The GR127935-sensitive 5-HT(1) receptors mediating canine internal carotid vasoconstriction: resemblance to the 5-HT(1B), but not to the 5-HT(1D) or 5-ht(1F), receptor subtype

This study has further investigated the pharmacological profile of the GR127935-sensitive 5-HT(1) receptors mediating vasoconstriction in the internal carotid bed of anaesthetized vagosympathectomized dogs. One-minute intracarotid infusions of the agonists 5-hydroxytryptamine (5-HT; 0.1 - 10 microg min(-1); endogenous ligand) and sumatriptan (0.3 - 10 microg min(-1); 5-HT(1B/1D)), but not PNU-142633 (1 - 1000 microg min(-1); 5-HT(1D)) or LY344864 (1 - 1000 microg min(-1); 5-ht(1F)), produced dose-dependent decreases in internal carotid blood flow without changing blood pressure or heart rate. The responses to 5-HT were apparently resistant to blockade by i.v. administration of the antagonists SB224289 (300 microg kg(-1); 5-HT(1B)), BRL15572 (300 microg kg(-1); 5-HT(1D)) or ritanserin (100 microg kg(-1); 5-HT(2)). In contrast, the responses to sumatriptan were antagonized by SB224289, but not by BRL15572. In the animals receiving SB224289, but not those receiving BRL15572, the subsequent administration of ritanserin abolished the 5-HT-induced vasoconstriction and unmasked a vasodilator component. Similarly, in ritanserin-treated animals, the subsequent administration of SB224289, but not BRL15572, completely blocked the 5-HT-induced vasoconstriction, revealing vasodilatation. In animals receiving initially BRL15572, the subsequent administration of SB224289 did not affect (except at 10 microg min(-1)) the vasoconstrictor responses to 5-HT. Notably, in animals pretreated with 1000 microg kg(-1) of mesulergine, a 5-HT(2/7) receptor antagonist, 5-HT produced a dose-dependent vasoconstriction, which was practically abolished by SB224289. After BRL15572, no further blockade was produced and the subsequent administration of ritanserin was similarly inactive. These results suggest that the GR127935-sensitive 5-HT(1) receptors mediating canine internal carotid vasoconstriction resemble the 5-HT(1B) but not the 5-HT(1D) or 5-ht(1F), receptor subtype.     

Mediation of 5-HT-induced internal carotid vasodilatation in GR127935- and ritanserin-pretreated dogs by 5-HT7 receptors

The vasoconstrictor effects of 5-hydroxytryptamine (5-HT) in the internal carotid bed of anaesthetised dogs with bilateral vagosympathectomy are mainly mediated by both 5-HT1B and 5-HT2 receptors. The blockade of this vasoconstrictor effect of 5-HT by the combined use of the antagonists, GR127935 (5-HT1B/1D) and ritanserin (5-HT2), unmasks a dose-dependent vasodilator effect of 5-HT, but not of sumatriptan. Therefore, the present study set out to analyse the pharmacological profile of this vasodilator 5-HT receptor in the internal carotid bed of vagosympathectomized dogs systematically pretreated with intravenous (i.v.) injections of GR127935 (30 microg/kg) and ritanserin (100 microg/kg). One-minute (1-min) intracarotid (i.c.) infusions of 5-HT (0.1-10 microg/min), 5-carboxamidotryptamine (5-CT; 0.01-0.3 microg/min), 5-methoxytryptamine (5-MeO-T; 1-100 microg/min) and acetylcholine (ACh; 0.003-0.1 microg/min) resulted in dose-dependent increases in internal carotid blood flow (without changes in blood pressure or heart rate) with a rank order of agonist potency of ACh > 5-CT > 5-HT > or =5-MeO-T. The internal carotid vasodilator responses to 5-HT, 5-CT and 5-MeO-T, which remained unaffected after saline (0.03 ml/kg and 0.1 ml/kg, i.v.), were specifically and dose-dependently blocked by i.v. administration of lisuride (10 microg/kg and 30 microg/kg), clozapine (1000 microg/kg), mesulergine (300 microg/kg and 1000 microg/kg) and LY215840 (300 microg/kg and 1000 microg/kg) with the following apparent rank order of potency: lisuride > mesulergine = LY215840 > or = clozapine. The above results suggest that the 5-HT receptor mediating internal carotid vasodilatation in vagosympathectomized dogs pretreated with GR127935 and ritanserin is operationally similar to other 5-HT7 receptors mediating vascular and non-vascular responses.     

Canine external carotid vasoconstriction to methysergide, ergotamine and dihydroergotamine: role of 5-HT1B/1D receptors and alpha2-adrenoceptors

