine (40 nmol) reduced c-Jun protein expression in rostral ventrolateral medulla and blood pressure and both responses were antagonized by selective imidazoline I1 receptor (efaroxan, 4 nmol), but not alpha2-adrenoceptor (SK&F 86466, 10 nmol) blockade. These results suggest: (1) the c-jun containing neurons in the brainstem are involved in the centrally mediated hypotension elicited by centrally acting antihypertensive agents, and (2) the alpha2-adrenoceptor modulates c-jun gene expression in the NTS and rostral ventrolateral medulla implicated in centrally mediated hypotension, and (3) the imidazoline I1 receptor mediated inhibition of c-jun gene expression in the rostral ventrolateral medulla, but not in the NTS, contributes to the centrally mediated hypotension by the second generation drugs.

Topics: Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzazepines; Benzofurans; Blood Pressure; Brain Stem; Cerebrospinal Fluid; Clonidine; Consciousness; Gene Expression; Heart Rate; Hydralazine; Hypotension; Imidazoles; Imidazoline Receptors; Immunohistochemistry; Male; Medulla Oblongata; Neurons; Nordefrin; Oxazoles; Proto-Oncogene Proteins c-jun; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2; Receptors, Drug; Rilmenidine; RNA, Messenger; Time Factors; Vasoconstrictor Agents; Vasodilator Agents

We tested the hypothesis that the I1 receptor mediates the reduction in rostral ventrolateral medulla (RVLM) neuronal norepinephrine (NE; index of sympathetic activity) that leads to hypotension independent of other brainstem areas or the alpha2-adrenergic receptor. To this end, we developed a model that permitted measurement of real-time changes in neuronal NE in the RVLM or nucleus tractus solitarius (NTS) along with blood pressure and heart rate in the conscious SHR in response to localized microinjections of selective I1 (rilmenidine) or alpha2-adrenergic (alpha-methylnorepinephrine; alpha-MNE) agonist versus the mixed I1/alpha2 agonist clonidine. To further support the hypothesis, we investigated the effects of localized selective alpha2- (SK&F86466) or I1 (efaroxan) blockade on the reductions in neuronal NE and blood pressure elicited by intra-RVLM rilmenidine. In the latter experiment, changes in RVLM neuronal c-Fos (another marker of sympathetic neural activity) were also investigated. Intra-RVLM rilmenidine (40 nmol) or clonidine (1 nmol) similarly reduced RVLM NE and blood pressure; these responses were approximately 2-fold greater than those elicited by the pure alpha2-adrenergic agonist alpha-MNE (10 nmol). By contrast, intra-NTS rilmenidine or clonidine had no effect on NTS NE or blood pressure versus significant reductions in both parameters by alpha-MNE. Intra-RVLM rilmenidine decreased c-Fos expression, and these responses were abolished by efaroxan but not by SK&F 86466. These findings suggest: (1) in the RVLM, I1-receptor signaling suppresses cardiovascular neuron activity, which leads to lowering of blood pressure; (2) although the alpha2-adrenergic receptor in the RVLM serves a similar role, it does not exert a tonic neuronal inhibitory effect and is not essential, as a downstream signaling entity, for the I1-evoked neurobiological effects in the brainstem. The potential confounding effects of anesthetics on the I1 and/or alpha2 receptor-mediated neuronal and cardiovascular responses were circumvented in the present study.

Topics: Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Area Postrema; Benzazepines; Benzofurans; Blood Pressure; Clonidine; Consciousness; Heart Rate; Hypotension; Imidazoles; Interleukin-1; Male; Motor Activity; Neurons; Nordefrin; Norepinephrine; Oxazoles; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2; Rilmenidine; Solitary Nucleus

Previous studies using the novel imidazoline1-binding site ligand 1-(4,5-dihydro-1H-imidazol-2-yl)isoquinoline hydrochloride, BU98008, have shown it induces a hypothermic response in rodents following intraperitoneal administration. Radioligand binding data has shown that BU98008 is a highly selective imidazoline1-binding site ligand with 300 fold selectively for the imidazoline1-binding site relative to alpha2-adrenoceptors. However, alpha2-adrenoceptor agonists are known to induce hypothermia, therefore, the present study has investigated the ability of the selective alpha2-adrenoceptor antagonist, RX811059 (2-ethoxy idazoxan) and the mixed imidazoline1-binding site/alpha2-adrenoceptor antagonist, efaroxan, to attenuate the BU98008-induced hypothermia. Preliminary experiments confirmed that BU98008 induced a dose-dependent decrease in body temperature in mice at 10 and 20 mg/kg. The response was not affected by pre-treatment with RX811059 but was significantly attenuated following pre-treatment with efaroxan. These data suggest that BU98008-induced hypothermia is mediated by activation of imidazoline1-binding site. Body temperature may therefore provide a novel assay for investigating agonist and antagonist action at the imidazoline1-binding site.

Topics: Animals; Benzofurans; Binding Sites; Body Temperature; Dose-Response Relationship, Drug; Idazoxan; Imidazoles; Imidazoline Receptors; Isoquinolines; Ligands; Male; Mice; Receptors, Drug

The neuronal pathways involved in the muscle relaxant effect of tizanidine were examined by measurement of spinal reflexes in rats. Tizanidine (i.v. and intra-4th ventricular injection) decreased the mono- and disynaptic (the fastest polysynaptic) reflexes (MSR and DSR, respectively) in non-spinalized rats. Depletion of central noradrenaline by 6-hydroxydopamine abolished the depressant effect of tizanidine on the MSR almost completely and attenuated the effect on the DSR. Co-depletion of serotonin by 5,6-dihydroxytryptamine and noradrenaline resulted in more prominent attenuation of tizanidine-induced inhibition of the DSR. Supraspinal receptors were then studied using yohimbine- and some imidazoline-receptor ligands containing an imidazoline moiety. Idazoxan (I1, I2, I3, and alpha2), efaroxan (I1, I3, and alpha2), and RX821002 (I3 and alpha2), but not yohimbine, an alpha2-adrenergic receptor antagonist with no affinity for I receptors, antagonized the inhibitory effects of tizanidine. Thus, supraspinal I receptors (most likely I3) and descending monoaminergic influences are necessary for tizanidine-induced inhibition of spinal segmental reflexes.

Topics: 5,6-Dihydroxytryptamine; Adrenergic Agents; Adrenergic alpha-Antagonists; Animals; Benzofurans; Clonidine; Dose-Response Relationship, Drug; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Muscle Relaxants, Central; Norepinephrine; Oxidopamine; Rats; Rats, Wistar; Receptors, Drug; Reflex, Monosynaptic; Serotonin; Serotonin Agents; Spinal Cord; Sympathetic Nervous System; Time Factors

Imidazolines are a class of investigational antidiabetic drugs. It is still unclear whether the imidazoline ring is decisive for insulinotropic characteristics.. We studied the imidazoline efaroxan and its imidazole analogue, KU14R, which is currently classified as an imidazoline antagonist. The effects of both on stimulus secretion-coupling in normal mouse islets and beta cells were compared by measuring KATP channel activity, plasma membrane potential, cytosolic calcium concentration ([Ca2+]c) and dynamic insulin secretion.. In the presence of 10 mmol/l but not of 5 mmol/l glucose, efaroxan (100 micromol/l) strongly enhanced insulin secretion by freshly isolated perifused islets, whereas KU14R (30, 100 or 300 micromol/l) was ineffective at both glucose concentrations. Surprisingly, the insulinotropic effect of efaroxan was not antagonised by KU14R. KATP channels were blocked by efaroxan (IC50 8.8 micromol/l, Hill slope -1.1) and by KU14R (IC50 31.9 micromol/l, Hill slope -1.5). Neither the KATP channel-blocking effect nor the depolarising effect of efaroxan was antagonised by KU14R. Rather, both compounds strongly depolarised the beta cell membrane potential and induced action potential spiking. However, KU14R was clearly less efficient than efaroxan in raising [Ca2+]c in single beta cells and whole islets at 5 mmol/l glucose. The increase in [Ca2+]c induced by 10 mmol/l glucose was affected neither by efaroxan nor by KU14R. Again, KU14R did not antagonise the effects of efaroxan.. The presence of an imidazole instead of an imidazoline ring leads to virtually complete loss of the insulinotropic effect in spite of a preserved ability to block KATP channels. The imidazole compound is less efficient in raising [Ca2+]c; in particular, it lacks the ability of the imidazoline to potentiate the enhancing effect of energy metabolism on Ca2+-induced insulin secretion.

Topics: Action Potentials; Animals; Benzofurans; Calcium; Cell Line; Cytosol; Imidazoles; Imidazolines; Insulin; Insulin Secretion; Insulin-Secreting Cells; Membrane Potentials; Mice; Mice, Mutant Strains; Patch-Clamp Techniques; Potassium Channel Blockers; Potassium Channels

We previously reported that imidazoline receptors in the central nervous system are involved in modulation of halothane-epinephrine arrhythmias. These receptors have been subclassified as I1 and I2 subtypes, but it is not known which receptor subtype is involved in halothane-epinephrine-induced arrhythmias. We designed the present study to clarify the involvement of central imidazoline receptor subtype in the modulation of halothane-epinephrine-induced arrhythmias. Rats were anesthetized with halothane and monitored continuously for systemic arterial blood pressure and premature ventricular contractions. The arrhythmogenic dose of epinephrine was defined as the smallest dose that produces three or more premature ventricular contractions within a 15-s period. Intracisternal moxonidine dose-dependently inhibited the epinephrine-induced arrhythmias during halothane anesthesia. Intracisternal efaroxan, a selective I1 antagonist with little affinity for I2 subtype, but not rauwolscine, an alpha2 antagonist without affinity for imidazoline receptors, blocked the antiarrhythmic effect of moxonidine. Intracisternal BU 224 and 2-BFI, selective I2 ligands, also inhibited the epinephrine-induced arrhythmias dose-dependently; however, these effects were abolished by efaroxan. We conclude that central I1, but not I2, receptors play an important role in inhibition of halothane-epinephrine arrhythmia.

Topics: Animals; Arrhythmias, Cardiac; Benzofurans; Blood Pressure; Dose-Response Relationship, Drug; Epinephrine; Halothane; Heart Rate; Imidazoles; Imidazoline Receptors; Male; Rats; Rats, Sprague-Dawley; Receptors, Drug

Imidazoline receptors are divided into I(1) and I(2) subtypes. I(1)-imidazoline receptors are distributed in the heart and are upregulated during hypertension or heart failure. The aim of this study was to define the possible role of I(1)-imidazoline receptors in the regulation of atrial natriuretic peptide (ANP) release in hypertrophied atria. Experiments were performed on isolated, perfused, hypertrophied atria from remnant-kidney hypertensive rats. The relatively selective I(1)-imidazoline receptor agonist moxonidine caused a decrease in pulse pressure. Moxonidine (3, 10, and 30 micromol/l) also caused dose-dependent increases in ANP secretion, but clonidine (an alpha(2)-adrenoceptor agonist) did not. Pretreatment with efaroxan (a selective I(1)-imidazoline receptor antagonist) or rauwolscine (a selective alpha(2)-adrenoceptor antagonist) inhibited the moxonidine-induced increases in ANP secretion and interstitial ANP concentration and decrease in pulse pressure. However, the antagonistic effect of efaroxan on moxonidine-induced ANP secretion was greater than that of rauwolscine. Neither efaroxan nor rauwolscine alone has any significant effects on ANP secretion and pulse pressure. In hypertrophied atria, the moxonidine-induced increase in ANP secretion and decrease in pulse pressure were markedly augmented compared with nonhypertrophied atria, and the relative change in ANP secretion by moxonidine was positively correlated to atrial hypertrophy. The accentuation by moxonidine of ANP secretion was attenuated by efaroxan but not by rauwolscine. These results show that moxonidine increases ANP release through (preferentially) the activation of atrial I(1)-imidazoline receptors and also via different mechanisms from clonidine, and this effect is augmented in hypertrophied atria. Therefore, we suggest that cardiac I(1)-imidazoline receptors play an important role in the regulation of blood pressure.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Atrial Function; Atrial Natriuretic Factor; Benzofurans; Blood Pressure; Cardiomegaly; Clonidine; Heart Atria; Hemodynamics; Imidazoles; Imidazoline Receptors; Male; Rats; Rats, Sprague-Dawley; Receptors, Drug; Yohimbine

Cholinergic agonists produce spinal antinociception via mechanisms involving an increased release of intraspinal acetylcholine. The cholinergic receptor system interacts with several other receptor types, such as alpha2-adrenergic receptors. To fully understand these interactions, the effects of various receptor ligands on the cholinergic system must be investigated in detail. This study was initiated to investigate the effects of the alpha2-adrenergic receptor agonists clonidine and rilmenidine and the alpha2-adrenergic receptor antagonists yohimbine and efaroxan on spinal cholinergic receptors in the rat. Spinal microdialysis was used to measure in vivo changes of acetylcholine after administration of the ligands, with or without nicotinic receptor blockade. In addition, in vitro binding properties of the ligands on muscarinic and nicotinic receptors were investigated. It was found that clonidine and rilmenidine increased, while yohimbine decreased spinal acetylcholine release. Efaroxan affected acetylcholine release differently depending on concentration. Nicotinic receptor blockade attenuated the effect of all ligands. All ligands showed poor binding affinity for muscarinic receptors. On the other hand, all ligands possessed affinity for nicotinic receptors. Clonidine and yohimbine binding was best fit to a one site binding curve and rilmenidine and efaroxan to a two site binding curve. The present study demonstrates that the tested alpha2-adrenergic receptor ligands affect intraspinal acetylcholine release in the rat evoked by nicotinic receptor mechanisms in vivo, and that they possess binding affinity to nicotinic receptors in vitro. The binding of alpha2-adrenergic receptor ligands to nicotinic receptors might affect the intraspinal release of acetylcholine.

Topics: Acetylcholine; Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-2 Receptor Antagonists; Animals; Benzofurans; Binding, Competitive; Clonidine; Dose-Response Relationship, Drug; Imidazoles; In Vitro Techniques; Injections, Spinal; Ligands; Male; Microdialysis; Oxazoles; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-2; Receptors, Cholinergic; Rilmenidine; Spinal Cord; Time Factors; Yohimbine

Glaucoma is the main cause of blindness in the developed countries. Its progress can be diminished by decreasing intraocular pressure (IOP) using pharmacological or surgical treatment. Antiglaucoma agents, alpha 2-adrenergic's receptor agonists have been known for several years as IOP lowering. Due to the fact that the majority of them turned out to be imidazoline receptor agonists, it is worth checking if selective imidazoline receptor (l1) agonists alter IOP. Preliminary animal experiments show that they lower IOP. In our study we examined the influence of rilmenidine, a potent l1 receptor agonist, on IOP in rabbits. Furthermore, we tried to find out whether l1 and alpha 2 receptor antagonists (efaroxan and rauwolscine) counteract the pharmacological effect of rilmenidine.. The study was conducted on adult male White New Zealand rabbits. All the substances were administered topically, and IOP was measured by applanation tonometry after topical anaesthesia before and 1, 3 and 5 hours after drug instillation.. Rilmenidine showed the lowering effect on IOP at the concentration of 0.4%. Efaroxan and rauwolscine partly inhibited rilmenidine effect.. Rilmenidine is a potential antiglaucoma agent, though further studies are necessary.

Topics: Administration, Topical; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Drug Interactions; Glaucoma; Imidazoles; Intraocular Pressure; Male; Oxazoles; Rabbits; Rilmenidine; Time Factors; Yohimbine

We studied the ability of moxonidine to interact with alpha2-adrenoceptors and Ii-imidazoline receptors in isolated mouse large intestine. Moxonidine caused contractions of longitudinal muscles in the large intestine, which depended on the dose of this preparation. Pretreatment with yohimbine (alpha2-adrenoceptor antagonist with low affinity for Ii-imidazoline receptors) and efaroxan (Ii-imidazoline receptor antagonist with low affinity for alpha2-adrenoceptors) abolished the effect of moxonidine. Antagonistic activity and relative selectivity of yohimbine and efaroxan suggest that the effects of moxonidine on mouse large intestine are realized via alpha2-adrenoceptors.

Topics: Animals; Benzofurans; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Intestine, Large; Mice; Muscle Contraction; Receptors, Adrenergic, alpha-2; Receptors, Drug; Yohimbine

The glucose dependence of the insulinotropic action of KATP channel-blocking imidazoline compounds was investigated. Administration of 100 micromol/l phentolamine, but not 100 micromol/l efaroxan, markedly increased insulin secretion of freshly isolated mouse islets when the perifusion medium contained 5 mmol/l glucose. When the glucose concentration was raised to 10 mmol/l in the continued presence of either imidazoline, a clear potentiation of secretion occurred as compared with 10 mmol/l glucose alone. In the presence of efaroxan, a brisk first-phase-like increase was followed by a sustained phase, whereas a more gradual increase resulted in the presence of phentolamine. Administration of 100 micromol/l phentolamine was somewhat more effective than 100 micromol/l efaroxan to inhibit KATP channel activity in intact cultured beta-cells (reduction by 96 vs. 83%). Both compounds were similarly effective to depolarize the beta-cells. When measured by the perforated patch-technique, the depolarization by efaroxan was often oscillatory, whereas that by phentolamine was sustained. In perifused cultured islets, both compounds increased the cytosolic calcium concentration ([Ca2+]c) in the presence of 5 and 10 mmol/l glucose. Efaroxan induced large amplitude oscillations of [Ca2+]c, whereas phentolamine induced a sustained increase. It appears that a KATP channel block by imidazolines is not incompatible with a glucose-selective enhancement of insulin secretion. The glucose selectivity of efaroxan may involve an inhibitory effect distal to [Ca2+]c increase and/or the generation of [Ca2+]c oscillations.

