Regional and functional differences of 5-hydroxytryptamine-receptor subtypes in guinea pig stomach

Functions and the presence of 5-hydroxytryptamine (5-HT) receptors in the fundus, corpus and antrum of the guinea pig stomach were examined by measuring contractile force and acetylcholine (ACh) release. Stimulation of the 5-HT1 receptor caused tetrodotoxin (TTX)-insensitive relaxations in the preparations from 3 regions. Stimulation of the 5-HT2 receptor caused TTX-insensitive contractions in the preparations of fundus and antrum. Stimulation of 5-HT3 receptors caused contractions that were sensitive to TTX and atropine and enhanced the outflow of [3H]ACh from preparations of only antrum. Stimulation of 5-HT4 receptors caused contractions of antral strips and decreased relaxations of corporal strips and enhanced the outflow of [3H]ACh from the preparations of both corpus and antrum. In the guinea pig stomach, the fundus possesses relaxant 5-HT1 receptor < contractile 5-HT2 receptors and caused the contractile response to 5-HT. The corpus possesses relaxant 5-HT1 receptors and relaxant receptors other than 5-HT1, 5-HT2, 5-HT3 and 5-HT4 receptors > contractile 5-HT4 receptor, and therefore 5-HT caused relaxations. The antrum possesses relaxant 5-HT1 receptor < contractile 5-HT2, 5-HT3 and 5-HT4 receptors, and thus 5-HT caused contractions.     

A paradox: the 5-HT2-receptor antagonist ketanserin restores the 5-HT-induced contraction depressed by methysergide in large coronary arteries of calf

Methysergide depresses the contractile effects of 5-hydroxytryptamine (5-HT) in bovine large coronary arteries devoid of endothelium. The IC50 of methysergide for depression of the response to 5-HT was (-log mol/l) 9.8. A low sensitivity contractile effect of 5-HT was not influenced by 1-1,000 nmol/l methysergide. The maximum force of this residual response is approximately 1/3 of the maximum force elicited by 5-HT in the absence of methysergide. Ketanserin restored the 5-HT-induced contraction depressed by methysergide. In the presence of 0.1 mumol/l ketanserin, methysergide caused depression of the 5-HT-induced effects with an IC50 (-log mol/l) of 6.5 without affecting the residual response. We propose that methysergide depresses 5-HT-induced contractions by acting on an allosteric site. The effect of binding of methysergide to the allosteric site would lead to a conformational change of the 5-HT2-receptor, thereby only allowing the production of a residual 5-HT-induced contraction. Ketanserin competes with high affinity not only with 5-HT for the 5-HT2-receptor but also with methysergide for the allosteric site, thus shifting the receptor back into its original conformation. The affinity estimate of ketanserin for the allosteric site yielded a KB (-log mol/l) of 10.3. Ketanserin (1-1,000 nmol/l) antagonized the contractile effects of 5-HT with a potency expected from its affinity for 5-HT2-receptors (-log KB, mol/l 9.4). However, micromolar concentrations of ketanserin antagonized the effects of 5-HT less than expected from its affinity for 5-HT2-receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characteristics of 5-hydroxytryptamine receptors involved in contraction of feline ileal longitudinal smooth muscle

A number of studies have demonstrated that 5-hydroxytryptamine (5-HT) can induce muscle contraction or relaxation response and enhance secretion in the gastrointestinal tract via a multiplicity of 5-HT receptor subtypes. In the present study, we investigated the pharmacological characterization of the 5-HT-induced contractile response in longitudinal smooth muscle isolated from the feline ileum. Addition of 5-HT into muscle chambers enhanced the basal tone and spontaneous activity in a concentration-dependent manner. The neurotoxin tetrodotoxin did not alter the 5-HT-induced contraction of the longitudinal muscles. Neither atropine nor guanethidine affected the contraction. The 5-HT agonists, 5-methylserotonin hydrochloride and mosapride, also evoked concentration-dependent contractions. The 5-HT-induced contraction was enhanced by the 5HT(2) receptor antagonist ketanserin and the 5-HT(3) receptor antagonist ondansetron but was inhibited by the 5-HT(1) receptor antagonist methysergide and 5-HT(4) receptor antagonist GR113808. These results indicate that 5-HT(1) and 5-HT(4) receptors may mediate the contraction of the 5-HT-induced response and 5-HT(2) and 5-HT(3) receptors may mediate 5-HT-induced relaxation in feline ileal longitudinal smooth muscles.     