The antimigraine drugs methysergide, ergotamine and dihydroergotamine (DHE) produce selective vasoconstriction in the external carotid bed of vagosympathectomized dogs anaesthetized with pentobarbital and artificially respired, but the receptors involved have not yet been completely characterized. Since the above drugs display affinity for several binding sites, including alpha-adrenoceptors and several 5-HT1 and 5-HT2 receptor subtypes, this study has analysed the mechanisms involved in the above responses. Intracarotid (i.c.) infusions during 1 min of methysergide (31-310 microg min(-1)), ergotamine (0.56-5.6 microg min(-1)) or DHE (5.6-31 microg min(-1)) dose-dependently reduced external carotid blood flow (ECBF) by up to 46+/-4, 37+/-4 and 49+/-5%, respectively. Blood pressure and heart rate remained unchanged. The reductions in ECBF by methysergide were abolished and even reversed to increases in animals pre-treated with GR127935 (10 microg kg(-1), i.v.). The reductions in ECBF by ergotamine and DHE remained unchanged in animals pre-treated (i.v.) with prazosin (300 microg kg(-1)), but were partly antagonized in animals pre-treated with either GR127935 (10 or 30 microg kg(-1)) or yohimbine (1000 microg kg(-1)). Pre-treatment with a combination of GR127935 (30 microg kg(-1)) and yohimbine (1000 microg kg(-1)) abolished the responses to both ergotamine and DHE. The above doses of antagonists were shown to produce selective antagonism at their respective receptors. These results suggest that the external carotid vasoconstrictor responses to methysergide primarily involve 5-HT1B/1D receptors, whereas those to ergotamine and DHE are mediated by 5-HT1B/1D receptors as well as alpha2-adrenoceptors.     

Unravelling the pharmacological profile of the canine external carotid vasodilator '5-HT1-like' receptors: coexistence of sympatho-inhibitory 5-HT1B and postjunctional 5-HT7 receptors

It has been suggested that the external carotid vasodilatation produced by serotonin (5-hydroxytryptamine; 5-HT) in anaesthetised dogs with intact vagosympathetic trunks is mediated by sympatho-inhibitory '5-HT1D' receptors and musculotropic '5-HT1-like' receptors. The present study has re-analysed this suggestion with regard to the classification schemes recently proposed by the NC-IUPHAR subcommittee on 5-HT receptors. In pentobarbital-anaesthetised dogs with intact vagosympathetic trunks, 1-min intracarotid (i.c.) infusions of 5-carboxamidotryptamine (5-CT; 0.01-0.3 microg/min), 5-HT (0.3-30 microg/ min), 5-methoxytryptamine (5-MeO-T; 1-100 microg/min) or sumatriptan (1-100 microg/min) dose-dependently increased the external carotid blood flow without affecting blood pressure or heart rate. The selective 5-HT1D receptor agonist, PNU-142633 (1-1000 microg/min), was essentially inactive. After mesulergine (300 microg/kg, i.v.), an antagonist at cardiovascular 5-HT7 receptors, the above responses to 5-HT, 5-CT and 5-MeO-T were blocked, whilst those to sumatriptan remained unaffected. In contrast, after the 5-HT1B/1D receptor antagonist, GR127935 (10 microg/kg, i.v.), the responses to 5-HT, 5-CT and 5-MeO-T were not affected, but those to sumatriptan were abolished. Furthermore, after the selective 5-HT1B receptor antagonist, SB224289 (300 microg/kg, i.v.), the responses to 5-HT, 5-CT and 5-MeO-T were significantly enhanced, whereas those to sumatriptan were abolished. Interestingly, the responses to all these agonists remained unmodified after the selective 5-HT1D receptor antagonist, BRL15572 (300 microg/kg, i.v.). The above results suggest that the '5-HT1-like' receptors, which mediate canine external carotid vasodilatation, display the pharmacological profile of sympatho-inhibitory 5-HT1B receptors and musculotropic 5-HT7 receptors, and confirm the existence of vasoconstrictor 5-HT1B receptors.     

Pharmacological profile of the 5-HT-induced inhibition of cardioaccelerator sympathetic outflow in pithed rats: correlation with 5-HT1 and putative 5-ht5A/5B receptors

Continuous infusions of 5-hydroxytryptamine (5-HT) inhibit the tachycardiac responses to preganglionic (C7-T1) sympathetic stimulation in pithed rats pretreated with desipramine. The present study identified the pharmacological profile of this inhibitory action of 5-HT. The inhibition induced by intravenous (i.v.) continuous infusions of 5-HT (5.6 microg x kg-1x min-1) on sympathetically induced tachycardiac responses remained unaltered after i.v. treatment with saline or the antagonists GR 127935 (5-HT1B/1D), the combination of WAY 100635 (5-HT1A) plus GR 127935, ritanserin (5-HT2), tropisetron (5-HT3/4), LY215840 (5-HT7) or a cocktail of antagonists/inhibitors consisting of yohimbine (alpha2), prazosin (alpha1), ritanserin, GR 127935, WAY 100635 and indomethacin (cyclooxygenase), but was abolished by methiothepin (5-HT1/2/6/7 and recombinant 5-ht5A/5B). These drugs, used in doses high enough to block their respective receptors/mechanisms, did not modify the sympathetically induced tachycardiac responses per se. I.v. continuous infusions of the agonists 5-carboxamidotryptamine (5-CT; 5-HT1/7 and recombinant 5-ht5A/5B), CP 93129 (r5-HT1B), sumatriptan (5-HT1B/1D), PNU-142633 (5-HT1D) and ergotamine (5-HT1B/1D and recombinant 5-ht5A/5B) mimicked the above sympatho-inhibition to 5-HT. In contrast, the agonists indorenate (5-HT1A) and LY344864 (5-ht1F) were inactive. Interestingly, 5-CT-induced cardiac sympatho-inhibition was abolished by methiothepin, the cocktail of antagonists/inhibitors, GR 127935 or the combination of SB224289 (5-HT1B) plus BRL15572 (5-HT1D), but remained unchanged when SB224289 or BRL15572 were given separately. Therefore, 5-HT-induced cardiac sympatho-inhibition, being unrelated to 5-HT2, 5-HT3, 5-HT4, 5-ht6, 5-HT7 receptors, alpha1/2-adrenoceptor or prostaglandin synthesis, seems to be primarily mediated by (i). 5-HT1 (probably 5-HT1B/1D) receptors and (ii). a novel mechanism antagonized by methiothepin that, most likely, involves putative 5-ht5A/5B receptors.     