Topics: Animals; Benzofurans; Cells, Cultured; Glucose; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Membrane Potentials; Mice; Mice, Inbred Strains; Potassium Channels

To determine the involvement of central imidazoline receptors in the cardiovascular actions of the chronically administered antihypertensive agents moxonidine, rilmenidine and clonidine.. In 21 rabbits with implanted fourth-ventricular catheters, we investigated the central effects of three cumulative doses of an I(1)-imidazoline/alpha(2)-adrenoceptor antagonist, efaroxan, and of an alpha(2)-adrenoceptor antagonist, 2-methoxyidazoxan (2-MI), on the changes in blood pressure and heart rate (HR) elicited by chronic subcutaneous administration of moxonidine, rilmenidine and clonidine, after 1 and 3 weeks of treatment. A low, medium and high dose of 2-MI was matched to three doses of efaroxan, such that each produced equal reversal of the hypotension induced by fourth-ventricular alpha-methyldopa and hence produced a similar degree of alpha(2)-adrenoceptor blockade.. Clonidine and moxonidine, at doses of 1 mg/kg per day, and rilmenidine at 5 mg/kg per day, produced sustained reductions in mean arterial pressure of 13 +/- 3, 15 +/- 2 and 13 +/- 2 mmHg, respectively over the 3-week treatment period, but did not alter HR. Central administration of efaroxan on day 9 and day 23 of treatment produced a greater increase in blood pressure than did 2-MI with all three antihypertensive agents. Blood pressure reached levels that were significantly above the original control values. By contrast, the alpha(2)-adrenoceptor antagonist 2-MI only induced a rebound blood pressure effect in clonidine- and to a lesser extent in rilmenidine-treated rabbits. Both efaroxan and 2-MI produced a similar degree of tachycardia in moxonidine-, rilmenidine- and clonidine-treated animals.(2). The greater effect of efaroxan compared to the alpha(2)-adrenoceptor antagonist 2-MI suggests that the hypotension induced by chronic subcutaneous administration of moxonidine, rilmenidine and clonidine is mediated predominantly via an action on central imidazoline receptors. Furthermore, all agents showed a propensity to produce rebound hypertension with imidazoline receptor blockade. However, only clonidine showed a rebound phenomenon when challenged by acute central alpha(2)-adrenoceptor blockade

Topics: Adrenergic alpha-2 Receptor Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Cardiovascular System; Clonidine; Dose-Response Relationship, Drug; Drug Administration Schedule; Female; Fourth Ventricle; Heart Rate; Idazoxan; Imidazoles; Imidazoline Receptors; Injections, Intraventricular; Male; Oxazoles; Rabbits; Receptors, Drug; Rilmenidine

It is known that moxonidine acts as an agonist at presynaptic alpha(2)-adrenoceptors of the postganglionic sympathetic nerve terminals and leads to a reduction in noradrenaline release. In addition, it is conceivable that I(1)-binding sites located in other regions of the pre- and postganglionic sympathetic neurons are involved in this effect. Our aim was to investigate whether and to what extent activation of the I(1)-binding sites contributes to the moxonidine-induced inhibition of noradrenaline release. Noradrenaline release was induced in pithed spontaneously hypertensive rats (pretreated with phenoxybenzamine/desipramine at 10/0.5 mg/kg) by stimulation of sympathetic overflow from the spinal cord. Noradrenaline overflow was reduced using moxonidine (0.18, 0.6, and 1.8 mg/kg) by 39.4, 70.4, or 78.7%, respectively, even when all alpha(1)-/alpha(2)-adrenoceptors were blocked effectively by phenoxybenzamine. In contrast, the I(1)-antagonist efaroxan (0.1, 1, and 3 mg/kg) increased noradrenaline overflow from 453 (control) to 1710, 1999, or 2754 pg/ml, suggesting an autoreceptor-like function of I(1)-binding sites. In consequence, moxonidine (0.18, 0.6, and 1.8 mg/kg) reduced the increase in noradrenaline overflow in efaroxan-treated animals (1 mg/kg) by 22.7, 41.7, and 50.5%, respectively. Agmatine (6 and 60 mg/kg), an endogenous agonist at I(1)-binding sites, reduced noradrenaline overflow (-36 or 53%), even under alpha(2)-adrenoceptor blockade. When 2-endo-amino-3-exo-isopropylbicyclo[2.2.1]heptane (AGN192403) (10 mg/kg) was injected, a selective blocker of I(1)-binding sites, noradrenaline overflow was not influenced by agmatine. It is concluded that moxonidine reduces noradrenaline overflow by acting at I(1)-binding sites in addition to its agonistic property at alpha(2)-adrenoceptors. The exact location of the I(1)-binding sites on the pre- or postsynaptic sympathetic neurons is unknown, but the location in the pre- or postsynaptic membrane of the sympathetic ganglion is the most plausible explanation.

Topics: Adrenergic alpha-Antagonists; Agmatine; Animals; Antihypertensive Agents; Benzofurans; Binding Sites; Blood Pressure; Disease Models, Animal; Heart Rate; Idazoxan; Imidazoles; Male; Norepinephrine; Phenoxybenzamine; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2

Certain imidazoline drugs stimulate insulin secretion acutely but their longer term effects on the viability of pancreatic beta-cells are less well characterised. Indeed, some reports have suggested that imidazolines can be toxic to beta-cells while others have reported protective effects against other cytotoxic agents.. In order to address these discrepancies, the effects of two structurally related imidazolines, efaroxan and idazoxan, on the viability of clonal BRIN-BD11 beta-cells, were compared.. BRIN-BD11 cells were exposed to test reagents and their viability monitored by measuring cellular reducing ability and DNA fragmentation. Nitric oxide was measured indirectly via medium nitrite formation.. Efaroxan (up to 100 micro M) did not directly affect BRIN-BD11 cell viability in the absence of other agents and it did not protect these cells against the cytotoxic effects of interleukin-1beta. Indeed, analysis of DNA fragmentation in BRIN-BD11 cells revealed that efaroxan enhanced the level of damage caused by interleukin-1beta. Idazoxan caused a time- and dose-dependent loss of BRIN-BD11 cell viability in the absence of other ligands. This was associated with marked DNA degradation but was not associated with formation of nitric oxide. The effects of idazoxan were insensitive to blockade of alpha(2)-adrenoceptors or 5-HT(1A) (5-hydroxytryptamine; serotonin) receptors.. The results confirm that idazoxan is cytotoxic to beta-cells but show that efaroxan is better tolerated. However, since efaroxan enhanced the cytotoxic effects of interleukin-1beta, it appears that this imidazoline may sensitise BRIN-BD11 cells to the damaging effects of certain cytokines.

Topics: Animals; Benzofurans; Binding Sites; Cell Line; Cell Survival; DNA; DNA Fragmentation; Dose-Response Relationship, Drug; Idazoxan; Imidazoles; Insulin; Insulin Secretion; Interleukin-1; Islets of Langerhans; Nitric Oxide; Time Factors

(+/-)-Efaroxan 1 is a selective antagonist at the imidazoline I1 receptor. [3H] (+/-)-Efaroxan was required to explore its mechanism of action via receptor binding assay, and the radioligand was prepared by means of catalytic dehalogenation of a dibrominated precursor with tritium.

Topics: Benzofurans; Imidazoles; Imidazoline Receptors; Isotope Labeling; Magnetic Resonance Spectroscopy; Protons; Radioligand Assay; Receptors, Drug; Tritium

The pancreatic beta-cell expresses an imidazoline-binding site that is involved in the regulation of insulin secretion. This site is pharmacologically atypical in comparison with the I(1) and I(2) sites described in other tissues, and it has been classified as I(3). The structural requirements for binding of ligands to the I(3) site have not been fully defined, although a range of synthetic I(3) ligands have been characterized in functional terms. Evidence has been presented that an endogenous I(3) ligand may exist, because extracts of brain contain an active principle that stimulates insulin secretion in a manner consistent with the involvement of I(3) sites. The active component has not been identified but has been equated with the long-sought clonidine displacing substance (CDS) that is proposed as the endogenous ligand for imidazoline-binding sites. Recent evidence has indicated that one active component of CDS may be a beta-carboline, but it is not known whether beta-carbolines can stimulate insulin secretion. Thus, we have studied the effects of beta-carbolines on insulin secretion and cytosolic Ca(2+) levels in rodent and human islet cells. The results reveal that harmane, pinoline, and norharmane cause a dose- and glucose-dependent increase in insulin secretion but show that this response differs in a number of ways from that elicited by the well-characterized I(3)-agonist, efaroxan. Thus, beta-carbolines represent a new class of insulin secretagogues, although it remains unclear whether their action is mediated solely by I(3) sites in the beta cell.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Binding Sites; Calcium; Carbolines; Cells, Cultured; Diazoxide; Glucose; Humans; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Ligands; Male; Rats; Rats, Wistar

Hypertension is commonly accompanied by obesity, hyperlipidemia, and insulin resistance in humans, a cluster of abnormalities known as metabolic syndrome X. With the notable exception of inhibitors of the renin-angiotensin system, which have mildly beneficial effects on insulin resistance, most antihypertensive agents worsen one or more components of metabolic syndrome X. Second-generation centrally acting antihypertensive agents such as rilmenidine and moxonidine have mixed effects on components of metabolic syndrome X, which might reflect in part actions on two different receptors: I(1)-imidazoline and alpha(2)-adrenergic. Using a rat model of metabolic syndrome X, we sought to separate the influence of these two receptors on glucose and lipid metabolism by using selective antagonists. Rilmenidine and moxonidine acutely raised glucose and lowered insulin, thereby further worsening glucose tolerance. These effects were entirely mediated by alpha(2)-adrenergic receptors. Rilmenidine and moxonidine also lowered glucagon, an effect that was mediated solely by I(1)-imidazoline receptors since it was potentiated by alpha(2)-blockade, but eliminated in the presence of I(1)-antagonists. Lowering of triglyceride and cholesterol levels followed the same pattern as glucagon, implicating I(1)-imidazoline receptors in lipid-lowering actions. Chronic treatment with moxonidine reproduced the beneficial effects on glucagon and lipids while the acute hyperglycemic response did not persist. Thus, alpha(2)-adrenergic receptors mediate an acute deterioration of glucose tolerance, whereas in contrast I(1)-imidazoline receptors appear to mediate the persistent long-term improvements in glucose tolerance. The therapeutic action of I(1)-imidazoline agonists may be primarily mediated through reduced glucagon secretion.

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Disease Models, Animal; Female; Glucagon; Glucose; Glucose Tolerance Test; Humans; Imidazoles; Imidazoline Receptors; Lipid Metabolism; Male; Metabolic Syndrome; Obesity; Oxazoles; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2; Receptors, Drug; Rilmenidine; Yohimbine

The function of presynaptic imidazoline-1 receptors (I1-R) in the heart remains unclear. In rat hearts, UK14.304 and moxonidine reduced norepinephrine (NE) release. AGN192403 had no influence on NE, whereas rilmenidine, agmatine, rauwolscine, and efaroxan increased NE. These effects of moxonidine and rilmenidine were not affected by AGN192403 adminstration. Conversely, after pretreatment with UK14.304, only moxonidine displayed a pronounced inhibitory action on NE release (sensitive to AGN192403), indicating a synergistic inhibitory action at I1-R under conditions of a stimulated alpha2-adrenergic autoinhibition.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Agmatine; Animals; Benzofurans; Bridged Bicyclo Compounds; Brimonidine Tartrate; Drug Synergism; Heart; Heptanes; Imidazoles; Imidazoline Receptors; Norepinephrine; Oxazoles; Quinoxalines; Rats; Rats, Wistar; Receptors, Adrenergic, alpha-2; Receptors, Drug; Rilmenidine; Sympathetic Nervous System; Sympatholytics; Yohimbine

In pithed spontaneous hypertensive rats, noradrenaline overflow was diminished by moxonidine even when alpha(2)-adrenoceptors were blocked quantitatively using phenoxybenzamine, suggesting an I(1)-receptor-mediated mechanism of noradrenaline release. This hypothesis was confirmed, since the noradrenaline overflow was (1) increased under alpha(2)-adrenoceptors blockade by the mixed I(1)/alpha(2)-antagonists efaroxan or idazoxan, (2) still reduced by moxonidine when both alpha(2)- and I(1)-receptors were blocked, and (3) diminished by agmatine after pretreatment with phenoxybenzamine, but not with AGN192403. An indirect ganglionic I(1)-receptor-mediated mechanism of noradrenaline release is supposed.

Topics: Adrenergic alpha-Antagonists; Agmatine; Animals; Benzofurans; Blood Pressure; Bridged Bicyclo Compounds; Electric Stimulation; Heptanes; Hypertension; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Norepinephrine; Phenoxybenzamine; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2; Receptors, Drug; Spinal Cord

The putative imidazoline I(3) receptor antagonist 2-(2-ethyl-2,3-dihydrobenzo[b]furan-2-yl)-1H-imidazole (KU14R) has been shown to block the effects of the atypical I(3) agonist efaroxan at the level of the ATP-sensitive K(+) (K(ATP)) channel in isolated pancreatic islet beta cells, but its effects in vivo are not known. We have therefore investigated the effects of KU14R on blood glucose and insulin level in vivo. When KU14R was administered before or after a hypoglycaemic dose of efaroxan, the fall in blood glucose was at least additive. When the antihyperglycaemic imidazoline ligand S22068 was administered after a dose of KU14R, it did not alter the hypoglycaemic response. In the mouse isolated vas deferens preparation, neither rauwolscine (at concentrations which competitively antagonised the inhibitory response to 5-bromo-6-(2-imidazolin-2-ylamino)-quinoxaline (UK14304)) nor KU14R affected inhibition produced by S22068. At 10(-4) M, KU14R had weak alpha(2)-adrenoceptor antagonist activity. We conclude that KU14R does not act as an antagonist of either efaroxan or S22068 at an imidazoline site in vivo.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Blood Glucose; Brimonidine Tartrate; Dose-Response Relationship, Drug; Drug Interactions; Electric Stimulation; Homeostasis; Hypoglycemic Agents; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Insulin; Ligands; Male; Mice; Mice, Inbred CBA; Piperazines; Quinoxalines; Receptors, Drug; Time Factors; Vas Deferens

A novel ATP-sensitive potassium channel (K(ATP)) channel agonist, BPDZ 154 (6,7-dichloro-3-isopropylamino-4H-1,2,4-benzothiadiazine 1,1-dioxide), was synthesized, and its effects on insulin-secreting cells were evaluated using electrophysiology, (86)Rb(+) and (45)Ca(2+) efflux, and RIA determinations of insulin secretion. BPDZ 154, an analog of diazoxide, inhibited both glucose-induced insulin secretion from isolated perifused islets and the secretion of insulin induced by glucose and tolbutamide. These effects were mediated by the activation of ATP-sensitive potassium channels because BPDZ 154 induced a concentration-dependent increase in channel activity that was inhibited by the sulfonylurea tolbutamide and the imidazoline efaroxan. In beta-cells isolated from patients with either nontypical hyperinsulinism (preserved K(ATP) channel function) or from the control areas of the pancreas of patients with focal hyperinsulinism, BPDZ 154 activated K(ATP) channels and was found to be more effective and less readily reversible than diazoxide. By contrast, it was not possible to activate K(ATP) channels by either diazoxide or BPDZ 154 in beta-cells from patients with hyperinsulinism as a consequence of defects in K(ATP) channel function. In beta-cells isolated from a patient with pancreatic insulinoma, K(ATP) channels were readily recorded and modulated by BPDZ 154. These data suggest that BPDZ 154 or BPDZ 154-like compounds may have therapeutic potential in the treatment of certain forms of hyperinsulinism.

Topics: Adenosine Triphosphate; Adolescent; Adrenergic alpha-Antagonists; Animals; Benzofurans; Benzothiadiazines; Calcium Radioisotopes; Cell Line; Child, Preschool; Cyclic S-Oxides; Female; Glucose; Humans; Hyperinsulinism; Hypoglycemic Agents; Imidazoles; Infant; Insulin; Insulin Secretion; Insulinoma; Islets of Langerhans; Male; Pancreatic Neoplasms; Potassium Channels; Rats; Rats, Wistar; Rubidium Radioisotopes; Tolbutamide

Imidazoline and guanidiniun-substituted isoindoline compounds have been reported to demonstrate affinity for the putative imidazoline receptors (I(1)) and alpha-2 (alpha(2)) adrenoceptors. The purpose of this study was to determine the relative contribution of I(1) receptors to ocular actions of moxonidine (MOX) and brimonidine (BRIM) by utilizing relatively selective alpha(2) and I(1) antagonists. MOX, an alpha(2)/I(1) receptor agonist, BRIM, a selective alpha( 2) agonist, efaroxan (EFA), an I(1)/alpha(2) antagonist and rauwolscine (RAU), a relatively selective alpha(2) antagonist, were utilized to study alterations in sympathetically evoked contractions of the cat nictitating membrane (CNM). MOX (1-10 microg) suppressed, dose dependently, contractions of the CNM elicited by electrically stimulating the cervical preganglionic sympathetic trunk. The suppressive effect of MOX was antagonized more effectively by EFA (333 microg) than by rauwolscine (333 microg). In contrast, RAU, but not EFA, completely reversed the suppressive effects of BRIM on electrically induced contractions of the CNM. In conclusion, these in vivo data suggest that I(1) receptors are involved in the pre-junctional (neuronal) modulation of contractions in the CNM (Supported by NIH grant EY06338).

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Brimonidine Tartrate; Cats; Dose-Response Relationship, Drug; Electric Stimulation; Imidazoles; Imidazoline Receptors; Nictitating Membrane; Ocular Physiological Phenomena; Platelet Aggregation Inhibitors; Quinoxalines; Receptors, Adrenergic, alpha-2; Receptors, Drug; Yohimbine

Efaroxan induces membrane depolarization by interaction with the pore forming subunit of the ATP-sensitive potassium channel, Kir6.2. However, this effect is not responsible for its full secretory activity. In this study we have used an anti-idiotypic approach to generate antibodies that recognize additional proteins that may be regulated by efaroxan in pancreatic beta-cells. Using these antisera in an expression cloning strategy we have identified a monomeric GTP-binding protein, Rhes, as a potential target for regulation by imidazoline ligands. Rhes is shown to be expressed in beta-cells and its expression is regulated by efaroxan under conditions when a structurally related molecule, KU14R, is ineffective. The results reveal that beta-cells express Rhes and suggest that changes in the expression of this molecule may regulate the sensitivity of beta-cells to imidazoline secretagogues.

Topics: Animals; Antibody Specificity; Benzofurans; Cells, Cultured; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Fluorescent Antibody Technique; GTP-Binding Proteins; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Rabbits; Rats; Reverse Transcriptase Polymerase Chain Reaction

The objective of this study was to examine the ocular hydrodynamic effects of topically and centrally administered naphazoline, alone and following pretreatment with pertussis toxin (PTX) and alpha(2)/I(1)receptor antagonists. Topically and intracisternally administered naphazoline was examined for its ability to alter intraocular pressure (IOP) of rabbits in the absence and presence of receptor antagonists (rauwolscine, efaroxan) and a G(i/o)ribosylating agent PTX. In addition, the topical effects of naphazoline on pupil diameter and aqueous humor flow rate were evaluated. Topical unilateral application of naphazoline (7.5, 25 and 75 micro g; 25 micro l) elicited an ipsilateral dose-dependent mydriasis (2, 4 and 5.5 mm) that peaked at 2 hr with a duration of up to 5 hr. The IOP decreases induced by naphazoline were bilateral and dose-dependent (3, 6 and 10 mmHg); the response peaked at 1 hr and lasted for up to 5 hr. Pretreatment with efaroxan (250 micro g) elicited significantly greater antagonism of the ocular hypotensive response to naphazoline than did rauwolscine (250 micro g) suggesting an involvement of imidazoline (I(1)) receptors. Intracisternal application of naphazoline (3.3 micro g) also produced bilateral reductions (6 mmHg) of IOP that were immediate (10 min post drug) and lasted for approximately 2 hr. In PTX-pretreated (2.5 micro g kg(-1), i.a.) rabbits, the ocular hypotensive effects of naphazoline by both routes (topically and centrally) were attenuated by 50--65%. In addition to producing ocular hypotension, topical application of naphazoline (75 micro g; 25 micro l) caused significant reduction, from 2.8 to 1.5 micro l min(-1), in aqueous humor flow. These in vivo data indicate that, regardless of route of administration, alteration of aqueous humor flow by naphazoline was induced by the activation of alpha(2)and I(1)receptors. The ocular hypotensive effects produced by central administration did not result in sedation, therefore, there is the suggestion that central alpha(2)adrenergic receptors were stimulated minimally by naphazoline. Thus, these data suggest that ocular hypotensive effects and suppression of aqueous humor flow rate by naphazoline are mediated, in part, by alpha(2)and/or central I(1)at both central (brain) and peripheral (eye) sites. Moreover, these data indicate that the receptors are linked to PTX-sensitive G((i/o))proteins.

