nitroarginine and 7-nitroindazole

Nitric oxide (NO) is produced in physiological and pathophysiological conditions by three distinct isoforms of NO synthase (NOS): endothelial NOS (ecNOS), inducible NOS (iNOS), and brain NOS (bNOS). Selective inhibition of iNOS may be beneficial in various forms of shock and inflammation, whereas inhibition of bNOS may protect against neuroinjury. This article surveys the enzymatic mechanism of NO production, lists the strategies and pharmacological tools for selective inhibition of distinct NOS isoforms, and considers the side-effects of the various approaches. Selective inhibition of NOS isoforms is achieved by: (a) targeting the differential co-factor (calmodulin or tetrahydrobiopterin) requirement of various NOS isoforms, and NOS; (b) targeting the differential substrate requirements of cells expressing various isoforms of NOS (L-arginine uptake blockers or arginase); (c) the use of pharmacological agents that are selectively taken up by cells expressing various isoforms of NOS (7-nitroindazole); or (d) developing pharmacological NOS inhibitors with isoform specificity. The amino acid-based NOS inhibitor, NG-nitro-L-arginine, shows a preference for ecNOS and bNOS over iNOS, whereas L-N6-(1-iminoethyl)lysine is selective for iNOS over bNOS. Certain non-amino acid-based small molecules, such as aminoguanidine and certain S-alkylated isothioureas, also express selectivity towards iNOS and have anti-inflammatory and anti-shock properties. 7-nitroindazole, a bNOS-selective inhibitor, protects in central nervous system injury. Clearly, there are a number of distinct approaches that are worthy of further research efforts in order to achieve even more selective targeting of various NOS isoforms

Topics: Animals; Arginase; Arginine; Brain; Endothelium, Vascular; Enzyme Inhibitors; Humans; Indazoles; Isoenzymes; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine

Four inhibitors of nitric oxide synthase (NOS), administered as acute pretreatments, attenuated several signs of naloxone-precipitated opioid withdrawal in morphine-dependent rats. Profiles of these drugs for inhibiting the expression of withdrawal were similar to that of clonidine, a drug used clinically to treat opioid withdrawal. The nonselective NOS inhibitors, NG-nitro-L-arginine and NG-nitro-L-arginine methyl ester, and N(5)-(1-iminoethyl)-L-ornithine, a selective inhibitor of endothelial NOS, Increased blood pressure in awake, morphine-naive and morphine-dependent rats not undergoing withdrawal. 7-Nitroindazole, a selective inhibitor of neuronal NOS, did not elevate blood pressure. Insofar as hypertension is a component of opioid withdrawal in humans, the ability of 7-nitroindazole to attenuate morphine withdrawal in rats without eliciting a vasopressor response suggests that 7-nitroindazole may have human therapeutic potential. Research directions for the continued development of 7-nitroindazole as a therapeutic modality are discussed with respect to issues of physical dependence, tolerance, and safety.

Topics: Adrenergic alpha-Agonists; Animals; Arginine; Blood Pressure; Clonidine; Depression, Chemical; Enzyme Inhibitors; Heart Rate; Indazoles; Morphine; Narcotics; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Opioid-Related Disorders; Ornithine; Substance Withdrawal Syndrome

Application of mild irritants (1 M NaCl; pH 2.0) on the gastric mucosa potentiates the protective secretion of bicarbonates by epithelial cells. This response is mainly mediated by capsaicin-sensitive afferent nerve endings located in the submucosa. It was shown that activation of vanilloid type 1 receptors (TRPV1) induced by exogenous acidification of GM is not sufficient to potentiate the production of HCO

Topics: Amiloride; Animals; Bicarbonates; Capsaicin; Gastric Mucosa; Gene Expression Regulation; Hydrogen-Ion Concentration; Indazoles; Male; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type III; Nitroarginine; Osmolar Concentration; Perfusion; Phrenic Nerve; Rats; Rats, Sprague-Dawley; Signal Transduction; Sodium Chloride; Stomach; TRPV Cation Channels; Vagotomy

Roles of isoforms of constitutive synthase of nitric oxide, neuronal or endothelial (nNOS or eNOS), in control of gastric bicarbonate secretion induced by mild irritation of the gastric mucosa was assessed at the normoacid state or after blockade of gastric acid secretion with omeprazole. In anesthetized rats, the concentration of HCO3- in luminal perfusate was calculated basing on measurements of pH/PCO2. Mucosal irritation during 20 min with acidic hypertonic solution (1 M NaCl, pH 2.0) caused marked and omeprazole-independent increase of HCO-secretion. Selective blocker ofnNOS in vivo 7-nitroindazole (7-NI), and the nonselective blocker ofnNOS and eNOS, N(G)-nitro-L-arginine (L-NNA), were applied either intravenously (10 mg/kg), or locally via retrograde injection into the splenic artery (1 mg/kg). At the normo-acid state, the irritation-induced secretion of was suppressed by 7-NI, but was not affected by L-NNA. After administration of omeprazole, both 7-NI and L-NNA equally inhibited HCO3- output. The effect of 7-NI (but not L-NNA) was abolished by cyclooxygenase (COX) inhibitor, indomethacin, which by itself suppressed irritation-induced secretion of HCO3-. Additionally, bicarbonate output was substantially reduced by the blocker of soluble guanylate cyclase (GC), methylene blue. We conclude that irritation-induced secretion of HCO3- is largely mediated by intramural nNOS and depends on GC-COX interaction. As it was theoretically estimated, eNOS activity caused a reduction of HCO3- output in the normo-acid stomach. Omeprazole abolished the effect of eNOS.

Topics: Animals; Bicarbonates; Enzyme Inhibitors; Gastric Mucosa; Indazoles; Indomethacin; Irritants; Male; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type III; Nitroarginine; Omeprazole; Rats, Sprague-Dawley

Zinc is an important trace element in biological systems; however, excessive extracellular zinc could lead to neuronal cell death following ischemia, seizures, and brain trauma. In this study, we investigated whether the intracortical injection of zinc sulphate (200 μg/kg, i.c.) changes total number of Purkinje cells in the cerebellum and whether different types nitric oxide synthase inhibitors, N-(G)-nitro-L-arginine methyl ester (L-NAME), N(omega)-nitro-L-arginine (L-NNA), aminoguanidine and 7-nitroindazole (7-NI), have protective effects against zinc neurotoxicity in Wistar albino rats. Animals were divided into 6 groups: control, zinc, zinc+L-NAME (100 mg/kg, i.p.), zinc+L-NNA (100 mg/kg, i.p.), zinc+7-NI (100 mg/kg, i.p.) and zinc+aminoguanidine (100 mg/kg, i.p.) groups. Total number of Purkinje cells in the cerebellum was estimated using unbiased stereological technique as 318,947 ± 20,549, 123,483 ± 23,762, 206,537 ± 43,128, 178,135 ± 26,635, 193,148 ± 46,104 and 212,910 ± 26,399 in the control, zinc, zinc+L-NAME, zinc+L-NNA, zinc+7-NI and zinc+aminoguanidine groups, respectively (mean ± SD). The number of Purkinje cells in zinc group was significantly lower than that of the other groups (P<0.001). It was found that the nitric oxide synthase inhibitors have neuroprotective effect against zinc neurotoxicity on Purkinje cells. These data show that the inhibition of the nitric oxide synthase could prevent some of the deleterious effects of zinc on Purkinje cells.

Topics: Animals; Cell Count; Cell Death; Cerebellum; Enzyme Inhibitors; Guanidines; Indazoles; Neuroprotective Agents; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Purkinje Cells; Rats; Rats, Wistar; Zinc Sulfate

Cognitive and attentional deficits in schizophrenia include impairment of the sensorimotor filter as measured by prepulse inhibition (PPI). In this way, the study of animals that naturally present low PPI responses could be a useful approach for screening new antipsychotic drugs. Several pieces of evidence suggest that dopamine and nitric oxide (NO) can modulate PPI but their role in those animals is unknown.. The aim of this study was to investigate the role of dopamine and NO in Wistar rats with naturally low PPI response.. Male Wistar rats with low PPI responses received an i.p. injection of the antipsychotics haloperidol (0.1, 0.3 or 1mg/kg) or clozapine (0.5, 1.5 or 5mg/kg), the anxiolytic diazepam (1 or 3mg/kg) or the NO synthase (NOS) inhibitors, N(G)- nitro-l-arginine (l-NOARG; 40mg/kg, acutely or sub-chronically) or 7-Nitroindazole (7-NI; 3, 10 or 30mg/kg). All animals were submitted to the PPI test 1h after injection. Striatal and cortical dopamine, DOPAC, and noradrenaline levels of rats with low PPI responses were compared to rats with normal PPI responses.. We found increased levels of catecholamines on the striatum and prefrontal cortex of Wistar rats with low PPI. In these animals, both antipsychotics, typical and atypical, and NOS inhibitors significantly increased PPI.. Taken together, our findings suggest that the low PPI phenotype may be driven by an overactive catecholamine system. Additionally, our results corroborate the hypothesis of dopamine and NO interaction on PPI modulation and suggest that Wistar rats with low PPI may represent an interesting non-pharmacological model to evaluate new potential antipsychotics.

Topics: Acoustic Stimulation; Analysis of Variance; Animals; Anticonvulsants; Behavior, Animal; Biogenic Monoamines; Brain; Diazepam; Dopamine Antagonists; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Exploratory Behavior; Haloperidol; Indazoles; Inhibition, Psychological; Male; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Wistar; Reflex, Startle

The objective of this study was to determine the effects of 7-nitroindazole (7NI--a preferential neuronal nitric oxide synthase (NOS) inhibitor) and NG-nitro-L-arginine (NNA--a non-selective NOS inhibitor) on the anticonvulsant action of pregabalin (PGB--a third-generation antiepileptic drug) in the maximal electroshock (MES)-induced seizure model in mice. Electroconvulsions were produced in mice by means of an alternating current (50 Hz, 500 V, 25 mA, ear-clip electrodes, 0.2 s stimulus duration, tonic hindlimb extension taken as the endpoint). The anticonvulsant action of PGB in the MES test was expressed as median effective doses (ED50 values) of the drug, protecting 50% of animals tested against MES-induced seizures. The acute adverse-effect potentials of PGB in combination with 7NI and NNA were evaluated in the chimney test (motor coordination), step-through passive avoidance task (long-term memory) and grip-strength test (skeletal muscular strength) in mice. 7NI (50 mg/kg, ip) significantly enhanced the anticonvulsant action of PGB by reducing the ED50 value of PGB from 145.0 mg/kg to 74.4 mg/kg (p<0.01). Similarly, 7NI at the lower dose of 25 mg/kg also potentiated the anticonvulsant action of PGB by lowering the ED50 value of PGB from 145.0 mg/kg to 117.9 mg/kg, although the results did not attain statistical significance. In contrast, NNA (40 mg/kg, ip) had no impact on the anticonvulsant effects of PGB. Moreover, none of the examined combinations of PGB with 7NI and NNA affected motor coordination, long-term memory and skeletal muscular strength in mice. Based on this preclinical study, one can conclude that 7NI significantly enhanced and NNA had no effect on the anticonvulsant activity of PGB against MES-induced seizures in mice.

Topics: Animals; Anticonvulsants; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Synergism; Drug Therapy, Combination; Electroshock; Enzyme Inhibitors; gamma-Aminobutyric Acid; Indazoles; Male; Mice; Neuroprotective Agents; Nitroarginine; Pregabalin; Seizures; Toxicity Tests, Acute

The influence of an irreversible inhibitor of constitutive NO synthase (L-NOArg; 1.0 mg/kg ip), a relatively selective inhibitor of inducible NO synthase (L-NIL; 1.0 mg/kg ip) and a relatively specific inhibitor of neuronal NO synthase (7-NI; 0.1 mg/kg ip), on antihyperalgesic action of selective antagonists of B2 and B1 receptors: D-Arg-[Hyp3,Thi5,D-Tic7,Oic8] bradykinin (HOE 140; 70 nmol/kg ip) or des Arg10 HOE 140 (70 nmol/kg ip) respectively, in model of diabetic (streptozotocin-induced) and toxic (vincristine-induced) neuropathy was investigated.. The changes in pain thresholds were determined using mechanical stimuli--the modification of the classic paw withdrawal test described by Randall-Selitto.. The results of this paper confirm that inhibition of bradykinin receptors and inducible NO synthase but not neuronal NO synthase activity reduces diabetic hyperalgesia. Pretreatment with L-NOArg and L-NIL but not 7-NI, significantly increases antihyperalgesic activity both HOE 140 and des Arg10 HOE 140. It was also shown that both products of inducible NO synthase and neuronal NO synthase activation as well as bradykinin are involved in hyperalgesia produced by vincristine. Moreover, L-NOArg and 7-NI but not L-NIL intensify antihyperalgesic activity of HOE 140 or des-Arg10HOE 140 in toxic neuropathy.. Results of these studies suggest that B1 and B2 receptors are engaged in transmission of nociceptive stimuli in both diabetic and toxic neuropathy. In streptozotocin-induced hyperalgesia, inducible NO synthase participates in pronociceptive activity of bradykinin, whereas in vincristine-induced hyperalgesia bradykinin seemed to activate neuronal NO synthase pathway. Therefore, concomitant administration of small doses of bradykinin receptor antagonists and NO synthase inhibitors can be effective in alleviation of neuropathic pain, even in hospital care.

Topics: Animals; Bradykinin; Bradykinin Receptor Antagonists; Diabetes Mellitus, Experimental; Diabetic Neuropathies; Drug Evaluation, Preclinical; Enzyme Inhibitors; Hyperalgesia; Hypoglycemic Agents; Indazoles; Lysine; Male; Nitric Oxide Synthase; Nitroarginine; Pain Measurement; Rats; Rats, Wistar; Receptors, Bradykinin; Streptozocin; Tetrahydroisoquinolines; Vincristine

Chronic L-DOPA pharmacotherapy in Parkinson's disease is often accompanied by the development of abnormal and excessive movements known as L-DOPA-induced dyskinesia. Rats with 6-hydroxydopamine lesion of dopaminergic neurons chronically treated with L-DOPA develop a rodent analog of this dyskinesia characterized by severe axial, limb, locomotor and orofacial abnormal involuntary movements. While the mechanisms by which these effects occur are not clear, they may involve the nitric oxide system. In the present study we investigate if nitric oxide synthase inhibitors can prevent dyskinesias induced by repeated administration of L-DOPA in rats with unilateral 6-hydroxydopamine lesion. Chronic L-DOPA (high fixed dose, 100 mg/kg; low escalating dose, 10-30 mg/kg) treatment induced progressive dyskinesia changes. Two nitric oxide synthase inhibitors, 7-nitroindazole (1-30 mg/kg) and NG-nitro-L-arginine (50 mg/kg), given 30 min before L-DOPA, attenuate dyskinesia. 7-Nitroindazolee also improved motor performance of these animals in the rota-rod test. These results suggest the possibility that nitric oxide synthase inhibitors may be useful to treat L-DOPA-induced dyskinesia.

Topics: Animals; Antiparkinson Agents; Corpus Striatum; Disease Models, Animal; Dyskinesia, Drug-Induced; Enzyme Inhibitors; Indazoles; Levodopa; Male; Motor Activity; Nitric Oxide Synthase; Nitroarginine; Oxidopamine; Parkinson Disease; Rats; Rats, Wistar; Substantia Nigra

Drugs that facilitate dopaminergic neurotransmission induce cognitive and attentional deficits which include inability to filter sensory input measured by prepulse inhibition (PPI). Methylphenidate, an amphetamine analog is used in the treatment of attention deficit hyperactivity disorder. Given that nitric oxide (NO) modulates dopamine effect our aim is to analyze the nitric oxide synthase (NOS) and soluble guanylate cyclase (sGC) inhibitors effect on PPI disruption induced by methylphenidate. The inhibitors effects were compared to those produced by haloperidol and clozapine. Male Swiss mice received a first i.p. injection (one hour before testing), of either saline, or N(G) nitro l-arginine (10, 40 or 90 mg/kg), or 7-Nitroindazole (3, 10, 30 or 60 mg/kg), or oxadiazolo-quinoxalin (5 or 10 mg/kg), or haloperidol (1 mg/kg), or clozapine (5 mg/kg). Thirty min later mice received the second injection of either saline or methylphenidate (20 or 30 mg/kg) or amphetamine (5 or 10 mg/kg). One group of mice received intracerebroventricular 7-Nitroindazole (50 or 100 nM) followed by systemic administration of saline or methylphenidate (30 mg/kg). The results revealed a methylphenidate dose-dependent disruption of PPI comparable to amphetamine. The effect was prevented by either nitric oxide synthase or guanilate cyclase inhibitors or clozapine or haloperidol. In conclusion, methylphenidate induced a dose-dependent PPI disruption in Swiss mice modulated by dopamine and NO/sGC. The results corroborate the hypothesis of dopamine and NO interacting to modulate sensorimotor gating through central nervous system. It may be useful to understand methylphenidate and other psychostimulants effects.

