guanosine-5--o-(3-thiotriphosphate) and Hypothermia

Rat lines that were selectively bred for high (high DPAT-sensitive, HDS) or low (low DPAT-sensitive, LDS) hypothermic responses to the specific 5-HT(1A) receptor agonist, 8-hydroxy-di-n-propylaminotetralin (8-OH-DPAT), differ in receptor binding and certain behaviors related to anxiety and depression. After reviewing this literature, the present communication summarizes new experiments designed to clarify and extend the nature of the pharmacological and biochemical differences between the lines. A challenge with the 5-HT(2) receptor agonist, DOI, produced similar degrees of head shakes and skin crawls in the HDS and LDS rats, suggesting similar sensitivity of 5-HT(2A) and 5-HT(2C) receptors. In contrast, DOI-induced flat body posture (FBP), which has been linked to 5-HT(1A) receptor stimulation, was observed more readily in the HDS rats. The HDS and LDS rats exhibited similar degrees of increase in 8-OH-DPAT-stimulated [35S]GTPgammaS binding in several brain regions. This result suggests that the dramatic differences in hypothermia in HDS and LDS rats cannot be related to 5-HT(1A) receptor-mediated action on G proteins. Overall, these findings indicate that the selective breeding for 5-HT(1A)-mediated hypothermia has been fairly selective, and that differences in emotionally relevant behaviors between these two rat lines can strongly be associated with an unidentified component of the 5-HT(1A) receptor signaling pathway.

Topics: 8-Hydroxy-2-(di-n-propylamino)tetralin; Amphetamines; Animals; Behavior, Animal; Breeding; Emotions; Guanosine 5'-O-(3-Thiotriphosphate); Hypothermia; Mice; Mice, Knockout; Rats; Rats, Mutant Strains; Receptors, Serotonin; Receptors, Serotonin, 5-HT1; Serotonin Antagonists; Serotonin Receptor Agonists

Preclinical studies support a critical role of 5-HT

Topics: 8-Hydroxy-2-(di-n-propylamino)tetralin; Anhedonia; Animals; Antidepressive Agents, Second-Generation; Anxiety; Autoreceptors; Axl Receptor Tyrosine Kinase; Brain-Derived Neurotrophic Factor; Depression; Fluoxetine; Guanosine 5'-O-(3-Thiotriphosphate); Hippocampus; Hypothermia; Mice; Mice, Knockout; Neuronal Plasticity; Olfactory Bulb; Proto-Oncogene Proteins; Receptor Protein-Tyrosine Kinases; Receptor, Serotonin, 5-HT1A; Receptor, trkB; Receptors, Serotonin, 5-HT4

The purpose of this study was to examine the role of the prodynorphin gene in alcohol sensitivity, preference and vulnerability to alcohol consumption. Handling-induced convulsion (HIC) associated to alcohol, alcohol-induced loss of righting reflex (LORR), hypothermic effects in response to acute ethanol challenge, blood ethanol levels (BELs), conditioned place preference, voluntary ethanol consumption and preference, tyrosine hydroxylase (TH), dopamine transporter (DAT) and proenkephalin (PENK) gene expression, and µ-, δ- and κ-opioid agonist-stimulated [S(35) ]- guanosine 5'-triphosphate-binding autoradiography were studied in prodynorphin knockout (PDYN KO) and wild-type (WT) mice. There were no differences in HIC, LORR or the decrease in body temperature in response to acute ethanol challenge between PDYN KO and WT mice. PDYN KO mice presented higher BEL, higher ethanol-conditioned place preference and more ethanol consumption and preference in a two-bottle choice paradigm than WT mice. These findings were associated with lower TH and higher DAT gene expression in the ventral tegmental area and substantia nigra, and with lower PENK gene expression in the caudate-putamen (CPu), accumbens core (AcbC) and accumbens shell (AcbSh) in PDYN KO. The functional activity of the µ-opioid receptor was lower in the CPu, AcbC, AcbSh and cingulate cortex (Cg) of PDYN KO mice. In contrast, δ- and κ-opioid receptor-binding autoradiographies were increased in the CPu and Cg (δ), and in the CPu, AcbC and Cg (κ) of PDYN KO. These results suggest that deletion of the PDYN gene increased vulnerability for ethanol consumption by altering, at least in part, PENK, TH and DAT gene expression, and µ-, δ- and κ-opioid receptor functional activity in brain areas closely related to ethanol reinforcement.