Further pharmacological analysis of the orphan 5-HT receptors mediating feline vasodepressor responses: close resemblance to the 5-HT7 receptor

It has been suggested that the late hypotensive response to serotonin (5-hydroxytryptamine; 5-HT) in vagosympathectomised cats, being potently mimicked by 5-carboxamidotryptamine (5-CT), not modified by ketanserin and blocked by methiothepin or methysergide, is mediated by '5-HT1-like' receptors. Nevertheless, current guidelines for 5-HT receptor classification refer to this receptor as an orphan receptor. Thus, the present study set out to reanalyse the above suggestion in terms of the classification schemes proposed in 1994 and 1998 by the NC-IUPHAR subcommittee on the classification of 5-HT receptors. Intravenous (i.v.) bolus injections of 5-CT (0.003-0.3 microg/kg), 5-HT (1-100 microg/kg) and 5-methoxytryptamine (5-MeO-T; 1-100 microg/kg) produced dose-dependent vasodepressor responses with a rank order of agonist potency of 5-CT > 5-HT = 5-MeO-T with sumatriptan (10-300 microg/kg) virtually inactive. The vasodepressor responses to 5-HT, 5-CT and 5-MeO-T were not attenuated following i.v. administration of the antagonists GR127935 (5-HT(IB/ID); 30 microg/kg), tropisetron (5-HT3/4; 3000 microg/kg), (+/-)-pindolol (beta-adrenergic and 5-HT1A; 4000 microg/kg) or equivalent volumes of physiological saline. In contrast, the above vasodepressor responses were markedly and specifically antagonised by i.v. methiothepin (100 microg/kg), lisuride (30 microg/kg and 100 microg/kg), mesulergine (300 microg/kg and 1000 microg/kg) or LY215840 (300 microg/kg and 1000 microg/kg). The above lines of evidence, therefore, indicate that the orphan receptors mediating the vasodepressor responses to 5-HT in vagosympathectomised cats are pharmacologically similar to other 5-HT7 receptors mediating vascular and non-vascular responses (e.g. relaxation of the canine external carotid artery and guinea-pig ileum as well as feline tachycardia).     

The atypical 5-HT2 receptor mediating tachycardia in pithed rats: pharmacological correlation with the 5-HT2A receptor subtype

1. In pithed rats, 5-HT mediates tachycardia both directly (by 5-HT(2) receptors) and indirectly (by a tyramine-like effect). The receptor mediating tachycardia directly has been classified as an 'atypical' 5-HT(2) receptor since it was 'weakly' blocked by ketanserin. Moreover, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), a 5-HT(2) agonist, failed to mimic 5-HT-induced tachycardia. Since 5-HT(2) receptors consist of 5-HT(2A), 5-HT(2B) and 5-HT(2C) subtypes, this study investigated if these subtypes mediate the above response. 2. In pithed rats, intraperitoneally (i.p.) pre-treated with reserpine (5 mg kg(-1)), intravenous (i.v.) administration of 5-HT, 5-methoxytryptamine (5-MeO-T), 1-(3-chlorophenyl) piperazine (mCPP) and 5-carboxamidotryptamine (5-CT) (10, 30, 100 and 300 microg kg(-1) each), produced dose-dependent tachycardic responses. Interestingly, DOI (10 - 1000 microg kg(-1), i.v.) induced only slight, dose-unrelated, tachycardic responses, whilst the 5-HT(2C) agonist, Ro 60-0175 (10 - 1000 microg kg(-1), i.v.), produced a slight tachycardia only at 300 and 1000 microg kg(-1). In contrast, sumatriptan and 1-(m-trifluoromethylphenyl)- piperazine (TFMPP) were inactive. The rank order of potency was: 5-HT > or =5-MeO-T> mCPP > or =5-CT > or =DOI > Ro 60-0175. 3. The tachycardic responses to 5-HT, which remained unaffected after i.v. saline (0.3 and 1 ml kg(-1)) or propranolol (3 mg kg(-1)), were selectively blocked by the 5-HT(2A) antagonists ketanserin (30 and 100 microg kg(-1)) or spiperone (10 and 30 microg kg(-1)) as well as by the non-selective 5-HT(2) antagonists, ritanserin (10 and 30 microg kg(-1)) or mesulergine (100 microg kg(-1)). Remarkably, these responses were unaffected by the antagonists rauwolscine (5-HT(2B)), SB204741 (5-HT(2B/2C)) or Ro 04-6790 (5-ht(6)) (300 and 1000 microg kg(-1) each). 4. These results suggest that the 'atypical' 5-HT(2) receptors mediating tachycardia in reserpinized pithed rats are pharmacologically similar to the 5-HT(2A) receptor subtype.     

