5-HT-INDUCED NEUROGENIC RELAXATIONS OF THE GUINEA-PIG PROXIMAL COLON - INVESTIGATION INTO THE ROLE OF ATP AND VIP IN ADDITION TO NITRIC-OXIDE

In the guinea-pig proximal colon, 5-hydroxytryptamine (5-HT) relaxes t he longitudinal muscle by stimulating neuronal 5-HT receptors, which i nduces the release of nitric oxide (NO). It was investigated whether t he inhibitory neurotransmitters adenosine 5'-triphosphate (ATP) and/or vasoactive intestinal polypeptide (VIP) could be involved as well. An tagonists to block the contractile response to 5-HT via 5-HT2, 5-HT3 o r 5-HT4 receptors were present throughout the experiments and methacho line was administered to precontract the strips. ATP, VIP and 5-HT ind uced concentration-dependent relaxations, in the case of 5-HT yielding a non-monophasic concentration-response curve. Tetrodotoxin (TTX; 300 nM), N-G-nitro-L-arginine (L-NNA, 100 mu M) and their combination did not inhibit the relaxations induced by VIP (up to 0.3 mu M) or 0.3 - 3 mu M ATP but reduced those by 10 mu M ATP. Suramin (300 mu M) strong ly inhibited the relaxations to ATP and VIP. L-NNA and suramin also in hibited the relaxations to 5-HT. In the presence of L-NNA (100 mu M), suramin did not significantly inhibit the relaxations to 5-HT. Suramin did not affect the relaxations to isoprenaline, nitroglycerin or exog enous NO (1 mu M), demonstrating its specificity. Apamin (30 nM) inhib ited both the relaxations to ATP (by 70-100%) and to 5-HT; relaxations to isoprenaline were partially inhibited, indicating a non-specific c omponent in the inhibitory action of apamin. However, relaxations to e xogenous VIP (up to 0.3 mu M), NO (1 mu M) and to nitroglycerin were n ot inhibited. In the presence of L-NNA (100 mu M), apamin inhibited th e relaxations to 5-HT only at 30 mu M. alpha,beta-methylene-ATP (alpha ,beta-Me-ATP; 100 mu M) did not desensitize the responses to ATP. Reac tive blue 2 affected the relaxations to isoprenaline at concentrations necessary to significantly inhibit the relaxations to ATP (i.e. from 10 mu M onwards). Thus, it was not possible to test either alpha,beta Me-ATP or reactive blue 2 against the relaxations to 5-HT. alpha-Chymo trypsin (0.015 mg.ml(-1)) and trypsin (0.005 mg.ml(-1)) almost abolish ed the relaxations to VIP, but did not affect those to isoprenaline an d 5-HT. The VIP receptor antagonists [p-Cl-D-Phe(6) Leu(17)]VIP (1 mu M) and VIP10-28 (1 and 3 mu M) did not affect the concentration-respon se curve to VIP and were hence not tested against 5-HT. Phosphoramidon (1 mu M) had no effect on the relaxations to VIP or 5-HT. It can be c oncluded that in the guinea-pig colon longitudinal muscle, VIP and ATP induce relaxation via a direct effect on the smooth muscle, not invol ving NO. 5-HT-induced relaxations are mediated by NO as well as by a s ubstance which is sensitive to inhibition by suramin but not apamin. I t is suggested that this substance is ATP and not a peptide.