The role of the agonist binding site in Ca2+ inhibition of the recombinant 5-HT3A receptor

The mechanism and site of action of Ca2+ at the recombinant murine 5-hydroxytryptaminee (5-HT),, receptor were investigated using whole-cell voltage clamp, radioligand binding and single-cell Ca2+ imaging. Inhibition of the 5-HT (3 muM)-induced response by 10 muM Ca 2+ reached a plateau at 68.5% inhibition, with half-maximal effect at 2.6 mM, This was due to an increase in EC., from 2.35 to 3.87 LM and a 30% reduction in I-max. Ca2+ also resulted in the inhibition of binding of both 5-HT3 receptor agonist [H-3]m-clorophenylbi [3 2 +) to guanide and antagonist [H-3]granisetron due to an increase in K-d with no change in B (max). An increase in EC50, from 2.6 0 mM Ca 4.7 PM (10 inM Ca2'), with no change in maximal [Ca 2+]i, was observed from Ca2' imaging studies. Largely similar effects were observed with Mg2'. The combined data suggest that Ca2+ acting at a site that directly or indirectly influences the agonist binding site plays a significant role in its inhibitory effect at the 5-HT3 receptor. (9 2001 Published by Elsevier Science B.V.