Pharmacological evidence that alpha1-and alpha2-adrenoceptors mediate vasoconstriction of carotid arteriovenous anastomoses in anaesthetized pigs.

Vasoconstriction of carotid arteriovenous anastomoses may be involved in the therapeutic action of acutely acting anti-migraine agents, including the triptans and ergot alkaloids. While 5-HT1B/1D receptors mediate the effect of triptans, ergotamine and dihydroergotamine also interact with alpha-adrenoceptors. In the present study, we investigated the potential role of alpha1- and alpha2-adrenoceptors in mediating vasoconstriction of carotid arteriovenous anastomoses in anaesthetized pigs. Ten minute intracarotid infusions of phenylephrine (1, 3 and 10 microg kg(-1) min(-1)) or BHT 933 (3, 10 and 30 microg kg(-1) min(-1)) produced dose-dependent decreases in total carotid and arteriovenous anastomotic conductances; no changes were observed in the capillary fraction. The carotid vascular effects of phenylephrine and BHT 933 were selectively abolished by prazosin (100 microg kg(-1), i.v.) and rauwolscine (300 microg kg(-1), i.v.), respectively. The responses to phenylephrine and BHT 933 were not affected by the selective 5-HT1B/1D receptor antagonist GR127935 (500 microg kg(-1), i.v.). These results show that both alpha1- and alpha2-adrenoceptors can mediate vasoconstriction of carotid arteriovenous anastomoses in anaesthetized pigs. Since vasoconstrictor activity in this in vivo model is predictive of anti-migraine activity, an agonist activity at particularly the alpha2-adrenoceptor subtypes, in view of their less ubiquitous nature, could provide migraine abortive potential. Thus, the present results may aid further understanding of the mode of action of some current anti-migraine agents and may eventually be helpful in the development of future treatment in migraine.     

5—羟色胺受体介导迷走交感神经切断犬颈外血管收缩

One specific example reflecting the complexity of cardiovascular responses induced by serotonin (5-hydroxytryptamine; 5-HT) and the progress achieved in the pharmacological characterization of the receptors involved can be illustrated by the effects of 5-HT on the canine external carotid artery bed. Within this framework, it has been shown that the external carotid vasoconstrictor response to 5-HT in the dog is mediated by '5-HT1-like' receptors, which being blocked by the 5-HT1B/1D receptor antagonist GR127935, resemble 5-HT1B/1D (previously called 5-HT1D beta/1D alpha) receptors. It was proposed that these receptors could belong to the 5-HT1B, rather than the 5-HT1D, subtype on the basis of their resistance to blockade by a high dose of ritanserin (a potential 5-HT1D receptor antagonist) and the presence of mRNA for 5-HT1B(5-HT1D beta) receptors, but not for 5-HT1D(5-HT1D alpha) receptors, in vascular smooth muscle. With the advent of subtype-selective antagonists it was subsequently shown that external carotid vasoconstriction to 5-HT and sumatriptan is dose-dependently antagonized by the selective 5-HT1B receptor antagonist SB224289 (2,3,6,7-tetrahydro-1'-methyl-5-[2'-methyl-4' (5-methyl-1,2,4-oxadiazol-3-yl) biphenyl-4-carbonyl] furo [2,3-f] indole-3-spiro-4'-piperidine hydrochloride), whereas the selective 5-HT1D receptor antagonist BRL15572 (1-(3-chlorophenyl)-4-[3,3-diphenyl (2-(S,R) hydroxypropanyl) piperazine] hydrochloride) was ineffective. These findings represent the first in vivo evidence showing that vascular constriction induced by 5-HT and sumatriptan is mediated primarily via 5-HT1B, but not 5-HT1D receptors. The pharmacological profile of these receptors could be similar (isolated human temporal artery and porcine carotid arteriovenous anastomoses) to other putative 5-HT1B receptors mediating vasoconstrictor responses. In view of the putative pathophysiologic role of external carotid (and extracerebral) vasodilation in migraine, the constriction of these blood vessels by sumatriptan via 5-HT1B receptors may be, at least partly, responsible for its therapeutic efficacy in migraine.     

Operational characteristics of the 5-HT1-like receptors mediating external carotid vasoconstriction in vagosympathectomized dogs