Topics: Animals; Auditory Perception; Clozapine; Dopamine Antagonists; Dopamine Uptake Inhibitors; Dose-Response Relationship, Drug; Enzyme Inhibitors; GABA Antagonists; Haloperidol; Indazoles; Male; Methylphenidate; Mice; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oxadiazoles; Quinoxalines; Reflex, Startle

Nitric oxide (NO) is an intercellular messenger that, among other things, plays an important role in the nervous system as a gaseous neurotransmitter, modulating long-term potentiation (LTP) induction of synaptic transmission. LTP has been suggested to be the basis of memory formation. On the other hand NO also participates in excitotoxic processes which play an important role in many neuropathological states. The aim of this work was to observe the effect of two NO synthase (NOS) inhibitors (N omega-Nitro-L-arginine, NA; 7-nitroindazole, NI) on spontaneous behaviour, spatial learning and motor functions in Lurcher (+/Lc) and wild type (+/+) mice, derived from the B6CBA strain. Heterozygous Lurcher mutant mice represent a natural model of the olivocerebellar degeneration. They suffer from postnatal, practically total, extinction of cerebellar Purkinje cells (due to the excitotoxic apoptosis) and a partial decrease of granule cells and inferior olive neurons (ION) because of the lost target of their axons. +/+ animals are healthy littermates of +/Lc. NA is a nonselective NOS inhibitor which influences, except neuronal (n), also endothelial (e) NOS with an impact on blood pressure, NI is a selective nNOS inhibitor without any circulatory effect. The adult animals of both types (+/Lc; +/+) were influenced by acute administration of both inhibitors (25 mg/kg i.p. 30 min. before experiments) and newborns only by both acute and long-term administration of NI (1 month, starting from postnatal day 2, P2). Control solutions - saline or solvents of both NA and NI inhibitors--diluted 1M HCl and dimethyl sulfoxide (DMSO) respectively, were given at a relevant volume in the same way. The effect of both inhibitors and control solutions on motor functions was tested using four standard procedures (horizontal wire, slanting ladder, rotating cylinder, foot-bridge); in newborns at the age of 14 days. Spatial learning ability was examined in five-day long procedure in the Morris water maze (MWM) (in newborns started on P21). Spontaneous behaviour was studied only in adult animals (after acutely influencing them) employing the open field method. The results showed, that neither the Lurcher mutant, nor wild type mice derived from the B6CBA strain were significantly affected by NOS inhibitors NA and NI in spatial learning after both the acute and long-term application. Only significant decrease of swimming speed was found in both types of mice after the acute administration of

Topics: Animals; Enzyme Inhibitors; Indazoles; Maze Learning; Mice; Mice, Neurologic Mutants; Motor Activity; Nitric Oxide Synthase; Nitroarginine; Olivopontocerebellar Atrophies

To identify a possible role for nitric oxide (NO) in acute hypoxic tolerance (HT) we measured hypoxic survival time (HST), effect of hypoxic conditioning (HC), and survival following hypoxic conditioning while blocking or mimicking the action of nitric oxide synthase (NOS). To inhibit NOS, CD-1 mice were given supplemental endogenous NOS inhibitor asymmetrical dimethylarginine (ADMA) or a synthetic NOS inhibitor N(omega)-nitro-L-arginine (L-NNA), both of which nonselectively inhibit three of the isoforms of NOS [inducible (iNOS), neuronal (nNOS), and endothelial NOS (eNOS)]. ADMA (10 mg/kg i.p.) or saline vehicle was given 5 min before HST testing. L-NNA was given orally at 1 g/l in drinking water with tap water as the control for 48 h before testing. Both ADMA and L-NNA significantly increased HST and augmented the HC effect on HST. Neither the nNOS selective inhibitor 7-nitroindazole (7-NI) nor the iNOS selective inhibitor N-{[3-(aminomethyl)phenyl]methyl}-enthanimidamide (1400W) had a statistically significant effect on HST or HT. The NO donor, 3-morpholinosydnoeimine, when given alone did not significantly decrease HT, but it did mitigate the increased HT effect of L-NNA. These data confirm that acute hypoxic conditioning increases HT and that NOS inhibition by endogenous (ADMA) and a synthetic NOS inhibitor (L-NNA) further increases HT, whereas iNOS and nNOS inhibition does not, suggesting that it is the inhibition of eNOS that mediates enhancement of HT.

Topics: Acute Disease; Animals; Arginine; Endothelium, Vascular; Enzyme Inhibitors; Hypoxia; Imines; Indazoles; Male; Mice; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Nitroarginine; Time Factors

Conflicting results have been reported regarding the role of the nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway in the hippocampus on anxiety modulation.. To investigate the effects of intrahippocampal injections of drugs that modify the NO-cGMP pathway in rats submitted to two animal models that are sensitive to anxiolytic drugs, the elevated plus-maze and the Vogel punished licking test.. Male Wistar rats with cannulae aimed at the dentate gyrus of the dorsal hippocampus received microinjections of the NO synthase (NOS) inhibitors N (G)-nitro-L: -arginine methyl ester (LNAME, 15-300 nmol/0.2 microl), N (G)-nitro-L: -arginine (LNOARG, 50-300 nmol/0.2 microl), 7-nitroindazole (7NI, 10-100 nmol/0.2 microl), or the soluble guanylate cyclase inhibitor 1H-oxadiazolo-quinoxalin-1 one (ODQ, 10-100 nmol/0.2 microl), and were submitted to the elevated plus-maze. In a second group, the animals received 7NI, LNAME, or ODQ and were submitted to the Vogel punished licking test. To control for drug-induced changes in locomotor behavior, the animals were submitted to an open arena or to the Rota-rod test.. All drugs increased the exploration of the open arms of the elevated plus-maze. They also increased the number of punished licks in the Vogel test, indicating an anxiolytic effect. The anxiolytic effect of LNAME was prevented by previous treatment with L: -arginine (300 nmol/0.2 microl). Except for the lower dose of LNAME (15 nmol), administration of the NOS inhibitors or ODQ did not change exploratory activity in the open field nor cause any gross locomotor impairment in the Rota-rod test.. The results suggest that NO plays an anxiogenic role in the dentate gyrus of the dorsal hippocampus.

Topics: Analysis of Variance; Animals; Anxiety; Cyclic GMP; Dentate Gyrus; Dose-Response Relationship, Drug; Guanylate Cyclase; Hippocampus; Indazoles; Male; Maze Learning; Microinjections; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oxadiazoles; Punishment; Quinoxalines; Rats; Rats, Wistar; Receptors, Cytoplasmic and Nuclear; Soluble Guanylyl Cyclase

Intracerebral microdialysis/HPLC studies in Sprague-Dawley rats showed that the acquisition and execution of an emotional conditioned reflex was accompanied by an increase in the extracellular citrulline level in the nucleus accumbens; citrulline is a co-product of nitric oxide synthesis. The increase in the citrulline level evoked by execution of this reflex decreased after injection of 7-nitroindazole (0.5 mM), a selective inhibitor of neuronal NO synthase, into the nucleus accumbens, and was completely blocked by injection of N-nitroarginine (0.5 mM), a non-selective inhibitor NO synthase. The increase in the nucleus accumbens citrulline level seen during execution of the emotional conditioned reflex was prevented by administration of both of these NO synthase inhibitors. These data suggest that during the acquisition and execution of the emotional conditioned reflex, there is an increase in nitric oxide production in the nucleus accumbens, which arises predominantly as a result of activation of neuronal NO synthase.

Topics: Animals; Citrulline; Conditioning, Classical; Emotions; Enzyme Inhibitors; Extracellular Space; Indazoles; Male; Microdialysis; Microinjections; Nitric Oxide Donors; Nitric Oxide Synthase Type I; Nitroarginine; Nucleus Accumbens; Rats; Rats, Sprague-Dawley

Brief ischemia was reported to protect retinal cells against injury induced by subsequent ischemia-reperfusion with de novo protein synthesis, and this phenomenon is known as late ischemic preconditioning. The aims of the present study were to determine whether nitric oxide synthase (NOS) was involved in the mechanism of late ischemic preconditioning in rat retina using pharmacological tools. Under anesthesia with pentobarbital sodium, male Sprague-Dawley rats were subjected to 60 min of retinal ischemia by raising intraocular pressure to 130 mm Hg. Ischemic preconditioning was achieved by applying 5 min of ischemia 24 hrs before 60 min of ischemia. Retinal sections sliced into 5 microm thick were examined 7 days after ischemia. Additional groups of rats received NG-nitro-L-arginine and NG-monomethyl-L-arginin, non-selective NO synthase inhibitors, 7-nitroindazole, a neuronal NOS inhibitor, and aminoguanidine and L-N6-(1-iminoethyl) lysine, inducible NO synthase (iNOS) inhibitors before preconditioning, and were subjected to 60 min of ischemia. In the non-preconditioned group, cell loss in the ganglion cell layer and thinning of the inner plexiform and inner nuclear layer were observed 7 days after 60 min of ischemia. Ischemic preconditioning for 5 min completely protected against the histological damage induced by 60 min of ischemia applied 24 hrs thereafter. Treatment of rats with aminoguanidine and L-N6-(1-iminoethyl) lysine, but not NG-nitro-L-arginine, NG-monomethyl-L-arginine or 7-nitroindazole, wiped off the protective effect of ischemic preconditioning. The inhibitory effect of aminoguanidine was abolished by L-arginine, but not D-arginine. The results in the present study suggest that NO synthesized by iNOS is involved in the histological protection by late ischemic preconditioning in rat retina.

Topics: Animals; Enzyme Inhibitors; Guanidines; Indazoles; Ischemia; Ischemic Preconditioning; Lysine; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Nitroarginine; Rats; Rats, Sprague-Dawley; Retina; Retinal Vessels

Melatonin, the major hormone produced by the pineal gland, is shown to have anticonvulsant effects. Nitric oxide (NO) is a known mediator in seizure susceptibility modulation. In the present study, the involvement of NO pathway in the anticonvulsant effect of melatonin in pentylenetetrazole (PTZ)-induced clonic seizures was investigated in mice. Acute intraperitoneal administration of melatonin (40 and 80 mg/kg) significantly increased the clonic seizure threshold induced by intravenous administration of PTZ. This effect was observed as soon as 1 min after injection and lasted for 30 min with a peak effect at 3 min after melatonin administration. Combination of per se non-effective doses of melatonin (10 and 20 mg/kg) and nitric oxide synthase (NOS) substrate L-arginine (30, 60 mg/kg) showed a significant anticonvulsant activity. This effect was reversed by NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME, 30 mg/kg), implying an NO-dependent mechanism for melatonin effect. Pretreatment with L-NAME (30 mg/kg) and N(G)-nitro-L-arginine (L-NNA, 10 mg/kg) inhibited the anticonvulsant property of melatonin (40 and 80 mg/kg) and melatonin 40 mg/kg, respectively. Specific inducible NOS (iNOS) inhibitor aminoguanidine (100 and 300 mg/kg) did not affect the anticonvulsant effect of melatonin, excluding the role of iNOS in this phenomenon, while pretreatment of with 7-NI (50 mg/kg), a preferential neuronal NOS inhibitor, reversed this effect. The present data show an anticonvulsant effect for melatonin in i.v. PTZ seizure paradigm, which may be mediated via NO/L-arginine pathway by constitutively expressed NOS.

Topics: Animals; Anticonvulsants; Arginine; Convulsants; Dose-Response Relationship, Drug; Drug Therapy, Combination; Enzyme Inhibitors; Guanidines; Indazoles; Injections, Intraperitoneal; Kindling, Neurologic; Male; Melatonin; Mice; Mice, Inbred Strains; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitroarginine; Pentylenetetrazole; Seizures

Oxygen free radicals have been implicated in the pathogenesis of acoustic injury of the cochlea. The purpose of this study was to evaluate the effects of tempol (a superoxide anion scavenger), 3-aminobenzamide (a poly (ADP-ribose) synthetase (PARS) inhibitor), N-nitro-l-arginine (a non-selective nitric oxide synthase (NOS) inhibitor), 7-nitroindazole (a selective neuronal NOS inhibitor) and aminoguanidine (a selective inducible NOS inhibitor) on acoustic injury. Mice were exposed to a 4 kHz pure tone of 110-128 dB SPL for 4h. Tempol, 3-aminobenzamide or N-nitro-l-arginine was intraperitoneally administered immediately before the onset of acoustic overexposure, while 7-nitroindazole or aminoguanidine was intraperitoneally administered every 12h starting immediately before the onset of acoustic overexposure. The threshold shift of the auditory brainstem response (ABR) and hair cell loss were then evaluated one and two weeks after acoustic overexposure. Tempol and 3-aminobenzamide significantly protected the cochlea against acoustic injury, whereas the NOS inhibitors did not exert any protective effect. These findings suggest that reactive oxygen species and PARS are involved in acoustic injury of the cochlea. However, further study is necessary to elucidate the roles of nitric oxide and nitric oxide synthase in acoustic injury.

Topics: Analysis of Variance; Animals; Antioxidants; Auditory Threshold; Benzamides; Cochlea; Cyclic N-Oxides; Enzyme Inhibitors; Evoked Potentials, Auditory, Brain Stem; Female; Free Radical Scavengers; Guanidines; Hearing Loss, Noise-Induced; Indazoles; Mice; Neuroprotective Agents; Nitric Oxide Synthase; Nitroarginine; Noise; Poly(ADP-ribose) Polymerase Inhibitors; Spin Labels

Drugs like haloperidol (Hal) that decrease dopamine (DA) neurotransmission in the striatum induce catalepsy in rodents and Parkinson disease-like symptoms in humans. Nitric oxide synthase (NOS) inhibitors interfere with motor activity, disrupting rodent exploratory behavior and inducing catalepsy. Catalepsy induced by NOS inhibitors probably involves striatal DA-mediated neurotransmission. Antioxidants such as ascorbic acid (vitamin C) and alpha-tocopherol (vitamin E) have also been shown to interfere with movement modulation and the DA system.. The objective of the study is to investigate if the antioxidants vitamins C and E would influence the catalepsy produced by Hal and NOS inhibitors.. The effects of the following treatments on catalepsy were examined using the hanging-bar test on male Swiss mice (25-30 g): (1) vitamin C (30-1,000 mg/kg)xHal (1 mg/kg); (2) vitamin C (90-1,000 mg/kg)xN (G)-nitro-L: -arginine (LNOARG, 10 and 40 mg/kg); (3) vitamin C (300 mg/kg)xN (G)-nitro-L: -arginine methylester (LNAME, 20-80 mg/kg); (4) vitamin C (300 mg/kg) x 7-nitroindazole (7NI, 3-50 mg/kg); (5) vitamin C (90 mg/kg i.p.) x LNOARG [40 mg/kg twice a day during 4 days (subchronic treatment)]; (7) vitamin E (3-100 mg/kg) x Hal (1 mg/kg); and (6) vitamin E (3-100 mg/kg) x LNOARG (40 mg/kg).. Vitamin C enhanced the catalepsy produced by NOS inhibitors and Hal. Treatment with vitamin C did not affect tolerance to LNOARG cataleptic effect induced by subchronic treatment. Vitamin E potentiated the catalepsy induced by LNOARG at all doses tested; in contrast, catalepsy induced by Hal was enhanced only by the dose of 100 mg/kg.. Results support an involvement of dopaminergic and nitrergic systems in motor behavior control and provide compelling evidence that combined administration of the antioxidants vitamins C and E with either Hal or NOS inhibitors exacerbates extrapyramidal effects. Further studies are needed to assess possible clinical implications of these findings.

