The role of the agonist binding site in Ca(2+) inhibition of the recombinant 5-HT(3A) receptor.

The mechanism and site of action of Ca(2+) at the recombinant murine 5-hydroxytryptamine (5-HT)(3A) receptor were investigated using whole-cell voltage clamp, radioligand binding and single-cell Ca(2+) imaging. Inhibition of the 5-HT (3 microM)-induced response by 10 mM 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(50) from 2.35 to 3.87 microM and a 30% reduction in I(max). Ca(2+) also resulted in the inhibition of binding of both 5-HT(3) receptor agonist [3H]m-chlorophenylbiguanide and antagonist [3H]granisetron due to an increase in K(d), with no change in B(max). An increase in EC(50) from 2.6 (1 mM Ca(2+)) to 4.7 microM (10 mM Ca(2+)), with no change in maximal [Ca(2+)](i), was observed from Ca(2+) imaging studies. Largely similar effects were observed with Mg(2+). The combined data suggest that Ca(2+) acting at a site that directly or indirectly influences the agonist binding site plays a significant role in its inhibitory effect at the 5-HT(3) receptor.