It has recently been shown that the external carotid vasoconstrictor response to 5-HT in the dog is primarily mediated by sumatriptan-sensitive 5-HT₁-like receptors; however, the fact that these receptors are not blocked by metergoline, a 5-HT_1D ligand, raises questions about their possible correlation with the 5-HT_1D receptor subtype. Since a number of drugs display high affinity for the 5-HT_1D (GR127935) and 5-HT_1F (e.g. methysergide and oxymetazoline) receptor subtypes, in this study we have used these drugs to determine whether the above vasoconstrictor 5-HT₁-like receptors correlate with the 5-HT_1D and/or 5-HT_1F receptor subtypes.One-minute intracarotid infusions of 5-HT (0.3–30 g/min), sumatriptan (1–30 g/min), oxymetazoline (0.03–3 g/min) and noradrenaline (0.3–3 g/min) resulted in dose-dependent decreases in external carotid blood flow without changes in arterial blood pressure or heart rate. These vasoconstrictor responses remained unaltered after i.v. administration of physiological saline (0.015, 0.05 and 0.15 ml/kg; n = 4) or ritanserin (1 mg/kg; n = 5). In contrast, GR127935 (1, 3 and 10 g/kg, n = 6) potently blocked the responses to 5-HT (unmasking a dose-dependent vasodilator component) and sumatriptan without affecting those to oxymetazoline or noradrenaline. Interestingly, methysergide (10, 30 and 100 g/kg, n = 5) also blocked the vasoconstrictor responses to 5-HT and sumatriptan, but unlike GR127935, did not revert the vasoconstrictor response to 5-HT; the responses to oxymetazoline remained unaffected, but those to noradrenaline were apparently attenuated by the highest dose.Taken together, the above findings suggest that the sumatriptan-sensitive 5-HT₁-like receptors mediating canine external carotid vasoconstriction resemble 5-HT_1D receptors, probably of the 5-HT_1D subtype on the basis of the resistance to blockade by ritanserin. The pharmacological profile of these receptors could be similar (bovine and human cerebral arteries, porcine carotid arteriovenous anastomoses and human coronary arteries) to other putative 5-HT_1D receptors mediating vascular responses.     

The anxiolytic-like effect of 5-HT1B receptor ligands in rats: a possible mechanism of action

We have examined the effect of lesions of 5-hydroxytryptamine (5-HT) neurons, produced by p-chloroamphetamine (p-CA; 2 x 10 mg kg(-1)), and the influence of flumazenil (Ro 15-1788, 10 mg kg(-1)), a benzodiazepine receptor antagonist, on the anxiolytic-like activity of CP 94253 (5-propoxy-3-(1,2,3,6-tetrahydro-4-pyridinyl)-1H-pyrrolo[3,2-b]pyridine), a 5-HT1B receptor agonist, SB 216641 (N-[3-[3-(dimethylamino)ethoxy]-4-methoxyphenyl]-2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)-[1,1'-biphenyl]-4-carboxamide), a 5-HT1B receptor antagonist, and GR 127935 (N-[4-methoxy-3-(4-methyl-l-piperazinyl)phenyl]-2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)-l, l'-biphenyl-4-carboxamide), a 5-HT1B/1D receptor antagonist, in the Vogel conflict drinking test in rats. Diazepam was used as a reference compound. CP 94253 (2.5 mg kg(-1)), SB 216641 (2.5 mg kg(-1)), GR 127935 (10 mg kg(-1)) and diazepam (5 mg kg(-1)) significantly increased the number of shocks accepted during experimental sessions in the conflict drinking test in vehicle- and p-CA-pretreated rats. Flumazenil did not change the anxiolytic-like effect of CP 94253 (2.5 mg kg(-1)), but wholly blocked the anxiolytic-like effects of SB 216641 (2.5 mg kg(-1)), GR 127935 (10 mg kg(-1)) and diazepam (5 mg kg(-1)). p-CA and flumazenil alone were inactive in the conflict drinking test. The results suggested that the anxiolytic-like effect of the 5-HT1B receptor ligands CP 94253, SB 216641 and GR 127935 was possibly linked to the postsynaptic 5-HT1B receptors or/and 5-HT1B heteroreceptors. The results suggested also that benzodiazepine receptors were indirectly involved in the effects of SB 216641 and GR 127935 (but not of CP 94253), which might have been due to a possible interaction between the 5-HT and the GABA/benzodiazepine systems.     

m-CPP, a 5-HT2C receptor agonist that modifies the perfusion pressure of the hindquarter vascular bed of anesthetized rat

In the present work we studied the actions of the intra-arterial administration of meta-chlorophenylpiperazine (m-CPP - a 5-HT(2C) receptor agonist) in the hindquarters of the anesthetized rat. The lowest doses used (0.001, 0.01, 0.1, 0.25 and 0.5 microg/kg) induced vasodilatation whereas the highest doses produced vasoconstriction (1, 6.25, 12.5 and 25 microg/kg). Both vasodilatation and vasoconstriction were inhibited by the 5-HT(1,2 )receptor antagonist methiothepin, whereas the 5-HT(2 )receptor antagonist ritanserin blocked only the vasoconstrictor responses. 1-[4-(1-Adamantanecarboxamido)butyl]-4-(2-methoxyphenyl)piperazine (a 5-HT(1A) receptor antagonist) and ICI 118,551 (a beta(2)-receptor antagonist) failed to modify the vasodilator responses of m-CPP. Both BRL 15572 (a 5-HT(1D) receptor antagonist) and GR 55562 (a 5-HT(1B) receptor antagonist) only partially inhibited this action. Our data reveal that m-CPP induces the 5-HT(1 )and/or non-specific vasodilator effect and 5-HT(2) vasoconstrictor effects in the hindquarter vascular bed of the rat.     