Characterization of the receptors involved in the 5-HT-induced excitation of canine antral longitudinal muscle

1. We aimed to characterize the 5-HT receptors involved in the 5-HT-induced effect on electrically induced contractions of dog antrum longitudinal muscle in vitro. 2. In the presence of L-NOARG (0.1 mM), electrical field stimulation (EFS) induced atropine- and tetrodotoxin-sensitive contractions. Tetrodotoxin or atropine left any agonist tested ineffective. These EFS-induced contractions were on average enhanced by 5-HT (0.3 microM), however, pronounced variation in the response to 5-HT was observed. There were non-significant trends of the selective 5-HT3 receptor antagonist granisetron (1 microM), and methysergide (1 microM; preventing interactions of 5-HT with 5-HT1, 5-HT2, 5-ht5, 5-HT6 and 5-HT7 receptors) to increase the response to 5-HT. The selective 5-HT4 receptor antagonist GR 113808 (0.1 microM) displayed a non-significant trend to inhibit the 5-HT-induced increase. 3. Combination experiments with methysergide (1 microM), granisetron (1 microM) and GR 113808 (0.1 microM) revealed that the 5-HT (0.3 microM)-induced response consisted of (1) an excitatory component blocked by GR 113808, (2) excitatory and inhibitory components both blocked by methysergide. 4. The selective 5-HT4 receptor agonist prucalopride (0.3 microM) increased EFS-induced contractions, an effect prevented by GR 113808 (0.1 microM). 5. The increase of EFS-induced contractions by the preferential 5-HT2 receptor agonist alpha-Me-5-HT (0.3 microM) was antagonized by 5-HT2B receptor antagonists. 6. The 5-HT1/5-HT7 receptor agonist 5-carboxamidotryptamine (5-CT; 0.3 microM) inhibited EFS-induced contractions. This was prevented by methysergide (1 microM), the 5-HT7 receptor antagonist mesulergine (0.3 microM) and the selective 5-HT7 receptor antagonist SB-269970 (0.3 microM). 7. In the presence of GR 113808 (0.1 microM), alpha-Me-5-HT (1 microM) increased EFS-induced contractions. The 5-HT (0.3 microM)-induced inhibition of the stimulation by alpha-Me-5-HT was prevented by SB-269970 (0.3 microM). 8. In conclusion, dog antral longitudinal muscle is endowed with (1) excitatory neuronal 5-HT4 receptors and 5-HT2B receptors and (2) inhibitory smooth muscle 5-HT7 receptors.     

Pharmacological characterization of the 5-HT receptors mediating contraction and relaxation of canine isolated proximal stomach smooth muscle

1. We aimed to characterize 5-HT receptors mediating contraction and relaxation to 5-HT in dog proximal stomach longitudinal muscle (LM) strips. 2. Of the tryptamine analogues tested, 5-HT was the most potent contractile agent at basal length, while 5-CT was the most potent relaxant of PGF(2alpha)-induced contraction. Neither the contractions to 5-HT, nor the relaxations to 5-CT were influenced by tetrodotoxin, illustrating that action potential propagation is not involved. 3. The 5-HT-induced contraction was antagonized by mesulergine (0.03 to 0.3 microM) and ketanserin (2 - 20 nM), but the antagonism was not of a simple competitive nature, indicating multiple receptor involvement. Ketanserin (3 to 30 nM) and mesulergine (30 nM) competitively antagonized the alpha-Me-5-HT-induced contraction (pK(B): 8.83+/-0.09 and pA(2): 8.25+/-0.06 respectively). These affinity values are in line with literature affinities of ketanserin and mesulergine at 5-HT(2A) receptors in various bioassays. 4. The 5-CT-induced inhibition of PGF(2alpha)-induced contraction was competitively antagonized by mesulergine (pK(B) estimate: 8.52+/-0.12) and by the selective 5-HT(7) receptor antagonist SB-269970 (pK(B) estimate: 9.36+/-0.14). Both pK(B) estimates are in line with literature affinities of these compounds for 5-HT(7) receptors. Mesulergine (30 nM) and SB-269970 (10 nM) shifted the relaxant curve to 5-HT parallel to the right in the presence of ketanserin (0.3 microM) (pA(2) estimates of 8.08+/-0.10 and 8.75+/-0.14 respectively), indicative of 5-HT(7) receptor involvement. 5. It is concluded that 5-HT induces dog proximal stomach (LM) contraction via smooth muscle 5-HT(2A) receptors and relaxation via smooth muscle 5-HT(7) receptors.     