Topics: alpha-Tocopherol; Animals; Ascorbic Acid; Catalepsy; Disease Models, Animal; Dopamine Antagonists; Dose-Response Relationship, Drug; Drug Synergism; Drug Tolerance; Enzyme Inhibitors; Haloperidol; Indazoles; Male; Mice; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitroarginine; Time Factors; Vitamins

We investigated whether chronic ouabain treatment changes the vasoconstrictor responses induced by electrical field stimulation (EFS) in endothelium-denuded rat superior mesenteric arteries and a possible role of neuronal nitric oxide (NO).. Mesenteric arteries from untreated and ouabain-treated rats (approximately equal to 8.0 microg/kg per day, for 5 weeks) were used in this study. Vascular reactivity was analyzed by isometric tension recording. Expression of the neuronal NO synthase isoform was analyzed by Western blot. Noradrenaline release was evaluated in segments incubated with [H]noradrenaline.. Systolic (SBP) and diastolic (DBP) blood pressure were higher in ouabain-treated rats than in untreated rats (SBP, untreated: 120 +/- 3.5 mmHg versus ouabain-treated: 150 +/- 4.7 mmHg, P < 0.01; DBP, untreated: 87 +/- 3.0 mmHg versus ouabain-treated: 114 +/- 2.6 mmHg, P < 0.001). EFS-induced vasoconstrictions were smaller in arteries from ouabain-treated rats than in those from untreated animals, while the EFS-induced [H]noradrenaline release and the vasoconstriction induced by exogenous noradrenaline (1 nmol/l-10 micromol/l) remained unmodified. The non-selective NO synthase (NOS) inhibitor, N-nitro-L-arginine methyl ester (100 micromol/l), increased the EFS-induced vasoconstriction in mesenteric arteries from both groups, although the effect was more pronounced in segments from ouabain-treated rats. The selective neuronal NOS inhibitor, 7-nitroindazole (7-NI; 100 micromol/l) increased EFS-induced contraction only in segments from ouabain-treated rats. Neuronal NOS expression was greater in the mesenteric arteries from ouabain-treated rats than in those from untreated animals. Sodium nitroprusside (0.1 nmol/l-10 micromol/l) induced a similar vasodilatation in segments from both groups.. These results suggest that chronic ouabain treatment is accompanied by an increase in neuronal NO release that reduces EFS-induced vasoconstriction.

Topics: Animals; Electric Stimulation; Enzyme Inhibitors; Hypertension; Indazoles; Male; Mesenteric Arteries; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Ouabain; Rats; Rats, Wistar; Tritium; Vasoconstriction

We previously showed that pancreatic beta-cells express a neuronal isoform of nitric oxide synthase (nNOS) that controls insulin secretion by exerting two enzymatic activities: nitric oxide (NO) production and cytochrome c reductase activity. We now bring evidence that two inhibitors of nNOS, N-omega-nitro-l-arginine methyl ester (l-NAME) and 7-nitroindazole (7-NI), increase glucose-induced insulin secretion but affect beta-cell function differently. In the presence of l-NAME, insulin response is monophasic, whereas 7-NI preserves the normal biphasic secretory pattern. In addition, the alterations of beta-cell functional response induced by the inhibitors also differ by their sensitivity to a substitutive treatment with sodium nitroprusside, a chemical NO donor. These differences are probably related to the nature of the two inhibitors. Indeed, using low-temperature SDS-PAGE and real-time analysis of nNOS dimerization by surface plasmon resonance, we could show that 7-NI, which competes with arginine and tetrahydrobiopterin (BH(4)), an essential cofactor for nNOS dimer formation, inhibits dimerization of the enzyme, whereas the substrate-based inhibitor l-NAME stabilizes the homodimeric state of nNOS. The latter effect could be reproduced by the two endogenous inhibitors of NOS, N-omega-methyl-l-arginine and asymmetric dimethylarginine, and resulted interestingly in a reduced ability of the protein inhibitor of nNOS (PIN) to dissociate nNOS dimers. We conclude that intracellular factors able to induce abnormalities in the nNOS monomer/dimer equilibrium could lead to pancreatic beta-cell dysfunction.

Topics: Animals; Arginine; Carrier Proteins; Cell Line; Dimerization; Drosophila Proteins; Dyneins; Enzyme Inhibitors; Glucose; Indazoles; Insulin; Insulin Secretion; Kinetics; Male; Miconazole; NG-Nitroarginine Methyl Ester; Nitric Oxide Donors; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Nitroprusside; Rats; Rats, Wistar; Surface Plasmon Resonance

The problem of antidepressant-resistant depression has necessitated finding ways of augmenting the actions of currently existing antidepressants. The present studies investigate the possibility of synergistic interactions between nitric oxide (NO) synthase inhibitors and antidepressants in the mouse forced swim test (FST), a pre-clinical test of antidepressant activity. Treatment with a behaviourally subactive dose of the NO synthase inhibitor NG-nitro-L-arginine (L-NA) (3 mg/kg) augmented the behavioural effect of the tricyclic antidepressant imipramine. In a similar fashion L-NA (3 mg/kg) augmented the effect of the selective serotonin re-uptake inhibitor (SSRI) fluoxetine but not the noradrenaline re-uptake inhibitor, reboxetine in the FST. The interaction observed between L-NA and fluoxetine generalised to other selective serotonin re-uptake inhibitors, namely, sertraline and citalopram in the FST. Treatment with a subactive dose of the neuronally selective NO synthase inhibitor, 7-nitroindazole (30 and 50 mg/kg), augmented the behavioural effects of imipramine and fluoxetine, respectively. Thus inhibition of NO synthase enhances the activity of antidepressants that work via a serotonergic mechanism in the FST. The results of the present investigation support a view that antidepressant effects, or enhancement of such effects in the FST, may be elicited via NO synthase inhibition. Furthermore, these data raise the possibility that inhibition of NO synthase could be used as a strategy to enhance the clinical efficacy of serotonergic antidepressants.

Topics: Analysis of Variance; Animals; Antidepressive Agents; Behavior, Animal; Dose-Response Relationship, Drug; Drug Synergism; Escape Reaction; Fluoxetine; Imipramine; Immobilization; Indazoles; Male; Mice; Motor Activity; Nitric Oxide Synthase; Nitroarginine; Selective Serotonin Reuptake Inhibitors; Swimming

The aim of the present study was to evaluate the role of nitric oxide (NO) of sensory neural origin in neurogenic inflammatory response in the trigeminovascular system. Antidromic vasodilatation and plasma extravasation in response to electrical stimulation (15 V, 5 Hz, 0.5 ms, 100 impulses) of the trigeminal ganglion were investigated in the dura mater and nasal mucosa/upper eyelid by laser Doppler flowmetry and [(125)I]-labelled bovine serum albumin, respectively. Electrical stimulation of the trigeminal ganglion of rats elicited a reproducible ipsilateral enhancement of both meningeal and nasal mucosal blood flow. N(omega)-nitro-L-arginine (L-NNA; 4, 8, and 16 mg/kg, i.v.), a nonselective inhibitor of nitric oxide synthase (NOS), inhibited antidromic vasodilatation both in the dura mater (15.86+/-2.05%, 22.82+/-2.51%, and 36.28+/-4.37%) and nasal mucosa (35.46+/-8.57%, 58.72+/-9.2%, and 89.99+/-8.94%) in a dose-dependent manner. Specific inhibitors of neuronal NOS, 7-nitroindazole (7-NI; 20 mg/kg, i.v.) and 3-bromo-7-nitroindazole (3Br-7NI; 10 mg/kg, i.v.) were administered to assess the possible role of NO released from the trigeminal sensory fibres. The meningeal vasodilatation was inhibited by both 3Br-7NI and 7-NI (63.36+/-7.7% and 49+/-6.5%, respectively). The nasal hyperaemic response was also reduced by 3Br-7NI (78.26+/-8.7%). Plasma extravasation in the dura mater and upper eyelid evoked by electrical stimulation of the trigeminal ganglion (25 V, 5 Hz, 0,5 ms, 5 min), expressed as extravasation ratios (ERs) of the stimulated vs. nonstimulated sides, was 1.80+/-0.8 and 4.63+/-1.24, respectively. This neurogenic oedema formation was not inhibited by neither L-NNA nor 3Br-7NI. It is concluded that neural nitrergic mechanisms are involved in the meningeal vasodilatation evoked by electrical stimulation of the trigeminal ganglion.

Topics: Animals; Capillary Permeability; Dose-Response Relationship, Drug; Dura Mater; Electric Stimulation; Enzyme Inhibitors; Indazoles; Male; Meningeal Arteries; Meninges; Nasal Mucosa; Nitrergic Neurons; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Rats; Rats, Wistar; Trigeminal Ganglion; Vasodilation

We examined the contribution of nitric oxide (NO) on the contractile impairment in diaphragm muscles of endotoxemic rats. Force-frequency relationship was depressed 24 h after lipopolysaccharide administration. 7-Nitroindazole, aminoguanidine and 1H-[1,2,4]Oxadiazole (4,3-a)quinoxalin-1-one (ODQ) partially restored the contractile impairment, Nomega-Nitro-L-Arginine (L-NNA) was ineffective. K+ contractions were reduced by 50% in endotoxemic muscles, 7-nitroindazole partially recovered, while aminoguanidine and L-NNA were ineffective. Verapamil reduced contractility to a greater extent in endotoxemic muscles. Caffeine and ryanodine contractions were augmented during endotoxemia without NOS contribution. L-NNA, 7-nitroindazole, ODQ and hemoglobin did not affect, but aminoguanidine completely restored partially inhibited neurotransmission by d-tubocurarine. Endotoxemia did not change membrane potentials and neurotransmitter release but slightly increased excitability. At this stage of endotoxemia, (1) constitutive NOS appears to be the dominant isoform, (2) NO does not have a major role on contractile dysfunction and (3) impairment could be explained by altered sensitivity of the voltage sensor. (4) NO does not substantially modulate neuromuscular transmission in normal and endotoxemic rats.

Topics: Animals; Caffeine; Diaphragm; Endotoxemia; Enzyme Inhibitors; Guanidines; In Vitro Techniques; Indazoles; Lipopolysaccharides; Male; Muscle Contraction; Neuromuscular Nondepolarizing Agents; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oxadiazoles; Quinoxalines; Rats; Rats, Wistar; Ryanodine; Tubocurarine; Vasodilator Agents; Verapamil

Recent studies indicate that nitric oxide (NO) synthase inhibitors have antidepressant-like potential in various animal models. In the present study the behavioural activity of the NO synthase inhibitors, N(G)-nitro-L-arginine (L-NA) and 7-nitroindazole (7-NI), were assessed in a modified rat forced swimming test (FST). Both L-NA and 7-NI, dose dependently reduced immobility and increased swimming behaviour in the rat FST. This behavioural profile parallels the one previously shown with selective serotonin re-uptake inhibitors and serotonergic agonists. Thus, we examined the role of serotonin mediating the behavioural effects of L-NA and 7-NI in the rat FST. Depletion of endogenous serotonin using para-chlorophenylalanine (pCPA; 3 x 150 mg/kg, i.p.) completely blocked L-NA (20 mg/kg, i.p.) and 7-NI (20 mg/kg, i.p.)-induced reductions in immobility and increases in swimming behaviour during the FST. In conclusion these observations suggest that NO synthase inhibitors elicit their antidepressant-like activity in the modified swimming test through a serotonin dependent mechanism.

Topics: Animals; Antidepressive Agents; Depression; Dose-Response Relationship, Drug; Enzyme Inhibitors; Immobilization; Indazoles; Male; Motor Activity; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Sprague-Dawley; Serotonin

Nitric oxide (NO) has been implicated in some of the central pathways engaged in the regulation of the sleep-wake cycle. The existence of nitric oxide synthase (NOS) in the cholinergic basal forebrain (BF) cells projecting to the cortex suggests a role for NO in the activation induced by the BF during arousal. We tested, in the anesthetized cat, the hypothesis that inhibition of NOS would decrease the ability of BF cholinergic fibers to induce cortical activation. In control conditions, BF stimulation evoked an awake-like EEG pattern (i.e., a decrease in the low-frequency-high-amplitude oscillatory activity and an increase in the high-frequency-low-amplitude activity). After blocking NOS activity, the capacity of BF stimulation to induce cortical activation was strongly impaired. Furthermore, voltammetric measurements of NO levels revealed an increase in cortical NO after BF stimulation, also blocked by systemic NOS inhibition. These results indicate that the blockade of NOS activity significantly reduces the ability of BF stimulation to induce changes in the EEG pattern and suggest a role for NO in the BF-cholinergic system implicated in arousal mechanisms.

Topics: Animals; Arousal; Basal Nucleus of Meynert; Cats; Cerebral Cortex; Electric Stimulation; Electrodes, Implanted; Electroencephalography; Female; Indazoles; Male; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Time Factors

The effect of homocysteine (HCY) on lipid peroxidation (LP), a current mechanism of oxidative neurotoxicity, was investigated in rat brain synaptosomes. LP was assessed by measuring the amount of thiobarbituric acid-reactive substances (TBARS) formed from synaptosomal fractions following HCY treatment. Increasing HCY concentrations (5-1000 micro M) enhanced the TBARS formation in brain synaptosomes in a concentration-dependent manner. When compared at equimolar concentrations (100 micro M), the oxidative potency of HCY was lower than that of the oxidant ferrous sulfate, similar to that produced by glutamate (Glu) and the mitochondrial toxin 3-nitropropionic acid, and higher than that of the Glu agonists, kainate and quinolinate. The N-methyl-D-aspartate receptor (NMDAr) antagonist dizocilpine (MK-801) completely blocked the HCY-induced LP at concentrations between 5 to 1000 micro M, whereas the well-known antioxidant N-acetylcysteine (NAC) was less effective, but still protective against the HCY oxidative toxicity at higher concentrations (400 and 1000 micro M). Three nitric oxide synthase (NOS) inhibitors, 7-nitroindazole (7-NI), Nomega-nitro-L-arginine (L-NARG) and Nomega-nitro-L-arginine methyl ester (L-NAME), were also tested on HCY-induced LP at increasing concentrations. Both nonspecific NOS inhibitors (L-NARG and L-NAME) decreased more effectively the HCY-induced LP than did the selective neuronal NOS inhibitor, 7-NI. These results show that submillimolar concentrations of HCY can induce oxidative injury to nerve terminals, and this effect involves NMDAr stimulation, NOS activation, and associated free radicals formation.

Topics: Acetylcysteine; Animals; Antioxidants; Brain; Dizocilpine Maleate; Dose-Response Relationship, Drug; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Free Radical Scavengers; Homocysteine; Indazoles; Lipid Peroxidation; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Thiobarbituric Acid Reactive Substances

To assess the systemic and nociceptive effects of nitric oxide synthase (NOS) inhibitors in the modulation of acute pain in rats subjected to the formalin test.. Formalin 5% was injected in the hind paw in the presence and absence of NOS inhibitors (e.g., 7-nitro indazole, N-nitro-L-arginine and aminoguanidine). Catheters were chronically implanted to continuously record mean arterial blood pressure (MAP) and heart rate (HR). MAP, HR and paw lifting time were recorded at control and every five minutes for 35 min following formalin and NOS inhibitors.. Formalin injected into the rat hind paw induced a biphasic nociceptive behaviour: an initial acute phase (phase 1: during zero to five minutes after the formalin injection) followed by a prolonged tonic response (phase 2: beginning about ten minutes after the formalin injection). Aminoguanidine, an inhibitor of the inducible NOS and 7-nitro indazole, an inhibitor of the neuronal NOS, did not affect phase 1, whereas N-nitro-L-arginine, a non-selective NOS inhibitor decreased it (49%). All three NOS inhibitors diminished nociceptive behaviours during phase 2. L-arginine reversed antinociceptive effects of N-nitro-L-arginine in phase 1 and in phase 2. Pressor effects induced by formalin in phase 1 were abolished following all three NOS inhibitors. During phase 2, formalin-induced pressor effects remained unaffected by N-nitro-L-arginine and aminoguanidine but were inhibited by 7-nitro indazole.. Our data demonstrate that NO is predominantly generated by vascular endothelial NOS in phase 1 and phase 2, whereas the neuronal NOS and the inducible NOS exhibit antinociceptive effects through a non-NO related pathway in phases 1 and 2 in rats subjected to the formalin test.