Topics: Alcohol Drinking; Animals; Autoradiography; Basal Ganglia; Body Temperature; Dopamine; Dopamine Plasma Membrane Transport Proteins; Enkephalins; Ethanol; Gene Expression; Guanosine 5'-O-(3-Thiotriphosphate); Hypothermia; In Situ Hybridization; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Nucleus Accumbens; Protein Precursors; Receptors, Opioid; Reflex, Righting; Reinforcement, Psychology; Reward; Tyrosine 3-Monooxygenase; Unconsciousness

Cannabinoid CB(1) receptors (CB(1)Rs) mediate the effects of ▵(9)-tetrahydrocannabinol (THC), the psychoactive component in marijuana. Repeated THC administration produces tolerance and dependence, which limit therapeutic development. Moreover, THC produces motor and psychoactive side effects. β-arrestin2 mediates receptor desensitization, internalization, and signaling, but its role in these CB(1)R effects and receptor regulation is unclear.. CB(1)R signaling and behaviors (antinociception, hypothermia, catalepsy) were assessed in β-arrestin2-knockout (βarr2-KO) and wild-type mice after THC administration. Cannabinoid-stimulated [(35)S]GTPγS and [(3)H]ligand autoradiography were assessed by statistical parametric mapping and region-of-interest analysis.. β-arrestin2 deletion increased CB(1)R-mediated G-protein activity in subregions of the cortex but did not affect CB(1)R binding, in vehicle-treated mice. βarr2-KO mice exhibited enhanced acute THC-mediated antinociception and hypothermia, with no difference in catalepsy. After repeated THC administration, βarr2-KO mice showed reduced CB(1)R desensitization and/or downregulation in cerebellum, caudal periaqueductal gray, and spinal cord and attenuated tolerance to THC-mediated antinociception. In contrast, greater desensitization was found in hypothalamus, cortex, globus pallidus, and substantia nigra of βarr2-KO compared with wild-type mice. Enhanced tolerance to THC-induced catalepsy was observed in βarr2-KO mice.. β-arrestin2 regulation of CB(1)R signaling following acute and repeated THC administration was region-specific, and results suggest that multiple, overlapping mechanisms regulate CB(1)Rs. The observations that βarr2-KO mice display enhanced antinociceptive responses to acute THC and decreased tolerance to the antinociceptive effects of the drug, yet enhanced tolerance to catalepsy, suggest that development of cannabinoid drugs that minimize CB(1)R interactions with β-arrestin2 might produce improved cannabinoid analgesics with reduced motor suppression.

Topics: Animals; Arrestins; Autoradiography; beta-Arrestins; Catalepsy; Central Nervous System; Down-Regulation; Dronabinol; Drug Tolerance; G-Protein-Coupled Receptor Kinases; Guanosine 5'-O-(3-Thiotriphosphate); Hypothermia; Mice; Mice, Knockout; Nociception; Receptor, Cannabinoid, CB1; Signal Transduction; Sulfur Radioisotopes

Salvinorin A, the primary psychoactive derivative of the hallucinogenic herb Salvia divinorum, is a potent and highly selective kappa-opioid receptor (KOR) agonist. Several recent studies, however, have suggested endocannabinoid system mediation of some of its effects.. This study represents a systematic examination of this hypothesis.. Salvinorin A was isolated from S. divinorum and was evaluated in a battery of in vitro and in vivo procedures designed to detect cannabinoid activity, including CB(1) receptor radioligand and [(35)S]GTPgammaS binding, calcium flux assay, in vivo cannabinoid screening tests, and drug discrimination.. Salvinorin A did not bind to nor activate CB(1) receptors. In vivo salvinorin A produced pronounced hypolocomotion and antinociception (and to a lesser extent, hypothermia). These effects were blocked by the selective KOR antagonist, JDTic, but not by the CB(1) receptor antagonist rimonabant. Interestingly, however, rimonabant attenuated KOR activation stimulated by U69,593 in a [(35)S]GTPgammaS assay. Salvinorin A did not substitute for Delta(9)-tetrahydrocannabinol (THC) in mice trained to discriminate THC.. These findings suggest that similarities in the pharmacological effects of salvinorin A and those of cannabinoids are mediated by its activation of KOR rather than by any direct action of salvinorin A on the endocannabinoid system. Further, the results suggest that rimonabant reversal of salvinorin A effects in previous studies may be explained in part by rimonabant attenuation of KOR activation.