Topics: Administration, Topical; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Analysis of Variance; Animals; Aqueous Humor; Benzofurans; Dose-Response Relationship, Drug; Drug Synergism; Female; Imidazoles; Injections, Intraventricular; Intraocular Pressure; Male; Naphazoline; Pertussis Toxin; Pupil; Rabbits; Receptors, Adrenergic, alpha-2; Virulence Factors, Bordetella

Functional effects of prolonged exposure to the sulfonylurea, tolbutamide, were examined in the clonal electrofusion-derived BRIN-BD11 cell line. In acute 20-min incubations, 50-400 microM tolbutamide stimulated a dose-dependent increase (P < 0.01) in insulin release at both non-stimulatory (1.1 mM) and stimulatory (8.4 mM) glucose. Culture with 100 microM tolbutamide (18 hr) caused a marked (67%) decrease in subsequent insulin-secretory responsiveness to acute challenge with 200 microM tolbutamide, though notably, tolbutamide culture exerted no influence on 200 microM efaroxan-induced insulin secretion. Duration of exposure (3-18 hr) to 100 microM tolbutamide in culture also time-dependently influenced subsequent responsiveness to acute tolbutamide challenge, with progressive 47-58% decreases from 6-18 hr (P < 0.001). Similarly, 6- to 18-hr culture with 100 microM efaroxan specifically desensitized efaroxan-induced insulin release. Tolbutamide- and efaroxan-induced desensitization exhibited a time-dependent reversibility, with a sustained return to full insulin-secretory responsiveness by 12 hr. Notably, 18-hr culture with tolbutamide or efaroxan did not significantly affect insulinotropic responses to 16.7 mM glucose, 10 mM 2-ketoisocaproic acid, 10 mM alanine, 10 mM arginine, or 30 mM KCl. Diverse inhibitory actions of tolbutamide or efaroxan culture on late events in stimulus-secretion coupling reveal that drug desensitization is both a specific and important phenomenon. As such, the model system described could prove an important tool in determining the complex modes of action of established and novel clinically useful insulinotropic compounds.

Topics: Animals; Benzofurans; Cell Line; Drug Interactions; Glucose; Hypoglycemic Agents; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Rats; Sulfonylurea Compounds; Time Factors; Tolbutamide

Efaroxan, like several other imidazoline reagents, elicits a glucose-dependent increase in insulin secretion from pancreatic beta-cells. This response has been attributed to efaroxan-mediated blockade of KATP channels, with the subsequent gating of voltage-sensitive calcium channels. However, increasing evidence suggests that, at best, this mechanism can account for only part of the secretory response to the imidazoline. In support of this, we now show that efaroxan can induce functional changes in the secretory pathway of pancreatic beta-cells that are independent of KATP channel blockade. In particular, efaroxan was found to promote a sustained sensitization of glucose-induced insulin release that persisted after removal of the drug and to potentiate Ca2+-induced insulin secretion from electropermeabilized islets. To investigate the mechanisms involved, we studied the effects of the efaroxan antagonist KU14R. This agent is known to selectively inhibit insulin secretion induced by efaroxan, without altering the secretory response to glucose or KCl. Surprisingly, however, KU14R markedly impaired the potentiation of insulin secretion mediated by agents that raise cAMP, including the adenylate cyclase activator, forskolin, and the phosphodiesterase inhibitor isobutylmethyl xanthine (IBMX). These effects were not accompanied by any reduction in cAMP levels, suggesting an antagonistic action of KU14R at a more distal point in the pathway of potentiation. In accord with our previous work, islets that were exposed to efaroxan for 24 h became selectively desensitized to this agent, but they still responded normally to glucose. Unexpectedly, however, the ability of either forskolin or IBMX to potentiate glucose-induced insulin secretion was severely impaired in these islets. By contrast, the elevation of cAMP was unaffected by culture of islets with efaroxan. Taken together, the data suggest that, in addition to effects on the KATP channel, imidazolines also interact with a more distal component that is crucial to the potentiation of insulin secretion. This component is not required for Ca2+-dependent secretion per se but is essential to the mechanism by which cAMP potentiates insulin release. Overall, the results indicate that the actions of efaroxan at this distal site may be more important for control of insulin secretion than its effects on the KATP channel.

Topics: 1-Methyl-3-isobutylxanthine; Adenosine Triphosphate; Animals; Benzofurans; Bucladesine; Cyclic AMP; Diazoxide; Glucose; Imidazoles; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Male; Permeability; Phosphodiesterase Inhibitors; Potassium Channels; Rats; Rats, Wistar

Imidazoline compounds have been considered for the treatment of type 2 diabetes. We have now investigated the effects of imidazolines on interleukin (IL)-1beta-induced beta-cell apoptosis and the signal transduction pathways involved. Inhibition of Ca2+ influx into beta-cells by D-600, a blocker of voltage-gated L-type Ca2+ channels, suppressed IL-1beta-induced apoptosis. Our data show that calcineurin, Ca2+/calmodulin-dependent serine/threonine protein phosphatase 2B, is responsible for the effect of Ca2+ on beta-cell apoptosis. We also demonstrate that IL-1beta-mediated apoptosis correlates with expression of inducible nitric oxide synthase (iNOS) and the increase in intracellular production of nitric oxide. An inhibitor of cGMP-dependent protein kinase (PKG), KT5823, suppressed IL-1beta-induced apoptosis, suggesting the involvement of a PKG-dependent pathway in the apoptotic process. One of the major findings in this study is that imidazoline compounds RX871024 and efaroxan, suggested as prototypes of a new generation of drugs against type 2 diabetes, can protect against IL-1beta-induced apoptosis in pancreatic beta-cells, possibly by their inhibition of the expression of iNOS, a key element in the IL-1beta-induced apoptotic pathway in pancreatic beta-cells. These data suggest that imidazoline compounds should be explored as a potential therapeutic agent for the treatment of both type 1 and type 2 diabetes.

Topics: Animals; Apoptosis; Benzofurans; Calcineurin; Calcineurin Inhibitors; Calcium Channel Blockers; Calcium Channels; Cells, Cultured; Dose-Response Relationship, Drug; Enzyme Inhibitors; Gallopamil; Imidazoles; Indoles; Interleukin-1; Islets of Langerhans; Membrane Potentials; Mice; Mice, Obese; Models, Biological; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitriles; Patch-Clamp Techniques; Pyrethrins

In the present study, we tested the hypothesis that the activation of imidazoline I(1)-receptor, which is coupled to phosphatidylcholine-specific phospholipase C, results in downstream activation of mitogen-activated protein kinase (p42(mapk) and p44(mapk) isoforms) in PC12 cells. PC12 cells pretreated with nerve growth factor (50 ng/ml, 48 h) to initiate neuronal differentiation were incubated with [methyl-3H]choline and [3H]myristate. Activation of imidazoline I(1) receptor by rilmenidine (10 microM) caused time-dependent increases in diacylglycerol accumulation and phosphocholine release. The Western blotting analysis showed that rilmenidine (10 microM) produced a time-dependent activation of p42(mapk) and p44(mapk) that reached its maximum at 15 min and returned to control levels after 30 min. This finding was confirmed by immunofluorescence labeling of activated mitogen-activated protein kinase in the same model system. Efaroxan (imidazoline I(1)-receptor antagonist) or tricyclodecan-9-yl-xanthogenate (D609, phosphatidylcholine-specific phospholipase C inhibitor) attenuated the phosphorylation of p42(mapk) and p44(mapk) induced by rilmenidine. Nerve growth factor-induced phosphorylation of both mitogen-activated protein kinase isoforms was not affected by D609. These results support the hypothesis that the activation of the imidazoline I(1) receptor coupled phosphatidylcholine-specific phospholipase C results in the downstream activation of mitogen-activated protein kinase.

Topics: Acetylcholine; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Choline; Diglycerides; Enzyme Activation; Imidazoles; Imidazoline Receptors; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Nerve Growth Factor; Oxazoles; PC12 Cells; Phosphorylation; Phosphorylcholine; Rats; Receptors, Drug; Rilmenidine; Signal Transduction; Type C Phospholipases

The blockade of exocytosis induced by the putative endogenous ligand for imidazoline receptors, agmatine, was studied by using on-line measurement of catecholamine release in bovine adrenal medullary chromaffin cells. Agmatine inhibited the acetylcholine-evoked release of catecholamines in a concentration-dependent manner (IC(50)=366 microM); the K(+)-evoked release of catecholamines was unaffected. Clonidine (100 microM) and moxonidine (100 microM) also inhibited by 75% and 50%, respectively, the acetylcholine-evoked response. In cells voltage-clamped at -80 mV, the intermittent application of acetylcholine pulses elicited whole-cell inward currents (I(ACh)) that were blocked 63% by 1 mM agmatine. The onset of blockade was very fast (tau(on) = 31 ms); the recovery of the current after washout of agmatine also occurred very rapidly (tau(off = 39 ms). Efaroxan (10 microM) did not affect the inhibition of I(ACh) elicited by 1 mM agmatine. I(ACh) was blocked 90% by 100 microM clonidine and 50% by 100 microM moxonidine. The concentration-response curve for acetylcholine to elicit inward currents was shifted to the right in a non-parallel manner by 300 microM agmatine. The blockade of I(ACh) caused by agmatine (100 microM) was similar at various holding potentials, around 50%. When intracellularly applied, agmatine did not block I(ACh). At 1 mM, agmatine blocked I(Na) by 23%, I(Ba) by 14%, I(K(Ca)) by 16%, and I(K(VD)) by 18%. In conclusion, agmatine blocks exocytosis in chromaffin cells by blocking nicotinic acetylcholine receptor currents. In contrast to previous views, these effects seem to be unrelated to imidazoline receptors.

Topics: Acetylcholine; Agmatine; Animals; Benzofurans; Binding Sites; Binding, Competitive; Calcium Channels; Catecholamines; Cattle; Cells, Cultured; Chromaffin Cells; Clonidine; Dose-Response Relationship, Drug; Electric Stimulation; Imidazoles; Imidazoline Receptors; Membrane Potentials; Potassium; Potassium Channels; Receptors, Drug; Receptors, Nicotinic; Sodium Channels

Our previous studies have shown that ethanol counteracts centrally mediated hypotensive responses to clonidine. In this study, we investigated the relative roles of central alpha2-adrenergic and I1 imidazoline receptors in the antagonistic ethanol-clonidine hemodynamic interaction. The effects of selective blockade of alpha2- or I1 receptor by 2-methoxyidazoxan and efaroxan, respectively, on the blood pressure and heart rate responses to clonidine and subsequent ethanol administration were evaluated in conscious spontaneously hypertensive rats. Intracisternal administration of clonidine (1.5 microg/kg) produced significant (30 mm Hg; p < 0.05) and sustained (at least 60 min) decreases in blood pressure and heart rate. Systemic ethanol (1 g/kg), administered 10 min after clonidine, counteracted the hypotensive response and restored blood pressure to the preclonidine levels. Treatment with 2-methoxyidazoxan (0.16 microg/kg, intracisternal) or efaroxan (0.45 microg/kg, intracisternal) produced similar attenuation of the hypotensive and bradycardic responses to clonidine. The ability of ethanol to counteract the hypotensive action of clonidine was significantly (p < 0.05) attenuated in rats pretreated with efaroxan. The pressor response to ethanol lasted only 10 min compared with at least 60 min in the absence of efaroxan. In contrast, ethanol counteraction of clonidine-evoked hypotension was not altered when alpha2-adrenoceptors were blocked by 2-methoxyidazoxan. These findings suggest that centrally mediated hypotensive and bradycardic effects of clonidine in conscious spontaneously hypertensive rats involve activation of both alpha2-adrenergic and I1 imidazoline receptors. Furthermore, the findings suggest the dependence of a fully expressed ethanol counteraction of the hypotensive action of clonidine on functional I1 receptor within the central nervous system.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Catheterization; Clonidine; Drug Interactions; Ethanol; Heart Rate; Hypertension; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2; Receptors, Drug

Local injection of 4 nmol moxonidine (unilaterally) into the rostroventrolateral medulla of spontaneously hypertensive rats (SHR-SP) decreased mean blood pressure and heart rate by 24+/-3 and 3+/-4%, respectively. Pretreatment with the I1/alpha2-receptor antagonist efaroxan abolished the moxonidine-induced decrease in mean blood pressure, but had no effect on heart rate. Yohimbine blocked hypotension, delayed bradycardia (8 nmol), or completely inhibited the effects of moxonidine (16 nmol). Our results indicate that both I1-imidazoline receptors and alpha2-adrenoceptors of the rostroventrolateral medulla are involved in the realization of moxonidine-induced changes.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Heart Rate; Hemodynamics; Imidazoles; Imidazoline Receptors; Male; Medulla Oblongata; Rats; Rats, Inbred SHR; Receptors, Adrenergic, alpha-2; Receptors, Drug; Yohimbine

The purpose of this work is to determine the relative contributions of central imidazoline (I(1)) receptors to the ocular hydrodynamic action of moxonidine. Moxonidine (MOX), an alpha(2) and I(1) receptor agonist, and efaroxan (EFA), a relatively selective I(1) antagonist, were utilized to study alterations in intraocular pressure (IOP) and aqueous flow in New Zealand white rabbits subjected to intracerebroventricular (i.c.v.) cannulation and sympathectomy. Intracerebroventricular administration of MOX (0.033, 0.33 and 3.33 microg) to normal rabbits produced dose-dependent, bilateral IOP decreases of 3, 6, and 8 mmHg, respectively. The ocular hypotensive response to MOX was immediate (10 min. post drug), lasted for one hour, and was inhibited by prior administration of efaroxan (3.33 microg i.c.v.). In unilaterally sympathectomized (SX) rabbits, the ocular hypotensive response induced by i.c.v MOX in the denervated eye was attenuated approximately 50%, but the duration of ocular hypotension in the surgically altered eye was longer than that of the normal eye. MOX (0.33 microg i.c.v.), caused a statistically significant decrease (2.24 to 1.59 ml/min.) in aqueous flow in normal eyes. In SX eyes, there was no change in aqueous flow by MOX, suggesting that IOP effect in i.c.v. MOX observed in the SX eye might be mediated by changes in outflow resistance. Sedation was observed in all the rabbits treated with MOX (i.c.v.) and was dose-dependent. These in vivo data support the suggestion that centrally located I(1) receptors modulate the early contralateral response to topically administered MOX and are involved in lowering of IOP and aqueous flow in rabbit. In addition, expression of the full ocular hypotensive effect of centrally applied MOX depends on intact sympathetic innervation. Ocular hypotension induced by MOX in the SX eye may involve an effect on uveoscleral outflow.

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Aqueous Humor; Benzofurans; Brain; Dose-Response Relationship, Drug; Female; Imidazoles; Imidazoline Receptors; Injections, Intraventricular; Intraocular Pressure; Male; Rabbits; Receptors, Drug; Sympathectomy; Sympathetic Nervous System

The effect of alpha-2 adrenoceptor antagonists, yohimbine and efaroxan, on the plasma glucose and insulin levels was studied in non-diabetic control, type-I (insulin-dependent) and type-II (non-insulin-dependent) diabetic rats. Pretreatment with either yohimbine or efaroxan potentiated glucose-induced insulin release in non-diabetic control rats and produced an improvement of the oral glucose tolerance and potentiated glucose-induced insulin release in type-II but not in type-I diabetic rats. Treatment with either yohimbine or efaroxan reduced the plasma glucose level and increased the plasma insulin level of non-diabetic control and type-II diabetic rats but not of type-I diabetic rats. Effects of efaroxan were more marked. Pretreatment of non-diabetic control and type-II diabetic rats with either yohimbine or efaroxan inhibited clonidine-induced hyperglycaemia and suppressed or reversed clonidine-induced hypoinsulinaemia. Also, pretreatment of these animals with either yohimbine or efaroxan enhanced the hypoglycaemic and insulinotropic effects of glibenclamide. The combination of glibenclamide and efaroxan led to a synergistic increase in insulin secretion, while that of glibenclamide and yohimbine led to an additive increase. The hyperglycaemic effect of diazoxide in non-diabetic control and type-II diabetic rats was inhibited by pretreatment with either yohimbine or efaroxan. The hypoinsulinaemic effect of diazoxide in these animals was antagonized and reversed by pretreatment with yohimbine and efaroxan, respectively. In type-I diabetic rats, there was no change in the plasma glucose and insulin levels induced by the treatment of animals with each of clonidine or diazoxide alone or in combination with either yohimbine or efaroxan. Glibenclamide produced a slight decrease in the plasma glucose level of type-I diabetic rats, at the end of the 120 min period of investigation but there was no change in the plasma insulin level. Pretreatment of these animals with either yohimbine or efaroxan produced no change in glibenclamide effects. Additionally, bath application of efaroxan or glibenclamide inhibited the relaxant effects of different concentrations of diazoxide on the isolated norepinephrine-contracted aortic strips, while the application of yohimbine produced insignificant changes. The combination of glibenclamide and efaroxan led to complete inhibition of the relaxant effects of different concentrations of diazoxide, while that of glibenclamide and

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Aorta, Thoracic; Benzofurans; Blood Glucose; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Dose-Response Relationship, Drug; Drug Therapy, Combination; Hypoglycemic Agents; Imidazoles; In Vitro Techniques; Insulin; Male; Rabbits; Rats; Rats, Wistar; Vasodilation; Yohimbine

Pancreatic beta-cells express imidazoline binding sites which play a role in the regulation of insulin secretion, but it is not known whether ligands for these sites also affect other aspects of beta-cell physiology. In the present study, we have investigated the effects of a range of imidazoline reagents on the growth and viability of clonal pancreatic beta-cells (RINm5F and HIT-T15). Three imidazoline compounds (idazoxan, phentolamine and antazoline) were found to cause marked inhibition of beta-cell growth in a time and concentration dependent manner. Idazoxan was the most potent of these with as little as 0.5 microM causing a significant decrease in beta-cell viability (EC50 approximately 10 microM). All three imidazolines also decreased the viability of clonal beta-cells in parallel with their inhibitory effects on cell growth. These effects were not reproduced by any of a wide-range of other imidazoline compounds, including effective insulin secretagogues such as efaroxan and RX821002. The effects of the three ligands did not correlate with their relative potencies for binding to any of the well-characterised imidazoline binding sites nor to alpha2-adrenoceptors. In addition, the inhibitory responses were not antagonised by other imidazoline binding site ligands. The inhibitory effects of idazoxan on the growth of RINm5F and HIT-T15 beta-cells required as little as 3-h exposure to the imidazoline and were not readily reversible when the reagent was removed. Reductions in growth rate were accompanied by marked alterations in the morphology of the cells, which could be detected before loss of viability. Cells exposed to phentolamine showed the characteristic features of apoptosis in that the nuclei were condensed (as judged by acridine orange staining) and electrophoresis of DNA revealed the presence of oligonucleosomal fragmentation. These changes could not be detected in cells exposed to idazoxan despite the more profound reduction in viability induced by this agent. We conclude that a sub-group of imidazoline compounds can exert profoundly detrimental effects on the growth and viability of clonal beta-cells but that these effects do not correlate with their binding affinity at imidazoline binding sites or alpha2-adrenoceptors.