Topics: Analgesics; Animals; Arginine; Blood Pressure; Enzyme Inhibitors; Formaldehyde; Guanidines; Heart Rate; Indazoles; Nitric Oxide Synthase; Nitroarginine; Pain; Pain Measurement; Rats; Rats, Sprague-Dawley; Sodium Chloride; Time Factors

The possible protection against the toxicity of 1-methyl-4-phenylpyridinium (MPP(+)) afforded by inhibitors of nitric oxide synthase (NOS) and the antagonist of N-methyl-D-aspartate receptor function, MK-801, was studied in a brain-slice superfusion system. Significant decreases in levels of dopamine and its metabolites 3,4-dihyroxyphenylacetic acid (DOPAC) and homovanillic acid were observed following incubation of slices with 25 microM MPP(+). The activity of intracellular lactate dehydrogenase (LDH), a marker of cell viability, was also significantly decreased. These effects were attenuated by preincubation with I mM 7-nitroindazole (7NI), a selective inhibitor of the neuronal isoform of nitric oxide synthase (NOS). In contrast, the nonspecific NOS inhibitor N(omega)-nitro-L-arginine, also at 1 mM, had no effect on levels of dopamine metabolites but did show a small attenuation of the levels of dopamine. 7NI alone caused some increase in levels of dopamine and a decrease in the metabolite DOPAC, which is consistent with it also acting as an inhibitor of monoamine oxidase-B. MK-801 afforded no significant protection of aminergic cells, although changes in LDH activity suggested that there may have been some protection of non-aminergic neurons affected by this, relatively high concentration of MPP(+).

Topics: 1-Methyl-4-phenylpyridinium; 3,4-Dihydroxyphenylacetic Acid; Animals; Brain; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Homovanillic Acid; In Vitro Techniques; Indazoles; L-Lactate Dehydrogenase; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Rats; Rats, Wistar

We investigated the role of central endothelin-A (ET(A)) receptors and nitric oxide (NO) in regulating arterial pressure during restraint stress and recovery from stress. Rats received intracerebroventricular (icv) injections of the ET(A) receptor antagonist BQ123 (24 microg/kg) and were then subjected to two restraint-rest cycles (1 h of restraint and 1 h of rest/cycle). Although mean arterial pressure (MAP) values in BQ123-treated and control rats increased at the onset of restraint and remained elevated during restraint, MAP values in BQ123-treated rats were consistently greater than in control rats. During rest periods, MAP values in control rats decreased to below baseline levels, whereas those in BQ123-treated rats remained significantly higher. NO content was decreased in the brain stems of BQ123-treated compared with control rats after the 4-h protocol. Injections (icv) of the NO synthase (NOS) inhibitor N(G)-nitro-L-arginine (L-NNA) eliminated the decreases in MAP values during rest periods in both BQ123-treated and control rats. Inhibition of neuronal NOS with icv injection of 7-nitroindazole sodium salt resulted in MAP values intermediate between control rats and rats receiving L-NNA. These results support the hypothesis that endothelin acts through ET(A) receptors in the brain, possibly via release of NO, to decrease arterial pressure during restraint and recovery from restraint.

Topics: Animals; Antihypertensive Agents; Blood Pressure; Endothelin Receptor Antagonists; Enzyme Inhibitors; Indazoles; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type III; Nitroarginine; Peptides, Cyclic; Rats; Rats, Sprague-Dawley; Receptor, Endothelin A; Receptors, Endothelin; Recovery of Function; Restraint, Physical; Stress, Physiological

We examined modulation by nitric oxide (NO) of sympathetic neurotransmitter release and vasoconstriction in the isolated pump-perfused rat kidney. Electrical renal nerve stimulation (RNS; 1 and 2 Hz) increased renal perfusion pressure and renal norepinephrine (NE) efflux. Nonselective NO synthase (NOS) inhibitors [N(omega)-nitro-L-arginine methyl ester (L-NAME) or N(omega)-nitro-L-arginine], but not a selective neuronal NO synthase inhibitor (7-nitroindazole sodium salt), suppressed the NE efflux response and enhanced the perfusion pressure response. Pretreatment with L-arginine prevented the effects of L-NAME on the RNS-induced responses. 2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO), which eliminates NO by oxidizing it to NO(2), suppressed the NE efflux response, whereas the perfusion pressure response was less susceptible to carboxy-PTIO. 8-Bromoguanosine cGMP suppressed and a guanylate cyclase inhibitor [4H-8-bromo-1,2,4-oxadiazolo(3,4-d)benz(b)(1,4)oxazin-1-one] enhanced the RNS-induced perfusion pressure response, but neither of these drugs affected the NE efflux response. These results suggest that endogenous NO facilitates the NE release through cGMP-independent mechanisms, NO metabolites formed after NO(2) rather than NO itself counteract the vasoconstriction, and neuronal NOS does not contribute to these modulatory mechanisms in the sympathetic nervous system of the rat kidney.

Topics: Animals; Benzoates; Cyclic GMP; Electric Stimulation; Enzyme Inhibitors; Imidazoles; In Vitro Techniques; Indazoles; Kidney; Male; Nervous System; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitroarginine; Norepinephrine; Oxadiazoles; Oxazines; Perfusion; Pressure; Rats; Rats, Wistar

Systemic injections of nitric oxide synthase (NOS) inhibitors have been shown to decrease exploratory behavior in rats. This effect may be related to motor impairments since these drugs can induce catalepsy in rodents.. To compare the effects of two NOS inhibitors in tests aimed to investigate exploratory behavior and to assess motor control.. The acute effects of the NOS inhibitors NG-nitro- L-arginine ( L-NOARG, 10-80 mg/kg IP) and 7-nitroindazole (7-NIO, 3-30 mg/kg IP) on exploratory activity were analyzed in an open field arena. Drug effects on catalepsy were examined in the hanging-bar and wire-ring test. Footprint pattern after treatment with the two NOS inhibitors was evaluated and the results compared with those obtained with the dopamine D2 receptor antagonist haloperidol (1-2 mg/kg IP). Sub-chronic (twice a day for 4 days) effects of L-NOARG (40 mg/kg) or 7-NIO (30 mg/kg) were also tested in the open field arena and catalepsy test.. L-NOARG and 7-NIO decreased locomotion and rearing in the open field arena. Both drugs induced catalepsy in the hanging-bar test but did not change footprint pattern. The cataleptic effect of L-NOARG in the hanging bar and wire-ring tests were highly correlated ( r=0.927). The exploratory and cataleptic effects of L-NOARG and 7-NIO provided evidence for tolerance after sub-chronic treatment.. These results confirm that inhibition of neuronal NO formation induces impairment of exploratory behavior. This effect does not seem to involve aspects evaluated by footprint analysis, such as weight support, trunk stability and foot placement. They could, however, be related to drug-induced catalepsy.

Topics: Animals; Catalepsy; Dose-Response Relationship, Drug; Enzyme Inhibitors; Exploratory Behavior; Indazoles; Male; Mice; Nitric Oxide Synthase; Nitroarginine

Microtubule-associated protein MAP-2 is a neuronal phosphoprotein which modulates microtubule stability and spatial organization of signal transduction pathways. The functions of MAP-2 are modulated by phosphorylation. We studied the modulation of MAP-2 phosphorylation using the N-methyl- D-aspartate (NMDA) type of glutamate receptors and the signal transduction pathways mediating this modulation in primary cultures of rat cerebellar neurons. NMDA induced a rapid increase (330% of basal at 5 min) in MAP-2 phosphorylation which was not prevented by KN-62, indicating that it is not mediated by activation of Ca-calmodulin-dependent protein kinase. NMDA-induced phosphorylation of MAP-2 was inhibited by the nitric oxide synthase inhibitors nitroarginine and 7-nitroindazole and by PD098059 (an inhibitor of MAP kinase kinase), but was only slightly reduced by calphostin C or U-73122, inhibitors of protein kinase C and of phospholipase C, respectively. This indicates that the main pathway mediating NMDA-induced phosphorylation of MAP-2 is activation of nitric oxide synthase and subsequent activation of MAP kinase. We show that activation of NMDA receptors induces an activation of MAP kinase which is prevented by nitroarginine. The nitric oxide-generating agent (+/-)-S-nitroso-N-acetylpenicillamine (SNAP) also induced activation of MAP kinase and increased phosphorylation of MAP-2. Other nitric oxide-generating agents (NOC-18 and NOR-3) also increased MAP-2 phosphorylation. The interplay between NMDA receptors-associated signal transduction pathways and MAP-2 may be involved in the modulation of neuronal responses to extracellular signals and in the regulation of neuronal function.

Topics: Animals; Cells, Cultured; Cerebellum; Enzyme Activation; Enzyme Inhibitors; Estrenes; Excitatory Amino Acid Agonists; Flavonoids; Indazoles; MAP Kinase Signaling System; Microtubule-Associated Proteins; Mitogen-Activated Protein Kinases; N-Methylaspartate; Naphthalenes; Neurons; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroarginine; Penicillamine; Phosphodiesterase Inhibitors; Phosphorylation; Pyrrolidinones; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

The aim of this study was to examine by electrophysiological techniques whether nitric oxide (NO) is involved in the development of desensitization to the opioid agonist Met5-enkephalin (ME) in locus coeruleus neurons from rat brain slices. Bath perfusion with ME (0.05-1.6 microM) caused a concentration-dependent reduction in the firing rate of locus coeruleus cells, whereas perfusion with a high concentration of ME (10 microM) desensitized the inhibitory effect of subsequent ME (0.8 microM) applications. However, in slices perfused with the NO synthase inhibitors 7-NI (100 microM), L-NAME (100 microM) or L-NA (100 microM) the ME (10 microM)-induced opioid desensitization was strongly attenuated. The effect of L-NAME was prevented by administration of L-arginine (100 microM). These results suggest that nitric oxide may contribute to opioid desensitization in locus coeruleus neurons.

Topics: Animals; Electrophysiology; Enkephalin, Methionine; Enzyme Inhibitors; In Vitro Techniques; Indazoles; Locus Coeruleus; Male; Neurons; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Sprague-Dawley; Receptors, Opioid, mu

Many properties of nitric oxide, NO, (localization, diffusiveness, half-life, vasodilatory affects) have supported its potential role in mediating the link between local cerebral activity and blood flow. However, evidence that both supports and refutes a role for NO in functional hyperemia have been presented. The present study employed multiple nitric oxide synthase inhibitors, two anesthetic regimes and laser-Doppler flowmetry to test the hypothesis that NO is critically involved in mediating the functional hyperemic response within rodent whisker-barrel cortex (WBC). In urethane anesthetized animals, functional hyperemic responses were obtained both before and after 1 mg/kg atropine infusion, 30 mg/kg i.v. L-NAME (N-Nitro-L-arginine methylester) infusion, 30 mg/kg L-NA (N-Nitro-L-arginine) infusion or 25 mg/kg 7-NI (7-nitroindazole). L-NAME was also tested in a group of animals pretreated with halothane before urethane anesthesia. Neither the magnitude of the blood flow response nor its time course was altered by NO blockade or atropine administration when compared to pre-infusion controls in urethane anesthetized rats. In contrast, animals that were pretreated with halothane exhibited a 33% inhibition of functional hyperemia after L-NAME administration. Taken together, these data do not support a primary role for NO in rat WBC functional hyperemia and suggest that previous reports of inhibition may have been secondary to the anesthesia employed.

Topics: Anesthesia, General; Anesthetics, Inhalation; Anesthetics, Intravenous; Animals; Atropine; Cerebrovascular Circulation; Enzyme Inhibitors; Halothane; Hyperemia; Indazoles; Male; Muscarinic Antagonists; Nerve Tissue Proteins; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Sprague-Dawley; Receptors, Muscarinic; Somatosensory Cortex; Touch; Urethane; Vasodilation; Vibrissae

The role of nitric oxide formation in the vasodilation in the eye and other orbital tissues caused by pre-ganglionic stimulation of the facial nerve was studied in cats under alpha-chloralose anaesthesia. Regional blood flows were determined with radioactive microspheres during unilateral stimulation of the facial nerve before and after inhibition of nitric oxide synthase (NOS), alone or in combination with muscarinic blockade.N(omega)-nitro-L-arginine (L-NA), a non-selective NOS-inhibitor, caused a significant increase in mean arterial blood pressure (MABP) and a decrease in cardiac output (CO). Concomitantly, local blood flows on the non-stimulated control side were reduced in most of the investigated tissues, indicating marked vasoconstriction. An inhibitor selective for neuronal NOS, 7-nitro-indazole (7-NI), had no significant effect on MABP, CO or local blood flows. During facial nerve stimulation at 5 Hz (n =6), choroidal blood flow on the stimulated side was 108+/-41% (P

Topics: Animals; Atropine; Blood Pressure; Cardiac Output; Cats; Choroid; Electric Stimulation; Enzyme Inhibitors; Facial Nerve; Female; Indazoles; Male; Microspheres; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Orbit; Parasympatholytics; Regional Blood Flow; Vasodilation

Nitric oxide (NO) is produced by NO synthase (NOS) and contributes to the regulation of vascular tone in the perinatal lung. Although the neuronal or type I NOS (NOS I) isoform has been identified in the fetal lung, it is not known whether NO produced by the NOS I isoform plays a role in fetal pulmonary vasoregulation. To study the potential contribution of NOS I in the regulation of basal fetal pulmonary vascular resistance (PVR), we studied the hemodynamic effects of a selective NOS I antagonist, 7-nitroindazole (7-NINA), and a nonselective NOS antagonist, N-nitro-L-arginine (L-NNA), in chronically prepared fetal lambs (mean age 128 +/- 3 days, term 147 days). Brief intrapulmonary infusions of 7-NINA (1 mg) increased basal PVR by 37% (P < 0.05). The maximum increase in PVR occurred within 20 min after infusion, and PVR remained elevated for up to 60 min. Treatment with 7-NINA also increased the pressure gradient between the pulmonary artery and aorta, suggesting constriction of the ductus arteriosus (DA). To test whether 7-NINA treatment selectively inhibits the NOS I isoform, we studied the effects of 7-NINA and L-NNA on acetylcholine-induced pulmonary vasodilation. The vasodilator response to acetylcholine remained intact after treatment with 7-NINA but was completely inhibited after L-NNA, suggesting minimal effects on endothelial or type III NOS after 7-NINA infusion. Western blot analysis detected NOS I protein in the fetal lung and great vessels including the DA. NOS I protein was detected in intact and endothelium-denuded vessels, suggesting that NOS I is present in the medial or adventitial layer. We conclude that 7-NINA, a selective NOS I antagonist, increases basal PVR, systemic arterial pressure, and DA tone in the late-gestation fetus and that NOS I protein is present in the fetal lung and great vessels. We speculate that NOS I may contribute to NO production in the regulation of basal vascular tone in the pulmonary and systemic circulations and the DA.

