PHARMACOLOGICAL CHARACTERIZATION OF 5-HYDROXYTRYPTAMINE(3) RECEPTORS IN MURINE BRAIN AND ILEUM USING THE NOVEL RADIOLIGAND [H-3] RS-42358-197 - EVIDENCE FOR RECEPTOR HETEROGENEITY

Previous studies have demonstrated species-specific differences in 5-hydroxytryptamine3 (5-HT3) receptors, but unequivocal evidence of 5-HT3 receptor subtypes, within a species, has not yet been obtained. The purpose of the current study was to test for heterogeneity in 5-HT3 receptors in murine tissues. 5-HT3 receptors in membranes derived from brain cerebral cortex of CD-1, C57Bl/6, and Swiss Webster mice and ileum of CD-1 mice were labeled with the 5-HT3 receptor antagonist [H-3]RS-42358-197. Structurally diverse competing ligands were then used to characterize the binding site. [H-3]RS-42358-197 bound with similar affinity in each of the cortical tissues (mean K(D) = 0.14 nM; range, 0.06-0.32 nM) but bound with lower affinity in ileal tissue (2.5 nM). The density of sites labeled with [H-3]RS-42358-197 ranged from 10.4 fmol/mg of protein in Swiss Webster mouse cortex to 44.2 fmol/mg of protein in Sprague-Dawley rat cortex. Displacing ligands produced a pharmacologic profile of the [H-3]RS-42358-197 binding site consistent with it being a 5-HT3 receptor: (R)-YM060 > (S)-zacopride > (R)-zacopride > MDL 72222 > 2-methyl-5-HT. However, greater-than-or-equal-to 10-fold differences in the affinity of certain ligands were found when comparing 5-HT3 binding sites in membranes from cerebral cortex of the different strains of mice and when comparing 5-HT3 binding sites in brain and ileal membranes prepared from the CD-1 mouse strain. Ligands demonstrating selectivity included RS-42358-197, (R)-zacopride, 1-(m-chlorophenyl)biguanide, (R)-YM060, and MDL 72222. These studies demonstrate tissue- and strain-dependent differences in murine 5-HT3 binding sites. This suggests that 5-HT3 receptors exist as multiple subtypes within species and that subtype-selective 5-HT, ligands may be identified.