piperidines and hemopressin

The endocannabinoid system (ECS) is activated when natural arachidonic acid derivatives (endogenous cannabinoids or endocannabinoids) bind as lipophilic messengers to cannabinoid receptors CB1 and CB2. The ECS comprises many hydrolytic enzymes responsible for the endocannabinoids cleavage. These hydrolases, such as fatty acid amide hydrolase (FAAH) and monoacylglyceride lipase (MAGL), are possible therapeutic targets for the development of new drugs as indirect cannabinoid agonists. Recently, a new family of endocannabinoid modulators was discovered; the lead structure of this family is the nonapeptide hemopressin produced from enzymatic cleavage of the α-chain of hemoglobin and acting as negative allosteric modulator of CB1. Hemopressin shows several physiological effects, e.g., antinociception, hypophagy, and hypotension. However, it is still a matter of debate whether this peptide, isolated from the brain of rats, is a real neuromodulator of the ECS. Recent evidence indicates that hemopressin could be a by-product formed by chemical degradation of a longer peptide RVD-hemopressin during the extraction from the brain homolysate. Indeed, RVD-hemopressin is more active than hemopressin in certain biological tests and may bind to the same subsite as Rimonabant, which is an inverse agonist of CB1 and a μ-opioid receptor antagonist. These findings have stimulated several studies to verify this hypothesis and to evaluate possible therapeutic applications of hemopressin, its peptidic derivatives, and synthetic analogues, opening new perspectives to the development of novel cannabinoid drugs.

Topics: Analgesics; Animals; Cannabinoids; Endocannabinoids; Hemoglobins; Hypotension; Peptide Fragments; Piperidines; Protein Binding; Pyrazoles; Rats; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Receptors, Opioid, mu; Rimonabant

Alzheimer's disease (AD) is a serious neurodegenerative disease. Senile plaques (SPs) composed of amyloid-β (Aβ) are typical features of AD. Aβ plays a key role in the disease and has the ability to induce other pathological characteristics of AD, including oxidative stress injury. (m)VD-hemopressin (VD), a peptide derived from mouse brain extracts, can bind cannabinoid 1 receptor (CB1R) as an agonist. Our previous report indicated that VD reverses memory impairment induced by Aβ1-42 in mice. This study aimed to clarify the mechanism by which VD protects hippocampal neurons against Aβ1-42-induced impairment. Our results showed that VD inhibited oxidative stress injury induced by Aβ1-42, as demonstrated by the VD-induced reversal of the upregulation of reactive oxygen species (ROS) and the intracellular lipid peroxidation product malondialdehyde (MDA) and the downregulation of the activities of the antioxidative enzymes catalase (CAT) and glutathione peroxidase (GSH-PX) in mouse hippocampal neurons. We also found that VD restored the decrease in cell growth and viability induced by Aβ1-42 and reversed Aβ1-42-induced apoptosis mediated by the apoptosis-associated proteins Bcl-2 and Bax. However, cotreatment with AM251 (an antagonist of CB1R) blocked the effects of VD. In brief, this study suggested that through CB1R, VD reversed the impairment of cell growth and viability, oxidative stress injury and apoptosis induced by Aβ1-42. Therefore, VD may be a promising agent for the treatment of diseases that involve oxidative stress injury and apoptosis induced by Aβ1-42, such as AD.

Topics: Amyloid beta-Peptides; Animals; Apoptosis; bcl-2-Associated X Protein; Catalase; Cell Survival; Cells, Cultured; Hemoglobins; Hippocampus; Malondialdehyde; Mice, Inbred C57BL; Neurons; Neuroprotective Agents; Oxidative Stress; Peptide Fragments; Piperidines; Pyrazoles; Reactive Oxygen Species; Receptor, Cannabinoid, CB1

The cannabinoid system plays an important role in memory processes, many studies have indicated that cannabinoid receptor ligands have ability to modulate memory in rodents. A nonapeptide hemopressin (Hp) derived from rat brain, acts as a peptide antagonist or selective inverse peptide agonist of cannabinoid 1 (CB1) receptor. N-terminally extended forms of Hp isolated from mouse brain, (m)RVD-hemopressin(α) (RVD) and (m)VD-hemopressin(α) (VD) also bind CB1 receptor, however, as peptide agonists. Here, we investigated the roles of Hp, RVD, and VD on memory in mice using novel object recognition (NOR) and object location recognition (OLR) tasks. In normal young mice, intracerebroventricular (i.c.v.) infusion of Hp before training not only improved memory formation, but also prolonged memory retention in the tasks, these effects could be inhibited by RVD or VD at the same dose and intraperitoneal (i.p.) injection of a small molecule agonist of CB1 receptor WIN55, 212-2 15min before administration of Hp inhibited the memory-improving effect of Hp. In addition, under the same experimental conditions, i.c.v. RVD or VD displayed memory-impairing effects, which could be prevented by Hp (i.c.v.) or AM251 (i.p.), a small molecule antagonist of CB1 receptor. Infusion of amyloid-β (1-42) (Aβ1-42) 14days before training resulted in impairment of memory in mice which could be used as animal model of Alzheimer's disease (AD). In these mice, RVD or VD (i.c.v.) reversed the memory impairment induced by Aβ1-42, and the effects of RVD and VD could be suppressed by Hp (i.c.v.) or AM251 (2mg/kg, i.p.). Separate administration of Hp had no effect in Aβ1-42-treated mice. The above results suggested that Hp, RVD and VD, as CB1 receptor peptide ligands, may be potential drugs to treatment of the memory deficit-involving disease, just as AD.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Behavior, Animal; Benzoxazines; Cannabinoid Receptor Agonists; Cannabinoid Receptor Antagonists; Disease Models, Animal; Hemoglobins; Infusions, Intraventricular; Male; Memory Disorders; Mice; Morpholines; Naphthalenes; Oligopeptides; Peptide Fragments; Piperidines; Pyrazoles; Receptor, Cannabinoid, CB1; Recognition, Psychology