Topics: Acetylcholine; Animals; Blotting, Western; Ductus Arteriosus; Enzyme Inhibitors; Fetus; Gestational Age; Hemodynamics; Indazoles; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Pulmonary Circulation; Sheep; Vasodilation; Vasomotor System

The effects of nitric oxide synthase (NOS) inhibitors, N(omega)-nitro-L-arginine and 7-nitroindazole, and the NOS substrate L-arginine on kainic acid (KA)-induced microglial reactivity and stress response were studied in the hippocampus 7 and 1 days after KA, respectively. Density of peripheral-type benzodiazepine receptors was measured as an index of microglial reactivity. Histological damage in hippocampus was evaluated at 7 days by neuronal counting. KA increased the maximal number of binding sites (B(max)) versus controls. Administration of either 7-nitroindazole (25 mg/kg) or N(omega)-nitro-L-arginine (20 and 50 mg/kg) 24 hr before KA, further increased B(max). This later effect was abolished by L-arginine (1 g/kg), which given 24 hr before KA decreased B(max) to control values. Also, KA-induced HSP72 stress response was attenuated by pre-treatment with L-arginine. Histological evaluation showed reduced cell numbers in the pyramidal cell layer of the hippocampus in groups receiving KA, either alone or in combination with 7-nitroindazole. Administration of L-arginine before KA attenuated neuronal loss in CA3 but not CA1. A clear protective effect was observed, however, in CA1 and CA3, in rats receiving both L-arginine plus 7-nitroindazole before KA. The results show that the combination of a NO substrate with a NOS inhibitor reduces the neurotoxic effects of KA in the rat hippocampus. This study suggests that extremely fine regulation of NO levels in the different neural cell types can modulate excitotoxicity.

Topics: Animals; Antineoplastic Agents; Blotting, Western; Enzyme Inhibitors; Excitatory Amino Acid Agonists; Heat-Shock Proteins; Hippocampus; HSP72 Heat-Shock Proteins; Indazoles; Intracellular Signaling Peptides and Proteins; Isoquinolines; Kainic Acid; Male; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroarginine; Protein Serine-Threonine Kinases; Radioligand Assay; Rats; Rats, Sprague-Dawley; Receptors, GABA-A

We have found that paraquat (PQ), a widely used herbicide, causes wet dog shakes (WDS), which involve the central opioid system, in rats. A non-selective nitric oxide (NO) synthase (NOS) inhibitor, N(omega)-nitro-L-arginine (L-NA), but not its less active enantiomer, N(omega)-nitro-D-arginine, decreased the PQ-induced WDS in a dose-related manner. A selective neuronal NOS inhibitor in vivo, 7-nitroindazole, also decreased the PQ-induced WDS. Although an opioid receptor antagonist, naloxone, reversed the suppressive effect of these NOS inhibitors on the PQ-induced WDS, L-arginine, an NO precursor, had no effect on it. These findings suggest that the suppression of the PQ-induced WDS by NOS inhibition is associated with the central opioid system and is insusceptible to exogenous L-arginine.

Topics: Animals; Arginine; Enzyme Inhibitors; Herbicides; Indazoles; Male; Naloxone; Narcotic Antagonists; Nitric Oxide Synthase; Nitroarginine; Paraquat; Rats; Rats, Sprague-Dawley; Reflex

Reperfusion injury is one of the factors that unfavorably affects stroke outcome and shortens the window of opportunity for thrombolysis. Surges of nitric oxide (NO) and superoxide generation on reperfusion have been demonstrated. Concomitant generation of these radicals can lead to formation of the strong oxidant peroxynitrite during reperfusion.. We have examined the role of NO generation and peroxynitrite formation on reperfusion injury in a mouse model of middle cerebral artery occlusion (2 hours) and reperfusion (22 hours). The infarct volume was assessed by 2,3,5-triphenyl tetrazolium chloride staining; blood-brain barrier permeability was evaluated by Evans blue extravasation. Nitrotyrosine formation and matrix metalloproteinase-9 expression were detected by immunohistochemistry.. Infarct volume was significantly decreased (47%) in animals treated with the nonselective nitric oxide synthase (NOS) inhibitor N(omega)-nitro-L-arginine (L-NA) at reperfusion. The specific inhibitor of neuronal NOS, 7-nitroindazole (7-NI), given at reperfusion, showed no protection, although preischemic treatment with 7-NI decreased infarct volume by 40%. Interestingly, prereperfusion administration of both NOS inhibitors decreased tyrosine nitration (a marker of peroxynitrite toxicity) in the ischemic area. L-NA treatment also significantly reduced vascular damage, as indicated by decreased Evans blue extravasation and matrix metalloproteinase-9 expression.. These data support the hypothesis that in addition to the detrimental action of NO formed by neuronal NOS during ischemia, NO generation at reperfusion plays a significant role in reperfusion injury, possibly through peroxynitrite formation. Contrary to L-NA, failure of 7-NI to protect against reperfusion injury suggests that the source of NO is the cerebrovascular compartment.

Topics: Animals; Biomarkers; Blood-Brain Barrier; Coloring Agents; Endothelium, Vascular; Enzyme Inhibitors; Evans Blue; Indazoles; Infarction, Middle Cerebral Artery; Matrix Metalloproteinase 9; Mice; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oxidants; Permeability; Reperfusion Injury; Tyrosine

In a hamster model (genetic symbol dt(sz)) of primary paroxysmal non-kinesiogenic dystonic choreoathetosis, recent studies have shown beneficial effects of glutamate and dopamine receptor antagonists. Nitric oxide (NO), synthesized from L-arginine by NO synthase in response to glutamate receptor activation, elicits cyclic GMP and modulates glutamate-mediated processes and striatal dopamine release. Therefore, the effects of NO synthase inhibitors and of L-arginine on severity of dystonia were investigated in dt(sz) hamsters in which dystonic attacks, characterized by twisting movements and postures, can be induced by stress. The NO synthase inhibitors N(G)-nitro-L-arginine (L-NNA), N(G)-nitro-L-arginine methyl ester (L-NAME) and 7-nitroindazole significantly reduced the severity of dystonia. At antidystonic effective doses neither L-NNA nor L-NAME caused observable side effects, whereas 7-nitroindazole exerted moderate reduction of locomotor activity. The antidystonic effect of L-NAME was reversed by co-administration of the NO precursor L-arginine. However, L-arginine administered alone did not exert any effect on severity of dystonia. Cerebellar cyclic GMP levels in brains of mutant hamsters in comparison to non-dystonic control hamsters did not significantly differ, but the cerebellar cyclic GMP levels tended to be increased in dt(sz) hamsters during a dystonic attack. L-NAME significantly decreased the cerebellar cyclic GMP levels in both dt(sz) and control hamsters. Although an overproduction of NO is probably not critically involved in the pathogenesis of paroxysmal dystonia, it may contribute to the manifestation of dystonic attacks, as indicated by the antidystonic effects of NO synthase inhibitors. Peripheral side effects may limit the clinical use of NO synthase inhibitors, but more selective inhibitors of the neuronal NO synthase should be considered as interesting candidates for the treatment of paroxysmal dystonia.

Topics: Animals; Brain Chemistry; Cricetinae; Cyclic GMP; Dopamine; Dystonia; Enzyme Inhibitors; Indazoles; Motor Activity; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitroarginine; Receptors, N-Methyl-D-Aspartate

Nicotine exerts its central actions by regulating cationic fluxes through nicotinic acetylcholine receptors (nAChRs). By this effect, the drug likely also modifies events occurring beyond the nAChR, including the regulation of nitric oxide (NO) synthesis. The present study was undertaken to assess the effects of acute and chronic nicotine administration (0.4 mg/kg, s.c.) on levels of NO(-)(2)+NO(-)(3), stable metabolites of NO, in brain regions of male and female rats. Nicotine increased levels of the metabolites, and therefore presumably of NO, with sex differences in the degree of stimulation, the brain regions affected, and the variance between the effects of acute and chronic administration. Prior inhibition of NO synthase eliminated the effect of nicotine in all regions studied. While nicotine appeared to increase NO indirectly via glutamate receptors in the cortex and hippocampus, this was not true of the corpus striatum, where blocking NMDA-type glutamate receptors with MK-801 had no effect. The findings support the view that NO is likely involved in some of the central effects of nicotine.

Topics: Analysis of Variance; Animals; Corpus Striatum; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Ganglionic Stimulants; Hippocampus; Indazoles; Male; Nicotine; Nitrates; Nitric Oxide; Nitrites; Nitroarginine; Rats; Rats, Sprague-Dawley; Sex Characteristics; Time Factors

Long-term potentiation (LTP) can be induced by giving only one burst (five stimuli at 200 Hz) on the positive phase of sensory-induced theta rhythm in awake or anaesthetized rats, a stimulation protocol that mimics naturally occurring neuronal activity. Nitric oxide has been discussed as an important neuronal messenger in the induction of LTP. However, experiments testing inhibitors of nitric oxide synthase (NOS) in vitro produced contradictory results. The non-specific NOS inhibitor Nitro-L-arginine (L-NARG) impaired LTP induced by high-frequency stimulation (HFS) [from 155 +/- 7% to 122 +/- 8%), but completely blocked theta-dependent LTP induction (161 +/- 8% to 102 +/- 5%). NOS inhibitors, e.g. 7-nitro indazole (7-NI) or 1-(2-trifluoromethylphenyl) imidazole (TRIM) that are more selective for neuronal NOS and affect blood pressure less also impaired HFS-induced LTP (186 +/- 11% to 135 +/- 9% for TRIM) but completely blocked theta-dependent LTP (154 +/- 7 to 91 +/- 8). L-Arginine reversed the effects of the NOS inhibitors tested. Therefore, NO appears to be a modulator that is important for synaptic plasticity in this more physiological stimulation technique in vivo. NO is not released in slice preparations in sufficient quantities or at the right timing. Instead, the unphysiologically strong HFS protocol appears to induce an NO-independent type of LTP in some cases.

Topics: Anesthesia; Animals; Electric Stimulation; Enzyme Inhibitors; Excitatory Postsynaptic Potentials; Hippocampus; Indazoles; Long-Term Potentiation; Male; Neurons; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Physical Stimulation; Polymethacrylic Acids; Rats; Rats, Wistar; Tail; Theta Rhythm; Wakefulness

Neuroprotection against cerebral ischemia can be realized if the brain is preconditioned by previous exposure to a brief period of sublethal ischemia. The present study was undertaken to test the hypothesis that nitric oxide (NO) produced from the neuronal isoform of NO synthase (NOS) serves as a necessary signal for establishing an ischemia-tolerant state in brain. A newborn rat model of hypoxic preconditioning was used, wherein exposure to sublethal hypoxia (8% oxygen) for 3 hours renders postnatal day (PND) 6 animals completely resistant to a cerebral hypoxic-ischemic insult imposed 24 hours later. Postnatal day 6 animals were treated 0.5 hour before preconditioning hypoxia with the nonselective NOS inhibitor L-nitroarginine (2 mg/kg intraperitoneally). This treatment, which resulted in a 67 to 81% inhibition of calcium-dependent constitutive NOS activity 0.5 to 3.5 hours after its administration, completely blocked preconditioning-induced protection. However, administration of the neuronal NOS inhibitor 7-nitroindazole (40 mg/kg intraperitoneally) before preconditioning hypoxia, which decreased constitutive brain NOS activity by 58 to 81%, was without effect on preconditioning-induced cerebroprotection, as was pretreatment with the inducible NOS inhibitor aminoguanidine (400 mg/kg intraperitoneally). The protective effects of preconditioning were also not blocked by treating animals with competitive [3-(2-carboxypiperazin-4-yl)propyl-1-phosphonate; 5 mg/kg intraperitoneally] or noncompetitive (MK-801; 1 mg/kg intraperitoneally) N-methyl-D-aspartate receptor antagonists prior to preconditioning hypoxia. These findings indicate that NO production and activity are critical to the induction of ischemic tolerance in this model. However, the results argue against the involvement of the neuronal NOS isoform, activated secondary to a hypoxia-induced stimulation of N-methyl-D-aspartate receptors, and against the involvement of the inducible NOS isoform, but rather suggest that NO produced by the endothelial NOS isoform is required to mediate this profound protective effect.

Topics: Animals; Animals, Newborn; Brain Ischemia; Calcium; Dizocilpine Maleate; Enzyme Inhibitors; Guanidines; Hypoxia; Indazoles; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Nitroarginine; Oxygen; Piperazines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Release of nitric oxide occurs as a consequence of glutamate stimulation of NMDA receptors and is dependent upon calcium-calmodulin activation of the enzyme nitric oxide synthase. Since nitric oxide may serve as an intracellular messenger for NMDA glutamatergic neurons, it could be hypothesized that blockade of its synthesis may produce pharmacological effects similar to those of NMDA receptor antagonists. The purpose of the present study was to compare the effects of nitric oxide synthase inhibitors to those of the high affinity NMDA open channel blocker phencyclidine in drug discrimination, a pharmacologically selective procedure in which phencyclidine produces distinctive effects. Rats were trained to discriminate 2 mg/kg phencyclidine from saline in a standard two-lever discrimination task with food reward. Whereas phencyclidine dose-dependently substituted for itself, 7-nitroindazole, L-NAME (N(G)-nitro-L-arginine methyl ester), and L-NOARG (N(G)-nitro-L-arginine) failed to substitute for phencyclidine when administered intraperitoneally. L-NAME and 7-nitroindazole were tested up to doses that disrupted responding, providing evidence that a behaviorally-relevant dosage range was evaluated. Although these results conflict with those of a previous study which found that nitric oxide synthase inhibitors substituted for phencyclidine and produced phencyclidine-like catalepsy in pigeons, they are consistent with research showing that these drugs did not produce phencyclidine-like pharmacological effects in behavioral procedures in rats.

Topics: Animals; Discrimination Learning; Discrimination, Psychological; Dose-Response Relationship, Drug; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Indazoles; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitroarginine; Phencyclidine; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Sodium Chloride

Excess neuronal nitric oxide (NO) production might cause adenosine triphosphate loss and cellular damage in hypoxic brain parenchyma. 31P nuclear magnetic resonance spectroscopy was used to study hypoxic intracellular responses in perfused respiring cerebrocortical slices, in which NO scavenging by hemoglobin is absent, during NO synthase blockade and NO augmentation.. Adenosine triphosphate concentrations were monitored at 4.7 Tesla in respiring slices before, during, and after 60 min of hypoxia (oxygen tension < 5 mmHg). Slices were not treated or were pretreated with 27 microM L-nitroarginine methyl ester (L-NAME), 27 microM 7-nitroindozole (7-NI), or 27 microM L-nitroarginine. Nitrotyrosine:tyrosine ratios of slice extracts were measured using high-performance liquid chromatography. Cresyl violet-stained sections (2 microm) from random slices were examined histologically.. After 60 min of hypoxia, adenosine triphosphate decreased to < or = 3, < or = 3, 65 +/- 6, and 25 +/- 4% of control in slices that were untreated or treated with L-nitroarginine, L-NAME, and 7-NI, respectively. After 120 min of hyperoxic recovery, adenosine triphosphate levels returned to control values in slices pretreated with L-NAME and 7-NI, but to only 30% of control in untreated or L-nitroarginine-treated slices. Nitric oxide donors administered during posthypoxic recovery partially antagonized the adenosine triphosphate recovery found with L-NAME and 7-NI. Nitric oxide synthase activity in slice homogenates, assayed via conversion of L-arginine to citrulline, was < or = 2% of control after all inhibitory treatments. The nitrotyrosine:tyrosine ratio increased by 52% in slices treated with 7-NI and by 200-300% in all other groups. Pretreatment with L-NAME and 7-NI reduced histologic evidence of cell swelling.. Neuronal NO is associated with rapid adenosine triphosphate reductions and peroxynitrite formation in acutely hypoxic cerebrocortical slices.

Topics: Adenosine Triphosphate; Animals; Cerebral Cortex; Enzyme Inhibitors; Hypoxia; Indazoles; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Sprague-Dawley

The effects of the nitric oxide-generating compounds hydroxylamine, sodium nitroprusside and S-nitroso-N-acetylpenicillamine, and the nitric oxide synthase inhibitors nitroarginine and 7-nitroindazole on taurine release from hippocampal slices from adult (three-month-old) and developing (seven-day-old) mice were characterized using a superfusion system. The basal release of [3H]taurine was enhanced when the nitric oxide donors were added at the beginning of superfusion, more markedly in the adult than in the immature hippocampus. The effect of hydroxylamine was clearly concentration-dependent. Hydroxylamine also markedly enhanced the release of endogenous taurine. The K+-stimulated (50 mM) release of taurine was generally inhibited by the nitric oxide-generating compounds in both age groups. Nitric oxide is thus able to act directly at presynaptic terminals, modulating taurine release as a retrograde messenger. The N-methyl-D-aspartate-evoked taurine release was reduced by the nitric oxide donors, particularly by sodium nitroprusside, in the adult hippocampus, while hydroxylamine and S-nitroso-N-acetylpenicillamine markedly potentiated the release in developing mice. In the immature hippocampus the hydroxylamine-enhanced taurine release seems to involve a Ca2+-independent, Na+-dependent and carrier-mediated process while in adult mice only a part of the hydroxylamine-enhanced release is mediated by the same mechanism. The results show that nitric oxide-generating compounds modify the basal, K+- and N-methyl-D-aspartate-evoked releases of taurine in both adult and immature hippocampus. The enhanced N-methyl-D-aspartate receptor-evoked release may be an important mechanism protecting the immature brain against excitotoxicity.

