Classification and nomenclature of somatostatin receptors.

There is considerable controversy about the classification and nomenclature of somatostatin receptors. To date, five distinct receptor genes have been cloned and named chronologically according to their respective publication dates, but two were unfortunately given the same appellation (SSTR4). Consensually, a nomenclature for the recombinant receptors has been agreed according to IUPHAR guidelines (sst1, sst2, sst3, sst4, and sst5). However, a more informative classification is to be preferred for the future, employing all classification criteria in an integrated scheme. It is already apparent that the five recombinant receptors fall into two classes or groups, on the basis of not only structure but also pharmacological characteristics. One class (already referred to by some as SRIF1) appears to comprise sst2, sst3 and sst5 receptor subtypes. The other class (SRIF2) appears to comprise the other two recombinant receptor subtypes (sst1 and sst4). This promising approach is discussed but it is acknowledged that much more data from endogenous receptors in whole tissues are needed before further recommendations on somatostatin receptor nomenclature can be made.

Serotonin receptors Therapeutic prospects in cardiovascular disease.

Serotonin is widely distributed throughout the body, can have potent and profound effects on the cardiovascular system, and may be involved in the pathophysiology of several cardiovascular disease states. Recent advances in the classification of serotonin receptors, along with the identification of selective drug tools for some of these receptor types, enables the putative role of different serotonin receptor subtypes in both the etiology and treatment of disease states to be explored. This review considers the potential therapeutic application of selective serotonin receptor agonists and antagonists in cardiovascular disease.

Effect of sumatriptan, a selective 5-HT1-like receptor agonist, on pial vessel diameter in anaesthetised cats.

The action of sumatriptan, a selective 5-HT1-like receptor agonist that is effective for the acute treatment of migraine, was compared on pial vessel diameter following perivascular or intravenous administration to anaesthetised cats. Sumatriptan (0.01-10 microM), when microinjected perivascularly, caused a decrease in pial artery diameter (maximum change of -19 +/- 9%; mean +/- SD) but had no effect on the diameter of pial veins. Sumatriptan (1 microM)-induced pial artery vasoconstriction was unaffected by coadministration of ketanserin (1 microM) or ondansetron (1 microM) but was significantly (p less than 0.01) attenuated by methiothepin (1 microM). Intravenous infusion of a clinically effective dose of sumatriptan (64 micrograms/kg/10 min) caused selective carotid vasoconstriction (22 +/- 6% increase in carotid vascular resistance with little or no change in blood pressure or heart rate) and no change in pial artery diameter, although sumatriptan (1 microM) administered perivascularly in these animals before and after the infusion caused pial artery vasoconstriction. These results demonstrate that perivascularly administered sumatriptan causes pial artery vasoconstriction via activation of 5-HT1-like receptors. However, intravenously administered sumatriptan does not cause pial artery vasoconstriction, which suggests that sumatriptan does not readily penetrate the cerebrovascular intima.

Effects of the 5-HT1D receptor antagonist GR127935 on extracellular levels of 5-HT in the guinea-pig frontal cortex as measured by microdialysis.

The involvement of 5-HT1D receptors in the regulation of 5-HT release in the guinea-pig brain was examined using the novel 5-HT1D receptor blocking drug GR127935. Levels of 5-HT were measured in frontal cortex of anaesthetized guinea-pigs using microdialysis. The infusion of GR127935 (100 nM) through the dialysis probe into frontal cortex caused a significant increase (61 +/- 8%) in cortical extracellular levels of 5-HT. The increase was transient (approximately 40 min) even in the continuous presence of GR127935. The transient increase was abolished by tetrodotoxin (1 microM). The 5-HT1 receptor agonist GR46611 (10 mg/kg s.c.) caused a significant and sustained (> 100 min) reduction in extracellular levels of 5-HT (65 +/- 5%). This response was abolished in animals pre-treated with GR127935, 0.05 mg/kg i.p. Paradoxically, systemic administration of higher doses of GR127935 (0.1-1 mg/kg i.p.) in naive anaesthetized guinea-pigs caused significant and sustained (> 120 min) decreases (> 65%) in cortical levels of 5-HT. The increase in extracellular 5-HT seen following infusion of GR127935 into frontal cortex may be due to GR127935 blocking 5-HT terminal autoreceptors causing a subsequent increase in the outflow of 5-HT from pre-synaptic terminals. This conclusion is supported by the ability of GR127935 to block the decrease in 5-HT induced by the 5-HT1 receptor agonist GR46611.(ABSTRACT TRUNCATED AT 250 WORDS)

Selective somatostatin sst(2) receptor blockade with the novel cyclic octapeptide, CYN-154806.