5-HT1-like receptors requiring functional cyclo-oxygenase and 5-HT2 receptors independent of cyclo-oxygenase mediate contraction of the human umbilical artery.

1. The interactions between 5-hydroxytryptamine (5-HT) and the antagonists ketanserin, methysergide and phentolamine were studied in isolated preparations of human umbilical artery (HUA) at physiological oxygen tension (Po2 approximately 15 mmHg) and at high PO2 (approximately 120 mmHg). 2. At physiological Po2 ketanserin, methysergide and phentolamine behaved as silent competitive antagonists of the 5-HT-induced contraction of HUA. pA2 values calculated by Schild analysis were 8.92, 8.52 and 6.37, respectively. 3. At high Po2, 5-HT-induced contractions were antagonised in a biphasic manner by ketanserin (0.1 microM); the response to low but not to high concentrations of 5-HT was resistant to blockade by ketanserin. The ketanserin-resistant component was abolished following cyclo-oxygenase inhibition by indomethacin (1 microM). 4. At high Po2, methysergide behaved as a partial agonist. Methysergide-induced contractions were inhibited but not abolished by indomethacin, and resistant to 5-HT2 receptor and alpha 1-adrenoceptor blockade. 5. At high Po2 the component of the response to 5-HT mediated by the ketanserin-resistant receptor was mimicked by the selective 5-HT1-like receptor agonist 5-carboxamidotryptamine (5-CT): 5-CT was 7 fold more potent than 5-HT. 6. At high Po2 the component of the response to 5-HT mediated by the ketanserin-resistant receptor was antagonised by phentolamine and the selective alpha 2-adrenoceptor antagonist Wy 26703. 7. These results suggest that (i) at physiological Po2 5-HT2 receptors almost exclusively mediate contractions induced by 5-HT, and (ii) at high Po2 the agonist potency order of 5-CT greater than 5-HT greater than methysergide suggests that ketanserin-resistant responses are mediated by 5-HT1-like receptors which require functional cyclo-oxygenase.     

No evidence for functional involvement of 5-HT2B receptors in serotonin-induced vasodilatation in the human forearm

The receptor involved in the serotonin (5-hydroxytryptamine [5-HT])-induced vasodilatation in the human forearm has not yet been identified. Experimental data point to the 5-HT2B receptor located on the endothelium. RS-127445 (2-amino-4-(4-fluoronaphthyl-1-yl)-6-isopropylpyrimidine) is a novel potent and selective 5-HT2B receptor antagonist. The effect of oral RS-127445 (500 mg) on 5-HT-induced vasodilatation was studied in a double-blind, randomized, placebo-controlled, crossover study in six healthy volunteers. On each study day 5-HT (0.5 ng/kg/min) was infused into the brachial artery for 8 min, before drug administration and at intervals of 20, 65, 110, 230, and 470 min after oral ingestion. At each infusion, plasma samples for study drug assay were taken and forearm blood flow was assessed using venous occlusion plethysmography. Although (log) drug concentrations exceeded pKi, there was no correlation between RS-127445 concentrations and 5-HT-induced vasodilatation. 5-HT-induced vasodilatation did not differ between treatments and time points. It appears that there is no functional involvement of 5-HT2B receptors in 5-HT-mediated vasodilatation in the human forearm.     

Allosteric properties of the 5-HT2 receptor system of the rat tail artery

Summary We present an analysis of the interactions of 5-hydroxytryptamine (5-HT) and antagonists (methysergide, ketanserin, ritanserin) with the 5-HT₂ receptor system of strips of rat tail artery. The mode of action of ritanserin was also studied on strips of calf coronary arteries. 1. Ketanserin competitively antagonized 5-HT-induced effects in rat tail artery with an affinity (pK_B = 9.4 nmol/l) consistent with the assumption of an interaction of 5-HT and ketanserin at 5-HT₂-receptors. 2. Methysergide reduced to 50–60% the maximum response to 5-HT in rat tail artery. Concentration-effect curves for 5-HT became biphasic in the presence of methysergide with quickly and slowly developing contractions at low and high concentrations of 5-HT, respectively. 100 nmol/l ketanserin completely restored effects of 5-HT depressed by low concentrations of methysergide (< 10 nmol/l). Higher concentrations of methysergide in the presence of 100 nmol/l ketanserin again depressed the effects of 5-HT. 3. Ritanserin resembles methysergide by causing insurmountable antagonism of 5-HT-induced contractions which can be prevented by ketanserin in both rat tail artery and calf coronary artery. These results are inconsistent with competition between ritanserin and 5-HT for the 5-HT₂ receptor. 4. The findings are consistent with the assumption of an interaction of ketanserin and methysergide or ritanserin with an allosteric site near the 5-HT₂-receptor. Both methysergide and ritanserin appear to antagonize the effects of 5-HT through an allosteric site which is distinct from the 5-HT₂ receptor. Send offprint requests to A. J. Kaumann at the above address     