Topics: Animals; Antazoline; Apoptosis; Benzofurans; Binding Sites; Cell Division; Cell Survival; Idazoxan; Imidazoles; Imidazoline Receptors; Insulin; Insulin Secretion; Islets of Langerhans; PC12 Cells; Phentolamine; Rats; Receptors, Drug

Moxonidine is a mixed I(1) imidazoline/alpha(2)moxonidine=morphine. The I(1) imidazoline preferring antagonist efaroxan produced a dose-dependent antagonism of both moxonidine (5.0 mg/kg) and clonidine (0.5 mg/kg). In addition, the alpha(2)-adrenergic receptor antagonist yohimbine produced a dose-related antagonism of moxonidine, but only partially antagonized clonidine. Prazosin failed to block the effects of either moxonidine or clonidine, indicating a lack of involvement of alpha(1) as well as alpha(2B) and alpha(2C) receptors. The present results suggest that alpha(2)-adrenergic receptors play an important role in mediating the effects of moxonidine in producing antinociception in the formalin test. Further, the present results demonstrate that the mechanism of action of moxonidine and clonidine differ in that clonidine, but not moxonidine, produces an antinociceptive effect through a yohimbine-insensitive mechanism in the formalin test.

Topics: Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Analgesics, Non-Narcotic; Animals; Benzofurans; Clonidine; Dose-Response Relationship, Drug; Formaldehyde; Imidazoles; Imidazoline Receptors; Injections, Subcutaneous; Male; Pain; Pain Measurement; Prazosin; Rats; Rats, Sprague-Dawley; Receptors, Drug; Time Factors; Yohimbine

The beta-carboline, harmane (0.1 - 1.0 nmol) produces dose dependent hypotension when microinjected unilaterally into the rostral ventrolateral medulla (RVLM) of the anaesthetized rat. The potency of harmane on blood pressure is similar to that of the imidazoline, clonidine. The hypotensive effects of both clonidine and harmane are reversed by microinjection of the relatively I(1)-receptor selective antagonist efaroxan (20 nmol). These results are consistent with harmane acting at an I(1)-receptor in the RVLM. This is the first report of an endogenous ligand for I(1)-receptors that has central effects on blood pressure.

Topics: Adrenergic alpha-Agonists; Animals; Benzofurans; Blood Pressure; Clonidine; Harmine; Heart Rate; Hypotension; Imidazoles; Imidazoline Receptors; Male; Medulla Oblongata; Microinjections; Rats; Rats, Sprague-Dawley; Receptors, Drug

K(ATP)-channel-dependent and K(ATP)-channel-independent insulin-releasing actions of the sulfonylurea, tolbutamide, were examined in the clonal BRIN-BD11 cell line. Tolbutamide stimulated insulin release at both nonstimulatory (1.1 mM) and stimulatory (16. 7 mM) glucose. Under depolarizing conditions (16.7 mM glucose plus 30 mM KCl) tolbutamide evoked a stepwise K(ATP) channel-independent insulinotropic response. Culture (18 h) with tolbutamide or the guanidine derivative BTS 67 582 (100 microM) markedly reduced (P < 0. 001) subsequent responsiveness to acute challenge with tolbutamide, glibenclamide, and BTS 67 582 but not the imidazoline drug, efaroxan. Conversely, 18 h culture with efaroxan reduced (P < 0.001) subsequent insulinotropic effects of efaroxan but not that of tolbutamide, glibenclamide, or BTS 67 582. Culture (18 h) with tolbutamide reduced the K(ATP) channel-independent actions of both tolbutamide and glibenclamide. Whereas culture with efaroxan exerted no effect on the K(ATP) channel-independent actions of sulfonylureas, BTS 67 582 abolished the response of tolbutamide and inhibited that of glibenclamide. These data demonstrate that prolonged exposure to tolbutamide desensitizes both K(ATP)-channel-dependent and -independent insulin-secretory actions of sulfonylureas, indicating synergistic pathways mediated by common sulfonylurea binding site(s).

Topics: Adenosine Triphosphate; Adrenergic alpha-Antagonists; Animals; Benzofurans; Cell Line; Dose-Response Relationship, Drug; Glucose; Guanidines; Hypoglycemic Agents; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Potassium Channels; Potassium Chloride; Rats; Tolbutamide

In the mouse medulla oblongata, we characterized binding properties and functional responses of two recognition sites for imidazoline compounds: I(1)-imidazoline and alpha(2)-adrenergic receptors. The mouse medulla expresses a higher density of I(1)-receptors than in the rat, whereas alpha(2)-receptor densities were similar between the two species. In anesthetized, ventilated and paralyzed mice, we tested the hypotensive actions of the I(1)/alpha(2) agonist moxonidine, determined its central site of its actions, and the relative roles of I(1) and alpha(2)-receptors. Experiments were performed in C(57)Bl(6) wild type and alpha(2A)-adrenergic receptor deficient mice. In both types of mice, neuronal activation within the rostral ventrolateral medulla (RVLM) region by glutamate microinjection elicited increases in arterial pressure. Moxonidine (0.5 nmol/site/10 nl) microinjected bilaterally into this vasopressor region decreased arterial pressure by 30% and heart rate by 11% in wild type mice. Efaroxan, the I(1)/alpha(2) antagonist (0.4 nmol) when microinjected into the RVLM elevated blood pressure itself and abolished the action of moxonidine, whereas alpha(2)-blockade with SK&F 86466 had no significant effect on blood pressure and did not attenuate moxonidine's effect. To more definitively test the role of alpha(2)-adrenergic receptors in the action of moxonidine, moxonidine was microinjected into the RVLM of alpha(2A)-adrenergic deficient mice. The decreases in arterial pressure were nearly identical to those of wild type mice, whereas bradycardia was attenuated. Thus, in the mouse moxonidine acts within the RVLM region to lower arterial pressure mainly through the I(1)-imidazoline receptor independent of alpha(2)-adrenergic receptors.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Amygdala; Animals; Antihypertensive Agents; Benzofurans; Binding, Competitive; Blood Pressure; Brain Chemistry; Glutamic Acid; Heart Rate; Hypertension; Imidazoles; Imidazoline Receptors; Injections, Intravenous; Medulla Oblongata; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Microinjections; Pons; Receptors, Adrenergic, alpha-2; Receptors, Drug

Moxonidine, an antihypertensive imidazoline compound, reduces blood pressure by selective activation of central imidazoline I(1)-receptors and inhibition of sympathetic nerve activity and by direct actions on the kidney, with both mechanisms resulting in diuresis and natriuresis. We hypothesized that the hypotensive and renal actions of moxonidine may be mediated by atrial natriuretic peptide (ANP), a cardiac peptide involved in pressure and volume homeostasis through its vasodilatory, diuretic, and natriuretic actions. Renal parameters were measured on an hourly basis over a period of 4 hours in conscious rats that received bolus intravenous injections of moxonidine (1 to 150 microg/300 microL saline). During the first hour, moxonidine dose-dependently stimulated diuresis, natriuresis, kaliuresis, and urinary cGMP, the index of ANP activity. Moxonidine (50 microg) significantly (P<0.001) stimulated urinary volume (0.35+/-0.04 versus 1.05+/-0.09 mL/h per 100 g), sodium (14. 3+/-2.5 versus 51.8+/-6.5 micromol/h per 100 g), potassium (10.5+/-2. 3 versus 32.3+/-3.2 micromol/h per 100 g), and cGMP (325+/-52 versus 744+/-120 pmol/h per 100 g). Pretreatment with a selective imidazoline receptor antagonist, efaroxan, dose-dependently inhibited moxonidine-stimulated renal parameters. Efaroxan (25 microg per rat) significantly inhibited moxonidine-stimulated diuretic and natriuretic effects and urinary cGMP excretion (744+/-120 versus 381+/-137 pmol/h per 100 g, P<0.02). The alpha(2)-adrenoceptor antagonist yohimbine (50 microg per rat) partially yet significantly inhibited moxonidine-stimulated diuresis and natriuresis but not cGMP excretion. Plasma ANP was dose-dependently increased by moxonidine and was inhibited by pretreatment with efaroxan (220.8+/-36.9 versus 100.3+/-31.7 pg/mL, P<0.03) but not by yohimbine. In conclusion, selective in vivo activation of imidazoline receptors by moxonidine is associated with dose-dependent diuresis, natriuresis, and kaliuresis as well as stimulated plasma ANP and urinary cGMP excretion, thus implicating ANP in the renal actions of moxonidine.

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Atrial Natriuretic Factor; Benzofurans; Cyclic GMP; Diuresis; Dose-Response Relationship, Drug; Female; Imidazoles; Imidazoline Receptors; Injections, Intravenous; Kidney; Natriuresis; Potassium; Rats; Rats, Sprague-Dawley; Receptors, Drug; Yohimbine

It is hypothesized that the locus coeruleus-noradrenergic system controls compensatory and repair mechanisms in the CNS, and that its dysfunction is a critical factor in the progression of central neurodegenerative diseases. Pharmacological activation of locus coeruleus neurons can be achieved with alpha2-adrenoceptor antagonists, and such compounds are protective in vivo in some models of brain injury where excitotoxicity is thought to be a causative factor. To further explore this neuroprotective potential, the effects of a 7-day treatment with the alpha2-antagonists, (+)-efaroxan and (+/-)-idazoxan, were evaluated in rats undergoing a unilateral lesioning of the striatum with the excitotoxin, quinolinic acid. The alpha2-antagonist treatments reduced both the ipsiversive circling response to apomorphine and the deficit of choline acetyltransferase in the lesioned animals. To elucidate the mechanisms underlying this neuroprotective effect, a modulation of the extracellular levels of amino acids within the striatum was investigated using in vivo microdialysis. Intrastriatal injection of quinolinic acid increased taurine and tyrosine levels by 2-2.5 fold, while most other amino acids were not significantly altered; the effect of (+)-efaroxan on these changes is being investigated. Further research is required to identify which of several possible mechanisms is involved in the neuroprotective action of alpha2-antagonists in vivo.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Behavior, Animal; Benzofurans; Corpus Striatum; Idazoxan; Imidazoles; Male; Microdialysis; Rats; Rats, Sprague-Dawley

The present study examined the influence of alpha-2 adrenoceptor ligands on circling behavior in rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal pathway. The alpha-2 adrenoceptor agonists, clonidine and UK 14304, inhibited both the ipsilateral rotation induced by the indirect dopaminergic agonist, methylphenidate, and the contralateral circling induced by the direct dopaminergic agonist, apomorphine. In contrast, the alpha-2 adrenoceptor antagonists, idazoxan and (+/-)-efaroxan, enhanced the circling induced by either methylphenidate or apomorphine. The facilitating activity of efaroxan was stereoselective because the (+)-enantiomer mimicked the effect of (+/-)-efaroxan, whereas the (-)-enantiomer was essentially inactive, thus indicating a mediation by alpha-2 adrenoceptors. Upon administration alone, the above-mentioned compounds did not modify spontaneous circling behavior, except for UK 14304, which decreased, and (+)-efaroxan, which slightly increased, the ipsilateral rotation. We conclude that activation and antagonism of alpha-2 adrenoceptors inhibit and enhance, respectively, the circling behavior evoked by both direct and indirect dopaminergic agonists. Although a modulation of dopamine release may be involved in some of these drug effects, the effects on apomorphine-induced circling indicate an influence of alpha-2 adrenoceptor compounds on nigrostriatal neurotransmission at sites downstream from the dopaminergic neurons themselves. These findings support the notion of a potential benefit of alpha-2 adrenoceptor antagonists in the treatment of Parkinson's disease.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Apomorphine; Behavior, Animal; Benzofurans; Brimonidine Tartrate; Clonidine; Dopamine Agonists; Idazoxan; Imidazoles; Ligands; Male; Methylphenidate; Oxidopamine; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-2; Stereoisomerism; Stereotyped Behavior; Substantia Nigra

The aim of this study was to determine whether alpha2-adrenoceptors or imidazoline I1-receptors are responsible for the central sympathoinhibition produced by rilmenidine and moxonidine, two clonidine-like antihypertensive drugs. Rilmenidine and moxonidine were compared with the indirectly acting alpha2-adrenoceptor agonist alpha-methyldopa. Three antagonists were used. Yohimbine and SK&F86466 were used as selective alpha2-adrenoceptor antagonists. They were compared with efaroxan which is also an alpha2-adrenoceptor antagonist, but, in addition, possesses affinity for imidazoline I1-receptors. According to some but not all studies, the affinity of efaroxan for I1-receptors is much higher than its affinity for alpha2-adrenoceptors. Drugs were administered into the cisterna cerebellomedullaris of conscious rabbits by a catheter implanted previously under halothane anaesthesia. Rilmenidine (10 microg kg(-1)), moxonidine (0.3 microg kg(-1)) and alpha-methyldopa (0.4 mg kg(-1)) lowered blood pressure and the plasma noradrenaline concentration; the degree of sympathoinhibition produced by the three agonists was very similar. When injected after the agonists, efaroxan (0.1-14 microg kg(-1); cumulative doses), yohimbine (0.4-14 microg kg(-1)) and SK&F86466 (0.4-44 microg kg(-1)) counteracted the effects of the agonists on blood pressure and the plasma noradrenaline concentration. Efaroxan was about tenfold more potent than yohimbine and SK&F86466 at antagonizing the hypotensive effects of alpha-methyldopa. Similarly, efaroxan was two- to tenfold more potent than yohimbine and SK&F86466 against rilmenidine and moxonidine. Finally, efaroxan was about as potent against alpha-methyldopa as against rilmenidine and moxonidine. The results confirm previous observations that selective alpha2-adrenoceptor antagonists are capable of completely antagonizing effects of rilmenidine and moxonidine. The effects of the alpha2-adrenoceptor antagonist with an additional high affinity for imidazoline I1-receptors, efaroxan, can also be explained by blockade of alpha2-adrenoceptors. Efaroxan was more potent against rilmenidine and moxonidine than the selective alpha2-adrenoceptor antagonists. This was probably due to the fact that the affinity of efaroxan for alpha2-adrenoceptors is higher than the affinity of yohimbine and SK&F86466, since efaroxan was also the most potent of the three antagonists against the indirectly acting alpha2adrenoceptor agonist alpha-methyldopa. The obser

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzazepines; Benzofurans; Blood Pressure; Dose-Response Relationship, Drug; Drug Interactions; Female; Heart Rate; Imidazoles; Imidazoline Receptors; Male; Methyldopa; Norepinephrine; Oxazoles; Rabbits; Receptors, Adrenergic, alpha-2; Receptors, Drug; Rilmenidine; Sympathetic Nervous System; Yohimbine

Acetylcholine release in the rat cortex in vivo has been shown to be modulated by alpha2-adrenoceptor ligands. We have previously reported that the systemic administration of selective alpha2-antagonists including (+)-efaroxan increase, while alpha2-adrenoceptor agonists such as UK-14304 reduce the release of acetylcholine in the medial prefrontal cortex of conscious rats as measured by microdialysis. To evaluate the extent to which noradrenergic afferent inputs are required for the expression of these different effects, the present study examined the drug-induced changes in cortical acetylcholine release in rats which had undergone prior noradrenergic deafferentation. Rats were pretreated with the noradrenergic neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (40 mg/kg, i.p.), which after three days had reduced noradrenaline levels in the medial prefrontal cortex by 84%. At that time, slices of cortex were incubated with [3H]choline, superfused and stimulated by consecutive exposures to increasing concentrations of K+. In N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine pretreated tissue, the [3H] outflows evoked by 20, 35 and 45 mM K+ were lower by 12%, 22% and 43%, respectively, in comparison to slices prepared from vehicle-pretreated control animals. For in vivo microdialysis experiments, rats were pretreated as above with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine, or prepared seven to eight days in advance with bilateral 6-hydroxydopamine lesions of the locus coeruleus. Neither of these lesioning procedures significantly affected the basal outflow of endogenous acetylcholine in the cortex. In control rats, cortical acetylcholine outflow was increased by up to 300% of baseline values by (+)-efaroxan (0.63 mg/kg, i.p.), and was reduced to 21% of baseline by UK-14304 (2.5 mg/kg, i.p.), confirming our previous findings. In N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine pretreated rats, the inhibitory effect of UK-14304 on acetylcholine outflow persisted, while the ability of (+)-efaroxan to increase outflow was essentially eliminated. In locus coeruleus-lesioned rats, where cortical noradrenaline levels were reduced by 64%, (+)-efaroxan still increased acetylcholine outflow, but this effect was significantly attenuated and less sustained in comparison to sham-operated control rats. Viewed together with complimentary biochemical, electrophysiological and neuroanatomical evidence in the literature, a model is presented to account for these findings, an

Topics: Acetylcholine; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Benzylamines; Brimonidine Tartrate; Cerebellum; Cerebral Cortex; Epinephrine; Globus Pallidus; Imidazoles; In Vitro Techniques; Kinetics; Parietal Lobe; Prefrontal Cortex; Quinoxalines; Rats; Receptors, Adrenergic, alpha-2; Substantia Innominata; Sympathomimetics

alpha2-Adrenergic receptor (AR)-selective compounds produce antihypertensive and antinociceptive effects. Moxonidine alleviates hypertension in multiple species, including humans. This study demonstrates that intrathecally administered moxonidine produces antinociception in mice. Antinociception was detected via the (52.5 degrees C) tail-flick and Substance P (SP) nociceptive tests. Moxonidine was intrathecally administered to ICR, mixed C57BL/6 x 129/Sv [wild type (WT)], or C57BL/6 x 129/Sv mice with dysfunctional alpha2aARs (D79N-alpha2a). The alpha2AR-selective antagonist SK&F 86466 and the mixed I1/alpha2AR-selective antagonist efaroxan were tested for inhibition of moxonidine-induced antinociception. Moxonidine prolonged tail-flick latencies in ICR (ED50 = 0.5 nmol; 0. 3-0.7), WT (0.17 nmol; 0.09-0.32), and D79N-alpha2a (0.32 nmol; 0. 074-1.6) mice. Moxonidine inhibited SP-elicited behavior in ICR (0. 04 nmol; 0.03-0.07), WT (0.4 nmol; 0.3-0.5), and D79N-alpha2a (1.1 nmol; 0.7-1.7) mice. Clonidine produced antinociception in WT but not D79N-alpha2a mice. SK&F 86466 and efaroxan both antagonized moxonidine-induced inhibition of SP-elicited behavior in all mouse lines. SK&F 86466 antagonism of moxonidine-induced antinociception implicates the participation of alpha2ARs. The comparable moxonidine potency between D79N-alpha2a and WT mice suggests that receptors other than alpha2a mediate moxonidine-induced antinociception. Conversely, absence of clonidine efficacy in D79N-alpha2a mice implies that alpha2aAR activation enables clonidine-induced antinociception. When clinically administered, moxonidine induces fewer side effects relative to clonidine; moxonidine-induced antinociception appears to involve a different alpha2AR subtype than clonidine-induced antinociception. Therefore, moxonidine may prove to be an effective treatment for pain with an improved side effect profile.