The cyclic octapeptide, CYN-154806, inhibited specific [(125)I]-[Tyr(11)]-SRIF binding to CHO-K1 cell membranes expressing human recombinant somatostatin (SRIF) sst(2) receptors (pIC(50) 8. 58) or rat sst(2(a)) and rat sst(2(b)) receptors (pIC(50) 8.35 and 8. 10, respectively). The affinity of CYN-154806 at other human somatostatin receptor types was at least 100 times lower (pIC(50) 5. 41-6.48). In functional studies, CYN-154806 inhibited SRIF-induced increases in extracellular acidification (EAR) in CHO-K1 cells expressing h sst(2) receptors (pK(B) 7.92) but had no effect on UTP-induced increases in EAR. CYN-154806 also blocked SRIF-induced increases [(35)S]-GTPgammaS binding in CHO-K1 cell membranes expressing h sst(2) receptors as well as rat sst(2(a)) and rat sst(2(b)) receptors (pK(B) 7.81, 7.68 and 7.96, respectively). In marked contrast, no blockade was observed at h sst(5) receptors in concentrations as high 10 microM. The antagonistic activity of CYN-154806 was also studied in isolated tissue preparations that are known to express endogenous SRIF receptors. Thus CYN-154806 blocked SRIF, but not DAMGO-induced inhibition of neurogenic contractions in rat isolated vas deferens and guinea-pig ileum (pK(B) 7.79 and 7.49, respectively). CYN-154806 had no effect on SRIF-28 induced inhibition of neurogenic contractions in guinea-pig vas deferens. The results demonstrate that CYN-154806 is a highly potent specific and selective SRIF sst(2) receptor blocking drug. Furthermore, sst(2) receptors mediate SRIF-induced inhibition of neurogenic contractions in rat vas deferens and guinea-pig ileum but not guinea-pig vas deferens which is thought to be mediated by sst(5) receptors.

A quantitative autoradiographical study on the distribution of somatostatin sst2 receptors in the rat central nervous system using [125I]-BIM-23027.

The kinetic properties, steady state binding characteristics and autoradiographic distribution of the somatostatin (SRIF) sst2 receptor-selective ligand, [125I]-BIM-23027, have been investigated in the rat central nervous system. Analysis of kinetic, saturation and competition binding data in rat hippocampal membranes was consistent with [125I]-BIM-23207 binding to a single population of non-interacting binding sites. Competition studies, using different SRIF ligands suggested that [125I]-BIM-23027 was binding to sites similar to that of the recombinant sst2 receptor. The rank order of affinity for displacing specific binding was BIM-23027 = SRIF > L-362855 > > BIM-23056. There was a widespread distribution of [125I]-BIM-23027 binding sites in the rat central nervous system. The highest density of binding was observed in the dentate gyrus, medial habenular, amygdala, claustrum and lateral septum as well as in the piriform, cingulate and parietal cortex. The cervical and lumbar spinal cord also displayed moderate levels of binding localized to the substantia gelatinosa. The cellular localization of [125I]-BIM-23027 binding was found to be associated with dendritic terminal fields. In contrast, the cellular signal for sst2 receptor mRNA was restricted to cell somata. The widespread distribution of [125I]-BIM-23027 binding sites within the brain suggests that receptors similar to the recombinant sst2 receptor may mediate a variety of different physiological effects within the central nervous system.

Proposals for the classification and nomenclature of functional receptors for 5-hydroxytryptamine.

