[3H]-Mesulergine labels 5-HT7 sites in rat brain and guinea-pig ileum but not rat jejunum

Abstract

1. The primary aim of this investigation was to determine whether binding sites corresponding to the 5-HT7 receptor could be detected in smooth muscle of the rat jejunum. Binding studies in rat brain (whole brain minus cerebellum) and guinea-pig ileal longitudinal muscle were also undertaken in order to compare the binding characteristics of these tissues. Studies were performed using [3H]-mesulergine, as it has a high affinity for 5-HT7 receptors. 2. In the rat brain and guinea-pig ileum, pKD values for [3H]-mesulergine of 8.0 +/- 0.04 and 7.9 +/- 0.11 (n = 3) and Bmax values of 9.9 +/- 0.3 and 21.5 +/- 4.9 fmol mg(-1) protein were obtained respectively, but no binding was detected in the rat jejunum. [3H]-mesulergine binding in the rat brain and guinea-pig ileum was displaced with the agonists 5-carboxamidotryptamine (5-CT) > 5-hydroxytryptamine (5-HT) > or = 5-methoxytryptamine (5-MeOT) > sumatriptan and the antagonists risperidone > or = LSD > or = metergoline > ritanserin > > pindolol. 3. Despite the lack of [3H]-mesulergine binding in the rat jejunum, functional studies undertaken revealed a biphasic contractile response to 5-HT which was partly blocked by ondansetron (1 microM). The residual response was present in over 50% of tissues studied and was found to be inhibited by risperidone > LSD > metergoline > mesulergine = ritanserin > pindolol, but was unaffected by RS 102221 (3 microM), cinanserin (30 nM), yohimbine (0.1 microM) and GR 113808 (1 microM). In addition, the agonist order of potency was 5-CT > 5-HT > 5-MeOT > sumatriptan. 4. In conclusion, binding studies performed with [3H]-mesulergine were able to detect 5-HT7 sites in rat brain and guinea-pig ileum, but not in rat jejunum, where a functional 5-HT7-like receptor was present.

Figure 1

Saturation curve of [³H]-mesulergine binding in the rat brain in the absence and presence of yohimbine (Y: 0.1 μM), prazosin (P: 0.1 μM) cinanserin (C: 30 nM) and RS 102221 (RS: 0.3 and 3 μM). All experiments were performed in the presence of raclopride (RC: 1 μM). Abscissa: Concentration of [³H]-mesulergine in nM; Ordinate: fmole mg of protein⁻¹. Each point is the mean±s.e.mean from three experiments.

Figure 2

(a, b) inhibition of [³H]-mesulergine binding to rat brain and (c, d) guinea-pig ileum by various ligands. Abscissa: Log molar concentration of the displacing drug, Ordinate: % of specific binding. The data represent the mean±s.e.mean per cent of maximum specific binding (defined with 1 μM risperidone) of at least three experiments.

Figure 3

Correlation between pK_i values obtained in recombinant 5-HT₇ receptors from rat or mouse and expressed in transfected cells (Roth et al., 1994; Plassat et al., 1993; Ruat et al., 1993; Shen et al., 1993) and pK_i or pK_B values obtained in rat brain, guinea-pig ileum and rat jejunum. Abscissa: pK_i values for recombinant 5-HT₇ receptors, Ordinate: pK_i or pK_B values obtained in rat brain and jejunum and guinea-pig ileum. The continuous line represents a line of identity.

Figure 4

Concentration-response curve for the contractile effect of 5-HT in the absence and presence of ondansetron (0.1, 1 and 10 μM) in isolated rat jejunum. Abscissa: Log molar concentration of 5-HT, Ordinate: % of maximum contraction obtained with acetylcholine (1 μM). The data represent the mean±s.e.mean of four paired experiments.

Figure 5

Concentration-response curves for 5-HT in the absence and presence of metergoline (10, 30 and 100 nM). Abscissa: Log molar concentration of 5-HT, Ordinate: % of maximum contraction obtained with acetylcholine (1 μM). The data represent the mean±s.e.mean of three paired experiments. All experiments were conducted in the presence of ondansetron (1 μM). The slope of the Schild regression (inset) was 0.8±0.09.