Topics: Adrenergic alpha-2 Receptor Agonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Analgesics, Non-Narcotic; Animals; Autoreceptors; Benzazepines; Benzofurans; Dose-Response Relationship, Drug; Female; Imidazoles; Imidazoline Receptors; Injections, Spinal; Male; Mice; Mice, Inbred C57BL; Mice, Inbred ICR; Nerve Endings; Norepinephrine; Pain Measurement; Receptors, Adrenergic, alpha-2; Receptors, Drug; Spinal Cord

Topics: Animals; Benzofurans; Blood Pressure; Female; Heart Rate; Idazoxan; Imidazoles; Imidazoline Receptors; Male; Phenoxybenzamine; Rats; Rats, Wistar; Receptors, Drug; Structure-Activity Relationship; Yohimbine

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Brain; Clonidine; Dihydroxyphenylalanine; Imidazoles; Male; Quinolines; Rats; Rats, Sprague-Dawley; Serotonin; Serotonin Antagonists; Tryptophan Hydroxylase; Tyrosine 3-Monooxygenase

We examined the role of I1-imidazoline (I1-IR) receptors in control of airway function, by testing the effects of systemic administration of the I1-IR agonist moxonidine on reflex responses of tracheal smooth muscle (TSM) tone to either lung deflation or mechanical stimulation of intrapulmonary rapidly adapting receptors. Experiments were performed in either alpha-chloralose anesthetized or decorticate, paralyzed, and mechanically ventilated beagle dogs. Moxonidine (10-100 micrograms/kg) administered via three different routes (femoral vein, muscular branch of superior thyroid artery, and vertebral artery) attenuated TSM responses to stimulation of airway sensory nerve fibers by two different ways and caused a decrease in arterial pressure and heart rate. These effects were dose dependent and were significantly reversed by efaroxan (an I1-IR and alpha 2-adrenergic blocker) administered via the vertebral artery. Intravertebral efaroxan abolished the hemodynamic effects of moxonidine. Intravenous moxonidine (10-100 micrograms/kg) did not alter airway smooth muscle responses to electrical stimulation of the peripheral vagus nerve. In addition, in vitro moxonidine (1-100 micrograms/ml) had no effect on contractile responses to increasing doses of acetylcholine. These findings indicate that moxonidine may act at a central site to suppress reflex airway constriction, even when given into the systemic circulation. Given the presence of I1-IR sites and alpha 2-adrenergic receptors in brain regions participating in airway reflexes, these receptor classes may be involved in brainstem control of the cholinergic outflow to the airways.

Topics: Acetylcholine; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Decerebrate State; Dogs; Efferent Pathways; Electric Stimulation; Heart Rate; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Lung; Muscle, Smooth; Nerve Fibers; Neurons, Afferent; Receptors, Drug; Reflex; Trachea; Vagus Nerve

The key step of the synthesis of efaroxan was the dihydrobenzofuran ring formation involving an intramolecular cyclization of the tertiary alcohol intermediate with the fluoroaromatic moiety in basic medium. This carbinol was prepared according to two routes, either from reaction of a benzyl Grignard reagent with an alpha-ketoester, or from a Darzens condensation.

Topics: Adrenergic alpha-Antagonists; Benzofurans; Imidazoles

Efaroxan was synthetised by cyclisation of the tertiary alcohol 2 which was prepared by the ring opening of the gem-disubstituted epoxide 3 with ortho-metallated fluorobenzene.

Topics: Adrenergic alpha-Antagonists; Benzofurans; Epoxy Compounds; Imidazoles

The properties of ATP-sensitive K+ (K(ATP)) channels were explored in the electrofusion-derived, glucose-responsive, insulin-secreting cell line BRIN-BD11 using patch-clamp techniques. In intact cells, K(ATP) channels were inhibited by glucose, the sulfonylurea tolbutamide, and the imidazoline compounds efaroxan and phentolamine. Each of these agents initiated insulin secretion and potentiated the actions of glucose. K(ATP) channels were blocked by ATP in a concentration-dependent manner and activated by ADP in the presence of ATP. In both intact cells and excised inside-out patches, the K(ATP) channel agonists diazoxide and pinacidil activated channels, and both compounds inhibited insulin secretion evoked by glucose, tolbutamide, and imidazolines. The mechanisms of action of imidazolines were examined in more detail. Pre-exposure of BRIN-BD11 cells to either efaroxan or phentolamine selectively inhibited imidazoline-induced insulin secretion but not the secretory responses of cells to glucose, tolbutamide, or a depolarizing concentration of KCl. These conditions did not result in the loss of depolarization-dependent rises in intracellular Ca2+ ([Ca2+]i), K(ATP) channel operation, or the actions of either ATP or efaroxan on K(ATP) channels. Desensitization of the imidazoline receptor following exposure to high concentrations of efaroxan, however, was found to result in an increase in SUR1 protein expression and, as a consequence, an upregulation of K(ATP) channel density. Our data provide 1) the first characterization of K(ATP) channels in BRIN-BD11 cells, a novel insulin-secreting cell line produced by electrofusion techniques, and 2) a further analysis of the role of imidazolines in the control of insulin release.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Adrenergic alpha-Antagonists; Animals; Benzofurans; Cell Fusion; Cell Line; Diazoxide; Glucose; Imidazoles; Insulin; Insulin Secretion; Insulinoma; Islets of Langerhans; Membrane Potentials; Pancreatic Neoplasms; Phentolamine; Pinacidil; Potassium Channels; Tolbutamide; Tumor Cells, Cultured

To study the effect of moxonidine (Mox) on carotid sinus baroreflex.. By perfusing the carotid sinus in anesthetized rats, the functional parameters of baroreflex were measured. The femoral artery was perfused with constant flow and the change of perfusing pressure was recorded to determine the effect of Mox on vascular tone.. Mox 32 and 100 mumol.L-1 shifted the functional curve of carotid sinus baroreflex to the right and upward, with the reduction in peak slope and in reflex decrease of mean arterial pressure, suggesting that Mox produced an inhibitory effect on baroreflex. The effect of Mox 100 mumol.L-1 on baroreflex was completely blocked by efaroxan 100 mumol.L-1. Mox increased vascular resistance.. Mox inhibits carotid baroreflex via its constrictive action on sinus wall.

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Baroreflex; Benzofurans; Blood Pressure; Carotid Sinus; Imidazoles; Male; Rats; Vascular Resistance

In previous reports, [3H]5-HT has been used to characterize the pharmacology of the rat and human 5-HT2B receptors. 5-HT, the native agonist for the 5-HT2B receptor, has a limitation in its usefulness as a radioligand since it is difficult to study the agonist low-affinity state of a G protein-coupled receptor using an agonist radioligand. When using [3H]5-HT as a radioligand, rauwolscine was determined to have relatively high affinity for the human receptor (Ki human = 14.3+/-1.2 nM, compared to Ki rat = 35.8+/-3.8 nM). Since no known high affinity antagonist was available as a radioligand, these studies were performed to characterize [3H]rauwolscine as a radioligand for the cloned human 5-HT2B receptor expressed in AV12 cells. When [3H]rauwolscine was initially tested for its usefulness as a radioligand, complex competition curves were obtained. After testing several alpha2-adrenergic ligands, it was determined that there was a component of [3H]rauwolscine binding in the AV12 cell that was due to the presence of an endogenous alpha2-adrenergic receptor. The alpha2-adrenergic ligand efaroxan was found to block [3H]rauwolscine binding to the alpha2-adrenergic receptor without significantly affecting binding to the 5-HT2B receptor and was therefore included in all subsequent studies. In saturation studies at 37 degrees C, [3H]rauwolscine labeled a single population of binding sites, Kd = 3.75+/-0.23 nM. In simultaneous experiments using identical tissue samples, [3H]rauwolscine labeled 783+/-10 fmol of 5-HT2B receptors/mg of protein, as compared to 733+/-14 fmol of 5-HT2B receptors/mg of protein for [3H]5-HT binding. At 0 degrees C, where the conditions for [3H]5-HT binding should label mostly the agonist high affinity state of the human 5-HT2B receptor, [3H]rauwolscine (Bmax = 951+/-136 fmol/mg), again labeled significantly more receptors than [3H]5-HT (Bmax = 615+/-34 fmol/mg). The affinity of [3H]rauwolscine for the human 5-HT2B receptor at 0 degrees C did not change, Kd = 4.93+/-1.27 nM, while that for [3H]5-HT increased greatly (Kd at 37 degrees C = 7.76+/-1.06 nM; Kd at 0 degrees C = 0.0735+/-0.0081 nM). When using [3H]rauwolscine as the radioligand, competition curves for antagonist structures modeled to a single binding site, while agonist competition typically resulted in curves that best fit a two site binding model. In addition, many of the compounds with antagonist structures displayed higher affinity for the 5-HT2B receptor when [3H]rauwolscin

Topics: Adrenergic alpha-Antagonists; Benzofurans; Binding, Competitive; Cell Line; Cloning, Molecular; Humans; Imidazoles; Radioligand Assay; Receptor, Serotonin, 5-HT2B; Receptors, Serotonin; Serotonin Antagonists; Transfection; Tritium; Yohimbine

The presence of alpha2-adrenoceptors in membranes from omental and s.c. adipose tissue from gilts and barrows was shown in saturation binding assays with [3H]yohimbine. Four trials tested effects of alpha2-adrenoceptor antagonists (A2AA) on plasma concentrations of NEFA and urea nitrogen (PUN). In Trial 1, barrows were given i.v. injections of saline, 200 microg/kg BW of one of three A2AA (efaroxan, idazoxan, or RX821002), or 25 microg/kg BW of isoproterenol. Concentrations of NEFA were measured in plasma harvested every 15 min from 1 h before to 2 h after treatment. Compared with results for saline-treated pigs, areas under the curve (AUC) for NEFA were increased (P < .05) by efaroxan, RX821002, and isoproterenol. In Trial 2, barrows received i.v. doses of saline, efaroxan (200 or 400 microg/kg BW), or RX821002 (200 or 400 microg/kg BW). Levels of NEFA were quantified in plasma obtained at 15-min intervals through 2 h after treatment. Among pigs treated with RX821002 at 400 microg/kg BW, mean NEFA AUC was more than three times greater (P < .05) than that for saline-treated animals. Trial 3 tested whether NEFA responses to A2AA were due to direct effects on alpha2-receptors or involved beta-adrenoceptor mediation. Pigs were first treated i.v. with saline or propranolol (1 mg/kg BW). One hour later, pigs were treated i.v. with RX821002 (400 microg/kg BW) or the beta-adrenoceptor agonist cimaterol (25 microg/kg BW). Compared to values for pigs treated with saline at both injections, NEFA AUC among pigs treated with saline at the first injection and RX821002 at the second doubled (P > .05). Plasma NEFA AUC among pigs treated with saline then cimaterol rose nearly fourfold (P < .05) compared with saline-treated controls. Mean NEFA AUC among propranolol-treated pigs was similar to values for saline-treated pigs, suggesting beta-adrenoceptor involvement in the effect of A2AA on NEFA. In Trial 4, pigs were treated s.c. 10 times at 8-h intervals with saline, RX821002 (400 microg/[kg BW x injection]), cimaterol (20 microg/[kg BW x injection]) or recombinant porcine somatotropin (rpST; 1 mg/[pig-injection]). After the 10th treatment, only cimaterol increased NEFA AUC compared to saline-treated controls (P < .05). Mean PUN AUC was reduced by RX821002 and rpST compared to controls; PUN among rpST-treated pigs was lower than that among RX821002-treated pigs (P < .05). In summary, A2AA increase lipolysis in swine by potentiating lipolytic effects of endogenous catechol

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Adrenergic beta-Agonists; Animals; Area Under Curve; Benzofurans; Blood Urea Nitrogen; Fatty Acids, Nonesterified; Female; Idazoxan; Imidazoles; Isoproterenol; Male; Random Allocation; Swine

One component of the mechanism by which imidazoline compounds promote insulin secretion involves closure of ATP-sensitive K+ channels in the beta-cell plasma membrane. Recently, however, it has also been proposed that these compounds may exert important effects on more distal effector systems. In the present work, we have investigated the contribution played by protein kinases A and C to the insulin secretory responses of isolated rat islets of Langerhans treated with efaroxan and RX871024 (1-phenyl-2-(imidazolin-2-yl) benzimidazole). Removal of extracellular Ca2+ or blockade of voltage-sensitive Ca2+ channels prevented stimulation of insulin secretion by efaroxan, confirming a critical role for increased Ca2+ influx in the secretory response. By contrast, inhibition of protein kinases A or C failed to alter efaroxan-induced insulin secretion. RX871024 dose-dependently increased insulin secretion from cultured islets incubated with 20 mM glucose. This effect was unaffected by modulation of protein kinase C, but was significantly attenuated by a selective inhibitor of protein kinase A (Rp-cAMPs). Measurements of cAMP revealed that RX871024 increased the islet cAMP content by more than 3-fold; reaching values similar in magnitude to those elicited by 50 microM 3-isobutyl-1-methyl xanthine. The results reveal that neither protein kinase A nor protein kinase C is obligatory for stimulation of insulin secretion by imidazolines. However, they suggest that a rise in cAMP may contribute to the amplified secretory response observed when cultured islets are incubated with RX871024 in the presence of a stimulatory glucose concentration.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Calcium; Calcium Channel Blockers; Cyclic AMP; Down-Regulation; Enzyme Inhibitors; Imidazoles; In Vitro Techniques; Indoles; Insulin; Insulin Secretion; Islets of Langerhans; Male; Protein Kinase C; Protein Kinase Inhibitors; Rats; Rats, Wistar

The effects of two potent sigma receptor agonists (+)-3-PPP ((R)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine) and DTG (N,N'-di-(o-tolyl)guanidine) on the insulin secretory responses in rat islets of Langerhans were investigated. Both sigma receptor ligands were able to potentiate the insulin secretory response of islets incubated at 6 mM glucose, in a dose-dependent manner and were also able to reverse the effects of diazoxide on insulin release. When islets were treated with efaroxan, a well-characterised imidazoline insulin secretagogue, and either (+)-3-PPP or DTG together, there was an unexpected and profound absence of stimulation of insulin release as compared to when islets were incubated with each compound alone. Experiments performed with islets where there was desensitization of DTG/sigma receptor or efaroxan/imidazoline binding site mediated responses suggest that at least two distinct receptor systems appear to be involved. The complex interactions of these two classes of drug require further investigation.

Topics: Adrenergic alpha-Antagonists; Animals; Azides; Benzofurans; Dopamine Agonists; Dose-Response Relationship, Drug; Guanidines; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Ligands; Male; Piperidines; Rats; Rats, Wistar; Receptors, Drug; Receptors, sigma

The insulin secretagogue activity of certain imidazoline compounds is mediated by a binding site associated with ATP-sensitive K+ (K(ATP)) channels in the pancreatic beta-cell. We describe the effects of a series of structural modifications to efaroxan on its activity at this site. Substitution of amino-, nitro- or azide- groups onto the 5-position of the benzene ring of efaroxan did not significantly affect the functional interaction of the ligand with the islet imidazoline binding site. Modification of the imidazoline ring to an imidazole to generate 2-(2-ethyl-2,3-dihydrobenzo[b]furan-2-yl)-1H-imidazole (KU14R) resulted in loss of secretagogue activity. Indeed, this reagent appeared to act as an imidazoline antagonist since it blocked the secretory responses to imidazoline compounds and also inhibited the blockade of beta-cell K(ATP) channels by efaroxan in patch clamp experiments. Application of KU14R alone resulted in a modest reduction in K(ATP) channel opening, suggesting that it may display weak partial agonism, at least in patch-clamp experiments.

Topics: Adenosine Triphosphate; Adrenergic alpha-Antagonists; Animals; Benzofurans; Cells, Cultured; Female; Imidazoles; Imidazoline Receptors; Islets of Langerhans; Male; Patch-Clamp Techniques; Potassium Channel Blockers; Rats; Rats, Wistar; Receptors, Drug; Structure-Activity Relationship

We examined the role of I1-imidazoline receptors in the control of airway function, by testing the effects of systemic administration of the I1-imidazoline agonist moxonidine on reflex responses of tracheal smooth muscle (TSM) tone to either lung deflation or mechanical stimulation of intrapulmonary rapidly adapting receptors. Experiments were performed in either alpha-chloralose anaesthetized or decorticate, paralyzed and mechanically ventilated beagle dogs. Moxonidine (10-100 microg/kg) administered via three different routes (the femoral vein, muscular branch of superior thyroid artery, and vertebral artery) attenuated TSM responses to stimulation of airway sensory nerve fibers by two different ways, and caused a decrease in arterial pressure and heart rate. These effects were dose-dependent, and were significantly reversed by efaroxan (an I1-imidazoline and alpha2-adrenergic blocker) administered via the vertebral artery. Intravertebral efaroxan abolished the hemodynamic effects of moxonidine. Intravenous moxonidine (10-100 microg/kg) did not alter airway smooth muscle responses to electrical stimulation of the peripheral vagus nerve. In addition, in vitro moxonidine (1-100 microg/ml) had no effect on contractile responses to increasing doses of acetylcholine. These findings indicate that moxonidine may act at a central site to suppress reflex airway constriction, even when given into the systemic circulation. Given the presence of I1-imidazoline sites and alpha2-adrenergic receptors in brain regions participating in airway reflexes, these receptor classes may be involved in brainstem control of the cholinergic outflow to the airways.