Topics: Animals; Calcium; Cannabinoid Receptor Modulators; Discrimination Learning; Diterpenes, Clerodane; Dronabinol; Guanosine 5'-O-(3-Thiotriphosphate); Hypothermia; Locomotion; Male; Mice; Mice, Inbred C57BL; Mice, Inbred ICR; Pain Measurement; Piperidines; Pyrazoles; Radioligand Assay; Receptor, Cannabinoid, CB1; Receptors, Opioid, kappa; Rimonabant; Salvia; Tetrahydroisoquinolines

Alcoholism is a complex disorder involving, among others, the serotoninergic (5-HT) system, mainly regulated by 5-HT(1A) autoreceptors in the dorsal raphe nucleus. 5-HT(1A) autoreceptor desensitization induced by chronic 5-HT reuptake inactivation has been associated with a decrease in ethanol intake in mice. We investigated here whether, conversely, chronic ethanol intake could induce 5-HT(1A) autoreceptor supersensitivity, thereby contributing to the maintenance of high ethanol consumption. C57BL/6J mice were subjected to a progressive ethanol intake procedure in a free-choice paradigm (3-10% ethanol versus tap water; 21 days) and 5-HT(1A) autoreceptor functional state was assessed using different approaches. Acute administration of the 5-HT(1A) receptor agonist ipsapirone decreased the rate of tryptophan hydroxylation in striatum, and this effect was significantly larger (+75%) in mice that drank ethanol than in those drinking water. Furthermore, ethanol intake produced both an increased potency (+45%) of ipsapirone to inhibit the firing of 5-HT neurons, and a raise (+35%) in 5-HT(1A) autoreceptor-mediated stimulation of [(35)S]GTP-gamma-S binding in the dorsal raphe nucleus. These data showed that chronic voluntary ethanol intake in C57BL/6J mice induced 5-HT(1A) autoreceptor supersensitivity, at the origin of a 5-HT neurotransmission deficit, which might be causally related to the addictive effects of ethanol intake.

Topics: 5-Hydroxytryptophan; Action Potentials; Alcohol Drinking; Alcoholism; Animals; Behavior, Animal; Body Temperature; Disease Models, Animal; Food Preferences; Guanosine 5'-O-(3-Thiotriphosphate); Hydroxyindoleacetic Acid; Hypothermia; In Vitro Techniques; Male; Mice; Mice, Inbred C57BL; Neurons; Protein Binding; Pyrimidines; Raphe Nuclei; Receptor, Serotonin, 5-HT1A; Serotonin; Serotonin Receptor Agonists; Tryptophan

The aim of this study was to examine the role of fatty acid amide hydrolase (FAAH) on ethanol sensitivity, preference, and dependence. The deletion of FAAH gene or the inhibition of FAAH by carbamoyl-biphenyl-3-yl-cyclohexylcarbamate (URB597) (0.1 mg/kg) markedly increased the preference for ethanol. The study further reveals that URB597 specifically acts through FAAH and that cannabinoid-1 (CB(1)) receptor is critical for N-arachidonoyl ethanolamide (AEA) mediated ethanol-reinforced behavior as revealed by lack of URB597 effect in both FAAH and CB(1)-/- mice compared with vehicle-treated -/- mice. The FAAH -/- mice displayed a lower sensitivity to hypothermic and sedative effects to acute ethanol challenge. The FAAH -/- mice also exhibited a reduction in the severity of handling-induced convulsions following withdrawal from chronic ethanol exposure. The CB(1) receptor and proenkephalin gene expressions, and CB(1) receptor and mu-opioid (MO) receptor-mediated G-protein activation were found to be significantly lower in the caudate-putamen, nucleus accumbens core and shell of FAAH -/- than +/+ mice. Interestingly, the MO receptor-stimulated G-protein signaling was greater in the striatum of FAAH -/- than +/+ mice following voluntary ethanol consumption. These findings suggest that an elevation in the AEA content and its action on the limbic CB(1) receptor and MO receptor might contribute to ethanol-reinforced behavior. Treatment with drugs that decrease AEA tone might prove useful in reducing excessive ethanol consumption.

