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Characterisation of the 5-HT4 receptor ligand, SL65.0155, in guinea-pig isolated colon
SL65.0155 is a partial agonist at the recombinant 5-HT4(b) and 4(d) receptor variants, shown to facilitate learning behaviours in rodents but antagonise at the 5-HT4 receptor on the muscle of rat esophagus (Moser et al., 2002). The latter preparation readily detects partial 5-HT4 receptor agonists. Therefore, to look for an ability to activate 5-HT4 receptors in the gut, we studied SL65.0155 in an enteric nerve model of the receptor, where the intrinsic activity of 5-HT4 receptor agonists is generally closer to that of 5-HT, perhaps because the process of receptor activation, neurotransmitter release and muscle contraction lends itself to amplification (Wardle et al., 1993). Distal colon longitudinal muscle-myenteric plexus preparations from male guinea-pigs (150-200 g) were suspended (0.5 g tension; isometric) in tissue baths containing Krebs solution (37 °C, 5 % CO2 in O2), methiothepin 0.1 µM and ondansetron 1 µM (to antagonise at 5-HT1, 2, 3 receptors). Consistent, tetrodotoxin (1 µM)-sensitive contractions were obtained using 5-HT 1 µM (~90 s contact every 15 min). Concentration-response curves were then constructed to 5-HT, prucalopride, tegaserod or SL65.0155 using single concentrations/ tissue (~90 s; 15 min). Contractions were expressed as a % of the previous 5-HT contractions. In the continued presence of SL65.0155 or vehicle (dH2O), 1 µM 5-HT was re-applied 15 min later to investigate any antagonist activity. The same protocol studied the ability of SB204070 (Wardle et al, 1994) to inhibit the effects of the 5-HT4 receptor agonists. The effects of SL65.0155 were studied on submaximal contractions evoked by carbachol 1 µM (~90 s, 15 min) instead of 5-HT. Statistical comparisons were made against time-matched vehicle controls using an unpaired two-tailed Student’s t-Test. 5-HT (0.1 nM – 1 µM) and prucalopride (1 nM - 10 µM) induced concentration-dependent contractions with similar maxima (108 ± 9 % of control 5-HT at 1 µM, pEC50: 8.0, n=4 and 104 ± 12 % at 10 µM, pEC 50: 7.3, n=5 respectively, P=0.8). These maximum responses were blocked by SB204070 1 µM (contraction: -0.8 ± 5.0 % of control 5-HT, P<0.001 and 9.5 ± 5.0 %, P<0.05 respectively, n=4). Tegaserod (1 nM – 30 µM) produced a bell-shaped concentration-response curve (maximum of 65 ± 17 % at 10 µM; n=6), blocked by SB204070 1 µM (contraction: -6.1 ± 19.8 % of control 5-HT, P<0.05, n=6). SL65.0155 (1 nM-10 µM) did not induce contractions but 1 & 10 µM reduced the response to subsequent 5-HT (contraction: 14.7 ± 3.8 % of control 5-HT, P<0.01 and -3.1 ± 2.0 %, P<0.001 respectively, n=4). Compared to time-matched vehicles (contraction: 123.2 ± 8.4 % of control carbachol, n=4), SL65.0155, 1 & 10 µM, reduced carbachol-induced contractions (97.6 ± 5.7 % of control carbachol, P<0.05 and 45.5 ± 10.4 %, P<0.001 respectively, n=5). In guinea-pig isolated colon, a well-coupled model of the 5-HT4 receptor, responsive to the 5-HT4 receptor agonists and prokinetic agents prucalopride and tegaserod, SL65.0155 inhibits 5-HT4 receptor mediated responses but at the high concentrations used, some additional, non-selective activity may be present.
Moser P.C., et al (2002) J. Pharmacol. Exp. Ther , 302, 731-741. |