Topics: Acetylcholine; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Cholinergic Fibers; Dogs; Electric Stimulation; Heart Rate; Imidazoles; Imidazoline Receptors; Mechanoreceptors; Muscle Contraction; Muscle, Smooth; Nerve Endings; Neurons, Afferent; Receptors, Drug; Trachea; Vagus Nerve; Vasodilator Agents

Rat PC 12 pheochromocytoma cells lack alpha2-adrenergic receptors but express plasma membrane I1-imidazoline receptors. In response to the I1-agonist moxonidine, diglycerides are generated via phosphatidylcholine-selective phospholipase C, and prostaglandin E2 is released. This report characterizes I-receptor-mediated release of arachidonic acid, the precursor to the prostaglandins. PC12 cells were incubated with [3H]arachidonic acid for 24 h and superfused with 0.01% bovine serum albumin in Krebs' physiological buffer at 1 ml/min. Calcium ionophore increased arachidonic acid release only marginally, implying that in PC12 cells arachidonic acid release is not driven by calcium. The I1-agonist moxonidine at concentrations between 10 nM and 1.0 microM rapidly elicited up to two-fold increases in [3H]arachidonic acid release. Guanabenz, a potent alpha2-agonist and I2-ligand, had no effect. The selective I1-antagonist efaroxan blocked the action of moxonidine. The phospholipase A2 inhibitor aristolochic acid had no effect, suggesting that arachidonic acid release may be through an indirect pathway, possibly involving diglycerides. Thus, I1-imidazoline receptors in PC12 cells are coupled to arachidonic acid release through an as yet unknown pathway.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Arachidonic Acid; Aristolochic Acids; Benzofurans; Enzyme Inhibitors; Guanabenz; Imidazoles; Imidazoline Receptors; PC12 Cells; Phenanthrenes; Phospholipases A; Phospholipases A2; Rats; Receptors, Drug

A deficient control of neuronal repair mechanisms by noradrenergic projections originating from the locus coeruleus may be a critical factor in the progression of neurodegenerative diseases. Blockade of presynaptic inhibitory alpha2-adrenergic autoreceptors can disinhibit this system, facilitating noradrenaline release. In order to test the neuroprotective potential of this approach in a model involving excitotoxicity, the effects of treatments with the alpha2-adreneceptor antagonists, (+)-efaroxan (0.63 mg/kg i.p., thrice daily for 7 days) or (+/-)-idazoxan (2.5 mg/kg i.p., thrice daily for 7 days), were evaluated in rats which received a quinolinic acid-induced lesion of the left striatum. Both drug treatments resulted in a reduced ipsiversive circling response to apomorphine and a reduced choline acetyltransferase deficit in the lesioned striatum. The mechanisms underlying this effect are not known for certain, but may include noradrenergic receptor modulation of glial cell function, growth factor synthesis and release, activity of glutamatergic corticostriatal afferents, and/or events initiated by NMDA receptor activation. These results suggest a therapeutic potential of alpha2-adrenoceptor antagonists in neurodegenerative disorders where excitotoxicity has been implicated.

Topics: Adrenergic alpha-Antagonists; Animals; Apomorphine; Behavior, Animal; Benzofurans; Choline O-Acetyltransferase; Corpus Striatum; Disease Models, Animal; Dopamine Agonists; Enzyme Activation; Huntington Disease; Idazoxan; Imidazoles; Male; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Quinolinic Acid; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-2

1. Imidazoline binding sites have been reported to be present in the locus coeruleus (LC). To investigate the role of these sites in the control of LC neuron activity, we studied the effect of imidazolines using in vivo and in vitro single-unit extracellular recording techniques. 2. In anaesthetized rats, local (27 pmoles) and systemic (1 mg kg(-1), i.v.) administrations of 2-(2-benzofuranyl)-2-imidazoline (2-BFI), a selective I-imidazoline receptor ligand, increased the firing rate of LC cells (maximal increase: 22+/-5%, P<0.001 and 16+/-7%, P<0.001 respectively). Chronic pretreatment with the irreversible monoamine oxidase inhibitor clorgyline (3 mg kg(-1), i.p., every 12 h for 14 days) abolished this effect. 3. In rat midpontine brain slices containing the LC, bath application (1 mM) of the imidazolines 2-BFI, 2-(4,5-dihydroimidaz-2-yl)-quinoline (BU224), idazoxan, efaroxan, phentolamine and (2-2-methoxy-1,4-benzodioxan-2-yl)-2-imidazoline (RX821002) reversibly stimulated LC cells. The maximal effect was approximately 90% except for RX821002 and efaroxan which induced smaller maximal effects (approximately 58% and approximately 35% respectively). Simultaneous application of idazoxan and 2BFI did not lead to additive effects. 4. Bath application of the alpha2-adrenoceptor antagonists, yohimbine (1 - 10 microM) and N-ethoxycarbonyl-1,2-dihydroquinoline (EEDQ) (10 microM), failed to modify LC activity. The irreversible blockade of alpha2-adrenoceptors with EEDQ (10 microM) did not alter the effect of idazoxan or that of efaroxan. Previous application of clorgyline (10 microM) did not modify the excitatory effect of 2-BFI or efaroxan. 5. Changes in the pH of the bathing solution (6.84-7.84) did not influence the effect caused by idazoxan. Bath application of 2-BFI (1 mM) reversed the inhibition induced by diazoxide (300 microM), an ATP-sensitive K+ channel opener, whereas application of glibenclamide (3 microM), an ATP-sensitive K+ channel blocker, partially blocked the effect of 2-BFI. 6. This study shows that imidazoline compounds stimulate the firing rate of LC neurons. This effect is not mediated by alpha2-adrenoceptors nor by I1 or I2-imidazoline receptors but involves a different subtype of imidazoline receptor. Our results indicate that this receptor is located extracellularly and modulates ATP-sensitive K+ channels.

Topics: Adenosine Triphosphate; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Diazoxide; Electrophysiology; Idazoxan; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Ligands; Locus Coeruleus; Male; Neurons; Potassium Channels; Rats; Receptors, Adrenergic, alpha-2; Receptors, Drug; Vasodilator Agents

Recent studies have suggested that a variety of ion channels possess a binding site for ligands such as phencyclidine (PCP), dizocilpine and certain sigma ligands and that some imidazoline compounds can also bind to this site. We have investigated whether interaction with this binding site could account for the ability of imidazolines to stimulate insulin secretion from rat islets. Neither PCP nor dizocilpine shared the insulin secretory activity of the imidazoline efaroxan in rat islets suggesting that they do not have similar actions in the pancreatic B-cell. Further, we were able to define a new antagonist, KU14R (2(2-ethyl 2,3-dihydro-2-benzofuranyl)-2-imidazole), which selectively blocks the insulin secretory response to imidazolines. The results suggest that imidazolines do not stimulate insulin secretion by causing physical blockade of the K(+)-ATP channel in pancreatic B-cells and show that their effects are not reproduced by PCP or sigma receptor ligands.

Topics: Adenosine Triphosphate; Animals; Benzofurans; Dizocilpine Maleate; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Ligands; Phencyclidine; Potassium Channels; Rats; Rats, Wistar; Receptors, Drug; Receptors, sigma

Although many of the behavioral effects of cocaine are widely believed to be mediated by blockade of dopamine transporters, recent studies suggest that norepinephrine (NE) may play a modulatory role. In this study, selective and nonselective beta-adrenergic receptor antagonists were administered alone or in combination with cocaine (2.5 mg/kg, i.p.) to rats trained to discriminate a low dose (2.5 mg/kg) from a high dose of cocaine (10 mg/kg) in a two-lever, FR10 drug discrimination procedure. The central beta 2/beta 1-adrenergic antagonists (-)-propranolol and tertatolol, and the beta 2-adrenergic antagonist, ICI 118,551, produced high-dose appropriate responding in a dose-related manner when administered (i.p.) in combination with the low training dose of cocaine. In contrast, neither the peripheral beta 2/beta 1-adrenergic antagonist, nadolol, nor the central beta 1-adrenergic antagonist, beta-xolol enhanced the behavioral effects of the low dose of cocaine in a manner comparable with that produced by compounds with central beta 2-adrenergic antagonist properties. Also in contrast to findings obtained using beta-adrenergic antagonists, neither the alpha 1-adrenergic agonist cirazoline, nor the alpha 2-adrenergic ligands (+/-)-efaroxan and UK-14304 enhanced the behavioral effects of the low dose of cocaine. Overall, these results suggest that central beta 2-adrenergic receptors may play a modulatory role in the discriminative stimulus effects of cocaine.

Topics: Adrenergic beta-Antagonists; Animals; Benzofurans; Betaxolol; Brimonidine Tartrate; Cocaine; Discrimination Learning; Discrimination, Psychological; Drug Synergism; Imidazoles; Male; Nadolol; Narcotics; Propanolamines; Propranolol; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha; Receptors, Adrenergic, beta-1; Receptors, Adrenergic, beta-2; Thiophenes

1. Imidazoline alpha 2-antagonist drugs such as efaroxan have been shown to increase the insulin secretory response to sulphonylureas from rat pancreatic B-cells. We have investigated whether this reflects binding to an islet imidazoline receptor or whether alpha 2-adrenoceptor antagonism is involved. 2. Administration of (+/-)-efaroxan or glibenclamide to Wistar rats was associated with a transient increase in plasma insulin. When both drugs were administered together, the resultant increase in insulin levels was much greater than that obtained with either drug alone. 3. Use of the resolved enantiomers of efaroxan revealed that the ability of the compound to enhance the insulin secretory response to glibenclamide resided only in the alpha 2-selective-(+)-enantiomer; the imidazoline receptor-selective-(-)-enantiomer was ineffective. 4. In vitro, (+)-efaroxan increased the insulin secretory response to glibenclamide in rat freshly isolated and cultured islets of Langerhans, whereas (-)-efaroxan was inactive. By contrast, (+)-efaroxan did not potentiate glucose-induced insulin secretion but (-)-efaroxan induced a marked increase in insulin secretion from islets incubated in the presence of 6 mM glucose. 5. Incubation of rat islets under conditions designed to minimize the extent of alpha 2-adrenoceptor signalling (by receptor blockade with phenoxybenzamine; receptor down-regulation or treatment with pertussis toxin) abolished the capacity of (+)- and (+/-)-efaroxan to enhance the insulin secretory response to glibenclamide. However, these manoeuvres did not alter the ability of (+/-)-efaroxan to potentiate glucose-induced insulin secretion. 6. The results indicate that the enantiomers of efaroxan exert differential effects on insulin secretion which may result from binding to effector sites having opposite stereoselectivity. Binding of (-)-efaroxan (presumably to imidazoline receptors) results in potentiation of glucose-induced insulin secretion, whereas interaction of (+)-efaroxan with a second site leads to selective enhancement of sulphonylurea-induced insulin release.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Drug Interactions; Glyburide; Hypoglycemic Agents; Idazoxan; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Male; Rats; Rats, Wistar; Stereoisomerism

The effects on arterial blood pressure and heart rate after an intracerebroventricular (i.c.v.) administration of clonidine were investigated using conscious normotensive cats. Injection of clonidine (5-10 microg; 5 microl; i.c.v.) elicited a decrease in mean arterial pressure (MAP) and heart rate (HR) in a dose-dependent manner. The highest dose of 10 microg of clonidine decreased MAP and HR by 39 +/- 3 mmHg and 74 +/- 5 b.p.m., respectively (n = 7). Pretreatment with yohimbine, the alpha2-adrenoceptor antagonist (8 microg; 5 microl; i.c.v.) blocked the cardiovascular responses to a subsequent i.c.v. injection of 10 microg clonidine (n = 7). Furthermore, preadministration of cimetidine (100 microg; 5 microl; i.c.v.), the H2 histamine receptor antagonist with imidazoline receptor activating properties, prevented the decreases in MAP and HR to a subsequent i.c.v. injection of 10 microg clonidine (n = 7). By contrast, pretreatment with the specific I1 imidazoline receptor blocker, efaroxan (100-500 microg; 5 microl; i.c.v.), failed to inhibit the cardiovascular effects of an i.c.v. administration of 10 microg clonidine (n = 7). These results suggest that the effects of centrally administered clonidine on MAP and HR are probably not mediated through activation of the I1 subtype of imidazoline receptors in conscious cats. However, the cardiovascular effects elicited by i.c.v. administration of clonidine appear to result from stimulation of central alpha2-adrenergic or the H2 histaminergic-like receptors.

Topics: Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Behavior, Animal; Benzofurans; Blood Pressure; Cardiovascular System; Cats; Cimetidine; Clonidine; Coloring Agents; Consciousness; Dose-Response Relationship, Drug; Evans Blue; Female; Heart Rate; Histamine H2 Antagonists; Imidazoles; Injections, Intraventricular; Yohimbine

In rabbit's aqueous humor, norepinephrine, epinephrine, dopamine and serotonin were detected simultaneously by a high performance liquid chromatography with electrochemical detection. Furthermore, the changes in catecholamine levels in aqueous humor were evaluated after topical application of moxonidine, an imidazoline1/alpha 2 receptor agonist, in the presence and absence of efaroxan. The level of norepinephrine in aqueous humor was reduced by moxonidine treatment. However, under the same set of conditions, there were no significant changes in the levels of dopamine, epinephrine or serotonin. Pretreatment with efaroxan antagonized moxonidine-induced suppression of norepinephrine levels. In other in vivo experiments, moxonidine caused a decrease in intraocular pressure which was antagonized by pretreatment with efaroxan. In the superior cervical ganglion preparation, norepinephrine release was increased 5-fold by the presence of a high K+ medium. The K(+)-evoked norepinephrine secretion was reduced by 55% by moxonidine. Pretreatment with efaroxan blocked the moxonidine-induced inhibition of norepinephrine release. It is concluded that inhibition of norepinephrine release from the superior cervical ganglion and suppression of aqueous norepinephrine levels contribute to the moxonidine-induced lowering of intraocular pressure. Moreover, the antagonism of moxonidine's in vivo and in vitro effects by efaroxan suggests the involvement of imidazoline1 receptors, but does not preclude activity on alpha 2 adrenoceptors.

Topics: Administration, Topical; Animals; Antihypertensive Agents; Aqueous Humor; Benzofurans; Dopamine; Female; Imidazoles; Intraocular Pressure; Male; Norepinephrine; Ocular Hypotension; Rabbits; Superior Cervical Ganglion

It is now well established that the imidazoline insulin secretagogue efaroxan mediates its effects by inducing closure of ATP-sensitive potassium channels in the pancreatic beta-cell, leading to membrane depolarisation, Ca2+ influx and increased insulin secretion. However, a recent study has shown that efaroxan may also act as a blocker of a second class of potassium channel (the Kmaxi channel) in red blood cells, raising the possibility that its effects in islets could be mediated by interactions with both types of channel. Since the antimycotic imidazole compound clotrimazole is a highly potent blocker of Kmaxi channels, we have studied the effects of this drug on insulin secretion. Clotrimazole stimulated insulin secretion from rat islets of Langerhans incubated in the presence of 6 mM glucose, in a dose-dependent manner. Experiments performed at different glucose concentrations showed that the actions of clotrimazole were most prominent at low glucose concentrations whereas it did not enhance secretion beyond the rate induced by 20 mM glucose. The insulinotropic action of clotrimazole was temperature dependent but was independent of extracellular calcium. Clotrimazole appeared to block ATP-sensitive potassium channels in islets since, like efaroxan and glibencamide, it was able to prevent the inhibitory effects of diazoxide on glucose-induced insulin secretion. However, neither the direct stimulatory effect of clotrimazole on insulin release nor the abilty of clotrimazole to reverse the inhibitory actions of diazoxide was sensitive to blockade by the imidazoline secretagogue antagonist KU14R. Overall, the results suggest that clotrimazole exerts an insulinotropic effect in pancreatic beta-cells that is distinct from the actions of imidazoline secretagogues such as efaroxan. Clotrimazole can increase insulin secretion at sub-maximal glucose concentrations by an action which appears to be independent of membrane ion channel events.

Topics: Animals; Benzofurans; Clotrimazole; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Male; Potassium Channel Blockers; Potassium Channels; Rats; Rats, Wistar

The I1-subtype of imidazoline binding sites has been characterized concerning binding specificity and tissue localization, and several physiological functions have been ascribed to it. However, the signaling pathways coupled to this putative receptor are not known. Pheochromocytoma PC12 cells express I1-imidazoline binding sites in plasma membrane and lack alpha2-adrenergic receptors, which recognize many I1-imidazoline ligands. In this cellular model, diacylglycerol (DAG), a second messenger, is generated in response to the putative I1-imidazoline agonist moxonidine. Using radioflux with [3H]myristate and direct measurements of DAG mass, we showed a rapid and transient peak of DAG in undifferentiated PC12 cells within the first 1 min of agonist exposure. In PC12 cells treated with nerve growth factor to initiate differentiation, DAG accumulation at 15 sec was facilitated, and the increase in DAG mass persisted throughout 10 min of agonist treatment. Efaroxan, a putative I1-antagonist, attenuated the effect of moxonidine on DAG accumulation in nerve growth factor-treated cells, as did D609, an inhibitor of phosphatidylcholine-selective phospholipase C. Phospholipase D did not seem to be involved in generation of DAG in response to I1-receptor activation, nor was there accumulation of phosphatidic acid. These findings suggest coupling of I1-imidazoline receptors to a phospholipase C to generate DAG as a second messenger, a process regulated by neuronal differentiation and possibly participating in the physiological responses to I1-imidazoline receptor activation.