As a result of controversy in the literature regarding the classification and nomenclature of functional receptors for 5-hydroxytryptamine (5-HT), a framework for classification is proposed. The formulation of these proposals has only been made possible by the recent advent of new drug tools. It is considered that there are three main types of 5-HT receptor, two of which have been well characterised pharmacologically, using selective antagonists, and which it is proposed to name 5-HT2 and 5-HT3. These two groups broadly encompass the "D" and "M" receptors, respectively, which Gaddum identified in the guinea-pig ileum (Gaddum and Picarelli, 1957). The 5-HT2 receptor, which mediates a variety of actions of 5-HT, has been definitively shown to correlate with the 5-HT2 binding site in the brain. No binding studies in brain tissue have yet been published with radiolabelled ligands specific for 5-HT3 receptors. A number of other actions of 5-HT appear to be mediated via receptors distinct from 5-HT2 or 5-HT3 receptors. Since selective antagonists are not yet available, these receptors cannot be definitively characterised, although in many cases they do have some similarities with 5-HT1 binding sites, which are a heterogeneous entity. Criteria are proposed for tentatively classifying these receptors as "5-HT1-like" (Table 1). Definitive characterisation of these receptors will await the identification of specific antagonists. This classification of 5-HT receptors into three main groups (Table 1) is based largely, but not exclusively, on data from studies in isolated peripheral tissues where definitive classification is possible. However, it is believed that this working classification will be relevant to functional responses to 5-HT in the central nervous system.

The effects of prostaglandins E1, E2 and F2alpha on vagal bradycardia in the anaesthetized mouse.

1 In anesthetized mice prostaglandins E1 and E2 reduced the bradycardia caused by electrical stimulation of the sectioned peripheral vagus nerve; prostaglandin F2alpha produced only a slight inhibition of the vagal response. 2 None of the prostaglandins studied affected acetylcholine-induced bradycardia. 3 Prostaglandins modify parasympathetic nerve activity in vivo presumably by a pre-synaptic action.

Influence of the endothelium on contractile effects of 5-hydroxytryptamine and selective 5-HT agonists in canine basilar artery.

1. We have investigated the influence of endothelial damage on the cerebrovascular reactivity to 5-hydroxytryptamine (5-HT) and some selective 5-HT agonists in canine basilar artery. 2. 5-HT, alpha-methyl 5-HT, GR 43175 (3-[2-dimethyl amino] ethyl-N-methyl-1H-indole-5-methane sulphonamide) and 5-carboxamidotryptamine (5-CT) produced concentration-dependent contractions of untreated dog basilar artery with a functional endothelium. Following endothelial damage by perfusion with Triton X-100 (0.1%), which abolished the relaxant response to substance P, the maximum contractile effect of 5-HT, alpha-methyl 5-HT, GR 43175 and 5-CT was markedly enhanced although there was little change in the EC50 values. Endothelial damage did not modify the vasoconstrictor effect of the thromboxane agonist, U46619, or potassium chloride. 3. Neither 5-HT nor 5-CT caused relaxation of untreated canine basilar arteries contracted with prostaglandin F2 alpha, U46619, uridine triphosphate or potassium chloride. 4. These results suggest that canine basilar artery spontaneously releases endothelium-derived relaxing factor which can attenuate the vasoconstrictor effect of 5-HT and selective 5-HT agonists. This effect appeared to be specific since the vasoconstrictor response to U46619 was not modified. 5. These results demonstrate that the cerebrovascular endothelium can markedly influence the reactivity of the vascular smooth muscle of canine basilar artery to 5-HT and 5-HT1-like receptor agonists. However we could find no evidence that 5-HT receptor activation stimulates endothelial cell function as it does in some other blood vessels.

Characterization of the prostanoid receptor types involved in mediating calcitonin gene-related peptide release from cultured rat trigeminal neurones.

1. Prostaglandins and the vasodilator neuropeptide, calcitonin-gene related peptide (CGRP), have both been implicated in the pathogenesis of migraine headache. We have used primary cultures of adult rat trigeminal neurones to examine the effects of prostanoids on CGRP release in vitro. 2. CGRP release was stimulated by prostaglandin E2 (PGE2) and the IP receptor agonist, carbaprostacyclin (cPGI2). These responses were extracellular calcium-dependent, and the PGE2-induced CGRP release was unaltered by inhibition of nitric oxide synthase (NOS), ATP receptor blockade, or the addition of adenosine deaminase. 3. Increases in CGRP levels were also observed in response to prostaglandin D2 (PGD2), and the EP2 receptor selective agonist, butaprost. No increases in CGRP release were observed in response to prostaglandin F2alpha (PGF2alpha) or the TP receptor selective agonist, U46619, or the EP3 receptor selective agonist, GR63799X. 4. The selective DP receptor antagonist, BWA868C, antagonized the PGD2-, but not PGE2- or cPGI2-induced release. Furthermore, the EP1 selective antagonist, ZM325802, failed to antagonize the PGE2-induced CGRP release from these cells. 5. These data indicate that activation of DP, EP and IP receptors can each cause CGRP release from trigeminal neurones, and suggest that the predominant EP receptor subtype involved may be the EP2 receptor. Together with evidence that the cyclo-oxygenase inhibitor, aspirin, particularly when administered intravenously is effective in treating acute migraine, these findings further suggest a role for prostaglandins in migraine pathophysiology.

