In the present study, well-resolved single-channel events of cloned human and murine homomeric 5-hydroxytryptamine type 3 (h5-HT(3A) and m5-HT(3A)) receptors, expressed in human embryonic kidney 293 cells, are reported for the first time. 5-HT (1 microM) applied in Ca(2+)- and Mg(2+)-free external solution to excised outside-out membrane patches induced non-rectifying single-channel inward currents. These currents were not observed in untransfected cells or in the presence of the antagonist ondansetron (1 microM). The mean single-channel conductance of the h5-HT(3A) and m5-HT(3A) receptors was 48+/-8 pS and 46+/-7 pS, the mean reversal potential was 10.3+/-1.8 mV and 4.7+/-5.3 mV, respectively. The analysis of single-channel open-times revealed for both h5-HT(3A) and m5-HT(3A) receptors only one type of open state, but different mean open-times (7.1+/-2.9 ms vs. 4.1+/-0.8 ms) indicating species-dependent gating mechanisms of 5-HT(3) receptors.