Topics: Alcohol Drinking; Amidohydrolases; Analgesics; Analgesics, Opioid; Animals; Behavior, Animal; Benzamides; Benzoxazines; Carbamates; Central Nervous System Depressants; Choice Behavior; Drug Interactions; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Ethanol; Guanosine 5'-O-(3-Thiotriphosphate); Hypothermia; Mice; Mice, Inbred C57BL; Mice, Knockout; Morpholines; Motor Activity; Naphthalenes; Protein Binding; Protein Precursors; Receptor, Cannabinoid, CB1

Chronic morphine administration produces tolerance in vivo and attenuation of mu opioid receptor (MOR)-mediated G-protein activation measured in vitro, but the relationship between these adaptations is not clear. The present study examined MOR-mediated G-protein activation in the CNS of mice with different levels of morphine tolerance.. Mice were implanted with morphine pellets, with or without supplemental morphine injections, to induce differing levels of tolerance as determined by a range of MOR-mediated behaviours. MOR function was measured using agonist-stimulated [(35)S]guanylyl-5'-O-(gamma-thio)-triphosphate ([(35)S]GTPgammaS) and receptor binding throughout the CNS.. Morphine pellet implantation produced 6-12-fold tolerance in antinociceptive assays, hypothermia and Straub tail, as measured by the ratio of morphine ED(50) values between morphine-treated and control groups. Pellet implantation plus supplemental injections produced 25-50-fold tolerance in these tests. In morphine pellet-implanted mice, MOR-stimulated [(35)S]GTPgammaS binding was significantly reduced only in the nucleus tractus solitarius (NTS) and spinal cord dorsal horn in tissue sections from morphine pellet-implanted mice. In contrast, MOR-stimulated [(35)S]GTPgammaS binding was significantly decreased in most regions examined in morphine pellet+morphine injected mice, including nucleus accumbens, caudate-putamen, periaqueductal gray, parabrachial nucleus, NTS and spinal cord.. Tolerance and the regional pattern of apparent MOR desensitization were influenced positively by the level of morphine exposure. These results indicate that desensitization of MOR-mediated G-protein activity is more regionally widespread upon induction of high levels of tolerance, suggesting that this response contributes more to high than low levels of tolerance to CNS-mediated effects of morphine.

Topics: Analgesics, Opioid; Animals; Binding Sites; Central Nervous System; Dose-Response Relationship, Drug; Drug Tolerance; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Hypothermia; Male; Mice; Morphine; Pain Measurement; Posterior Horn Cells; Receptors, Opioid, mu; Solitary Nucleus; Tail

In order to better define the role of 5-HT(1A) receptors in the modulation of extrapyramidal motor functions, we investigated the effect of 5-HT(1A) agonists on tacrine-induced tremulous jaw movements (TJM) in rats, a putative model of parkinsonian tremor. Acute injection of 5-HT(1A) agonists 8-OH-DPAT and buspirone dose-dependently counteracted the tacrine-induced oral movements (ED(50)=0.04 and 1.0mg/kg, respectively), an effect reversed by the selective 5-HT(1A) antagonist WAY 100,635. In contrast to classical antipsychotics, the atypical antipsychotics risperidone (ED(50)=0.3mg/kg) and clozapine (ED(50)=1.5mg/kg) blocked the oral movements induced by the cholinomimetic agent at or below the doses required for suppression of conditioned avoidance response. The compound F-97013-GD (6-methyl-2-[4-(naphtylpiperazin-1-yl)butyl]-3-(2H)-pyridazinone), a putative antipsychotic drug that in functional in vitro and in vivo assays behaved as a mixed dopamine D(2)-antagonist and 5-HT(1A)-partial agonist, also displayed a potent antitremorgenic effect in this paradigm (ED(50)=0.5mg/kg). Interestingly, pretreatment with WAY 100,635 blocked the inhibitory effect of F-97013-GD but not that of clozapine. The 5-HT depleting agent para-chlorophenylalanine (PCPA) partially attenuated tacrine-induced TJM but did not block the suppressive effect of 5-HT(1A) agonists. In addition, only high doses of F-97013-GD induced catalepsy in rodents and, like 8-OH-DPAT and clozapine, the compound reversed the haloperidol-induced catalepsy in rats. These results show that 5-HT(1A) receptors play a role in the regulation of tacrine-induced TJM and suggest that their activation by novel antipsychotics may not only reduce the extrapyramidal side effects EPS liability, but also be effective in the treatment of parkinsonian tremor.