The 5-HT(1) receptors inhibiting the rat vasodepressor sensory CGRPergic outflow: further involvement of 5-HT(1F), but not 5-HT(1A) or 5-HT(1D), subtypes

We have previously shown that 5-HT(1B) receptors inhibit prejunctionally the rat vasodepressor CGRPergic sensory outflow. Since 5-HT(1) receptors comprise 5-HT(1A), 5-HT(1B), 5-HT(1D) and 5-HT(1F) functional subtypes, this study has further investigated the role of 5-HT(1A), 5-HT(1D) and 5-HT(1F) receptor subtypes in the inhibition of the above vasodepressor sensory outflow. Pithed rats were pretreated with i.v. continuous infusions of hexamethonium and methoxamine, followed by 5-HT(1) receptor agonists. Then electrical spinal stimulation (T(9)-T(12)) or i.v. bolus injections of exogenous α-CGRP produced frequency-dependent or dose-dependent vasodepressor responses. The electrically-induced vasodepressor responses remained unchanged during infusions of the 5-HT(1A) receptor agonists 8-OH-DPAT and NN-DP-5-CT. In contrast, these responses were inhibited by the agonists sumatriptan (5-HT(1A/1B/1D/1F)), indorenate (5-HT(1A)), PNU-142633 (5-HT(1D)) or LY344864 (5-HT(1F)), which did not affect the vasodepressor responses to exogenous CGRP (implying a prejunctional sensory-inhibition). When analysing the effects of antagonists: (i) 310 μg/kg (but not 100 μg/kg) GR127935 (5-HT(1A/1B/1D/1F)) abolished the inhibition to sumatriptan, indorenate, PNU-142633 or LY344864; (ii) 310 μg/kg SB224289 (5-HT(1B)) or BRL15572 (5-HT(1D)) failed to block the inhibition to sumatriptan or PNU-142633, whereas SB224289+BRL15572 partly blocked the inhibition to sumatriptan; and (iii) 10 μg/kg WAY100635 (5-HT(1A)) failed to block the inhibition to indorenate. These results suggest that 5-HT(1F), but not 5-HT(1A) or 5-HT(1D), receptor subtypes inhibit the vasodepressor sensory CGRPergic outflow although, admittedly, no selective 5-HT(1F) receptor agonist is available yet. The pharmacological profile of these receptors resembles that shown in rat dorsal root ganglia by molecular biology techniques.     

Vasoconstriction in human isolated middle meningeal arteries: determining the contribution of 5-HT1B- and 5-HT1F-receptor activation

AIMS: Sumatriptan is a 5-HT1B/1D-receptor agonist which also has affinity for 5-HT1F-receptors. The vasoconstrictor effects of sumatriptan are thought to be 5-HT1B-receptor mediated and these receptors have been shown to be expressed in human cranial blood vessels. However, in the same tissue mRNA coding for 5-HT1F-receptors has also been identified and this study addresses the possibility of whether 5-HT1F-receptor activation contributes to vasoconstriction. METHODS: The ability of two selective 5-HT1B/1D-receptor antagonists (GR125,743 and GR127,935) with no affinity for 5-HT1F-receptors, to inhibit sumatriptan evoked contractions in human isolated middle meningeal artery was investigated. Using a series of 5-HT1B/1D-receptor agonists (sumatriptan, zolmitriptan, CP122,288, L-741,519 and L-741,604), some with high affinity for 5-HTIF-receptors and the non-selective 5-HT-receptor agonists 5-HT and 5-CT, we compared the vasoconstrictor potency of these drugs in human isolated middle meningeal artery with their affinities at cloned human 5-HT1B-, 5-HT1D-and 5-HT1F-receptors expressed in CHO cell lines. RESULTS: GR125,743 antagonized sumatriptan evoked contractions in a competitive manner (apparent pA2 9.1) and GR127,935 antagonized sumatriptan-induced responses in a non-competitive manner (reducing the maximum contraction to 27%). There was a significant correlation between vasoconstrictor potency and 5-HT1B-receptor affinity (r=0.93, P=0.002) but not with 5-HT1D- or 5-HT1F-receptor affinity (r=0.74, P=0.06; r= 0.31, P= 0.49, respectively). CONCLUSIONS: These experiments show that in human middle meningeal artery vasoconstriction to sumatriptan-like agents is 5-HT1B-receptor mediated with little if any contribution from 5-HT1F-receptor activation.     

Potential vascular alpha1-adrenoceptor blocking properties of an array of 5-HT receptor ligands in the rat

This study set out to analyse the potential ability of some 5-hydroxytryptamine (5-HT) receptor ligands widely used in cardiovascular experimental models to interact with vascular alpha1-adrenoceptors in the pithed rat. These ligands included: methiothepin, methysergide and metergoline (5-HT(1)/5-HT2); WAY-100635, buspirone, ipsapirone and 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) (5-HT(1A)); GR127935 (5-HT(1B/1D)); ketanserin, ritanserin, spiperone and pizotifen (5-HT2); granisetron and metoclopramide (5-HT3); tropisetron (5-HT3/5-HT4); ergotamine (5-HT(1B/1D), 5-ht(5A/5B)); clozapine (5-HT6/5-HT7); as well as LY215840 and mesulergine (5-HT2/5-HT7). For this purpose, the increases in diastolic blood pressure produced by the selective alpha1-adrenoceptor agonist, phenylephrine, were analysed before and after the above antagonists or saline. The adrenoceptor antagonist properties of prazosin (alpha1) and yohimbine (alpha2) were also analysed for comparison. Thus, the phenylephrine-induced vasopressor responses were dose-dependently antagonised with the following apparent rank order of potency by: prazosin > or = methiothepin > ketanserin > clozapine > or = lisuride > buspirone; this potency correlates with the affinity of these compounds for alpha1-adrenoceptor binding sites. In contrast, the other compounds were either devoid of any blocking effect on--or even potentiated (i.e. lisuride, methysergide, 8-OH-DPAT, granisetron and GR127935)--the responses to phenylephrine. These results show that methiothepin, ketanserin, clozapine, lisuride and buspirone can block alpha1-adrenoceptors in the rat systemic vasculature.     