Rabbit isolated renal artery contractions by some tryptamine derivatives, including 2-methyl-5-HT, are mediated by a 5-HT1-like receptor.

1. Despite the fact that 5-hydroxytryptamine (5-HT)-induced contractions of the rabbit isolated renal artery are mediated by a receptor belonging to the heterogeneous 5-HT1-like category, we observed that the so-called selective 5-HT3 receptor agonist, 2-methyl-5-HT, caused a concentration-dependent contraction of this vessel. This study was therefore undertaken to analyze the effects of 2-methyl-5-HT in the renal artery segments, either quiescent or precontracted with U46619 (10(-7) M). alpha-Methyl-5-HT and 5-methoxytryptamine, which have high affinities for 5-HT2 and 5-HT4 receptors, respectively, were used for comparison. 2. In the precontracted vessel segments, the maximum contractile responses obtained with 2-methyl-5-HT, alpha-methyl-5-HT, 5-methoxytryptamine and 5-HT were similar to those in the quiescent segments. However the pD2 values were higher in the precontracted segments, making them about 4-100 fold more sensitive. 3. Neither MDL 72222 (10(-6) M) nor tropisetron (3 x 10(-6) M) suppressed renal artery contractions elicited by 5-HT, 2-methyl-5-HT, alpha-methyl-5-HT or 5-methoxytryptamine, thus ruling out the involvement of 5-HT3 as well as 5-HT4 receptors. 4. On the other hand, both methiothepin (10(-8) and 10(-7) M) and ketanserin (10(-7) and 10(-6) M) caused a rightward shift of agonist concentration-effect curves.The two antagonists had similar pA2 values against the different agonists tested on either quiescent or precontracted vessels, but ketanserin (apparent pA2: 6.6 to 7.0) was between 20-100 fold less potent than methiothepin (apparent pA2: 8.4 to 8.8).5. The results of this functional study permit us to conclude that the contractile effects of 2-methyl-5-HT as well as ct-methyl-5-HT and 5-methoxytryptamine on the rabbit isolated renal artery are mediated by a 5-HT1-like receptor. Since, in addition, the reported ligand binding affinity of 2-methyl-5-HT at 5-HT3 receptors is similar to both the ligand binding affinity and the functional pD2 at 5-HTI sites, this compound cannot be regarded as a selective 5-HT3 receptor agonist. Similarly, a-methyl-5-HT and 5-methoxytryptamine have only a limited selectivity for 5-HT2 and 5-HT4 receptors, respectively.

Characterization of somatostatin receptors in guinea-pig isolated ileum, vas deferens and right atrium.