Topics: 5-Hydroxytryptophan; Animals; Antiparkinson Agents; Antipsychotic Agents; Apomorphine; Avoidance Learning; Binding, Competitive; Catalepsy; Dopamine Agonists; Dopamine D2 Receptor Antagonists; Dopamine Plasma Membrane Transport Proteins; Drug Interactions; Guanosine 5'-O-(3-Thiotriphosphate); Haloperidol; Head Movements; Hypothermia; Jaw; Male; Mice; Motor Activity; Piperazines; Pyridazines; Pyridines; Rats; Rats, Wistar; Receptor, Serotonin, 5-HT1A; Receptors, Dopamine D2; Serotonin Antagonists; Serotonin Receptor Agonists; Tremor

CT-3 (ajulemic acid) is a synthetic analogue of a metabolite of Delta9-tetrahydrocannabinol that has reported analgesic efficacy in neuropathic pain states in man. Here we show that CT-3 binds to human cannabinoid receptors in vitro, with high affinity at hCB1 (Ki 6 nM) and hCB2 (Ki 56 nM) receptors. In a functional GTP-gamma-S assay CT-3 was an agonist at both hCB1 and hCB2 receptors (EC50 11 and 13.4 nM, respectively). In behavioural models of chronic neuropathic and inflammatory pain in the rat, oral administration of CT-3 (0.1-1 mg/kg) produced up to 60% reversal of mechanical hyperalgesia. In both models the antihyperalgesic activity was prevented by the CB1-antagonist SR141716A but not the CB2-antagonist SR144528. In the tetrad of tests for CNS activity, CT-3 (1-10 mg/kg, po) produced dose-related catalepsy, deficits in locomotor performance, hypothermia, and acute analgesia. Comparison of 50% maximal effects in the tetrad and chronic pain assays produced an approximate therapeutic index of 5-10. Pharmacokinetic analysis showed that CT-3 exhibits significant but limited brain penetration, with a brain/plasma ratio of 0.4 measured following oral administration, compared to ratios of 1.0-1.9 measured following subcutaneous administration of WIN55,212-2 or Delta9-THC. These data show that CT-3 is a cannabinoid receptor agonist and is efficacious in animal models of chronic pain by activation of the CB1 receptor. Whilst it shows significant cannabinoid-like CNS activity, it exhibits a superior therapeutic index compared to other cannabinoid compounds, which may reflect a relatively reduced CNS penetration.

Topics: Analgesics; Animals; Benzoxazines; Cannabinoids; Catalepsy; Cell Line; Chromatography; Cricetinae; Cricetulus; Cyclohexanols; Disease Models, Animal; Dose-Response Relationship, Drug; Dronabinol; Drug Interactions; Freund's Adjuvant; Guanosine 5'-O-(3-Thiotriphosphate); Humans; Hypothermia; Inflammation; Ligation; Male; Morpholines; Motor Activity; Naphthalenes; Pain; Pain Measurement; Pain Threshold; Radioligand Assay; Rats; Rats, Wistar; Rotarod Performance Test; Sciatic Neuropathy; Sulfur Isotopes; Time Factors; Tritium

A series of new mixed benzimidazole-arylpiperazine derivatives were designed by incorporating in general structure III the pharmacophoric elements of 5-HT(1A) and 5-HT(3) receptors. Compounds 1-11 were synthesized and evaluated for binding affinity at both serotoninergic receptors, all of them exhibiting high 5-HT(3)R affinity (K(i)=10-62nM), and derivatives with an o-alkoxy group in the arylpiperazine ring showing nanomolar affinity for the 5-HT(1A)R (K(i)=18-150nM). Additionally, all the synthesized compounds were selective over alpha(1)-adrenergic and dopamine D(2) receptors (K(i)>1000-10,000nM). Compound 3 was selected for further pharmacological characterization due to its interesting binding profile as mixed 5-HT(1A)/5-HT(3) ligand with high affinity for both receptors (5-HT(1A): K(i)=18.0nM, 5-HT(3): K(i)=27.2nM). In vitro and in vivo findings suggest that this compound acts as a partial agonist at 5-HT(1A)Rs and as a 5-HT(3)R antagonist. This novel mixed 5-HT(1A)/5-HT(3) ligand was also effective in preventing the cognitive deficits induced by muscarinic receptor blockade in a passive avoidance learning test, suggesting a potential interest in the treatment of cognitive dysfunction.