Hemopressin (PVNFKFLSH; HP) is an orally active peptide derived from rat hemoglobin α-chain that could act as an inverse agonist at cannabinoid type 1 receptors (CB1). Here, we aim to investigate possible behavioral effects of HP in male Wistar rats tested in the elevated plus maze (EPM), following HP intraperitoneal (i.p., 0.05 mg/kg), oral (P.O., 0.05 and 0.5 mg/kg) or intracerebroventricular (I.C.V., 3 and 10 nmol) administration. HP induced a decrease in EPM open arm exploration, indicating an anxiogenic-like effect. However, i.p. administration of HP (1 mg/kg) followed by mass spectrometry analysis of brain-peptide extracts suggested that the intact HP does not cross the blood brain barrier. I.C.V. administrated HP produced anxiogenic-like effects that were prevented by Transient Receptor Potential Vanilloid Type 1 (TRPV1) antagonists, 6-iodonordihydrocapsaicin (1 nmol) or SB366791 (1 nmol), but not by the CB1 receptor antagonist AM251 (0.1 and 1 nmol). Altogether, these data suggest that I.C.V. administrated HP induces anxiogenic-like effects by activating TRPV1 receptors. The similar anxiogenic effects observed after i.p. or P.O. administration could be due to HP fragment(s) crossing the blood brain barrier. The present results advance our knowledge about HP pharmacology and suggest concerns in future clinical studies.

Topics: Anilides; Animals; Anxiety; Brain; Capsaicin; Cinnamates; Hemoglobins; Injections, Intraventricular; Male; Maze Learning; Peptide Fragments; Piperidines; Pyrazoles; Rats; Rats, Wistar

The function of the endocannabinoid system (ECS) in renal tissue is not completely understood. Kidney function is closely related to ion reabsorption in the proximal tubule, the nephron segment responsible for the re-absorption of 70-80% of the filtrate. We studied the effect of compounds modulating the activity of cannabinoid (CB) receptors on the active re-absorption of Na(+) in LLC-PK1 cells.. Changes in Na(+) /K(+) -ATPase activity were assessed after treatment with WIN55,212-2 (WIN), a non-selective lipid agonist, and haemopressin (HP), an inverse peptide agonist at CB1 receptors. Pharmacological tools were used to investigate the signalling pathways involved in the modulation of Na(+) transport.. In addition to CB1 and CB2 receptors and TRPV1 channels, the mRNAs encoding for enzymes of the ECS were also expressed in LLC-PK1. WIN (10(-7)  M) and HP (10(-6)  M) altered Na(+) re-absorption in LLC-PK1 in a dual manner. They both acutely (after 1 min) increased Na(+) /K(+) -ATPase activity in a TRPV1 antagonist-sensitive way. WIN's stimulating effect persisted for 30 min, and this effect was partially blocked by a CB1 antagonist or a PKC inhibitor. In contrast, HP inhibited Na(+) /K(+) -ATPase after 30 min incubation, and this effect was attenuated by a CB1 antagonist or a PKA inhibitor.. The ECS is expressed in LLC-PK1 cells. Both CB1 receptors and TRPV1 channels regulate Na(+) /K(+) -ATPase activity in these cells, and are modulated by lipid and peptide CB1 receptor ligands, which act via different signalling pathways.

Topics: Animals; Benzoxazines; Biological Transport; Cyclic AMP; Endocannabinoids; Hemoglobins; Kidney; LLC-PK1 Cells; Morpholines; Naphthalenes; Peptide Fragments; Piperidines; Pyrazoles; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Signal Transduction; Sodium; Sodium-Potassium-Exchanging ATPase; Swine; TRPV Cation Channels