Contractile effects of 5-hydroxytryptamine and 5-carboxamidotryptamine in the equine jejunum

The use of human prokinetic drugs in colic horses leads to inconsistent results. This might be related to differences in gastrointestinal receptor populations. The motor effects of 5-hydroxytryptamine (5-HT; serotonin) on the equine mid-jejunum were therefore studied. Longitudinal muscle preparations were set up for isotonic measurement. 5-HT induced tonic contractions with superimposed phasic activity; these responses were not influenced by tetrodotoxin and atropine, suggesting a non-neurogenic, non-cholinergic pathway. The 5-HT receptor antagonists GR 127935 (5-HT(1B,D)), ketanserin (5-HT(2A)), SB 204741 (5-HT(2B)), RS 102221 (5-HT(2C)), granisetron (5-HT(3)), GR 113808 (5-HT(4)) and SB 269970 (5-HT(7)) had no influence on the 5-HT-induced response; the 5-HT(1A) receptor antagonists NAN 190 (pK(b)=8.13+/-0.06) and WAY 100635 (pK(b)=8.69+/-0.07), and the 5-HT(1,2,5,6,7) receptor antagonist methysergide concentration-dependently inhibited the 5-HT-induced contractile response. The 5-HT(1,7) receptor agonist 5-carboxamidotryptamine (5-CT) induced a contractile response similar to that of 5-HT; its effect was not influenced by tetrodotoxin and atropine, and SB 269970, but antagonised by WAY 100635. 8-OHDPAT, buspiron and flesinoxan, which are active at rat and human 5-HT(1A) receptors, had no contractile influence. These results suggest that the contractile effect of 5-HT in equine jejunal longitudinal muscle is due to interaction with muscular 5-HT receptors, which cannot be characterised between the actually known classes of 5-HT receptors.     

Effect of mianserin, a potent anti-serotonin agent, on the isolated rat stomach fundus preparation

The influence of mianserin and cyproheptadine on the response of the rat stomach fundus in vitro to Ach (acetylcholine), 5-HT (serotonin) and tryptamine in the presence of the α-adrenolytic, piperoxan, were compared. Cyproheptadine and mianserin competitively antagonized the Ach-induced contractions, but cyproheptadine was more effective. Mianserin, a recognized antagonist of 5-HT on rat uterus and on bronchospasm of the guinea pig, did not affect 5-HT- and tryptamine-induced contractions of the isolated rate stomach fundus in concentrations up to 10^?6 M, while cyproheptadine proved highly effective. Higher doses of mianserin contracted the organ. A closed piperazine ring seems to be essential for such action. Although no anti-5-HT action could be found with mianserin on the rat fundus, it protected 5-HT-induced contractions against inhibition by phenoxybenzamine, cyproheptadine or methysergide. The α-adrenolytic piperoxan was more effective in protecting the response to 5-HT than to Ach against inhibition by phenoxybenzamine. It is concluded that differences might exist between D-tryptamine receptors in different organs and that the receptor protection by mianserin and pieperoxan towards the 5-HT-induced contractions may be related to an ability to interact at Ca²⁺-binding sites of the smooth muscle cell membrane. However, the possibility that piperoxan might in some way interfere with an inhibitory action of mianserin towards tryptamine receptors is not ruled out. Although both the response to 5-HT and tryptamine are affected by cyproheptadine, the 5-HT-induced contractions were more susceptible to inhibition by cyproheptadine than those of tryptamine. This possibly indicated a subsidiary action of tryptamine on other than the D-tryptamine receptors.     

Serotonin-induced vasoconstriction in dog kidney.