Topics: Animals; Benzofurans; Diglycerides; Dose-Response Relationship, Drug; Imidazoles; Imidazoline Receptors; Nerve Growth Factors; PC12 Cells; Phospholipase D; Rats; Receptors, Drug; Type C Phospholipases

Four antagonists were examined for their ability to differentiate alpha 2A-from the orthologous alpha 2D-adrenoceptors. The antagonists were (2S,12bS)1',3'-dimethylspiro(1,3,4,5',6,6',7,12b-octah ydro-2H- benzo[b]furo[2,3-a]quinolizine)-2,4'-pyrimidin-2'-one (MK912), 2-[2-(methoxy-1,4-benzodioxanyl)imidazoline (RX 821002), efaroxan and benoxathian. The alpha 2-autoreceptors in rabbit brain cortex were chosen as alpha 2A-and the alpha 2-autoreceptors in guinea-pig brain cortex as alpha 2D-adrenoceptors. Slices of the brain cortex were preincubated with 3H-noradrenaline and then superfused and stimulated electrically by brief pulse trains (4 pulses, 100 Hz) that led to little, if any, alpha 2-autoinhibition. 5-Bromo-6-(2-imidazolin-2-ylamino)-quinoxaline (UK 14,304) was used as an alpha 2-adrenoceptor agonist. UK 14, 304 decreased the stimulation-evoked overflow of tritium. The antagonists shifted the concentration-inhibition curve of UK 14, 304 to the right in an apparently competitive manner. Dissociation constants of the antagonists were calculated from the shifts. MK 912, RX 821002 and efaroxan had markedly higher affinity for (guinea-pig) alpha 2D-adrenoceptors (pKd values 10.0, 9.7 and 9.1, respectively) than for (rabbit) alpha 2A-adrenoceptors (pKd 8.9, 8.2 and 7.6, respectively). Benoxathian had higher affinity for alpha 2A-(pKd 7.4) than for alpha 2D-adrenoceptors (pKd 6.9). Ratios calculated from the Kd values of the four compounds differentiated between alpha 2A and alpha 2D up to 100 fold. It is concluded that MK 912, RX 821002, efaroxan and benoxathian are antagonists with high power to differentiate alpha 2A-from alpha 2D-adrenoceptors.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Autoreceptors; Benzofurans; Brimonidine Tartrate; Cerebral Cortex; Dioxanes; Electric Stimulation; Female; Guinea Pigs; Idazoxan; Imidazoles; Isotope Labeling; Male; Norepinephrine; Oxathiins; Quinolizines; Quinoxalines; Rabbits; Receptors, Adrenergic, alpha-2; Structure-Activity Relationship; Tritium

To determine the involvement of central imidazoline receptors in the cardiovascular actions of the antihypertensive agents moxonidine, rilmenidine and clonidine administered systemically.. We determined the relative potency of these drugs with respect to their effects on mean arterial pressure and heart rate by performing cumulative intravenous dose-response relationship studies in six conscious rabbits. In another eight rabbits with implanted fourth-ventricular catheters, we investigated the central effects of three cumulative doses of an l1-imidazoline/ alpha 2-adrenoceptor antagonist, efaroxan, and of an alpha 2-adrenoceptor antagonist, 2-methoxyidazoxan (2-Ml), on the hypotension and bradycardia elicited by a single intravenous dose of the above agents. The doses of antagonists were matched for an equal reversal of the hypotension induced by fourth-ventricular alpha-methyldopa (an alpha 2-adrenoceptor agonist) and hence for similar alpha 2-adrenoceptor blockade.. Moxonidine and rilmenidine were sevenfold and eightfold less potent, respectively, than was clonidine in eliciting hypotension. By comparison, moxonidine and clonidine were more potent than was rilmenidine in producing bradycardia. Efaroxan and 2-Ml reversed the hypotension and bradycardia induced by a single dose of all three agents dose-dependently. However, efaroxan was more effective than was 2-Ml at reversing the effects of rilmenidine and moxonidine. Complete reversal of their hypotensive effect was observed with the highest dose of efaroxan but the highest dose of 2-Ml reversed approximately 50% of that effect. In contrast, the two antagonists were equally effective at reversing the responses to clonidine.. These results suggest that the hypotension and bradycardia induced by intravenous administration of moxonidine and rilmenidine were mediated mainly by actions on central imidazoline receptors whereas clonidine appears to act predominantly on central alpha 2-adrenoceptors.

Topics: Adrenergic alpha-2 Receptor Antagonists; Animals; Antihypertensive Agents; Benzofurans; Blood Pressure; Cardiac Catheterization; Clonidine; Dose-Response Relationship, Drug; Female; Heart Rate; Idazoxan; Imidazoles; Imidazoline Receptors; Injections, Intravenous; Male; Oxazoles; Rabbits; Receptors, Drug; Rilmenidine; Time Factors

In order to determine whether agmatine could be a putative endogenous ligand for imidazoline receptors mediating insulin secretion and vasoconstriction, we compared its effects with those of the imidazoline, efaroxan. Agmatine exhibited a much lower potency and efficacy than efaroxan on insulin secretion from rat pancreas perfused with 8.3 mM glucose. On the other hand, in contrast to efaroxan (100 microM), agmatine (3 mM) did not increase arginine-induced insulin release. In addition, agmatine failed to reproduce the vasoconstrictor effect of efaroxan on pancreatic vessels. These results show that agmatine does not behave like efaroxan, an agonist for the imidazoline receptors mediating insulin secretion or vasoconstriction in the pancreas.

Topics: Agmatine; Animals; Benzofurans; Imidazoles; Imidazoline Receptors; Insulin; Insulin Secretion; Islets of Langerhans; Ligands; Male; Protein Binding; Rats; Rats, Wistar; Receptors, Drug

To examine ocular actions by rilmenidine, an imidazoline1 and alpha 2 adrenoceptor agonist.. Intraocular pressure was measured in normal and sympathetically denervated rabbits by pneumatonometry. Electrically stimulated 3H-norepinephrine release from sympathetic nerves was determined in isolated, perfused rabbit iris-ciliary bodies. cAMP levels were evaluated in rabbit iris-ciliary bodies by radioimmunoassay. Ca2+ concentrations were measured in rabbit transformed nonpigmented ciliary epithelial cells by fluorescence ratio microscopy.. Topical, unilateral administration of rilmenidine produced hypotensive responses in normal rabbits which were antagonized by either bilaterally administered efaroxan, an imidazoline receptor antagonist or rauwolscine, an alpha 2 receptor antagonist. Sympathectomy also eliminated the ocular hypotensive response. Rilmenidine (0.001, 0.01, 0.1, 1 microM) caused 5 +/- 1%, 18 +/- 5%, 35 +/- 10%, and 48 +/- 9% inhibition, respectively, of 3H-norepinephrine overflow whereas 10 microM efaroxan or rauwolscine caused enhancement of norepinephrine release by 102 +/- 23% or 86 +/- 25%, respectively. Furthermore, pretreatment with efaroxan or rauwolscine partially antagonized the inhibition of norepinephrine release induced by rilmenidine. In other experiments, rilmenidine (1 microM) inhibited isoproterenol-stimulated cAMP accumulation in rabbit iris-ciliary bodies by 43 +/- 9% which was antagonized by 10 microM efaroxan or rauwolscine. Rilmenidine induced large increases in [Ca2+]i in rabbit nonpigmented ciliary epithelial cells which were effectively antagonized by efaroxan or rauwolscine.. These in vivo and in vitro data suggest that the ocular hypotensive activity induced by rilmenidine is due, in part, to suppression of sympathetic neuroeffector function in the rabbit ciliary body and that alpha 2 adrenergic receptors and/or imidazoline1 receptors are involved.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antihypertensive Agents; Benzofurans; Calcium; Ciliary Body; Cyclic AMP; Female; Imidazoles; Imidazoline Receptors; Intraocular Pressure; Iris; Male; Norepinephrine; Ocular Hypotension; Oxazoles; Rabbits; Receptors, Adrenergic, alpha-2; Receptors, Drug; Rilmenidine; Sympathectomy; Sympathetic Nervous System; Yohimbine

1. Microfluorimetry techniques with fura-2 were used to characterize the effects of efaroxan (200 microM), phenotolamine (200-500 microM) and idazoxan (200-500 microM) on the intracellular free Ca2+ concentration ([Ca2+]i) in mouse isolated islets of Langerhans. 2. The imidazoline receptor agonists efaroxan and phentolamine consistently elevated cytosolic Ca2+ by mechanisms that were dependent upon Ca2+ influx across the plasma membrane; there was no rise in [Ca2+]i when Ca2+ was removed from outside of the islets and diazoxide (100-250 microM) attenuated the responses. 3. Modulation of cytosolic [Ca2+]i by efaroxan and phentolamine was augmented by glucose (5-10 mM) which both potentiated the magnitude of the response and reduced the onset time of imidazoline-induced rises in [Ca2+]i. 4. Efaroxan- and phentolamine-evoked increases in [Ca2+]i were unaffected by overnight pretreatment of islets with the imidazolines. Idazoxan failed to increase [Ca2+]i under any experimental condition tested. 5. The putative endogenous ligand of imidazoline receptors, agmatine (1 microM-1 mM), blocked KATP channels in isolated patches of beta-cell membrane, but effects upon [Ca2+]i could not be further investigated since agmatine disrupts fura-2 fluorescence. 6. In conclusion, the present study shows that imidazolines will evoke rises in [Ca2+]i in intact islets, and this provides an explanation to account for the previously described effects of imidazolines on KATP channels, the cell membrane potential and insulin secretion in pancreatic beta-cells.

Topics: Adrenergic alpha-Antagonists; Agmatine; Animals; Benzofurans; Calcium; Cytosol; Down-Regulation; Glucose; Idazoxan; Imidazoles; Imidazoline Receptors; Insulin; Islets of Langerhans; Mice; Phentolamine; Receptors, Drug; Signal Transduction

In the present report, we investigated the potential involvement of imidazoline I1 and I2 binding sites in the inhibition of the Ca(2+)-activated K+ channel (Gardos channel) by clotrimazole in human red cells. Ca(2+)-activated 86Rb influx was inhibited by clotrimazole and efaroxan but not by the imidazoline binding site ligands clonidine, moxonidine, cirazoline and idazoxan (100 microM). Binding studies with [3H]idazoxan and [3H]p-aminoclonidine did not reveal the expression of I1 and I2 binding sites in erythrocytes. These data indicate that the effects of clotrimazole and efaroxan on the erythrocyte Ca(2+)-activated K+ channel may be mediated by a 'non-I1/non-I2' binding site.

Topics: Adrenergic alpha-Antagonists; Benzofurans; Binding Sites; Calcium; Clotrimazole; Dose-Response Relationship, Drug; Erythrocytes; Humans; Imidazoles; Potassium Channels

Combinations of drugs are frequently used to achieve effective analgesia while minimizing side effects. Although the analgesic effects of morphine and clonidine seem to be synergistic, few studies have investigated other effects. Their role in inhibiting gastrointestinal transit was evaluated using different methods of analysis.. Percentage inhibition of transit induced by morphine, clonidine, or their combination was measured in mice that had been given an intragastric charcoal meal. Dose-response curves were obtained for each drug individually; for morphine:clonidine at 1:3, 1:1, and 1:0.33 ratios; and for morphine in the presence of 0.0138 mg/kg clonidine. The interaction was evaluated by isobolograms, combination indexes, and fixed-dose analysis.. Each drug and their combinations inhibited transit in a dose-related manner. Combinations of morphine and clonidine produced interaction that depended on the ratio and level of response. The interaction analyzed by isobolograms and combination indexes showed that combinations in 1:1 and 1:3 proportions were synergistic at the median effective doses or less and were antagonistic at larger doses. Fixed-dose analysis using different ratios showed similar results. The effects of the combination (median effective dose at 1:1 ratio) were antagonized by efaroxan but not by naloxone, suggesting a predominant role of alpha-2-mediated effects.. Investigations into drug interactions should include several levels of response and combinations at different ratios. Isobolographic analysis permits the statistical evaluation of results without making assumptions about mechanisms of action of the drugs or their interactions. In this study, the combination of morphine and clonidine should produce synergy, antagonism, or no interaction depending on the relative doses and the level of effect.

Topics: Adrenergic alpha-Antagonists; Analysis of Variance; Animals; Benzofurans; Clonidine; Constipation; Dose-Response Relationship, Drug; Drug Interactions; Gastrointestinal Transit; Imidazoles; Male; Mice; Morphine; Naloxone; Narcotic Antagonists; Narcotics; Sympatholytics

Noradrenergic modulation of the cortical cholinergic system in vivo was studied by examining the effect of the selective alpha 2-adrenoceptor antagonist (+)-efaroxan on cortical acetylcholine outflow in the conscious rat, using the microdialysis technique. (+)-Efaroxan produced a dose-dependent increase in acetylcholine outflow (up to 300% at 0.63 mg/kg) which persisted for up to 3 h and which was stereospecific. The results demonstrate that rat cortical acetylcholine release can be augmented by (+)-efaroxan and that alpha 2-adrenoceptors may be involved. (+)-Efaroxan may have therapeutic potential in disorders in which cortical acetylcholine release is deficient.

Topics: Acetylcholine; Adrenergic alpha-Antagonists; Animals; Benzofurans; Cerebral Cortex; Dose-Response Relationship, Drug; Imidazoles; Male; Microdialysis; Rats; Rats, Sprague-Dawley; Stereoisomerism

I1-imidazoline receptor activation by moxonidine has potent antigastric secretory and gastroprotective effects in rats. We therefore tested whether an imidazoline receptor antagonist, efaroxan, would influence gastric secretion and block the antisecretory and antiulcer effects of moxonidine. When given intracerebroventricularly (i.c.v.), moxonidine inhibited basal acid output in conscious rats to a maximum of 38%. Moxonidine given i.p. also significantly increased gastric adherent mucus levels in rats subjected to cold-restraint stress. Efaroxan alone given i.c.v., did not influence gastric secretion nor did it affect moxonidine's ability to decrease gastric secretion. Similarly, peripherally administered efaroxan did not block the antisecretory effect of moxonidine given i.c.v. However, when both compounds were given i.p., efaroxan pretreatment at all but the lowest doses significantly blocked the antigastric secretory effect of moxonidine. Efaroxan alone (i.p.) did not influence stress-induced gastric mucosal injury or adherent mucus levels. However, pretreatment of rats with efaroxan i.p. significantly blocked the mucus-preserving effect of i.p. moxonidine. These results demonstrate that central (i.c.v.) or peripheral (i.p.) administration of the I1-imidazoline receptor agonist moxonidine is associated with gastroprotection. The ability of i.p. efaroxan to block the effects of i.p. moxonidine but not i.c.v. moxonidine indicates that imidazoline receptors located centrally and peripherally may represent two unique sites associated with gastroprotection.

Topics: Animals; Benzofurans; Dose-Response Relationship, Drug; Gastric Acid; Gastric Mucosa; Imidazoles; Imidazoline Receptors; Male; Rats; Rats, Sprague-Dawley; Receptors, Drug

These experiments sought to: (1) determine if alpha 2/I1 agonists that are topically active on the eye have similar effects on intraocular pressure when applied to the CNS and (2) ascertain whether these agents lower IOP, in part, via central alpha 2 receptors and/or imidazoline (I1) receptors. New Zealand White rabbits were fitted with chronic indwelling stainless-steel guide cannulas in several brain regions including the lateral ventricle, third ventricle (3V), or medullary intermediate reticular zone. Animals were allowed 5 days' recovery time prior to experiments measuring the effects of drugs on IOP via applanation pneumatonometry. Some animals were also pretreated with 400 micrograms of 6-hydroxydopamine injected into the lateral ventricle to determine the site of action of these alpha 2/I1 agonists. In initial experiments involving microinjection into the lateral ventricle, UK-14,304-18 evoked ocular hypotension that was inhibited by the alpha 2-antagonist rauwolscine but not by the I1-receptor antagonist efaroxan. Conversely, moxonidine and oxymetazoline were preferentially inhibited by efaroxan rather than by rauwolscine. Subsequently, experiments have shown that moxonidine and oxymetazoline, but not UK-14,304-18 will lower intraocular pressure when microinjected into the medullary intermediate reticular zone region and that efaroxan, but not rauwolscine, will inhibit ocular hypotension induced by moxonidine and oxymetazoline. Pretreatment with 6-hydroxydopamine (48 hours) completely eliminated the ocular hypotension induced by moxonidine. These preliminary data demonstrate that alpha 2- and I1-receptors in the brain mediate ocular hypotension induced by UK-14,304-18 and moxonidine/oxymetazoline, respectively. Moreover, the medullary intermediate reticular zone area of the brain stem is the probable presynaptic site mediating ocular hypotension induced by moxonidine and oxymetazoline.

Topics: Administration, Topical; Animals; Benzofurans; Cornea; Imidazoles; Imidazoline Receptors; Injections, Intraventricular; Intraocular Pressure; Male; Medulla Oblongata; Ocular Hypertension; Rabbits; Receptors, Drug; Sympathectomy, Chemical; Yohimbine

The relative potencies of imidazoline compounds to induce insulin secretion and vascular resistance were compared in the isolated perfused rat pancreas. On insulin secretion, only the two imidazolines, antazoline and efaroxan, induced a concentration-dependent response, antazoline being 10 times more potent than efaroxan. In contrast, idazoxan, a blocker of imidazoline I1 sites, at concentrations up to 30 microM, antagonized the insulin response to 10 microM efaroxan (IC50 approximately equal to 14 +/- 2 microM) without affecting that to 3 microM tolbutamide. On pancreatic vessels, not only antazoline and efaroxan but also idazoxan induced a concentration-dependent vasoconstriction; the rank order of agonist potency was antazoline > efaroxan > idazoxan. In addition, cimetidine, an imidazole known to bind imidazoline I1 sites, ineffective per se, partially reversed the insulin stimulatory effect of efaroxan without affecting its vasoconstrictor effect. This study demonstrates that the insulin secretory and vasoconstrictor actions of imidazolines involve different imidazoline sites in rat pancreas. The results provide evidence for an I1 type mediating insulin secretion on B cells and an I2 type mediating vasoconstriction in vessels.

Topics: Adrenergic alpha-Antagonists; Analysis of Variance; Animals; Antazoline; Benzofurans; Binding, Competitive; Cimetidine; Dioxanes; Dose-Response Relationship, Drug; Idazoxan; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Male; Rats; Rats, Wistar; Receptors, Drug; Reference Standards; Tolbutamide; Vascular Resistance

In order to study the pharmacology of the putative imidazoline receptor involved in stimulation of insulin secretion, the potent and selective imidazoline I1 receptor agonist, moxonidine, was employed. Surprisingly, this agent caused a rapid and complete inhibition of glucose-induced insulin secretion in isolated rat islets of Langerhans. This response was reversible upon removal of the compound but was only partially attenuated under conditions of complete alpha 2 blockade, suggesting that it did not derive entirely from the weak alpha 2-adrenoceptor agonist activity of moxonidine. Furthermore, the response could not be attributed to activation of imidazoline I1 receptors since it was not reproduced by a second potent imidazoline I1 receptor agonist, cimetidine, and could not be alleviated by the imidazoline I1 receptor antagonist efaroxan. The results confirm that the imidazoline receptor involved in control of insulin secretion differs from the I1 subclass and suggest that moxonidine inhibits insulin secretion by a mechanism unrelated to imidazoline I1 receptor agonism.