1. Somatostatin14 (SS14) inhibits neurogenically mediated contractile responses in guinea-pig ileum and vas deferens and exerts a direct negative inotropic action in guinea-pig spontaneously beating right atrium. In this study, the receptors mediating these inhibitory effects have been characterized by comparing the potencies of several cyclic somatostatin analogues. 2. In the guinea-pig ileum, SS14, somatostatin28 (SS28), somatostatin25 (SS25) and several smaller cyclic somatostatin analogues including octreotide, angiopeptin and CGP 23996, inhibited neurogenically mediated contractile responses, each being of similar potency. 3. In contrast, in the guinea-pig vas deferens and right atrium, SS28 was about 30 times more potent than SS14. However, although angiopeptin was nearly as potent as SS14 as an agonist in the vas deferens, in guinea-pig atrium angiopeptin had low intrinsic activity and antagonized the negative inotropic action of both SS14 and SS28 (pKB values of 7.4 and 7.2, respectively). CGP 23996 was 2-7 times weaker than SS14 in guinea-pig vas deferens and atria. 4. Phosphoramidon (1 microM) and amastatin (10 microM) did not influence the potency of SS14 or SS28 in either the guinea-pig ileum or right atrium. In the guinea-pig vas deferens, phosphoramidon and amastatin did not affect the potency of SS28, but enhanced the potency of SS14 about 5 fold. Despite the presence of phosphoramidon and amastatin, SS28 was still more potent than SS14 in the vas deferens. 5. The putative somatostatin receptor blocking drug, cyclo(7-aminoheptanoyl Phe-D-Trp-Lys-Thr[Brl]) (CPP; 1 microM), did not antagonize the effects of either SS14 or SS28 in ileum, vas deferens or atrial preparations. 6. Somatostatin14 did not modify the contractile action of carbachol or alpha,beta-methylene ATP in the ileum and vas deferens respectively, suggesting that the site of the inhibitory effects on neurogenically mediated contractile responses in both preparations was pre-junctional. Consistent with this conclusion was the observation that the inhibitory effect of SS14 was markedly and inversely related to the external Ca2+concentration. The inhibitory effect of SS14 in guinea-pig atrium was only partly dependent on the external Ca2+ concentration.7. The somatostatin receptors mediating the inhibitory effect of SS14 in the ileum and vas deferens can be distinguished by the differential relative potencies of SS14 and SS28. In the former, SS14 and SS28 have similar potency whilst in the latter SS28 is much more potent. In this respect, the somatostatin receptor mediating negative inotropy in the guinea-pig right atrium appears similar to that identified in the vas deferens.8. We suggest that the somatostatin receptor mediating inhibition of neurogenic contraction in the ileum is similar to the recently cloned SSTR2 receptor. In contrast, the somatostatin receptor mediating negative inotropy in the atrium and inhibition of neurotransmission in the vas deferens appears similar to the SSTR4 receptor which recognises SS28 with higher affinity than SS14.

Antagonist effects of seglitide (MK 678) at somatostatin receptors in guinea-pig isolated right atria.

Somatostatin (SS) exerts a negative inotropic effect in isolated atria. Here we report that in guinea-pig isolated right atria, seglitide, a potent cyclic hexapeptide somatostatin agonist, behaves as a competitive somatostatin receptor antagonist with pA2 values against SS14, SS25 and SS28, of 6.50 +/- 0.40, 6.24 +/- 0.08 and 6.09 +/- 0.06, respectively. Seglitide had little or no effect on the negative inotropic action of carbachol or N6-cyclohexyladenosine. Our findings indicate that the receptor-response coupling characteristics of guinea-pig atria are such that in this preparation seglitide has low intrinsic activity and behaves specifically as a somatostatin receptor antagonist.

Characterization of human recombinant somatostatin sst5 receptors mediating activation of phosphoinositide metabolism.

1. We have functionally characterized the human recombinant somatostatin (SRIF) sst5 receptor in Chinese hamster ovary-K1 (CHOsst5) cells by measuring total [3H]-inositol phosphate ([3H]-InsPx) accumulation, in the presence of 10 mM LiCl, in cells labelled with [3H]-myo-inositol. 2. In CHOsst5 cells, SRIF, SRIF-28 and the cyclic hexapeptide, L-362,855, produced time- and concentration-related increases in [3H]-InsPx accumulation, with similar potency (pEC50 values of 6.5, 6.8 and 7.2, respectively). L-362,855 behaved as a partial agonist, producing approximately 30% of the SRIF maximum response. The other peptide analogues of SRIF, BIM-23027 and BIM-23056, were inactive as agonists. 3. Increasing concentrations of L-362,855 increased [3H]-InsPx accumulation and simultaneously produced rightward shifts of SRIF concentration-effect curves, with an estimated pKp value of 7.6, confirming that it was acting as a partial agonist. 4. BIM-23056, but not BIM-23027, potently antagonized SRIF-induced [3H]-InsPx accumulation, with an estimated pKB value of 7.4. BIM-23056 did not antagonize [3H]-InsPx accumulation induced by uridine 5'-triphosphate (UTP). 5. SRIF- but not UTP-induced [3H]-InsPx accumulation was inhibited by increasing concentrations of pertussis toxin (0.01-100 ng ml-1), indicating the involvement of pertussis toxin-sensitive G-proteins. 6. These findings show that the human recombinant sst5 receptor, when stably expressed in CHO-K1 cells, is able to mediate activation of phosphoinositide metabolism in a pertussis toxin-sensitive manner. In this system L-362,855 behaved as a partial agonist while BIM-23056 was a specific antagonist. These agents should provide useful tools for functionally characterizing endogenous SRIF receptors.

