Different effects of N-ethylmaleimide on the binding characteristics of 5-hydroxytryptamine receptor subtypes in rat brain membranes

Abstract

To characterize 5-hydroxytryptamine (5-HT) receptor subtypes in the CNS in terms of their binding and coupling properties with GTP-binding proteins (G proteins), we examined the effects of serotonergic agents GTP?S and N-ethylmaleimide (NEM), on the binding of [(3)H]8-OH-DPAT, [(125)I]iodo-cyanopindolol (ICYP), [(3)H]mesulergine, [(3)H]DOB and [(3)H]ketanserin in rat brain membranes. [(3)H]8-OH-DPAT, [(125)I]ICYP, [(3)H]mesulergine and [(3)H]ketanserin are radiolabeled ligands for 5-HT(1A), 5-HT(1B), 5-HT(1C) and 5-HT(2) sites, respectively. Correlation of the values of the dissociation constants (K(D)) of the serotonergic agonists or antagonists was evaluated in two categories of 5-HT receptor subtypes. Highly significant correlations were found among 5-HT(1A), 5-HT(1B) and 5-HT(1D) sites, and among 5-HT(1C) and 5-HT(2) sites. A significantly reversed correlation was observed between 5-HT(1A) and 5-HT(2). GTP?S caused a shift to the right on the 5-HT competition curve for the binding of [(125)I]ICYP, [(3)H]mesulergine or [(3)H]ketanserin. Although NEM-treatment partially induced a right shift on the agonist competition curve for [(125)I]ICYP binding, this treatment induced a left shift on the curve for [(3)H]mesulergine and [(3)H]ketanserin binding. In addition, the binding of [(3)H]DOB, a 5-HT(2) agonist, was increased by NEM-treatment. NEM seems to characterize agonist binding to the 5-HT receptor subtypes and the coupling of these subtypes to G proteins. These results suggest that 5-HT(1A), 5-HT(1B) and 5-HT(1D) receptors belong to one category, and 5-HT(1C) and 5-HT(2) receptors belong to another category.