Topics: Aging; Animals; Hippocampus; Hydroxylamine; In Vitro Techniques; Indazoles; Mice; Mice, Inbred Strains; N-Methylaspartate; Nitric Oxide Donors; Nitroarginine; Nitroprusside; Penicillamine; Potassium; S-Nitroso-N-Acetylpenicillamine; Taurine

Nitric Oxide (NO) mediates a series of physiological processes including regulation of vascular tone, macrophage-mediated cytotoxicity, platelet aggregation, learning and long-term potentiation, neuronal transmission. Although NO mediates several physiological functions, overproduction of NO can be detrimental and play multiple roles in the pathophysiology of focal cerebral ischemia. In the present study NOS activities were evaluated in cerebellum and cerebral cortex of ischemic and post-ischemic reperfused rats using an experimental model of partial cerebral ischemia; moreover, the effects of L-N(G)Nitroarginine (NA, nonselective NOS inhibitor) or 7-Nitroindazole (7-NI, selective neuronal NOS inhibitor) administration were assayed on percentage survival of ischemic rats. An increase of NOS activity in the cerebellum and in cerebral cortex of ischemic and post-ischemic reperfused rats was observed. NA administration failed to induce neuroprotective effects, by increasing percentage of mortality of treated ischemic rats with respect to control group. In contrast, the treatment with the selective neuronal NOS inhibitor, 7-NI, induced a significant neuroprotective effect.

Topics: Animals; Brain; Brain Ischemia; Cell Survival; Enzyme Inhibitors; Indazoles; Male; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Rats; Rats, Wistar; Reperfusion; Time Factors

The neurotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is dependent upon the MAO-B (monoamine oxidase type B)-catalyzed production of 1-methyl-4-phenylpyridinium ion (MPP(+)) and is likely to involve a perturbation of energy metabolism. Protection against MPTP neurotoxicity has been shown by treating mice with 7-nitroindazole (7-NI), a reversible inhibitor of both MAO-B and neuronal nitric oxide synthase (nNOS) activity. The objective of the present study was to evaluate (i) the relationship between the neuroprotective effect of 7-NI and MPTP-induced energy deficiency, and (ii) the role of nitric oxide production as a potential mechanism for energy perturbation after MPTP exposure. Maximum protection against striatal dopamine depletion and nigral neuronal loss was achieved when 7-NI (50 mg/kg, i.p.) was administered to C57BL/6 mice immediately before and after MPTP (50 mg/kg, s.c.). This short-term regimen of 7-NI administration parallels the time when MPTP exposure causes energy failure. 7-NI also completely prevented the loss of striatal ATP that occurs in mice during the initial hours after MPTP administration. In contrast, N(G)-nitro-L-arginine (two injections of 50 mg/kg each, given i.p. 20 and 4 h prior to MPTP), another NOS inhibitor, failed to affect MPTP-induced ATP depletion. Taken together, data indicate that (i) a temporal and causal relationship exists between the neuroprotective effect of 7-NI and its ability to counteract ATP reduction, and (ii) MAO-B rather than NOS inhibition is the mechanism by which 7-NI counteracts MPTP-induced ATP depletion.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Adenosine Triphosphate; Animals; Corpus Striatum; Energy Metabolism; Enzyme Inhibitors; Indazoles; Male; Mice; Mice, Inbred C57BL; Neuroprotective Agents; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine

Crus II is an area of the cerebellar cortex that receives trigeminal afferents from the perioral region. We investigated the mechanisms of functional hyperemia in cerebellum using activation of crus II by somatosensory stimuli as a model. In particular, we sought to determine whether stimulation of the perioral region increases cerebellar blood flow (BFcrb) in crus II and, if so, whether the response depends on activation of 2-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-kainate receptors and nitric oxide (NO) production. Crus II was exposed in anesthetized rats, and the site was superfused with Ringer. Field potentials were recorded, and BFcrb was measured by laser-Doppler flowmetry. Crus II was activated by electrical stimulation of the perioral region (upper lip). Perioral stimulation evoked the characteristic field potentials in crus II and increased BFcrb (34 +/- 6%; 10 Hz-25 V; n = 6) without changing arterial pressure. The BFcrb increases were associated with a local increase in glucose utilization (74 +/- 8%; P < 0.05; n = 5) and were attenuated by the AMPA-kainate receptor antagonist 2, 3-dihydroxy-6-nitro-7-sulfamoylbenzo-[f]quinoxaline (-71 +/- 3%; 100 microM; P < 0.01; n = 5). The neuronal NO synthase inhibitor 7-nitroindazole (7-NI, 50 mg/kg; n = 5) virtually abolished the increases in BFcrb (-90 +/- 2%; P < 0.01) but did not affect the amplitude of the field potentials. In contrast, 7-NI attenuated the increase in neocortical cerebral blood flow produced by perioral stimulation by 52 +/- 6% (P < 0.05; n = 5). We conclude that crus II activation by somatosensory stimuli produces localized increases in local neural activity and BFcrb that are mediated by activation of glutamate receptors and NO. Unlike in neocortex, in cerebellum the vasodilation depends almost exclusively on NO. The findings underscore the unique role of NO in the mechanisms of synaptic function and blood flow regulation in cerebellum.

Topics: Animals; Blood Pressure; Brain Stem; Carbon Dioxide; Cerebellar Cortex; Cerebellum; Cerebrovascular Circulation; Electric Stimulation; Enzyme Inhibitors; Glucose; Hyperemia; Indazoles; Laser-Doppler Flowmetry; Male; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oxadiazoles; Quinoxalines; Rats; Rats, Sprague-Dawley; Regional Blood Flow; Somatosensory Cortex; Tetrodotoxin

Topics: Animals; Cardiovascular Agents; Cerebrovascular Circulation; Enzyme Inhibitors; Indazoles; Indomethacin; Male; Nitric Oxide; Nitroarginine; Rats; Rats, Wistar; Thromboxane A2; Vascular Resistance; Vasoconstriction; Vasodilation

Nitric oxide (NO) is a unique interneuronal neurotransmitter and/or neuromodulator that is involved in a variety of physiological functions within the central nervous system (CNS). In neural tissue, NO is generated from an oxygen-dependent, constitutive NO synthase (NOS) by glutamatergic stimulation of N-methyl-D-aspartate (NMDA) receptors. Recent studies indicate that NO has excitatory effects on breathing within the CNS and mediates a central component of the hypoxic ventilatory reflex in mammals. Because NMDA receptors are important in central respiratory rhythmogenesis, we hypothesized that NO would have significant effects on the central pattern generator (CPG) for breathing in the brainstem. To test this hypothesis, the effects of NO on respiratory-related neural activity were investigated using an in vitro brainstem preparation from North American bullfrogs (Rana catesbeiana). Extracellular recordings of respiratory-related burst activity were made from cranial nerves V, X and XII before and during superfusion of the brainstem with NO-generating compounds, or inhibitors of NO synthesis. Addition of the NO donor, sodium nitroprusside (SNP; 0.1-1.0 mM), or the amino acid precursor for NO synthesis, L-arginine (L-Arg; 0.01-1.0 mM), caused significant increases in respiratory-related burst frequency. Inhibition of NOS with N omega-nitro-L-arginine (L-NA; 5-10 mM), a non-selective NOS inhibitor, caused a significant reduction in burst frequency or reversibly abolished neural activity. Brainstem perfusion with the specific neuronal NOS (nNOS) inhibitor, 7-nitro indazole (7-NI), produced significant, dose-dependent reversible reductions in burst frequency at concentrations of 0.1, 0.5 and 1.0 mM. These results suggest that production of NO, probably via nNOS, provides an excitatory input to the respiratory CPG in the amphibian brainstem. Our results suggest that NO may be a necessary inter- or intracellular messenger for neurotransmission and/or neuromodulation of central respiratory drive to motor effectors in the bullfrog.

Topics: Animals; Arginine; Brain Stem; Dimethyl Sulfoxide; Dose-Response Relationship, Drug; Enzyme Inhibitors; In Vitro Techniques; Indazoles; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Nitroprusside; Oxygen Consumption; Perfusion; Rana catesbeiana; Respiratory Mechanics

Nitric oxide synthases (NOS) have a bidomain structure comprised of an N-terminal oxygenase domain and a C-terminal reductase domain. The oxygenase domain binds haem, (6R)-5,6,7,8-tetrahydro-l-biopterin (tetrahydrobiopterin) and arginine, is the site where nitric oxide synthesis takes place and contains determinants for dimeric interactions. A novel scintillation proximity assay has been established for equilibrium and kinetic measurements of substrate, inhibitor and cofactor binding to a recombinant N-terminal haem-binding domain of rat neuronal NOS (nNOS). Apparent Kd values for nNOS haem-domain-binding of arginine and Nomega-nitro-L-arginine (nitroarginine) were measured as 1.6 microM and 25 nM respectively. The kinetics of [3H]nitroarginine binding and dissociation yielded an association rate constant of 1.3x10(4) s-1.M-1 and a dissociation rate constant of 1.2x10(-4) s-1. These values are comparable to literature values obtained for full-length nNOS, suggesting that many characteristics of the arginine binding site of NOS are conserved in the haem-binding domain. Additionally, apparent Kd values were compared and were found to be similar for the inhibitors, L-NG-monomethylarginine, S-ethylisothiourea, N-iminoethyl-L-ornithine, imidazole, 7-nitroindazole and 1400W (N-[3-(aminomethyl) benzyl] acetamidine). [3H]Tetrahydrobiopterin bound to the nNOS haem domain with an apparent Kd of 20 nM. Binding was inhibited by 7-nitroindazole and stimulated by S-ethylisothiourea. The kinetics of interaction with tetrahydrobiopterin were complex, showing a triphasic binding process and a single off rate. An alternating catalytic site mechanism for NOS is proposed.

Topics: Animals; Arginine; Binding Sites; Binding, Competitive; Biopterins; Enzyme Inhibitors; Heme; Indazoles; Isothiuronium; Kinetics; Nitric Oxide Synthase; Nitroarginine; Oxidoreductases; Oxygenases; Peptide Fragments; Protein Binding; Rats; Recombinant Fusion Proteins

Glutamate stimulation of N-methyl-D-aspartate (NMDA) receptors results in release of nitric oxide which may mediate the effects of NMDA receptor stimulation and/or may result in feedback inhibition of the presynaptic neuron. Results of a previous study showed that nitric oxide synthase (NOS) inhibitors blocked dizocilpine-induced behavior in mice. In the present study, NOS inhibitors were tested in combination with phencyclidine (PCP), a drug which typically dose-dependently disrupts prepulse inhibition of the acoustic startle response in rats. Alone, NOS inhibitors and promoters do not affect prepulse inhibition; however, when tested in combination with PCP, the NOS inhibitors, L-NOARG, 7-nitroindazole and arcaine--but not the NR2B-selective polyamine site NMDA antagonist, eliprodil--attenuated PCP-induced disruption of prepulse inhibition of the acoustic startle response. These effects are similar to those produced by many atypical antipsychotics and suggests that this class of drugs should be investigated further for their potential utility as antipsychotics and as treatments for PCP abuse.

Topics: Acoustic Stimulation; Animals; Biguanides; Biogenic Polyamines; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Indazoles; Inhibition, Psychological; Male; Nitric Oxide Synthase; Nitroarginine; Phencyclidine; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reflex, Startle

The role of nitric oxide (NO) in the long-term serotoninergic neurotoxicity induced by (+/-)3,4-methylenedioxymethamphetamine (MDMA) in rats was investigiated. Pretreatment with Nomega-nitro-L-arginine (L-NOARG) (10 mg kg-1), a nitric oxide synthase (NOS) inhibitor, partially protected against long-term serotonin (5-HT) depletion induced by MDMA (40 mg kg-1) in frontal cortex and parietal cortex, but not in other brain regions examined. Brain NOS activities in these two regions were significantly elevated at 6 h after MDMA administration. Moreover, L-NOARG pretreatment caused significant inhibition of brain NOS activity but did not affect the acute 5-HT and dopamine (DA) changes or the hyperthermia induced by MDMA. These results suggest that it is the NOS inhibitory properties of L-NOARG, rather than its effects on the acute monoamine changes or the hyperthermia induced by MDMA, that are responsible for the prevention of neurotoxicity. The regional differences on the protection of L-NOARG and on the activation of NOS by MDMA indicate the unequal role that NO may play in MDMA-induced neurotoxicity in different brain regions.

Topics: Animals; Body Temperature; Enzyme Activation; Enzyme Inhibitors; Hydroxyindoleacetic Acid; Indazoles; Male; N-Methyl-3,4-methylenedioxyamphetamine; Nerve Degeneration; Nerve Tissue Proteins; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroarginine; Rats; Rats, Sprague-Dawley; Serotonin; Serotonin Agents

The role of nitric oxide (NO) in the sympatho-vagal control of heart rate was investigated in the cardiac sympathectomized and vagotomized anaesthetised rabbit and in the isolated guinea-pig atria with intact vagus nerve. Specific inhibition of neuronal nitric oxide synthase (nNOS) with 1-(2-trimethylphenyl) imidazole (TRIM, 50 mg kg(-1) i.v. in vivo) significantly enhanced the magnitude of the change in heart rate (HR) with sympathetic nerve stimulation (SNS, 31.6+/-4.5 bpm control vs. 49.7+/-6.0 bpm in TRIM, P < 0.05, 10 Hz). This effect was reversed by L-arginine (deltaHR 37.2+/-4.1 bpm, 50 mg kg(-1) i.v.). An enhanced HR response to SNS was also seen with the non-isoform specific inhibitor, N-omega-nitro-L-arginine (L-NA, 50 mg kg(-1) i.v.). Infusing isoprenaline (0.2 microg kg(-1) min(-1)) did not mimic the change in HR response to SNS with TRIM. There was, however, no significant effect of inhibition of NOS with TRIM L-NA or NG-monomethyl-L-arginine (L-NMMA, 20 mg kg(-1) i.v.) on the magnitude of the change in HR with vagal nerve stimulation (5 Hz) in vivo. There was also no significant effect of NOS inhibition on the change in HR with vagal nerve stimulation in vivo in the presence of pre-adrenergic stimulation or in the presence of propranolol (0.5 mg kg(-1) i.v., 2, 5 and 10 Hz stimulation). This result was confirmed in the isolated guinea-pig atria with the specific nNOS inhibitor, 7-nitroindazole (7-NiNa, 100 microM) at 1, 2, 3 or 5 Hz stimulation frequency. Our data suggest that endogenous NO plays an inhibitory role in cardiac sympathetic neurotransmission, but there was no convincing evidence from our results for a major role for endogenous NO in vagal control of heart rate, with or without prior adrenergic stimulation.

Topics: Adrenergic Fibers; Animals; Arginine; Enzyme Inhibitors; Guinea Pigs; Heart Atria; Heart Rate; Indazoles; Isoproterenol; Male; Neural Inhibition; Nitric Oxide Synthase; Nitroarginine; omega-N-Methylarginine; Polymethacrylic Acids; Propranolol; Rabbits; Sympatholytics; Sympathomimetics; Vagus Nerve

Dopamine (DA) neurons in the ventral tegmental area and substantia nigra pars compacta were induced to fire in bursts with application of N-methyl-D-aspartate (NMDA, 20 microM) and apamin (100 nM) while recording intracellularly in the rat brain slice. L-Arginine (300 microM), a substrate for nitric oxide (NO) production, increased both the number of spikes per burst and the magnitude of interburst hyperpolarizations. Nitric oxide synthase inhibitors N-nitro-L-arginine methyl ester (L-NAME, 100 microM), N-nitro-L-arginine, and 7-nitroindazole inhibited NMDA-induced burst firing by reducing the number of spikes per burst. Moreover, L-arginine (100 microM) reversed the inhibition of burst firing produced by L-NAME. These findings suggest that NO facilitates NMDA-induced burst firing in DA neurons.