Serotonin (5-HT 0.1 and 0.3 micrograms/kg) was injected as a bolus into the renal artery of anesthetized dogs. Renal blood flow (RBF) decreased initially after 5-HT injection and then increased. The 5-HT-induced decrease in RBF was potentiated during intrarenal infusion of a 5-HT1 and 5-HT2 antagonist methysergide at 3 micrograms/kg/min or of a 5-HT2 antagonist ketanserin (3-30 micrograms/kg/min), but methysergide at 30 micrograms/kg/min attenuated the decrease in RBF. The delayed increase in RBF was suppressed during ketanserin or methysergide infusion. In ketanserin-pretreated dogs, methysergide (3-30 micrograms/kg/min) dose-dependently suppressed the 5-HT-induced decrease in RBF. A 5-HT3 antagonist, ICS 205-930 (3-30 micrograms/kg/min), did not affect the 5-HT-induced RBF responses. A Ca2+ entry blocker CD-349 (30 and 100 ng/kg/min) and a Ca2+ release inhibitor TMB-8 (30 and 100 micrograms/kg/min) suppressed the 5-HT-induced decrease in RBF. These results suggest that 5-HT activates 5-HT1 receptors to induce vasoconstriction by mobilization of extracellular and intracellular Ca2+, but simultaneous activation of 5-HT2 receptors attenuates the vasoconstriction, probably by causing vasodilation in dog kidney.     

Characterization of 5-HT receptor subtypes mediating contraction in human umbilical vein. 1. Evidence of involvement of 5-HT2A receptors using functional and radioligand binding assays

This study attempted to characterize pharmacologically the involvement of 5-HT(2A) receptors in 5-HT-induced contractile responses in human umbilical vein (HUV) rings employing functional and radioligand binding assays. In HUV rings, prazosin 1 micro M did not affect contractile responses elicited by 5-HT, ruling out the involvement of alpha(1)-adrenoceptors in contractile responses to 5-HT. 5-HT-induced contractions were competitively blocked by ketanserin, a 5-HT(2A)-selective antagonist. The apparent pA(2) value was 9.8 and the Schild slope significantly less than unity, suggesting that 5-HT-induced responses are mediated by a heterogeneous receptor population. Alpha-methyl-5-HT, a selective 5-HT(2) receptor agonist, induced contractions that were antagonized in a competitive manner by ketanserin. The slope regression was not significantly different from unity and the pA(2) value was 8.8. The selective 5-HT(2A) ligand spiperone produced a parallel rightward shift on 5-HT CRCs in HUV rings. The calculated pA(2) was 9.0, which is in accord for an interaction with the 5-HT(2A) receptor subtype. Alpha-methyl-5-HT CRCs were competitively blocked by spiperone treatment. The Schild analysis yielded a pA(2) of 9.1 with a slope not significantly different from unity. The 5-HT(2C/2A) antagonist mesulergine 10 nM did not affect 5-HT CRCs, suggesting that 5-HT(2C) receptors are not involved in 5-HT-elicited contractions. Higher concentrations of mesulergine showed a parallel rightward shift on 5-HT responses. The calculated pA(2) was 7.44, which suggests an interaction with the 5-HT(2A) receptor subtype. In addition, mesulergine competitively blocked alpha-methyl-5-HT CRCs. The Schild slope was not significantly different from unity and the p A(2) value was 7.98.The binding of [(3)H]ketanserin to HUV membranes was saturable and of high affinity. Ketanserin displayed a monophasic curve which was best fit with a single component of binding. Nonlinear least squares analysis of the binding curves revealed a high affinity K(d) of 0.30 nM and a B(max) of 134 fmol/mg protein.These findings provide strong pharmacological evidence of the involvement of 5-HT(2A) receptors in 5-HT-induced vasoconstriction in HUV. In addition, the contribution of another receptor population cannot be excluded. The results also suggest that this receptor population is neither an alpha(1)-adrenoceptor nor a 5-HT(2C) receptor subtype.     

Participation of 5-HT1-like and 5-HT2A receptors in the contraction of human temporal artery by 5-hydroxytryptamine and related drugs.