Mediation by SRIF1 receptors of the contractile action of somatostatin in rat isolated distal colon; studies using some novel SRIF analogues.

1. The motor effects of somatostatin-14 (SRIF), and several SRIF peptide analogues were investigated on the rat isolated distal colon. The objective of these studies was to characterize the receptor mediating the contractile action of SRIF by comparing the relative agonist potencies of a range of SRIF analogues. 2. SRIF (1 nM-1 microM) produced concentration-dependent contractions with an EC50 value of approximately 10 nM. Contractile responses induced by SRIF were insensitive to atropine (1 microM) or naloxone (1 microM) but abolished by tetrodotoxin (1 microM). Somatostatin-28 (SRIF28), also induced concentration-dependent contractions and was equipotent with SRIF. Phosphoramidon (1 microM) and amastatin (10 microM) did not increase the potency of either SRIF or SRIF28. 3. The SRIF peptide analogues, octreotide, SRIF25, seglitide, angiopeptin and CGP23996 (1 nM-1 microM) produced contractile responses in the rat distal colon, each having similar potency and maximal activity relative to SRIF. The SSTR2 receptor-selective hexapeptide, BIM23027 (0.1 nM-1 microM), and the SRIF stereoisomer, D-Trp8-SRIF (0.1 nM-1 microM), were the most potent agonists examined being approximately 12 and 7 times more potent than SRIF, respectively. In contrast, the SSTR5 receptor-selective analogue, L362,855, was approximately 120 times weaker than SRIF, whilst the SSTR3 receptor-selective analogue, BIM23056, was inactive at concentrations up to 3 microM. 4. The putative SRIF receptor antagonist, (cyclo(7-aminoheptanoyl Phe-D-Trp-Lys-Thr[Bzl]))(CPP) (1 microM), had no agonist activity and had no effect on contractions induced by SRIF. 5. The contractile actions of BIM23027 and seglitide were subject to pronounced desensitization. Desensitization of preparations by BIM23027 (0.3 JIM) abolished the contractile action of SRIF andSRIF28 but had no effect on contractions produced by acetylcholine (0.1 nM-I1M), suggesting thatBIM23027, SRIF and SRIF28 act via a common receptor mechanism.6. In conclusion, the rat isolated distal colon contracts in response to SRIF and a number of SRIF analogues. Seglitide and octreotide exhibited similar potency and maximal activity relative to SRIF,suggesting that in the rat colon the receptor mediating contraction belongs to the SRIF,-receptor group,of which the recombinant SSTR2, SSTR3 and SSTR5 receptors appear to be subtypes. The high potency of BIM23027, the weak agonist activity of L362,855 and the lack of activity exhibited by BIM23056suggests that the SRIF receptor mediating contraction in the rat distal colon is similar to there combinant SSTR2 receptor.

The selective carotid arterial vasoconstrictor action of GR43175 in anaesthetized dogs.

1. GR43175 is a highly selective agonist at 5-HT1-like receptors in the dog saphenous vein. This study describes the haemodynamic effects of GR43175 in barbitone-anaesthetized dogs. 2. GR43175 (1-1000 micrograms kg-1, i.v.) produced dose-dependent decreases in carotid arterial blood flow with little or no change in arterial blood pressure. The decrease in blood flow was associated with an increase in carotid arterial vascular resistance. In preliminary studies, the dose of GR43175 producing 50% of the maximum carotid vasoconstrictor response was 39 +/- 8 micrograms kg-1, i.v. 3. In comparative regional haemodynamic studies, GR43175 (1-1000 micrograms kg-1, i.v.) had little effect on total peripheral resistance or resistance in the mesenteric, vertebral and coronary arterial vascular beds. Low doses of GR43175 decreased, whilst high doses (100 micrograms kg-1, i.v. and above) increased femoral arterial vascular resistance. GR43175 (1-1000 micrograms kg-1, i.v.) had no effect on respiratory inflation pressure. In doses of 100 micrograms kg-1 i.v. and above, GR43175 caused small decreases in heart rate. 4. The carotid arterial vasoconstrictor action of GR43175 was resistant to antagonism by the 5-HT2 receptor, 5-HT3 receptor and alpha-adrenoceptor blocking drugs, ketanserin, MDL72222 and phentolamine respectively, but could be antagonized by the non-selective 5-HT1-like receptor blocking drug methiothepin. Methiothepin had no effect on the carotid vasoconstrictor action of the thromboxane A2 mimetic, U46619. 5. The results demonstrate that GR43175 produces a selective vasoconstriction in the carotid arterial circulation of anaesthetized dogs via activation of 5-HT1-like receptors, which appear similar to those mediating contraction of the dog isolated saphenous vein.