Topics: Animals; Apamin; Arginine; Dopamine; Drug Synergism; Electrophysiology; Enzyme Inhibitors; In Vitro Techniques; Indazoles; Male; Mesencephalon; N-Methylaspartate; Neurons; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitroarginine; Rats; Rats, Sprague-Dawley

The present study investigated the role of nitric oxide (NO) in epileptogenesis and whether this role correlated with ionotropic glutamate receptor (IGR). Using a self-constructed NO-sensitive microelectrode (SNM), we observed the effect of nitric oxide synthase (NOS) inhibitors, NMDA and non-NMDA selective antagonists on penicillin(PEN)-treated hippocampal slices by simultaneously recording evoked field potentials and nitric oxide release from CA1 pyramidal neurons. 7-nitroindazole (7-NI),Nomega-nitro-L-arginine (L-NNA) and DL-2-amino-phospho-novaleric acid (APV), but not 6,7-dinitroquinoxaline-2,3 (1h,4h)-dione(DNQX), depressed NO release and partly reversed PEN's epileptogenetic effect, while APV + 7-NI + L-NNA did not display a further inhibitory effect. These findings suggest both NOS inhibitor and NMDA antagonist involve as anticonvulsant factors in epileptogenesis, providing direct evidence for NO release in response to NMDA receptor activation. The anticonvulsant effect of NMDA antagonist may ascribe to its action on NO release.

Topics: 2-Amino-5-phosphonovalerate; Action Potentials; Animals; Anticonvulsants; Electrochemistry; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Hippocampus; In Vitro Techniques; Indazoles; Male; Microelectrodes; N-Methylaspartate; Nitric Oxide; Nitroarginine; Penicillins; Rats; Rats, Wistar

We investigated the enzymatic source, cellular production, and functional importance of nitric oxide (NO) in rat diaphragm. Neuronal and endothelial isoforms of constituitive nitric oxide synthase (nc-NOS, ec-NOS) were identified by immunostaining. NOS activity measured in diaphragm homogenates averaged 5.1 pmol/min/mg. Passive diaphragm fiber bundles produced NO derivatives (NOx) at the rate of 0.9 pmol/min/mg as measured by the cytochrome c reduction assay; NO production was confirmed by photolysis/ chemiluminescence measurements. Endogenous NO depressed diaphragm contractile function. The force of submaximal contraction was increased by NOS inhibitors, an effect that was stable for up to 60 min and was reversed by NO donors. We conclude that diaphragm muscle fibers express nc-NOS, ec-NOS, or both; passive myocytes produce NOx; and NO or NO-derivatives inhibit force production by modulating excitation-contraction coupling.

Topics: Animals; Diaphragm; Enzyme Inhibitors; Guanidines; Hindlimb; Indazoles; Male; Muscle Contraction; Muscle, Skeletal; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type III; Nitrites; Nitroarginine; Rats; Rats, Sprague-Dawley

Using a nitric oxide (NO)-sensitive platinum microelectrode (SNM) modified with chitosan nickel (II) complex and Nafion (Nafion-CTS (Ni)-Pt), we observed the effect of nitric oxide synthase (NOS) inhibitor 7-nitro-indazole (7-NI) and N omega-nitro-L-arginine (L-NNA) on penicillin-treated hippocampal slices by simultaneously recording the stimulus-evoked field potentials and nitric oxide release from CA1 pyramidal neurons. Our results show: (1) The linear relationship range and detection limit of SNM were 4.5 x 10(-4)-1.0 x 10(-8) mol/L and 5.0 x 10(-8) mol/L respectively. (2) Penicillin (PEN) could elevate NO release, the number and amplitude of stimulus-evoked field potentials spike (SEPS) in concentration-dependent manner; (3) Both 7-NI and L-NNA depressed NO release and partly reversed the effect of penicillin. The above findings suggest that convulsant provoking effect of NO could be inhibited by NOS inhibitors. The NO-sensitive electrode may provide an useful tool for continuous detection of NO in biological tissues.

Topics: Animals; Anticonvulsants; Epilepsy; Evoked Potentials; Hippocampus; In Vitro Techniques; Indazoles; Microelectrodes; Neurons; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Penicillin G; Rats

Previously, we have reported that intranigral infusions of malonate, an inhibitor of mitochondrial function, lead to the degeneration of the dopaminergic neurons of the nigrostriatal pathway that is mediated, at least in part, through NMDA receptor activation and nitric oxide formation. In the present study, unilateral focal infusions of malonate into the nucleus basalis magnocellularis (nbM) of male Sprague-Dawley rats (weighing 250-300 g) resulted in a dose-related depletion in ipsilateral cortical and amygdaloid choline acetyltransferase (ChAT) activity. Infusion of a 3 mumol dose of malonate into the nbM of vehicle-treated animals resulted in a 41 and 54% decrease in cortical and amygdaloid ChAT activity, respectively. Systemic pretreatment with lamotrigine (16 mg/kg, i.p.) and MK-801 (5 mg/kg, i.p.) attenuated the depletions in cortical and amygdaloid ChAT activity that resulted from an infusion of this dose of malonate into the nbM. Acetylcholinesterase (AChE) histochemistry of the nbM following focal infusion of malonate (3 mumol) showed a marked decrease in the number of AChE-positive neurons that was partially prevented by MK-801 pretreatment. Before examining the role of nitric oxide formation in malonate-induced toxicity, the ability of systemic administration of N omega-nitro-L-arginine (L-NA) to inhibit nitric oxide synthase (NOS) activity in the nbM and cerebellum was investigated. L-NA (2, 10, and 20 mg/kg, i.p.) produced a dose-related inhibition of nbM and cerebellar NOS activity that was maximal following a dose of 10 mg/kg L-NA. This level of NOS inhibition persisted for at least 13 h following L-NA (10 mg/kg) administration. Subsequently, the effect of L-NA pretreatment on malonate toxicity was evaluated. Following pretreatment with L-NA (2 and 10 mg/kg, i.p.), the toxic action of malonate on cortical and amygdaloid ChAT activity was not altered. In addition, infusion of a lower dose of malonate (2 mumol) into the nbM resulted in decreases in cortical and amygdaloid ChAT activity that were not altered by pretreatment with L-NA (2 and 10 mg/kg, i.p.). In 7-nitroindazole (7-NI; 25 and 50 mg/kg, i.p.)-pretreated animals, malonate (3 mumol) produced decreases in cortical and amygdaloid ChAT activity that were attenuated by both doses of 7-NI. Thus, malonate-induced destruction of the basal forebrain cholinergic neurons was attenuated by systemic pretreatment with lamotrigine, MK-801, and 7-NI but not by L-NA.

Topics: Animals; Dizocilpine Maleate; Enzyme Inhibitors; Indazoles; Lamotrigine; Male; Malonates; Nerve Degeneration; Neurons; Neuroprotective Agents; Nitric Oxide Synthase; Nitroarginine; Parasympathetic Nervous System; Prosencephalon; Rats; Rats, Sprague-Dawley; Triazines

Mechanisms of the modulation of sympathetic activity by neuronal NO were studied in vagotomized anesthetized pigs. Inhibition of neuronal NO synthase (nNOS) within the brain stem by intracerebroventricular (ICV) administration of 7-nitroindazole (7-NI, 1 mmol/L) or S-methyl-L-thiocitrulline (MeTC, 0.1 mmol/L) caused slight increases in renal sympathetic nerve activity (RSNA) but did not affect arterial blood pressure (BP) or cardiac output (CO). However, the sympathoexcitatory effects of glutamate (0.5 mL, 0.1 mol/L ICV) that were associated with marked increases in BP, CO, and heart rate were potentiated by both nNOS inhibitors. Furthermore, 7-NI and MeTC significantly enhanced the responses of RSNA, BP, and CO to activation of somatosympathetic reflexes via stimulation of the left greater sciatic nerve (nervus ischiadicus, 10 to 20 V, 30 Hz, 1-millisecond pulses). Subsequent systemic inhibition of either the neuronal (by 7-NI) or all isoforms of NOS by NG-nitro-L-arginine-methyl ester (20 mg/kg) had no significant additional effects on these responses. The effects of NOS inhibition were effectively counteracted by the endogenous NOS substrate L-arginine and by S-nitroso-N-acetyl-penicillamine (SNAP), a stable analogue of endogenous S-nitroso factors. Disruption of sympathoinhibitory baroreflex mechanisms by bilateral cutting of the carotid sinus nerves caused increases in RSNA and slightly increased responses to all excitatory stimuli but had no effects on the actions of the NOS inhibitors or SNAP. These results suggest that modulation of glutamate effects by nNOS-derived NO may be an important mechanism by which NO affects sympathetic activity in pigs.

Topics: Animals; Arginine; Brain Stem; Citrulline; Enzyme Inhibitors; Glutamic Acid; Heart Rate; Hemodynamics; Indazoles; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Penicillamine; S-Nitroso-N-Acetylpenicillamine; Swine; Sympathetic Nervous System; Thiourea; Vagotomy

Electrical stimulation of cerebellar parallel fibers (PF) increases cerebellar blood flow (BFcrb), a response that is attenuated by glutamate receptor antagonists and NO synthase (NOS) inhibitors. We investigated whether administration of NO donors could counteract attenuation by NOS inhibitors of vasodilation produced by PF stimulation. In halothane-anesthetized rats the cerebellar cortex was exposed and superfused with Ringer solution. PF were stimulated with microelectrodes (100 microA, 30 Hz), and BFcrb was recorded by a laser-Doppler probe. During Ringer superfusion, PF stimulation increased BFcrb by 56 +/- 7% and hypercapnia by 72 +/- 5% (n = 5). Superfusion with the nonselective NOS inhibitor N-nitro-L-arginine (L-NNA, 1 mM) reduced resting BFcrb and attenuated the response to PF stimulation (-47 +/- 5%) and hypercapnia (-46 +/- 7%; PCO2 = 50-60 mmHg). After L-NNA, superfusion with the NO donors 3-morpholinosydnonimine (100 microM, n = 5) or S-nitroso-N-acetyl-penicillamine (5 microM, n = 5) reestablished resting BFcrb (P > 0.05 vs. before L-NNA) and reversed L-NNA-induced attenuation of the response to hypercapnia (P > 0.05 vs. before L-NNA) but not PF stimulation (P > 0.05 vs. after L-NNA). Similar results were obtained when NOS activity was inhibited with the inhibitor of neuronal NOS 7-nitroindazole (50 mg/kg i.p.). Like NO donors, the guanosine 3',5'-cyclic monophosphate analog 8-bromoguanosine 3',5'-cyclic monophosphate (n = 5), administered after L-NNA, restored resting BFcrb and counteracted inhibition of the response to hypercapnia but not PF stimulation. In contrast to NO donors and 8-bromoguanosine 3',5'-cyclic monophosphate, the NO-independent vasodilator papaverine (100 microM, n = 5) had no effect on attenuation of responses to PF stimulation or hypercapnia. Thus NO donors are unable to reverse the effect of NOS inhibition on vasodilation produced by PF stimulation. The data support the hypothesis that the vascular response to PF stimulation, at variance with hypercapnia, requires NOS activation and NO production. Thus NO plays an obligatory role in vasodilation produced by increased functional activity in cerebellar cortex.

Topics: Animals; Blood Pressure; Carbon Dioxide; Cerebellum; Cyclic GMP; Electric Stimulation; Enzyme Inhibitors; Glutamic Acid; Hyperemia; Indazoles; Male; Molsidomine; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oxygen; Papaverine; Partial Pressure; Penicillamine; Rats; Rats, Sprague-Dawley; Regional Blood Flow; S-Nitroso-N-Acetylpenicillamine; Vasodilation

The nitric oxide-arginine pathway is intimately connected to the release of dopamine and glutamate, two neurotransmitter systems that may be dysfunctional in schizophrenia. In addition, nitric oxide synthase inhibitors share several behavioral effects with the psychotomimetic drug, phencyclidine. Previous research has found that phencyclidine-like drugs disrupt prepulse inhibition of the acoustic startle response, an animal model of sensorimotor gating, an attentional process that is disrupted in certain neuropsychiatric disorders in humans (e.g., acute schizophrenia). The purpose of the present study was to examine the effects of nitric oxide modulators in this model. Following injection with a nitric oxide modulator or phencyclidine, rats were placed in startle chambers in which they were exposed to acoustic pulses presented alone or preceded by a prepulse. As in previous reports, phencyclidine disrupted prepulse inhibition at doses that did not affect startle during pulse alone trials. In contrast, the nitric oxide synthase inhibitors, N(G)-nitro-L-arginine (L-NOARG) and N(G)-nitro-L-arginine methyl ester (L-NAME), dose-dependently decreased startle during pulse alone trials, but neither drug affected prepulse inhibition. A nitric oxide precursor, L-arginine, produced similar results. Sodium nitroprusside (a nitric oxide releaser) and 7-nitroindazole (a third nitric oxide synthase inhibitor) did not affect startle amplitudes during pulse alone or prepulse + pulse trials. The present results suggest that modulation of nitric oxide synthesis or availability does not disrupt sensorimotor gating of the acoustic startle response and is probably not involved in mediation of this type of attentional deficit in humans.

Topics: Acoustic Stimulation; Animals; Arginine; Excitatory Amino Acid Antagonists; Indazoles; Male; Neural Inhibition; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Nitroprusside; Phencyclidine; Rats; Rats, Sprague-Dawley; Reflex, Startle

We tested the hypothesis that nitric oxide (NO) plays a role in CBF autoregulation in the brain stem during hypotension. In anesthetized rats, local CBF to the brain stem was determined with laser-Doppler flowmetry, and diameters of the basilar artery and its branches were measured through an open cranial window during stepwise hemorrhagic hypotension. During topical application of 10(-5) mol/L and 10(-4) mol/L N(omega)-nitro-L-arginine (L-NNA), a nonselective inhibitor of nitric oxide synthase (NOS), CBF started to decrease at higher steps of mean arterial blood pressure in proportion to the concentration of L-NNA in stepwise hypotension (45 to 60 mm Hg in the 10(-5) mol/L and 60 to 75 mm Hg in the 10(-4) mol/L L-NNA group versus 30 to 45 mm Hg in the control group). Dilator response of the basilar artery to severe hypotension was significantly attenuated by topical application of L-NNA (maximum dilatation at 30 mm Hg: 16 +/- 8% in the 10(-5) mol/L and 12 +/- 5% in the 10(-4) mol/L L-NNA group versus 34 +/- 4% in the control group), but that of the branches was similar between the control and L-NNA groups. Topical application of 10(-5) mol/L 7-nitro indazole, a selective inhibitor of neuronal NOS, did not affect changes in CBF or vessel diameter through the entire pressure range. Thus, endothelial but not neuronal NO seems to take part in the regulation of CBF to the the brain stem during hypotension around the lower limits of CBF autoregulation. The role of NO in mediating dilatation in response to hypotension appears to be greater in large arteries than in small ones.

Topics: Animals; Basilar Artery; Blood Flow Velocity; Brain Stem; Enzyme Inhibitors; Hypotension; Indazoles; Laser-Doppler Flowmetry; Male; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Sprague-Dawley; Vasodilation

The involvement of nitric oxide in anoxia-induced long-term potentiation (anoxic LTP) of synaptic transmission was investigated in CA1 neurons of rat hippocampal slices using intracellular recording techniques in vitro. In response to superfusion of an anoxic artificial cerebral spinal fluid saturated with 95% N2-5% CO2, the excitatory postsynaptic potential (EPSP) generated in hippocampal CA1 neurons by stimulation of the Schaffer collateral/commissural afferent pathway was completely abolished within 10 min of anoxia. On return to reoxygenated medium, the EPSP returned to the control value within 10 min and was subsequently and progressively potentiated to reach a plateau 15-20 min after return to oxygen. This anoxia-induced persistent increase in synaptic transmission lasted for more than 1 h. Application of the nitric oxide synthase inhibitors 7-nitroindazole (7-NI) or L-N(G)-nitroarginine (NOARG) produced no effects on the baseline EPSP amplitude, but effectively attenuated the anoxic LTP. The inhibitory effects of both 7-NI and NOARG on the anoxic LTP were blocked by L-arginine, a substrate for nitric oxide synthase. These results suggest that nitric oxide is required for the generation of anoxia-induced LTP of glutamatergic synaptic transmission in the CA1 region of the rat hippocampus.