Topics: Animals; Avoidance Learning; Benzimidazoles; Cognition Disorders; Guanosine 5'-O-(3-Thiotriphosphate); Guinea Pigs; Hippocampus; Hypothermia; Ligands; Male; Mice; Muscle Contraction; Piperazines; Rats; Rats, Wistar; Serotonin 5-HT1 Receptor Antagonists; Serotonin 5-HT3 Receptor Antagonists; Serotonin Antagonists; Structure-Activity Relationship

1 (6-((R)-2-[2-[4-(4-Chloro-phenoxy)-piperidin-1-yl]-ethyl]-pyrrolidine-1-sulphonyl)-1H-indole hydrochloride) (SB-656104-A), a novel 5-hydroxytryptamine (5-HT(7)) receptor antagonist, potently inhibited [(3)H]-SB-269970 binding to the human cloned 5-HT(7(a)) (pK(i) 8.7+/-0.1) and 5-HT(7(b)) (pK(i) 8.5+/-0.2) receptor variants and the rat native receptor (pK(i) 8.8+/-0.2). The compound displayed at least 30-fold selectivity for the human 5-HT(7(a)) receptor versus other human cloned 5-HT receptors apart from the 5-HT(1D) receptor ( approximately 10-fold selective). 2 SB-656104-A antagonised competitively the 5-carboxamidotryptamine (5-CT)-induced accumulation of cyclic AMP in h5-HT(7(a))/HEK293 cells with a pA(2) of 8.5. 3 Following a constant rate iv infusion to steady state in rats, SB-656104 had a blood clearance (CL(b)) of 58+/-6 ml min(-1) kg(-1) and was CNS penetrant with a steady-state brain : blood ratio of 0.9 : 1. Following i.p. administration to rats (10 mg kg(-1)), the compound displayed a t(1/2) of 1.4 h with mean brain and blood concentrations (at 1 h after dosing) of 0.80 and 1.0 micro M, respectively. 4 SB-656104-A produced a significant reversal of the 5-CT-induced hypothermic effect in guinea pigs, a pharmacodynamic model of 5-HT(7) receptor interaction in vivo (ED(50) 2 mg kg(-1)). 5 SB-656104-A, administered to rats at the beginning of the sleep period (CT 0), significantly increased the latency to onset of rapid eye movement (REM) sleep at 30 mg kg(-1) i.p. (+93%) and reduced the total amount of REM sleep at 10 and 30 mg kg(-1) i.p. with no significant effect on the latency to, or amount of, non-REM sleep. SB-269970-A produced qualitatively similar effects in the same study. 6 In summary, SB-656104-A is a novel 5-HT(7) receptor antagonist which has been utilised in the present study to provide further evidence for a role for 5-HT(7) receptors in the modulation of REM sleep.

Topics: Animals; Cell Line; Cell Membrane; CHO Cells; Cricetinae; Cyclic AMP; Drug Administration Routes; Gene Expression Regulation; Guanosine 5'-O-(3-Thiotriphosphate); Guinea Pigs; Humans; Hypothermia; Phenols; Pyrrolidines; Radioligand Assay; Rats; Rats, Sprague-Dawley; Receptors, Serotonin; Serotonin; Serotonin Antagonists; Sleep, REM; Tritium