Characterization of 5-HT receptors mediating contraction of canine and primate basilar artery by use of GR43175, a selective 5-HT1-like receptor agonist.

1. The aim of this study was to characterize the 5-hydroxytryptamine (5-HT) receptor which mediates contraction of canine and primate isolated basilar artery by use of a variety of selective 5-HT agonists and antagonists. 2. 5-HT, alpha-methyl 5-HT and the selective 5-HT1-like receptor agonists, GR43175 and 5-carboxamidotryptamine (5-CT), each caused contraction of canine and primate basilar artery with a rank order of agonist potency of 5-CT greater than or equal to 5-HT greater than GR43175 greater than alpha-methyl 5-HT. The 5-HT1-like receptor agonists, GR43175 and 5-CT, produced maximum effects which were less than that produced by 5-HT or alpha-methyl 5-HT. 3. In canine basilar artery, ketanserin (0.1-1 microM) caused some depression of the maximum effect of 5-HT but produced little or no shift of the concentration-effect curve. The contractile effects of GR43175 were not modified by ketanserin (1 microM), MDL72222 (1 microM) or cyanopindolol (1 microM). However, the effects of 5-HT and GR43175 were specifically antagonized by methiothepin (0.1 microM); the mean agonist concentration-ratios were 33 and 48 respectively. 4. In primate basilar artery, ketanserin (1 microM) again caused a small depression of the 5-HT maximum response but had not effect against GR43175-induced contractions. In contrast, methiothepin (0.1 microM) antagonized both 5-HT- and GR43175-induced contractions; the mean agonist concentration-ratios were 35 for both. 5. These results demonstrate that a large component of the effects of 5-HT in canine and primate basilar artery is produced by stimulation of a 5-HT1-like receptor. This receptor can be characterized by the high potency of the novel, selective agonist, GR43175, and susceptibility to blockade by methiothepin. However, there also appears to be a population of 5-HT2 receptors in these prepAarations which contribute to the contractile effects of 5-HT.

Further characterization of the 5-HT receptor mediating vascular relaxation and elevation of cyclic AMP in porcine isolated vena cava.

1. 5-Hydroxytryptamine (5-HT) and 5-carboxamidotryptamine (5-CT) produce both smooth muscle relaxation and elevation of tissue adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels in isolated rings of neonatal porcine vena cava. We now present studies attempting to characterize in more detail the 5-HT receptor mediating these responses. 2. Both 5-HT and 5-CT relaxed porcine isolated vena cava rings (EC50 values 200 nM and 4 nM respectively) and elevated tissue cyclic AMP levels (EC50 values 1500 nM and 16 nM respectively). For both responses 5-CT was approximately 50-100 fold more potent than 5-HT. 3. Both 5-CT-induced smooth muscle relaxation and cyclic AMP elevation were potently and specifically antagonized to a similar extent by methiothepin, methysergide and spiperone. 4. At concentrations up to 1 microM, 8-hydroxy-2-(di-n-propylamino) tetralin, buspirone, ipsapirone, n,n-dipropyl-5-CT, cyanopindolol, RU24969, ketanserin, GR38032 and GR43175 were devoid of both agonist and antagonist activity for both responses. 5. These findings suggest that the same 5-HT1-like receptor mediates both smooth muscle relaxation and elevation of cyclic AMP. This receptor is unlike any known 5-HT1 ligand binding site or adenylate cyclase-coupled 5-HT receptor in brain tissues.

Presynaptic inhibitory action of 5-hydroxytryptamine in dog isolated saphenous vein.

