The modulation by Zn(2+) of recombinant murine 5-hydroxytryptamine(3A) (5-HT(3A)) receptor responses and its modification by Ca(2+) or Mg(2+) were studied using whole-cell voltage clamp and radioligand binding techniques. In the absence of other added divalent cations Zn(2+) enhanced the response to 5-HT by increasing maximum peak current (I(max)) to a maximum of 122.5%, decreasing the rate of desensitization (maximum t(1/2)=210%), and decreasing the EC(50) by approximately two fold. In the presence of Ca(2+) or Mg(2+), the effects of Zn(2+) on I(max) and t(1/2) were still manifest, although higher Zn(2+) concentrations were required; however, the effect on EC(50) was abolished. Zn(2+) also enhanced [3H]agonist but not [3H]antagonist binding. We propose there is more than one Zn(2+) binding site on the 5-HT(3) receptor molecule, and that one or more of these sites may also bind Ca(2+) and Mg(2+).