1. We investigated the hypothesis that, as in some other large human arteries, 5-HT-induced contraction of the temporal artery is mediated through two co-existing receptor populations, 5-HT1-like- and 5-HT2A. Temporal arterial segments were obtained from patients undergoing brain surgery and rings prepared set up to contract with 5-HT and related agents. Fractions of maximal 5-HT responses mediated through 5-HT1-like and 5-HT2A receptors, f1 and f2 = 1-f1, were estimated by use of the 5-HT2A-selective antagonist ketanserin. 2. In rings with intact endothelium 5-HT evoked contractions with a -log EC50, M of 7.0. Ketanserin (10-1000 nM) antagonized part of the 5-HT-induced contractions. Ketanserin-resistant components of 5-HT-induced contractions were found with -log EC50, M of 6.9 and f1 of 0.17 (100 nM ketanserin) and -log EC50, M of 6.4 and f1 of 0.20 (1000 nM ketanserin). 3. In rings with endothelial function attenuated by enzymatic treatment, 5-HT caused contractions with a -log EC50, M of 7.2 that were partially blocked by ketanserin. Ketanserin-resistant components of 5-HT-induced contractions were found with -log EC50, M 7.4 and f1 of 0.16 (100 nM ketanserin) and -log EC50, M of 7.5 and f1 of 0.14 (1000 nM ketanserin). 4. The ketanserin-resistant component of 5-HT-evoked contraction was blocked by methiothepin (100-1000 nM) consistent with mediation through 5-HT1-like receptors. 5. In rings with intact endothelium the 5-HT1-like-selective agonist, sumatriptan, caused small contractions with a -log EC50, M of 6.5 and intrinsic activity of 0.21 with respect to 5-HT that were resistant to blockade by 1000 nM ketanserin but antagonized by 100 nM methiothepin. 6. In rings with intact endothelium the 5-HT2A receptor partial agonist SK&F 103829 (2,3,4,5-tetrahydro-8[methyl sulphonyl]-1H3-benzazepin-7-ol methensulphonate) contracted rings with a -log EC50, M of 5.0 and an intrinsic activity of 0.49 with respect to 5-HT; the effects were antagonized by ketanserin 1000 nM. 7. We conclude that 80-86% of the maximum 5-HT-evoked contraction of human temporal artery is mediated through 5-HT2A receptors, the remainder through 5-HT1-like-receptors, regardless of whether or not endothelium is functional. The 5-HT1-like-receptors are more likely to be 5-HT1D beta receptors than 5-HT1D alpha receptors and sumatriptan is a full agonist for these receptors. As found in arteries of other species, SK&F 103829 is a partial agonist for 5-HT2A receptors of human temporal artery.     

Effects of the dopamine receptor agonists, fenoldopam and quinpirole, in the rat stomach

The effects of the DA1-receptor agonist, fenoldopam, and the DA2-receptor agonist, quinpirole, were studied with the longitudinal muscle of rat gastric fundus and circular muscle of rat gastric corpus, as there are contrasting reports about the receptors involved in the inhibitory effect of dopamine in these tissues. Quinpirole had no effect on basal tone in the longitudinal muscle of the rat gastric fundus and did not inhibit the sustained contractions induced by electrical field stimulation or by methacholine. Fenoldopam had no effect on the tone increased by methacholine but slightly potentiated the electrically induced contraction at the highest concentrations; it concentration dependently (10(-7)-3 X 10(-5) M) increased the basal tone. The contractile effect of fenoldopam was clearly antagonized by rauwolscine 10(-6) M, yohimbine 10(-6) M and phentolamine 3 X 10(-6) M plus propranolol 10(-5) M. The 5-HT receptor antagonist, methysergide, antagonized the fenoldopam-induced contractions in a non-competitive way. Fenoldopam and quinpirole had no effect on contractions induced in the circular muscle of the rat gastric corpus by methacholine or electrical field stimulation. They induced some contraction at basal tone, at their highest concentrations. As fenoldopam and quinpirole did not mimic the inhibitory effect observed with dopamine in the same models, no evidence was found for the presence of inhibitory dopamine receptors in rat gastric muscle. The contractile effect of fenoldopam in the longitudinal muscle of the fundus is probably due to an interaction with 5-HT receptors.     

A Pharmacological Comparison of the Receptors Mediating Contractile Responses to 5-Hydroxytryptamine in the Rat Isolated Caudal Artery and Fundic Strip