1 The effect of 5-hydroxytryptamine on contractile responses to sympathetic nerve stimulation has been studied in the dog isolated saphenous vein.2 Electrical stimulation (0.1 to 10 Hz) of dog saphenous vein strips produced frequency-dependent contractions. Contractions produced by stimulation at 2 Hz were almost completely blocked by tetrodotoxin (3.1 x 10(-8) mol/l) or phentolamine (5.0 x 10(-6) mol/l) but mecamylamine (5.0 x 10(-6) mol/l) had little effect. This suggests that the contractions were mediated predominantly through noradrenaline release from postganglionic noradrenergic nerves.3 Contractions produced by intermittent electrical stimulation at 2 Hz were inhibited by 5-hydroxy-tryptamine (1.0 x 10(-9) to 1.0 x 10(-7) mol/l) in a concentration-dependent manner whilst contractions induced by exogenous noradrenaline were not affected.4 The inhibitory action of 5-hydroxytryptamine was most marked at low frequencies of stimulation and with low pulse numbers.5 High external calcium concentrations (3.9 and 5.2 x 10(-3) mol/l) reduced the inhibitory action of 5-hydroxytryptamine.6 Cyproheptadine (1.0 x 10(-8) mol/l to 1.0 x 10(-6) mol/l) or morphine (1.0 x 10(-7) mol/l to 1.0 x 10(-5) mol/l) did not antagonize the inhibitory action of 5-hydroxytryptamine. Methysergide (1.0 x 10(-7) mol/l) slightly reduced the contractions produced by electrical stimulation and only weakly antagonized the action of 5-hydroxytryptamine.7 It is suggested that a 5-hydroxytryptamine receptor exists presynaptically in the dog isolated saphenous vein strip and that stimulation of this receptor by low concentrations of 5-hydroxytryptamine inhibits the release of noradrenaline from noradrenergic nerves. This receptor type is resistant to blockade by ;classical' 5-hydroxytryptamine antagonists.

Vascular 5-HT1-like receptors that mediate contraction of the dog isolated saphenous vein and carotid arterial vasoconstriction in anaesthetized dogs are not of the 5-HT1A or 5-HT1D subtype.

1. There is controversy about whether 5-HT1A receptors mediate contraction of isolated cerebral blood vessels. We have therefore compared the vascular actions of the 5-HT1A receptor agonist, 8-hydroxy-2-(di-n-propyl-amino)-tetralin (8-OH-DPAT) with those of the 5-HT1-like receptor agonist, sumatriptan, on the dog isolated saphenous vein, which contains a 5-HT1-like receptor similar to those on cerebral blood vessels, and in the carotid circulation of the anaesthetized dog. 2. 5-Hydroxytryptamine (5-HT), sumatriptan and 8-OH-DPAT each caused contraction of dog isolated saphenous vein with a rank order of agonist potency of 5-HT greater than sumatriptan greater than 8-OH-DPAT and EC50 values (95% confidence limits) of 0.06 (0.04-0.08), 0.3 (0.1-0.8) and 3.9 (2.0-7.5) microM respectively. The maximum contractile effect produced by each agonist was similar. 3. The contractile effects of 5-HT, sumatriptan and 8-OH-DPAT in the dog isolated saphenous vein were resistant to antagonism by the 5-HT1A receptor antagonists spiperone, spiroxatrine and pindolol (all 1 microM). The 5-HT1D receptor ligands, metergoline (0.1 microM) rauwolscine (1 microM) and yohimbine (1 microM) had little or no antagonist activity. In contrast, the non-selective 5-HT1-like receptor blocking drug, methiothepin (0.03-0.3 microM) potently antagonized the contractile effects of 5-HT, sumatriptan and 8-OH-DPAT to a similar degree, suggesting that all three agonists act at the same receptor. 4. In ganglion-blocked, anaesthetized dogs, intra-carotid administration of 8-OH-DPAT (0.3-3 pggkg-1) and sumatriptan (0.1l-1 pgkg -), caused dose-dependent carotid arterial vasoconstriction. The two agonists were approximately equipotent in this respect. 5. The carotid arterial vasoconstrictor actions of 8-OH-DPAT and sumatriptan were not modified by spiperone (1 mgkg- , i.v.) but were antagonized to a similar extent by the subsequent administration of methiothepin (1 mgkg- 1, i.v.). 6. These results suggest that 8-OH-DPAT contracts the dog isolated saphenous vein and constricts the carotid arterial circulation of anaesthetized dogs by activation of 5-HT1-like receptors which are not of the 5-HTlA subtype, nor, on the basis of data with metergoline in the dog isolated saphenous vein, of the 5-HTID subtype. The receptor involved in these actions appears to be the same as that mediating the vasoconstrictor effects of sumatriptan. This receptor does not appear to be like any known 5-HT1 ligand binding site; hence the current description, 5-HT1-like, remains the most appropriate.