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Tachykinin-induced contraction of the guinea-pig isolated oesophageal mucosa is mediated by NK(2) receptors.

ABSTRACT: 1. The tachykinin receptor present in the guinea-pig oesophageal mucosa that mediates contractile responses of the muscularis mucosae has been characterized, using functional in vitro experiments. 2. The NK(1) receptor-selective agonist, [Sar(9)(O(2))Met(11)]SP and the NK(3) receptor-selective agonists, [MePhe(7)]-NKB and senktide, produced no response at submicromolar concentrations. The NK(2) receptor-selective agonists, [Nle(10)]-NKA(4 - 10), and GR 64,349 produced concentration-dependent contractile effects with pD(2) values of 8.20+/-0.16 and 8.30+/-0.15, respectively. 3. The concentration-response curve to the non-selective agonist, NKA (pD(2)=8.13+/-0.04) was shifted significantly rightwards only by the NK(2) receptor-selective antagonist, GR 159,897 and was unaffected by the NK(1) receptor-selective antagonist, SR 140,333 and the NK(3) receptor-selective antagonist, SB 222,200. 4. The NK(2) receptor-selective antagonist, GR 159,897, exhibited an apparent competitive antagonism against the NK(2) receptor-selective agonist, GR 64,349 (apparent pK(B) value=9.29+/-0.16) and against the non-selective agonist, NKA (apparent pK(B) value=8.71+/-0.19). 5. The NK(2) receptor-selective antagonist, SR 48,968 exhibited a non-competitive antagonism against the NK(2) receptor-selective agonist, [Nle(10)]-NKA(4 - 10). The pK(B) value was 10.84+/-0.19.6. It is concluded that the guinea-pig isolated oesophageal mucosa is a useful preparation for studying the effects of NK(2) receptor-selective agonists and antagonists as the contractile responses to various tachykinins are mediated solely by NK(2) receptors.

SUBMITTER: Kerr KP

PROVIDER: S-EPMC1572466 | biostudies-other | 2000 Dec

REPOSITORIES: biostudies-other

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Project description:Background and purposeMelatonin is involved in the regulation of colonic motility, and sensation, but little is known about the influence of melatonin on 5-hydroxytryptamine (5-HT) release from colonic mucosa. A tachykinin NK₂ receptor-selective agonist, [β-Ala⁸]-neurokinin A(4-10) [βAla-NKA-(4-10)] can induce 5-HT release from guinea pig colonic mucosa via NK₂ receptors on the mucosal layer. The present study was designed to determine the influence of melatonin on 5-HT release from guinea pig colonic mucosa, evoked by the NK₂ receptor agonist, βAla-NKA-(4-10).Experimental approachThe effect of melatonin was investigated on the outflow of 5-HT and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) from muscle layer-free mucosal preparations of guinea pig colon, using high-performance liquid chromatography with electrochemical detection.Key resultsMelatonin caused a sustained decline in the βAla-NKA-(4-10)-evoked 5-HT outflow from the muscle layer-free mucosal preparations, but failed to affect its metabolite 5-HIAA outflow. The specific MT₃ receptor agonist, 5-methoxycarbonylamino-N-acetyltryptamine mimicked the inhibitory effect of melatonin on βAla-NKA-(4-10)-evoked 5-HT outflow. A MT₃ receptor antagonist prazosin shifted the concentration-response curve of melatonin to the right in a concentration-dependent manner and depressed the maximum effect, but neither a combined MT₁/MT₂ receptor antagonist luzindole, nor a MT₂ receptor antagonist N-pentanoyl-2-benzyltryptamine modified the concentration-response curve to melatonin.Conclusions and implicationsMelatonin inhibits NK₂ receptor-triggered 5-HT release from guinea pig colonic mucosa by acting at a MT₃ melatonin receptor located directly on the mucosal layer, without affecting 5-HT degradation processes. Possible contributions of MT₁/MT₂ melatonin receptors to the inhibitory effect of melatonin appear to be negligible. Melatonin may act as a modulator of excess 5-HT release from colonic mucosa.