Abstract— Contractile responses to 5-hydroxytryptamine (5-HT) and to a number of 5-HT-receptor agonists have been compared on the rat isolated caudal artery and stomach fundic strip. On the caudal artery, 5-HT was the most potent agonist tested. The 5-HT₁-like agonist, 5-carboxamidotryptamine (5-CT), was less potent than 5-HT and produced a lower maximum response. 8-Hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) and RU24969 (5-methoxy-3(1,2,3,6-tetrahydropyridin-4-yl)1H indole) were inactive as agonists and 8-OH-DPAT was not an antagonist. Ketanserin, ICI 169,369 (2-(2-dimethylaminoethyl-thio)-3-phenylquinoline hydrochloride) and ICI 170,809 (2-(2-dimethylamino-2-methylpropylthio)-3-phenylquinoline hydrochloride) were competitive antagonists of 5-HT on this preparation, indicating that 5-HT is acting via 5-HT₂ receptors. In contrast, all the agonists produced contractions of the fundic strip (rank order of potency, 5-HT = 5-CT > RU24969 > 8-OH-DPAT). The maximum response to RU24969 was significantly lower than the maximum responses to the other agonists. Ketanserin was only a weak antagonist of 5-HT in the fundic strip, demonstrating that 5-HT₂ receptors were not involved, but ICI 169,369 and ICI 170,809 were non-surmountable antagonists of 5-HT responses, as were methysergide and methiothepin. Since ICI 169,369 and ICI 170,809 are devoid of activity at 5-HT₃ and 5-HT₄ receptors, then these two subtypes would not appear to be implicated, a view that was confirmed in the case of 5-HT₃ receptors by experiments using ondansetron. Since radioligand binding studies have shown that ICI 169,369 and ICI 170,809 have high affinity for 5-HT_1C sites, α-methyl-5-HT, an agonist claimed to be selective for 5-HT_1C and 5-HT₂ receptors, was also tested on the fundic strip in the presence of ketanserin. This compound acted as an agonist and was antagonized in a competitive manner by ICI 170,809. Therefore, these results suggest that the receptors mediating contractile responses to 5-HT in the rat fundic strip are more similar to the 5-HT_1C subtype than any of the other subtypes that have been described previously.     

Evidence that hypophagia induced by d-fenfluramine and d-norfenfluramine in the rat is mediated by 5-HT2C receptors

The present series of studies is the first to investigate the pharmacological mechanisms underlying d-fenfluramine- and d-norfenfluramine-induced hypophagia in the rat using highly selective serotonin 5-HT2 receptor antagonists. Administration of d-fenfluramine, and its major metabolite d-norfenfluramine, suppresses food intake in animals. Both compounds stimulate the release of serotonin and are potent inhibitors of the re-uptake of 5-HT into nerve terminals. In addition, d-norfenfluramine also acts as a direct 5-HT(2B/2C) receptor agonist. Pre-treatment with the selective 5-HT2C receptor antagonist, SB-242084 (0.3-3 mg/kg), dose-dependently inhibited both d-fenfluramine- (3 mg/kg) and d-norfenfluramine-induced (2 mg/kg) hypophagia. In contrast, the hypophagic effect of d-fenfluramine and d-norfenfluramine was unaffected by prior treatment with the highly selective 5-HT2B receptor antagonists, SB-215505 (0.3-3 mg/kg) and RS-127445 (1-3 mg/kg) or the 5-HT2A receptor antagonists MDL 100,907 (0.003-0.03 mg/kg) and ketanserin (0.2, 0.5 mg/kg). In addition, the 5-HT1A receptor antagonist WAY-100635 (0.3, 1 mg/kg) and the 5-HT1B receptor antagonists GR-127935 (1, 2 mg/kg) and SB-224289 (2-10 mg/kg) did not affect d-fenfluramine-induced hypophagia. These data provide unequivocal evidence for an important role of the 5-HT2C receptor in the mediation of d-fenfluramine and d-norfenfluramine-induced hypophagia in the rat and do not support the involvement of 5-HT1A/1B/2A/2B receptors.     

Two types of receptors for 5-hydroxytryptamine on the cholinergic nerves of the guinea-pig myenteric plexus.

The effects of 5-hydroxytryptamine (5-HT) on spontaneous and electrically-evoked release of [3H]-acetylcholine (ACh) from guinea-pig myenteric plexus preparations preincubated with [3H]-choline have been investigated in the absence of cholinesterase inhibitors. 5-HT caused a transient increase in spontaneous release and an inhibition of the electrically-evoked release of [3H]-ACh. The 5-HT-induced contractions of the longitudinal muscle were clearly related to the increase in spontaneous release. The inhibitory effect was not due to activation of alpha-adrenoceptors since it was also observed in the presence of tolazoline and on strips from reserpine-pretreated guinea-pigs. After desensitization of the excitatory 5-HT receptors with 5-HT or metoclopramide the effects of 5-HT on spontaneous [3H]-ACh release were largely reduced. A variety of established antagonists at neuronal 5-HT receptors (i.e. metitepine 0.1-1 microM; methysergide 1 microM; ketanserin 0.1-1 microM; MDL 72222 0.1 microM; tropacocaine 1 microM) failed to block the excitation. The inhibition by 5-HT of the electrically evoked [3H]-ACh release was competitively antagonized by metitepine (pA2 7.6) and methysergide (pA2 7.0) but not by ketanserin. Tachyphylaxis to the inhibitory action of 5-HT did not occur. The results suggest that the excitatory 5-HT receptor ('M'-receptor) differs in its pharmacological properties from other neuronal 5-HT receptors. The presynaptically located inhibitory receptor may roughly correspond to the 5-HT1 receptor subtype but probably differs from the 5-HT autoreceptor.     