Hemopressin is the first peptide ligand to be described for the CB₁ cannabinoid receptor. Hemopressin acts as an inverse agonist in vivo and can cross the blood-brain barrier to both inhibit appetite and induce antinociception. Despite being highly effective, synthetic CB₁ inverse agonists are limited therapeutically due to unwanted, over dampening of central reward pathways. However, hemopressin appears to have its effect on appetite by affecting satiety rather than reward, suggesting an alternative mode of action which might avoid adverse side effects. Here, to resolve the neuronal circuitry mediating hemopressin's actions, we have combined blood-oxygen-level-dependent, pharmacological-challenge magnetic resonance imaging with c-Fos functional activity mapping to compare brain regions responsive to systemic administration of hemopressin and the synthetic CB₁ inverse agonist, AM251. Using these complementary methods, we demonstrate that hemopressin activates distinct neuronal substrates within the brain, focused mainly on the feeding-related circuits of the mediobasal hypothalamus and in nociceptive regions of the periaqueductal grey (PAG) and dorsal raphe (DR). In contrast to AM251, there is a distinct lack of activation of the brain reward centres, such as the ventral tegmental area, nucleus accumbens and orbitofrontal cortex, which normally form a functional activity signature for the central action of synthetic CB₁ receptor inverse agonists. Thus, hemopressin modulates the function of key feeding-related brain nuclei of the mediobasal hypothalamus, and descending pain pathways of the PAG and DR, and not higher limbic structures. Thus, hemopressin may offer behaviourally selective effects on nociception and appetite, without engaging reward pathways.

Topics: Animals; Appetite Depressants; Behavior, Animal; Cannabinoids; Hemoglobins; Hypothalamus, Middle; Injections, Intraperitoneal; Male; Mice; Mice, Knockout; Nerve Tissue Proteins; Neurons; Peptide Fragments; Periaqueductal Gray; Piperidines; Pyrazoles; Random Allocation; Raphe Nuclei; Rats; Rats, Sprague-Dawley; Receptor, Cannabinoid, CB1; Satiety Response

The current study evaluated the effects of hemopressin (HP) on pain modulation by endokinin A/B (EKA/B) and endokinin C/D (EKC/D) at the supraspinal level in mice. Intracerebroventricular administration of HP (10 nmol) fully antagonized the hyperalgesia induced by EKA/B (10, 30, and 100 pmol), and induced a dose-dependent potent analgesic effect. HP at different concentrations (10 pmol, 100 pmol, and 1 nmol) showed varying effects on the analgesic effect of EKA/B (3 nmol). HP extended the duration of the analgesic effect of EKC/D (3 nmol). Moreover, HP at different concentrations (10 pmol, 5 pmol, 1 pmol, and 100 fmol) co-administered with EKC/D (30 pmol) induced significant analgesia at two different time points: 5 min and 50 min. To investigate the antinociceptive mechanism, we used SR140333B and SR142801. HP (1 pmol) potentiated the analgesic effect of SR140333B (100 pmol)+EKA/B (30 pmol) in 5-10 min, while HP (100 pmol) had no effect in the analgesia induced by SR140333B (3 nmol)+EKA/B (3 nmol). HP (1 nmol) fully inhibited the analgesic effect of SR140333B (3 nmol)+EKC/D (3 nmol) or SR142801 (3 nmol)+EKC/D (3 nmol). HP (1 pmol) weakened the analgesic effect of SR142801 (100 pmol)+EKA/B (30 pmol), but HP (100pmol) strengthened the analgesic effect of SR142801 (3 nmol)+EKA/B (3 nmol). These findings may pave the way for a new strategy on investigating the interaction between tachykinins and opioids on pain modulation.

Topics: Analgesics; Animals; Dose-Response Relationship, Drug; Hemoglobins; Hyperalgesia; Injections, Intraventricular; Male; Mice; Neurokinin-1 Receptor Antagonists; Peptide Fragments; Piperidines; Receptors, Neurokinin-3; Tachykinins; Tropanes

Hemopressin is a short, nine amino acid peptide (H-Pro-Val-Asn-Phe-Lys-Leu-Leu-Ser-His-OH) isolated from rat brain that behaves as an inverse agonist at the cannabinoid receptor CB(1), and is shown here to inhibit agonist-induced receptor internalization in a heterologous cell model. Since this peptide occurs naturally in the rodent brain, we determined its effect on appetite, an established central target of cannabinoid signaling. Hemopressin dose-dependently decreases night-time food intake in normal male rats and mice, as well as in obese ob/ob male mice, when administered centrally or systemically, without causing any obvious adverse side effects. The normal, behavioral satiety sequence is maintained in male mice fasted overnight, though refeeding is attenuated. The anorectic effect is absent in CB(1) receptor null mutant male mice, and hemopressin can block CB(1) agonist-induced hyperphagia in male rats, providing strong evidence for antagonism of the CB(1) receptor in vivo. We speculate that hemopressin may act as an endogenous functional antagonist at CB(1) receptors and modulate the activity of appetite pathways in the brain.

Topics: Analysis of Variance; Animals; Behavior, Animal; Benzoxazines; Chlorocebus aethiops; Circadian Rhythm; COS Cells; Cyclohexanols; Dose-Response Relationship, Drug; Drinking Behavior; Dronabinol; Drug Administration Routes; Eating; Food Deprivation; Green Fluorescent Proteins; Hemoglobins; Hyperphagia; Leptin; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Morpholines; Naphthalenes; Peptide Fragments; Piperidines; Protein Transport; Psychotropic Drugs; Pyrazoles; Rats; Rats, Sprague-Dawley; Receptor, Cannabinoid, CB1; Rimonabant; Time Factors; Transfection