Project description:1. The effects of endothelin-1 (ET-1) on guinea-pig lower oesophageal sphincter (LOS) circular smooth muscle were investigated by using intracellular microelectrodes and isometric tension recording techniques. 2. ET-1 produced biphasic mechanical responses; an initial transient relaxation followed by a sustained contraction. The initial relaxation was not inhibited by either tetrodotoxin (TTX, 1 microM) or L-N(G)-nitroarginine (L-NOARG, 100 microM). The sustained contraction was greatly attenuated by nifedipine (1 microM). 3. ET-1 (1 - 30 nM) induced a concentration-dependent hyperpolarisation that was unaffected by TTX or L-NOARG. The ET(A) receptor antagonist, BQ123 (0.3 microM) abolished the ET-1-induced hyperpolarisation, whereas the ET(B) receptor antagonist, BQ788 (0.3 microM) had no detectable effect. Sarafotoxin S6c (10 nM) did not change the membrane potential. 4. The ET-1-induced hyperpolarisation was abolished by apamin (0.1 microM). Interestingly, apamin abolished the ET-1-induced transient relaxation but potentiated the sustained contraction. 5. In Ca(2+)-free Krebs solution, the ET-1-induced hyperpolarisation was greatly attenuated and returned to the control value when the tissue was reperfused with Krebs solution containing Ca(2+). The ET-1-induced hyperpolarisation was insensitive to nifedipine but was attenuated by SK&F 96365 (1 - [beta-[3-(4 - methoxy - phenyl)propoxy] - 4 - methoxyphenethyl] - 1H-imidazole hydrochloride, 50 microM), an inhibitor of receptor-mediated Ca(2+) entry. The residual component of the ET-1-induced hyperpolarisation was sensitive to thapsigargin (1 microM). 6. These results demonstrate that, in guinea-pig LOS circular smooth muscle, ET-1 hyperpolarizes the membrane by activating apamin-sensitive K(+) channels, mainly as a result of receptor-mediated Ca(2+) entry and partly by Ca(2+) release from intracellular stores. The hyperpolarisation triggers the initial transient relaxation, which acts to oppose the sustained contraction.

Project description:Much attention has focused on tachykinin receptors as therapeutic targets for neuropsychiatric disorders, although their expressional distributions in the primate central nervous system (CNS) remain unclear. We cloned the genes encoding the NK-1 and NK-3 tachykinin receptors (referred to as rmNK-1 and rmNK-3) from the rhesus monkey (Macaca mulatta) brain and examined their pharmacological profiles and regional distributions in the CNS. The deduced rmNK-1 amino-acid sequence differed by only two amino acids from the human NK-1 (hNK-1). The deduced rmNK-3 amino-acid sequence was two amino acids shorter than human NK-3 (hNK-3), with a seven-amino-acid difference in sequence. Ligand binding studies revealed that the affinity of rmNK-1 to substance P (SP) was comparable to that of hNK-1 in cell lines that expressed individual receptors stably. Nonpeptide antagonists had similar effects on the binding of rmNK-1 and hNK-1. Affinity of rmNK-3 for NKB was stronger than for SP and the IC50 value was comparable with that of hNK-3. Ca2+ imaging showed that activations of both rmNK-1 and rmNK-3 by specific ligands, SP and senktide, induced increased intracellular Ca2+ in cell lines that stably expressed individual primate tachykinin receptors. The amounts of rmNK-1 and rmNK-3 mRNAs were quantitatively determined in the monkey CNS. The expression of rmNK-1 was observed in all of the cortical and subcortical regions, including the hippocampus and the amygdala. The putamen contained the most NK-1 mRNA in the brain, with less rmNK-3 mRNA found in the cortex compared to rmNK-1 mRNA. In the monkey hippocampus and amygdala, rmNK-1 mRNA was present at markedly higher concentrations than rmNK-3 mRNA. The present results provide an insight into the distinct physiological nature and significance of the NK-1 and NK-3 tachykinin systems in the primate CNS. These findings are indispensable for establishing model systems in the search for a subtype-specific tachykinin receptor agonist and antagonist for the treatment of neuropsychiatric disorders.

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Tachykinin-induced contraction of the guinea-pig isolated oesophageal mucosa is mediated by NK(2) receptors.

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