Smooth muscle 5-HT2A receptors mediating contraction of porcine isolated proximal stomach strips

1. The aim of this study was to characterize the 5-HT receptors involved in the 5-HT-induced contraction of longitudinal muscle (LM) strips of porcine proximal stomach. This was done in a classical organ bath set-up for isotonic measurement. 2. The concentration-contraction curve to 5-HT was not modified by 5-HT(3) and 5-HT(4) receptor antagonism. Methysergide, ketanserin and mesulergine antagonized the curve to 5-HT. Concomitantly, increasing concentrations of ketanserin and mesulergine progressively revealed a biphasic nature of the 5-HT curve. Ketanserin antagonized the low-affinity receptor while it did not modify the high-affinity receptor. 3. Tetrodotoxin did not influence the concentration-contraction curve to 5-HT neither in the absence nor presence of ketanserin, indicating that nerves are not involved. 4. Ketanserin competitively antagonized the monophasic concentration-response curve to alpha-Methyl-5-HT, yielding a Schild slope that was not significantly different from unity. After constraining the Schild slope to unity, a pK(B) estimate of 8.23+/-0.90 was obtained. This affinity estimate of ketanserin closely approximates previously reported affinities at 5-HT(2A) receptors. 5. In the presence of ketanserin (0.1 microM; exposing the high-affinity receptor), a wide range of 5-HT receptor antagonists covering all 5-HT receptors known, was tested. Only methysergide and ritanserin inhibited the response to 5-HT, thus expressing affinity for the high-affinity receptor. This did not reveal the identity of the receptor involved. 6 It can be concluded that 5-HT induces pig proximal stomach (LM) contraction via 5-HT(2A) receptors located on smooth muscle. A ketanserin-insensitive phase of contractions could not be characterized between the actually known classes of 5-HT receptors with the pharmacological tools that were used.     

Ketanserin potentiates the prejunctional inhibitory effect of 5-hydroxytryptamine on rat vas deferens

5-Hydroxytryptamine (5-HT) slightly inhibited the twitch contractions of rat vas deferens caused by single pulse field stimulation at 0.1 Hz. The inhibitory effect of 5-HT was much less in the epididymal portion than in the prostatic portion of the vas deferens. Ketanserin potentiated the prejunctional inhibitory effect of 5-HT and attenuated its stimulatory effect. This potentiation was observable only in the epididymal portion, of the vas deferens. Cyproheptadine and mianserin, but not methysergide, had essentially similar potentiating effects to those of ketanserin. These results suggest that the 5-HT receptor that mediates prejunctional inhibition is not of the 5-HT2 type, and that ketanserin acts by suppressing the 5-HT-induced stimulatory effect, which is possibly mediated by a postjunctional 5-HT2 receptor, thus unmasking the inhibitory effect of 5-HT.     

Role of 5-HT2 receptors in serotonin-induced contraction in the human mammary artery

We studied the effects of serotonin (5-HT) on isolated human mammary arteries obtained from patients undergoing coronary by-pass grafting. 5-HT induced a concentration-dependent contractile response in the mammary artery, with an EC50 value of 0.34 microM. The 5-HT2 antagonist, ketanserin, reversed the contractions evoked by 5-HT in a competitive manner at a low concentration (10(-8) M), whereas non-competitive antagonism was apparent at higher concentrations (5 X 10(-8)-5 X 10(-7) M). To investigate whether the alpha 1-blocking component of ketanserin plays a role in the response observed in this vessel, we evaluated the effect of ketanserin on contractions induced by (-)-norepinephrine. Ketanserin, in concentrations up to 10(-7) M, did not influence the norepinephrine-induced contractions. Moreover, a threshold concentration of 5-HT (10(-7) M) amplified the contractile effect induced by norepinephrine (5 X 10(-8) M), and this response was inhibited by ketanserin (10(-7) M). The selective 5-HT3 antagonist, GR 38032F, did not affect the 5-HT-induced contractions. These findings indicate that the human mammary artery is a vascular tissue sensitive to 5-HT. The 5-HT2 receptor subtype appears to mediate the response.