Topics: Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Antagonists; Animals; Benzofurans; Cimetidine; Depression, Chemical; Female; Glucose; Histamine H2 Antagonists; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Male; Rats; Rats, Wistar; Receptors, Drug

The rostral ventrolateral medulla (RVLM) is the primary region maintaining vasomotor tone, and a site of action for central antihypertensive agents. In vitro [125I]p-iodoclonidine binding studies showed that moxonidine was selective for I1-imidazoline over alpha 2-adrenergic receptors in the RVLM. We identified efaroxan and SK&F 86466 as selective I1- and alpha 2-antagonists, respectively. We tested moxonidine's action within the RVLM of spontaneously hypertensive rats (SHRs) on I1-imidazoline or alpha 2-adrenergic receptors, and determined whether the RVLM mediates the action of systemic moxonidine. SHRs were anesthetized, paralyzed, and ventilated and the RVLM was localized by testing for a pressor response to 2 nmol glutamate. To test whether I1 or alpha 2 mediates hypotensive effects of moxonidine, the I1/alpha 2 antagonist efaroxan (4 nmol) or the alpha 2-blocker SK&F 86466 (10 nmol) was administered 15 min before 4 nmol moxonidine. Efaroxan elevated blood pressure and abolished the action of moxonidine, whereas alpha 2-blockade with SK&F 86466 slightly lowered blood pressure and only partially attenuated moxonidine's effect. The depressor effect of intravenous moxonidine (40 micrograms/kg) was reversed within 10 min by microinjection of 10 nmol efaroxan into the RVLM. Prior bilateral microinjections of efaroxan (10 nmol in 80 nl/site) into the RVLM prevented the hypotensive action of moxonidine given i.v. (40 micrograms/kg). Pharmacokinetic studies showed that at the peak vasodepressor response (8 min post-injection), [3H]moxonidine spread less than 1 mm from the injection site. Moxonidine is a centrally acting antihypertensive with a selective action on I1-imidazoline receptors in RVLM.

Topics: Adrenergic alpha-Antagonists; Affinity Labels; Animals; Antihypertensive Agents; Benzazepines; Benzofurans; Binding, Competitive; Blood Gas Analysis; Blood Pressure; Blood Pressure Determination; Cattle; Clonidine; Disease Models, Animal; Heart Rate; Hypertension; Imidazoles; Imidazoline Receptors; In Vitro Techniques; Medulla Oblongata; Microinjections; Radioligand Assay; Rats; Rats, Inbred SHR; Receptors, Drug

The effect on insulin release of efaroxan, an alpha 2-adrenoceptor antagonist and a highly potent drug at imidazoline I1 receptors, and the effects of seven other imidazoline compounds selective for the imidazoline I1 or I2 receptors, were studied in the rat insulinoma cell line RIN-5AH. The cells released insulin in response to glucose (0.3-10 mM), and efaroxan (100 microM) potentiated glucose-induced insulin release. (-)-Adrenaline completely displaced the binding of [125I]p-iodoclonidine to membranes of RIN-5AH cells, indicating that these cells do not express imidazoline I1 receptors. Cirazoline and idazoxan (100 microM), both highly potent drugs at imidazoline I2 receptors, and the guanidines guanoxan and amiloride (200 microM), also promoted insulin release from RIN-5AH cells. Irreversible blockade of imidazoline I2 receptors with 10 microM clorgyline did not prevent the stimulatory effects of cirazoline or idazoxan; however, these compounds completely reversed the inhibition by diazoxide (250 microM), an opener of ATP-dependent K+ channels (K+ATP channels), of glucose-induced insulin release. These data indicate that the imidazoline/guanidine compounds promote insulin release from RIN-5AH cells, by interacting with a novel binding site related to K+ATP channels that does not represent any of the known imidazoline I1 or I2 receptors.

Topics: Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Adrenergic beta-Agonists; Affinity Labels; Animals; Benzofurans; Binding, Competitive; Clonidine; Clorgyline; Diazoxide; Dioxanes; Guanidines; Idazoxan; Imidazoles; Imidazoline Receptors; Insulin; Insulinoma; Ligands; Pancreatic Neoplasms; Potassium Channels; Rats; Receptors, Drug; Regression Analysis; Software; Tumor Cells, Cultured

The effects of imidazolines and derivatives were studied on insulin secretion and vascular resistance in the isolated perfused rat pancreas. On insulin secretion, two imidazoline alpha 2-adrenoceptor antagonists, efaroxan (1-100 microM) and RX821002 (10 microM), had a stimulating response; however, idazoxan, like the non-imidazoline alpha 2-adrenoceptor antagonist yohimbine, was ineffective at 10 microM. The oxazoline rilmenidine with alpha 2-adrenergic activity at 10 microM), an imidazoline devoid of alpha 2-adrenergic activity, also had an insulin-releasing effect. On pancreatic vessels, all imidazolines tested (efaroxan, RX821002, antazoline and idazoxan), in contrast to yohimbine, induced vasoconstriction. Rilmenidine did not have a vasoconstrictor effect after blockade of alpha 2-adrenoceptors. Furthermore, the efaroxan-induced insulin release or vasoconstriction was not affected by the blockade of alpha 2- and alpha 1-adrenoceptors. This study shows that imidazolines and derivatives are able to stimulate insulin release and induce vasoconstriction in the rat pancreas. These effects cannot be ascribed to an interaction with alpha-adrenoceptors but may involve different types of imidazoline sites.

Topics: Adrenergic alpha-1 Receptor Antagonists; Adrenergic alpha-2 Receptor Antagonists; Adrenergic alpha-Agonists; Adrenergic alpha-Antagonists; Animals; Antazoline; Benzofurans; Imidazoles; In Vitro Techniques; Insulin; Insulin Secretion; Male; Pancreas; Rats; Rats, Wistar; Receptors, Adrenergic, alpha-1; Receptors, Adrenergic, alpha-2; Regional Blood Flow; Vascular Resistance; Vasoconstriction

1. The nature of the binding site mediating the insulin secretagogue activity of certain imidazoline compounds remains unclear and the pharmacology of the I1- and I2-imidazoline sites, described in many tissues, does not correlate with the observed responses to imidazolines in islets. In the present paper, we describe further results which support the concept that the islet imidazoline site may represent a novel subtype of imidazoline receptor. 2. Culture of rat isolated islets in the presence of imidazoline secretagogues (either efaroxan or phentolamine) resulted in loss of responsiveness on subsequent re-exposure to these agents. However, culture of islets with either idazoxan or UK14,304 (imidazoline ligands that do not stimulate insulin secretion) did not lead to any loss of response when the islets were subsequently exposed to efaroxan. By contrast, islets cultured with UK14,304 (a potent alpha 2-adrenoceptor agonist), displayed loss of sensitivity to noradrenaline, consistent with down-regulation of alpha 2-adrenoceptors. 3. In order to characterize the imidazoline site further, radioligand binding studies were performed in membranes from RINm5F insulinoma cells using [3H]-RX821002, an imidazoline insulin secretagogue that does not interact significantly with imidazoline sites in other tissues. [3H]-RX821002 labelled alpha 2-adrenoceptors with high affinity (2.01 +/- 0.7 nM) but also labelled a second, non-adrenoceptor site with much lower affinity. 4. Under conditions of alpha 2-adrenoceptor blockade (in the presence of adrenaline), efaroxan displaced [3H]-RX821002 binding to the low affinity site, in a dose-dependent manner. Competition studies employing additional imidazoline compounds of varying secretagogue activity revealed that the pharmacological profile of the low affinity site correlates well with that observed in secretion experiments.5. The results obtained from the down-regulation experiments with isolated islets and from the radioligand binding studies suggest that the low affinity [3H]-RX821002 binding site may represent the functional receptor responsible for the secretagogue activity of imidazoline compounds in the endocrine pancreas and that it has a pharmacological profile distinct from those of I,- and 12-sites.

Topics: Animals; Benzofurans; Binding Sites; Dioxanes; Down-Regulation; Idazoxan; Imidazoles; Imidazoline Receptors; Insulin; Insulin Secretion; Male; Rats; Rats, Wistar; Receptors, Drug

1) Moxonidine (MOX), injected icvt into the anterior lateral ventricle of NZW rabbits, induced bilateral, ocular hypotension (> 7.0 mmHg) that persisted for two hrs. 2) Oxymetazoline (OXY), injected icvt into the anterior lateral ventricle of NZW rabbits, induced bilateral ocular hypotension (> 7.0 mmHg) that peaked at two hrs. 3) Unilateral topical application of OXY induced maximal, bilateral ocular hypotension (> 12 mmHg), at 3 hrs, in both the contralateral and ipsilateral eyes, that persisted more than 12 hrs. 4) The putative imidazoline (I1) antagonist, efaroxan, injected icvt into the anterior lateral ventricle, inhibited significantly the ocular hypotension produced by icvt MOX, icvt OXY, and unilateral topical OXY. 5) Imidazoline (I1) receptors, located in the CNS, play a role in MOX- and OXY-induced ocular hypotension, as suggested by the ability of the putative imidazoline (I1) receptor antagonist efaroxan, to inhibit icvt MOX-, icvt OXY- and topical OXY-induced ocular hypotension.

Topics: Administration, Topical; Animals; Aqueous Humor; Benzofurans; Brain; Catheters, Indwelling; Imidazoles; Imidazoline Receptors; Injections, Intraventricular; Intraocular Pressure; Male; Ocular Hypotension; Oxymetazoline; Pupil; Rabbits; Receptors, Drug

1. The imidazoline alpha 2-adrenoceptor antagonist, efaroxan, stimulates insulin secretion from rat isolated islets and antagonizes the ability of diazoxide to inhibit glucose-induced insulin secretion. These effects result from closure of ATP-sensitive potassium channels although the mechanisms involved have not been elucidated. 2. In the present work, we have examined the effects of a close structural analogue of efaroxan, RX801080, in rat isolated islets of Langerhans. RX801080 was found to be ineffective as a stimulator of insulin secretion and did not prevent the inhibition of insulin secretion mediated by diazoxide. 3. RX801080 acted as an antagonist of the actions of several imidazolines (efaroxan, phentolamine and midaglizole) in rat islets. It dose-dependently inhibited the ability of efaroxan to antagonize the effects of diazoxide in islets and also completely inhibited the direct stimulation of insulin secretion mediated by efaroxan. 4. RX801080 also antagonized the effects of the non-imidazoline, ATP-sensitive potassium channel blocker, glibenclamide, in rat islets. It inhibited both the capacity of glibenclamide to stimulate insulin secretion and the ability of glibenclamide to overcome the inhibitory effects of diazoxide in rat islets. 5. Antagonism of glibenclamide responses by RX801080 was not due to inhibition of binding of the sulphonylurea to its receptor on the pancreatic beta-cell. 6. The results suggest that imidazoline compounds and sulphonylureas interact with distinct binding sites on islet cells, but that these sites can interact functionally to control islet cell ATP-sensitive potassium channel activity and insulin secretion.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Binding, Competitive; Cell Membrane; Drug Interactions; Glyburide; Imidazoles; Imidazoline Receptors; Insulin; Insulin Secretion; Islets of Langerhans; Kinetics; Male; Rats; Rats, Wistar; Receptors, Drug; Stimulation, Chemical

We have studied the stereospecificity of three responses mediated by the alpha 2-antagonist efaroxan in rat islets of Langerhans. alpha 2-Adrenergic inhibition of insulin secretion was relieved most effectively by the (+) enantiomer. In contrast, direct stimulation of insulin secretion and antagonism of the inhibitory effects of diazoxide were both preferentially mediated by the (-) enantiomer. Culture of islets in the presence of efaroxan resulted in loss of responsiveness to subsequent re-addition of the drug. On the basis of these results we propose the existence, in islets, of a novel 'non-adrenergic' binding site at which efaroxan acts as an agonist.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Culture Techniques; Dose-Response Relationship, Drug; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Male; Rats; Rats, Wistar; Stereoisomerism

Topics: Animals; Benzofurans; Diazoxide; Dioxanes; Idazoxan; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Rats

The effects of efaroxan (RX 821037A; 2-[2-(2-ethyl-2,3-dihydrobenzofuranyl)]-2-imidazoline HCl) at alpha 1- and alpha 2-adrenoceptors were investigated in isolated tissues, pithed rats and conscious rats. In isolated tissues, efaroxan competitively antagonised the inhibitory effects of p-aminoclonidine in the electrically stimulated (0.1 Hz) rat vas deferens, (pA2 = 8.89) and the contractile effects of phenylephrine on the rat anococcygeus muscle (pA2 = 6.03). Efaroxan had a selectivity ratio (alpha 2/alpha 1) of 724 compared to a value of 182 for idazoxan. In pithed rats, the i.v. doses of efaroxan (mumol/kg) producing 2-fold shifts in dose-response curves for UK-14,304 at prejunctional cardiac alpha 2-adrenoceptors and postjunctional vascular alpha 2-adrenoceptors, and for cirazoline at postjunctional vascular alpha 1-adrenoceptors, were 0.05, 0.13 and 2.96, respectively. In conscious fasted rats, prazosin (5 mg/kg p.o.) increased resting glucose levels and exacerbated the hyperglycaemic effects of UK-14,304 and adrenaline. In contrast, efaroxan (1-5 mg/kg p.o.) had little effect on resting plasma glucose but markedly antagonised the hyperglycaemic actions of UK-14,304 and adrenaline. Efaroxan increased resting plasma insulin levels and markedly potentiated the rise in insulin levels produced by adrenaline; this latter effect was prevented by the co-administration of propranolol. These results demonstrate that efaroxan is a potent and selective alpha 2-adrenoceptor antagonist and provide further support for the involvement of alpha 2-adrenoceptors in glucose homeostasis.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Blood Glucose; Brimonidine Tartrate; Clonidine; Decerebrate State; Dioxanes; Dose-Response Relationship, Drug; Epinephrine; Idazoxan; Imidazoles; Insulin; Male; Muscles; Prazosin; Quinoxalines; Rats; Rats, Inbred Strains; Vas Deferens

The effect of efaroxan (1 and 5 mg/kg p.o.; a selective alpha 2-adrenoceptor antagonist) was compared to glibenclamide (1 and 5 mg/kg p.o.; a standard sulphonylurea) on basal plasma glucose levels of fed and fasted rats. In addition, the effect of efaroxan (5 mg/kg p.o.) and glibenclamide (2 or 5 mg/kg p.o.), alone and in combination, on the hyperglycaemia and hyperinsulinaemia induced by glucose challenges, were investigated. An intra-arterial (250 mg/kg i.a.) and a subcutaneous (1 g/kg s.c.) glucose challenge were used to stimulate the fast and slow release phases of insulin secretion. Efaroxan increased plasma insulin levels in both conscious fed and fasted rats without greatly affecting plasma glucose levels. Glibenclamide also elevated insulin levels, but was associated with marked hypoglycaemia. Efaroxan and glibenclamide potentiated the slow and fast release of insulin secretion, but glibenclamide had a tendency to produce hypoglycaemia in these test situations, a property not shared by efaroxan. A combination of efaroxan and glibenclamide produced a greater elevation in the slow and fast insulin release phases than either compound alone, but did not enhance the hypoglycaemia seen with glibenclamide alone. These results provide further evidence that pancreatic alpha 2-adrenoceptors are involved in the regulation of insulin secretion.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Blood Glucose; Glucose; Glyburide; Imidazoles; Insulin; Male; Rats; Rats, Inbred Strains

The selective alpha 2-antagonist DG-5128 provoked a dose-dependent stimulation of insulin release from isolated rat islets. DG-5128 was only weakly effective as an antagonist of noradrenaline-induced inhibition of insulin secretion but, surprisingly, was able to reverse the suppression of secretion and increase in 86Rb efflux from preloaded islets, mediated by diazoxide. These effects were not reproduced with more effective alpha-antagonists, suggesting that stimulation of insulin secretion by DG-5128 is independent of alpha-receptor blockade.

Topics: Adrenergic alpha-Antagonists; Animals; Benzofurans; Diazoxide; Dioxanes; Dose-Response Relationship, Drug; Hypoglycemic Agents; Idazoxan; Imidazoles; In Vitro Techniques; Islets of Langerhans; Kinetics; Male; Phentolamine; Rats; Rats, Inbred Strains

The actions of efaroxan, a highly selective and potent alpha 2-adrenoceptor antagonist, on insulin secretion, cAMP levels, 86Rb+ efflux and ATP-regulated potassium (K+ATP) channels have been studied using isolated pancreatic islets of Langerhans and RINm5F cells. In the absence of an adrenoceptor agonist, efaroxan (1-100 microM) potentiated glucose-induced secretion over the range 4-10 mM glucose, but was without effect upon the maximal rate of secretion induced by 20 mM glucose. Efaroxan did not affect cAMP levels. Suppression of insulin release by the potassium channel opener diazoxide, was partially alleviated by efaroxan and was associated with an inhibition of the diazoxide-induced increase in the rate of 86Rb+ efflux. Using isolated patches of membrane we found efaroxan to be an effective blocker of K+ATP channels, with a KI value of 12 microM and a Hill coefficient of approximately 1. These data indicate that efaroxan promotes insulin secretion, in the absence of exogenous agonists, by a mechanism that involves inhibition of ATP-regulated K+ channels.

Topics: Adenosine Triphosphate; Adrenergic alpha-Antagonists; Animals; Benzofurans; Diazoxide; Drug Interactions; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Male; Potassium Channels; Rats; Rats, Inbred Strains

The imidazoline alpha 2-antagonist efaroxan was found to stimulate insulin secretion from isolated rat pancreatic islets incubated in glucose concentrations between 4 and 12 mM. This response could not be attributed to interaction of efaroxan with either classical alpha 2-receptors or with a B-cell 'imidazoline receptor', since the effect was not reproduced by the structural analogue idazoxan. Stimulation of insulin secretion by efaroxan correlated with the ability of the drug to reverse the inhibition of secretion mediated by the potassium channel agonist diazoxide. The data suggest that the capacity of efaroxan to stimulate insulin secretion may be related to an interaction with potassium channels in the pancreatic B-cell.

Topics: Animals; Benzofurans; Diazoxide; Glucose; Hypoglycemic Agents; Imidazoles; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Male; Potassium Channels; Rats; Rats, Inbred Strains

Sensitive and specific methods for the determination of efaroxan and idazoxan in blood plasma have been developed based on solvent extraction, chromatographic separation and quantification by selected-ion monitoring using a quadruple mass-selective detector. The use of a short non-polar bonded-phase capillary gas chromatography (GC) column facilitated rapid separation of the compounds of interest from internal standards, metabolites and endogenous material. Of equal significance was the ability to chromatograph these basic compounds without prior derivatisation. The application of bonded-phase capillary GC coupled to selected-ion monitoring resulted in robust analytical procedures with sub-ng ml(-1) sensitivity and high selectivity.

Topics: Adrenergic alpha-Antagonists; Benzofurans; Dioxanes; Dioxins; Gas Chromatography-Mass Spectrometry; Humans; Idazoxan; Imidazoles