Enhancement of 5-HT1B and 5-HT1D receptor antagonist effects on extracellular 5-HT levels in the guinea-pig brain following concurrent 5-HT1A or 5-HT re-uptake site blockade.

The effects of selective serotonin re-uptake inhibitor (SSRI), paroxetine, and 5-HT1A, 5-HT1B and 5-HT1B/1D receptor antagonists on in vivo extracellular 5-HT levels in the guinea-pig frontal cortex and dorsal hippocampus were investigated using the technique of microdialysis. The aim of the study was to further investigate the autoreceptor roles of the 5-HT1A, 5-HT1B and 5-HT1D receptors in the median vs dorsal raphe nuclei. In the frontal cortex, 5-HT1A (WAY 100635, 1 mg/kg i.p.) or 5-HT1B (SB-224289, 4 mg/kg i.p.) receptor antagonists had no effect on extracellular levels of 5-HT, whilst the mixed 5-HT1B/1D receptor antagonist (GR 127935, 0.3 mg/kg i.p) produced a significant decrease in extracellular 5-HT levels. Paroxetine (10 microM) significantly increased extracellular 5-HT levels when perfused locally into the cortex. Administration of SB-224289, followed 120 min later by WAY 100635, had no effect on extracellular 5-HT levels. In contrast, sequential administration of either WAY 100635 and GR 127935, or SB-224289 and paroxetine significantly increased extracellular 5-HT levels. In the dorsal hippocampus, whilst 5-HT1A receptor antagonism elicited by administration of WAY 100635 had no effect, both 5-HT1B and mixed 5-HT1B/1D receptor blockade significantly increased extracellular 5-HT levels. Administration of SB-224289 followed 120 min later with WAY 100635, or WAY 100635 followed 30 min later with GR 127935, potentiated the effect of the three compounds alone, significantly increasing extracellular 5-HT levels. These data demonstrate that to simultaneously increase extracellular 5-HT in both frontal cortex and dorsal hippocampus of the guinea-pig brain concurrent 5-HTA1A, 5-HT1B and 5-HT1D receptor blockade is required. Whereas in the dorsal hippocampus, 5-HT1B receptor blockade is sufficient to elicit an increase in extracellular 5-HT levels.     

Pharmacological studies of the acute effects of (+)-3,4-methylenedioxymethamphetamine on locomotor activity: role of 5-HT(1B/1D) and 5-HT(2) receptors.

The role of serotonin 5-HT(2) receptors (5-HT(2)R) in the hyperactivity induced by (+)-3,4-methylenedioxy-methamphetamine ((+)-MDMA; 3 mg/kg) was investigated. Hyperactivity induced by (+)-MDMA was robustly potentiated by the 5-HT(2B/2C)R antagonist SB 206553 (1.0, 2.0, and 4.0 mg/kg). Administration of the 5-HT(1B/1D)R antagonist GR 127935 (2.5 mg/kg) or the 5-HT(2A)R antagonist M100907 (1.0 mg/kg) partially suppressed the potentiated hyperactivity seen following SB 206553 plus (+)-MDMA; a blockade to activity levels seen with (+)-MDMA alone was observed following the combination of GR 127935 plus M100907. A modest potentiative interaction was seen when SB 206553 was combined with the DA releaser amphetamine (0.5 mg/kg) or amphetamine plus the 5-HT releaser fenfluramine (4.0 mg/kg). SB 206553 (1-4 mg/kg), GR 127935 (2.5 mg/kg) and M100907 (1 mg/kg) did not alter spontaneous activity upon administration singly or in combination. These data suggest that activation of 5-HT(2C)R exerts a strong inhibitory influence on the hyperactivity induced by (+)-MDMA, and that 5-HT(2C)R blockade unmasks hyperactivity mediated through several mechanisms.     

The in vivo pharmacological profile of eletriptan (UK-116,044): a potent and novel 5-HT(1B/1D) receptor agonist