Topics: Animals; Arginine; Enzyme Inhibitors; Excitatory Postsynaptic Potentials; Hippocampus; Hypoxia; In Vitro Techniques; Indazoles; Long-Term Potentiation; Male; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Sprague-Dawley; Reaction Time; Signal Transduction; Synaptic Transmission

Rat cerebellar nitric oxide synthase (NOS) purified from transfected human kidney cells catalyzes an NADPHdependent reduction of quinonoid dihydrobiopterin (qBH2) to tetrahydrobiopterin (BH4). Reduction of qBH2 at 25 microM proceeds at a rate that is comparable with that of the overall reaction (citrulline synthesis) and requires calcium ions and calmodulin for optimal activity; NADH has only 10% of the activity of NADPH. The reduction rate with the quinonoid form of 6-methyldihydropterin is approximately twice that with qBH2. 7,8-Dihydrobiopterin had negligible activity. Neither 7,8-dihydrobiopterin nor BH4 affected the rate of qBH2 reduction. Reduction is inhibited by the flavoprotein inhibitor diphenyleneiodonium, whereas inhibitors of electron transfer through heme (7-nitroindazole and N-nitroarginine) stimulated the rate to a small extent. Methotrexate, which inhibits a variety of enzymes catalyzing dihydrobiopterin reduction, did not inhibit. These studies provide the first demonstration of the reduction of qBH2 to BH4 by NOS and indicate that the reduction is catalyzed by the flavoprotein "diaphorase" activity of NOS. This activity is located on the reductase (C-terminal) domain, whereas the high affinity BH4 site involved in NOS activation is located on the oxygenase (N-terminal) domain. The possible significance of this reduction of qBH2 to the essential role of BH4 in NOS is discussed.

Topics: Animals; Arginine; Biopterins; Cell Line; Cerebellum; Enzyme Inhibitors; Humans; Indazoles; Kidney; Kinetics; Methotrexate; NADP; Nitric Oxide Synthase; Nitroarginine; Onium Compounds; Oxidation-Reduction; Rats; Recombinant Proteins; Transfection

Nitric oxide (NO) is produced by enzymes called nitric oxide synthases (NOS). At least three different isoforms of NOS have been identified in the kidney. This study examines the effects of selective inhibition of the inducible isoform (iNOS) and the neuronal isoform (bNOS) on the glomerular capillary pressure (PGC), through studies of the tubuloglomerular feedback (TGF) mechanism in anaesthetized rats. The proximal tubular stop-flow pressure (PSF) was measured to estimate changes in PGC obtained after activation of the TGF system by varying the loop of Henle perfusion rate with artificial ultrafiltrate including vehicle, NOS inhibition or L-arginine. Infusion of nonspecific NOS inhibition (N omega-Nitro-L-arginine) increased maximal TGF responses (delta PSF) by 84% and L-arginine decreased delta PSF by 37%. Aminoguanidine, a selective iNOS-inhibitor, failed to increase delta PSF, whereas the nonspecific NOS inhibitor methylguanidine increased delta PSF by 64%. 7-Nitro indazole (7-NI), a selective bNOS inhibitor, increased delta PSF by 57% when infused intratubularly, and intraperitoneal administration of 7-NI increased delta PSF by 78%, without any change in blood pressure. Since bNOS is exclusively located in the macula densa (MD) cells, these results confirm and strengthen the obligatory role of MD-produced NO in regulation of TGF and PGC, which has been suggested earlier. iNOS, widely expressed in the kidney, does not seem to play any important role in regulation of PGC.

Topics: Animals; Arginine; Capillary Resistance; Enzyme Inhibitors; Guanidines; Indazoles; Juxtaglomerular Apparatus; Kidney Glomerulus; Kidney Tubules, Proximal; Male; Methylguanidine; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pressure; Rats; Rats, Sprague-Dawley; Time Factors; Vasoconstriction

Inhibition of nitric oxide (NO) synthesis attenuates the hypercapnic cerebrovasodilation or the increases in cerebral blood flow (CBF) produced by acetylcholine (ACh), either topically applied or endogenously released in neocortex by stimulation of the basal forebrain cholinergic system. We investigated whether exogenous administration of NO, using NO donors, can reverse the attenuation of these responses by NO synthase (NOS) inhibitors. In halothane-anesthetized, ventilated rats the frontoparietal cortex was exposed and superfused with Ringer. CBF was monitored at the super fusion site by laser-Doppler flowmetry. The basal forebrain was stimulated (100 microA; 50 Hz) with microelectrodes stereotaxically implanted. Superfusion with the NOS inhibitor NG-nitro-L-arginine (L-NNA; 1 mM) reduced resting CBF (-38 +/- 2%; mean +/- SE) and attenuated the vasodilation elicited by hypercapnia (Pco2, 50-60 mmHg; -79 +/- 3%), ACh (10 microM; -83 +/- 7%), or basal forebrain stimulation (-44 +/- 2%) (P < 0.05, analysis of variance and Tukey's test). After L-NNA, topical application of 3-morpholinosydnonimine (SIN-1) (n = 7), S-nitroso-N-acetylpenicillamine (SNAP) (n = 6), or 8-bromoguanosine 3',5'-monophosphate (8-BrcGMP, n = 4) reestablished resting CBF (P > 0.05 from Ringer) and reversed the attenuation of the response to hypercapnia (P > 0.05 from Ringer). However, SIN-1 or SNAP failed to reverse the attenuation of the response to basal forebrain stimulation or topical ACh (P > 0.05 from L-NNA). After L-NNA, the NO-independent vasodilator papaverine (n = 4) reestablished resting CBF (P > 0.05 from Ringer) but failed to restore the hypercapnic vasodilation (P > 0.05 from L-NNA). The attenuation of hypercapnic response by the neuronal NOS inhibitor 7-nitroindazole was counteracted only partially by SIN-1 (n = 4) or 8-BrcGMP (n = 4). The data support the hypothesis that the vasodilation elicited by hypercapnia requires resting levels of NO for its expression, whereas the response to endogenous or exogenous ACh depends on agonist-induced NOS activation. In hypercapnia NO may act as a permissive factor by facilitating the action of other vasodilators, whereas in the vascular response initiated by ACh NO is likely to be the major mediator of smooth muscle relaxation.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Acetylcholine; Animals; Cerebrovascular Circulation; Dose-Response Relationship, Drug; Electric Stimulation; Enzyme Inhibitors; Hypercapnia; Indazoles; Male; Molsidomine; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Papaverine; Prosencephalon; Rats; Rats, Sprague-Dawley; Vasodilation

The present behavioral study was undertaken to investigate whether neuronal nitric oxide (NO) synthase mediates the abnormal consequences of increased NMDA receptor-mediated synaptic transmission in models of postural tremor, Parkinson's disease and epilepsy. We used 7-nitroindazole, a selective inhibitor of neuronal NO synthase, and NG-nitro-L-arginine (L-NAME), an unspecific NO synthase inhibitor, and compared their action with that of the competitive NMDA receptor antagonist 3-[(R)-2-carboxypiperazin-4-yl]-prop-2-enyl-1-phosphonic acid (D-CPPene). In both mice and rats, 7-nitroindazole, L-NAME and D-CPPene dose dependently reversed the harmaline-induced increase of cerebellar cyclic guanosine-5'-monophosphate (cGMP) levels. For subsequent behavioral experiments we used doses of 7-nitroindazole, L-NAME and D-CPPene which were equipotent in preventing harmaline-induced cGMP increase. Harmaline-induced tremor in mice and rats was suppressed by D-CPPene, but not by 7-nitroindazole or by L-NAME. This effect of D-CPPene was not due to unspecific suppression of motor activity, since D-CPPene did not affect locomotor activity at doses which reduced tremor. D-CPPene, but not 7-nitroindazole and L-NAME potentiated the antiparkinsonian action of the dopamine agonist lisuride in rats with unilateral 6-hydroxydopamine lesions of the substantia nigra. D-CPPene antagonized seizures induced by intracerebroventricular injection of NMDA in mice. In contrast, 7-nitroindazole and L-NAME had only a tendency to prevent seizures and to delay the latency to onset of seizures. We conclude from these results that neuronal NO synthase does not serve as a major mediator of increased NMDA receptor-mediated synaptic transmission in animal models of Parkinson's disease, postural tremor and epilepsy. The novel observation that D-CPPene suppresses harmaline-induced tremor leads us to suggest that NMDA receptor antagonists should be considered as novel therapeutics for postural tremor.

Topics: Animals; Anticonvulsants; Behavior, Animal; Brain; Cyclic GMP; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Harmaline; Hydroxydopamines; Indazoles; Lisuride; Locomotion; Male; N-Methylaspartate; Nitric Oxide Synthase; Nitroarginine; Piperazines; Rats; Receptors, N-Methyl-D-Aspartate; Seizures

We examined whether attenuation of the hypercapnic increase of cerebral blood flow (CBF) associated with nitric oxide synthase (NOS) inhibition is related to local neuronal or aortic endothelial NOS activity or local endothelial/neuronal NOS-dependent vasodilation. Halothane-anesthetized rats were ventilated, and CBF was measured by laser-Doppler flowmetry over the parietal and cerebellar cortex. Intravenous N omega-nitro-L-arginine (L-NNA; 30 mg/kg) inhibited brain and aortic NOS activity by 67-70%. Topical L-NNA (1 mM) inhibited brain NOS activity by 91-94%, whereas aortic NOS activity remained constant. In contrast, intravenous L-NNA attenuated the hypercapnic CBF rise much more efficiently than topical L-NNA. 7-Nitroindazole, another NOS inhibitor, attenuated endothelial and neuronal NOS activity equally well and inhibited the hypercapnic CBF increase as effectively as L-NNA. Topical L-NNA and 7-nitroindazole abolished local endothelial NOS-dependent vasodilation after 15 min, whereas hypercapnic CBF was only slightly reduced. L-NNA injected into the tissue abolished neuronal NOS-dependent vasodilation, whereas hypercapnic CBF was unchanged. The findings suggest that local NOS activity, whether neuronal or endothelial, is unimportant for the hypercapnic rise of CBF.

Topics: Acetylcholine; Administration, Topical; Animals; Aorta; Brain; Carbon Dioxide; Cerebrovascular Circulation; Endothelium, Vascular; Enzyme Inhibitors; Fluorescent Antibody Technique; Hypercapnia; Indazoles; Injections, Intravenous; Male; Nitric Oxide Synthase; Nitroarginine; Rats; Rats, Wistar

L-Glutamate and norepinephrine are examples of a major excitatory neurotransmitter and a neuromodulator in the cerebral cortex, respectively. Little is known of how chemical signaling between the anatomically distinct chemical pathways occurs. Specific activation of the N-methyl-D-aspartate (NMDA) class of glutamate receptor in synaptosomal preparations from guinea pig cerebral cortex caused release of both of these chemicals, and this release was blocked by agents that inhibit nitric oxide (NO) production or remove NO from the extracellular space. Furthermore, neurotransmitter release correlated with cortical NO production after NMDA receptor stimulation. These results suggest that NO production and its extracellular movement may be links in the pathway from NMDA receptor activation to changes in chemical signaling in surrounding synaptic terminals in the cerebral cortex.

Topics: Amino Acid Oxidoreductases; Animals; Arginine; Cerebral Cortex; Glutamates; Glutamic Acid; Guinea Pigs; In Vitro Techniques; Indazoles; Male; N-Methylaspartate; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Norepinephrine; omega-N-Methylarginine; Potassium Chloride; Receptors, N-Methyl-D-Aspartate; Synaptosomes

The nitric oxide (NO) synthase inhibitor N omega-nitro-L-arginine (NNA) and the putative brain-selective NO synthase inhibitor 7-nitroindazole (7-NI) were used to determine the role of endogenous NO on seizures induced by kainic acid (KA) in rats and KA, pilocarpine, bicuculline, picrotoxin and pentylenetetrazole (PTZ) in mice. Rats given a subconvulsant dose of KA (6 mg/kg, i.p.) had seizures after they had been pretreated with NNA (50 mg/kg, i.p.). With a higher dose of KA (12 mg/kg, i.p.), NNA caused an increase in wild running seizures and mortality. Unlike NNA, 7-NI had no effect on KA-induced seizures. Similarly, NNA but not 7-NI caused a worsening of seizures in mice measured as a shortening of seizure latency and an increase in wild running and mortality. The effect of NNA on seizure latency was completely reversed by the competitive substrate for NO synthase, L-arginine. NNA had no effect on seizure latency following any of the other convulsants and increased mortality following pilocarpine and picrotoxin alone. Our results indicate that NNA may enhance the severity of KA-induced seizures through suppression of NO synthase activity in the vascular endothelium. The resulting impairment of cerebrovascular autoregulation may cause a mismatch between metabolic demand and blood flow during seizures leading to facilitation of spread. The absence of a comparable effect of NNA on other seizure models may indicate differences in the degree to which seizure activity in different models is influenced by the metabolic impairment secondary to decreased blood flow.

Topics: Amino Acid Oxidoreductases; Animals; Arginine; Bicuculline; Cerebrovascular Circulation; Enzyme Inhibitors; Indazoles; Kainic Acid; Male; Mice; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pentylenetetrazole; Rats; Rats, Sprague-Dawley; Seizures

Topics: Acetylcholine; Adenosine Triphosphate; Amino Acid Oxidoreductases; Analgesics; Animals; Arginine; Blood Pressure; Blood Transfusion; Brain; Cats; Cerebral Arteries; Cerebral Cortex; Cerebrovascular Circulation; Electroencephalography; Hemorrhage; Hypotension; Indazoles; Male; Molsidomine; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Norepinephrine; Organ Specificity; Parietal Lobe; Pituitary Gland; Regional Blood Flow; Spinal Cord; Vasodilator Agents

7-Nitro-indazole (7-NI) has been described as novel nitric oxide synthase (NOS) inhibitor with in vivo selectivity for the neuronal isozyme [Moore et al. Br. J. Phaarmac. 110, 219-224 (1993)]. In the present study we have used purified porcine brain NOS to investigate the molecular mechanisms of enzyme inhibition by 7-NI. The drug was competitive with L-arginine, exhibited a kinetic KI of 2.8 microM, and additionally induced a slight reduction in Vmax. As a cytochrome P-450, NOS catalyzes a heme-mediated reduction of molecular oxygen, resulting in the formation of H2O2 in the absence of L-arginine. 7-NI turned out as a potent inhibitor of H2O2 formation (IC50 = 0.28 +/- 0.096 microM) but did not affect flavin-mediated electron transfer. Thus, 7-NI resembled imidazole, a known heme-site inhibitor of NOS. We found that imidazole was a purely competitive inhibitor of L-citrulline formation (KI = 263 microM) and blocked H2O2 formation at similar concentrations (IC50 = 280 +/- 38 microM). In accordance with their L-arginine-competitive effects in the citrulline assay, both drugs antagonized binding of radiolabeled NG-nitro-L-arginine (L-NNA), a high affinity probe for reversible labelling of the substrate site of NOS [Klatt et al., J. Biol. Chem. 269, 14781-14787 (1994)]. The calculated KI values for 7-NI and imidazole were 0.09 +/- 0.024 microM and 200 +/- 63 microM, respectively. Finally, binding of radiolabelled tetrahydrobiopterin, a NOS cofactor with unknown function, was also antagonized by 7-NI with a KI of 0.12 +/- 0.023 microM.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Amino Acid Oxidoreductases; Analgesics; Animals; Arginine; Binding, Competitive; Biopterins; Brain; Citrulline; Enzyme Inhibitors; Hydrogen Peroxide; Indazoles; Kinetics; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Protein Binding; Swine