F 11356 (4-[4-[2-(2-aminoethyl)-1H-indol-5-yloxyl]acetyl]piperazinyl-1-yl] ben zonitrile) was designed to take advantage of the superior potency and efficacy characteristics of 5-hydroxytryptamine (5-HT) compared with tryptamine at 5-HT1B/1D receptors. F 11356 has subnanomolar affinity for cloned human and nonhuman 5-HT1B and 5-HT1D receptors, and its affinity for 5-HT1A and other 5-HT receptors, including the 5-ht1F subtype, is 50-fold lower and micromolar, respectively. In C6 cells expressing human 5-HT1B or human 5-HT1D receptors, F 11356 was the most potent compound in inhibiting forskolin-induced cyclic AMP formation (pD2 = 8.9 and 9.6), and in contrast to tryptamine and derivatives, it produced maximal enhancement of [35S]guanosine-5'-O-(3-thio)triphosphate-specific binding equivalent to 5-HT. F 11356 was equipotent to 5-HT (pD2 = 7.1 versus 7.2) and more potent than tryptamine derivatives in contracting rabbit isolated saphenous vein. In isolated guinea pig trigeminal ganglion neurons, F 11356 was more potent (pD2 = 7.3 versus 6.7) and induced greater increases in outward hyperpolarizing Ca2+-dependent K+ current than sumatriptan. In anesthetized pigs, F 11356 elicited highly cranioselective, more potent (from 0.16 microgram/kg i.v.) and greater carotid vasoconstriction than tryptamine derivatives. Decreases in carotid blood flow were observed in conscious dogs from 0.63 mg/kg oral F 11356 in the absence of changes in heart rate or behavior. Oral activity was confirmed when hypothermic responses were elicited in guinea pigs (ED50 = 1.6 mg/kg), suggesting that F 11356 also accesses the brain. F 11356 thus is a selective, high-potency agonist at 5-HT1B/1D receptors, which distinguishes itself from tryptamine and derivatives in exerting high intrinsic activity at these receptors in vascular and neuronal models relevant to migraine.

Topics: Animals; Carotid Arteries; Colforsin; Cyclic AMP; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Guanosine 5'-O-(3-Thiotriphosphate); Guinea Pigs; Heart; Hemodynamics; Humans; Hypothermia; In Vitro Techniques; Male; Migraine Disorders; Muscle, Smooth, Vascular; Neurons; Nitriles; Piperazines; Rabbits; Radioligand Assay; Rats; Receptor, Serotonin, 5-HT1B; Receptor, Serotonin, 5-HT1D; Receptors, Serotonin; Saphenous Vein; Swine; Trigeminal Ganglion; Tryptamines

We replaced the coding region of the murine 5-hydroxytryptamine (5-HT)1B receptor by the human 5-HT1B receptor using homologous recombination in embryonic stem cells and generated and characterized homozygous transgenic mice that express only the human (h) 5-HT1B receptor. The distribution patterns of h5-HT1B and murine (m) 5-HT1B receptor mRNA and binding sites in brain sections of transgenic and wild-type mice were identical as measured by in situ hybridization histochemistry and radioligand receptor autoradiography. When measured in parallel under identical conditions, the h5-HT1B receptor expressed in mouse brain had the same pharmacological characteristics as that in human brain. Stimulation by 5-HT1B agonists of [35S]guanosine-5'-O-(3-thio)triphosphate binding in brain sections demonstrated the functional coupling of the h5-HT1B receptor to G proteins in mouse brain. In tissue slices from various brain regions, electrically stimulated [3H]5-HT release was not modified by 5-HT1B agonists in tissue from either transgenic and wild-type mice; a 5-HT1B antagonist enhanced electrically stimulated [3H]5-HT release in wild-type mouse brain, but was ineffective in the transgenics. The centrally active 5-HT1A/5-HT1B agonist RU24969 induced hypothermia but did not increase locomotor activity in the transgenic mice. The ineffectiveness of RU24969 in the transgenic mice could be due to the lower affinity of the compound for the h5-HT1B receptor compared with the m5-HT1B receptor. The present study demonstrates a complete replacement of the mouse receptor by its human receptor homolog and a functional coupling to G proteins. However, modulation of [3H]5-HT release could not be shown. Furthermore, behavioral effects were not clearly observed, which may be due to a lack of appropriate tools.

Topics: Animals; Benzamides; Benzopyrans; Binding Sites; Brain; Gene Expression; Guanosine 5'-O-(3-Thiotriphosphate); Humans; Hypothermia; In Situ Hybridization; Indoles; Mice; Mice, Transgenic; Motor Activity; Piperidones; Propylamines; Pyridines; Pyrimidines; Pyrroles; Receptor, Serotonin, 5-HT1B; Receptors, Serotonin; Recombination, Genetic; RNA, Messenger; Serotonin; Serotonin Antagonists; Serotonin Receptor Agonists; Spiro Compounds; Tritium