The anti-migraine drug, eletriptan [(R)-3-(1-methyl-2-pyrrolidinylmethyl)-5-[2-(phenylsulphonyl )ethyl]-1 H-indole; UK-116,044], is a novel 5-HT(1B/1D) receptor agonist. In this paper, the regional vasoconstrictor profile of eletriptan, in comparison with sumatriptan, was examined in the anaesthetised dog. The inhibitory actions of eletriptan on neurogenic inflammation in rat dura mater were also assessed. In the anaesthetised dog, eletriptan (1-1000 microg kg(-1) i.v.) produced a dose-dependent reduction of carotid arterial blood flow with a similar potency and maximum effect to sumatriptan (ED(50) values: eletriptan and sumatriptan, 12 and 9 microg kg(-1), i.v., respectively). However, eletriptan exhibited a significantly lower potency than sumatriptan in reducing coronary artery diameter (ED(50) values: 63 and 19 microg kg(-1), i.v., respectively, P<0.05). In the femoral circulation, sumatriptan caused a significant reduction in arterial blood flow (ED(50) 35 microg kg(-1) i.v.) whereas eletriptan (1-1000 microg kg(-1) i.v.) had no significant effect upon femoral arterial blood flow when compared to vehicle-treated animals. In rats, eletriptan (30-300 microg kg(-1) i.v.) administered prior to electrical stimulation of the trigeminal ganglion produced a dose-related and complete inhibition of plasma protein extravasation in the dura mater (mean extravasation ratio: control 1.9; eletriptan 1.0, minimum effective dose 100 microg kg(-1), P<0.05). The potency and maximum effect of eletriptan was identical to that of sumatriptan in this model. When administered during a period of continual stimulation of the trigeminal nerve, eletriptan (100 microg kg(-1) i.v.) produced a complete inhibition of plasma protein extravasation. The ability to reduce canine carotid arterial blood flow and inhibit neurogenic inflammation in rat dura mater suggests that vascular and neurogenic mechanisms may contribute to eletriptan's clinical efficacy in migraine patients. In addition, eletriptan exhibits some selectivity for reducing carotid arterial blood flow when compared with femoral arterial blood flow and coronary artery diameter, in the anaesthetised dog.     

Further evidence that the discriminative stimulus properties of indorenate are mediated by 5-HT 1A/1B/2C receptors

Indorenate (5-methoxytryptamine beta-methylcarboxylate, INDO) is a serotonin (5-hydroxytryptamine, 5-HT) agonist that has affinity for 5-HT(1A/1B/2C) receptors. Unlike other anxiolytics such as 5-HT receptor agonists, INDO may not share tolerance or dependency with the benzodiazepine anxiolytics. It has been reported that the discriminative stimulus properties of 5-HT(1A/1B/2C) agonists, but not those of 5-HT(3/4) agonists, generalize to INDO. Therefore, the aim of the present study was to obtain further evidence on the differential involvement of 5-HT(1A/1B/2C) receptors in the discriminative stimulus properties of INDO by evaluating its interactions with antagonists of the 5-HT(1A), 5-HT(1B), 5-HT(2C), and 5-HT(3/4) receptor subtypes. Rats were trained to discriminate INDO from saline in a conditioned taste aversion paradigm. For Group D(+)S(-), administration of INDO signalled that saccharin flavour was followed by LiCl, while injection of vehicle signalled safe consumption of saccharin solution. Group D(-)S(+) had the contingencies reversed. After this training, rats had generalization tests where INDO administration was preceded by different doses of the following antagonists: WAY100635 (5-HT(1A)), NAN190 (5-HT(1A)), methiothepin (5-HT(1A/1B/2C)), GR127935 (5-HT(1B/1D)), ketanserin (5-HT(2A/2C)), ritanserin (5-HT(2C/2A)), mesulergine (5-HT(2C/2A)), metergoline (5-HT(2C/2A)), SB206553 (5-HT(2B/2C)), and tropisetron (5-HT(3/4)). In Group D(+)S(-), the order of potency to block the discriminative stimulus properties of INDO was WAY100635>ketanserin>ritanserin>GR127935>mesulergine congruent with SB206553>metergoline>methiothepin>NAN190, while in Group D(-)S(+), the order was WAY100635>GR127935>ketanserin>ritanserin>mesulergine congruent with SB206553>metergoline>methiothepin>NAN190. Tropisetron did not produce any alteration of the discriminative control by INDO. These results suggest that the discriminative signal of INDO is mediated by 5-HT(1A/2C/1B) receptors and that blockade of any of its components produces a degradation of its discriminative effects.     

Characterization of lipopolysaccharide-induced emesis in conscious piglets: effects of cervical vagotomy, cyclooxygenase inhibitors and a 5-HT(3) receptor antagonist

The emetic response to intraperitoneal (i.p., 0.5, 2, 8 mg kg(-1)) and intravenous (i.v., 200 microg kg(-1)) administration of bacterial lipopolysaccharides (LPS) was characterized in conscious piglets observed for 4 h. The latencies and the incidence of the emetic response to LPS (i.p.) decreased and increased, respectively, in a dose-dependent manner. In 14 additional piglets, a bilateral vagotomy performed 4 h prior to LPS administration abolished the vomiting induced by i.p. LPS (2 mg kg(-1)), and decreased its incidence by 77% in the i.v. injected animals. Sham-operated animals (n=6) exhibited a similar emetic pattern to the controls injected intraperitoneally with LPS (2 mg kg(-1)). In 7 piglets, the administration of granisetron, a 5-HT(3) receptor antagonist (i.v., 2 mg kg(-1)), 30 min prior to the i.p. LPS injection (2 mg kg(-1)) failed to reduce significantly the emetic activity; whereas, in 6 animals, a combination of meloxicam (0.3 mg kg(-1)) and indomethacin (5 mg kg(-1)), two cyclooxygenase (COX) inhibitors, administered per os 1.5 h prior to the i.p. LPS (2 mg kg(-1)) abolished the emetic response to endotoxins. The present results show that the activation of the medullary "vomiting centre" in response to i.p. administration of LPS is mediated via vagal afferents and is likely to involve prostaglandins.