The effects of waglerin-I, a toxin from Trimeresurus wagleri, on neuromuscular (NM) transmission were studied on the phrenic nervediaphragm preparation and triangularis sterni nerve-muscle preparation of mice. The toxin (1.2–4.0 μM) reversibly inhibited the indirectly elicited twitch tension of the diaphragm and decreased the ACh-elicited muscle contracture of chronically denervated diaphragm, while the directly elicited twitch tension was not affected. The toxin reversibly decreased the amplitude of miniature endplate potential (MEPP) at 0.52 μM and endplate potential (EPP) at 1.2–4 nM. The toxin (120nM–0.4 μM) also decreased the quantal content of EPP. The perineural waveforms were recorded with an extracellular electrode placed into the perineural sheaths of motor nerves of M. triangularis sterni. The toxin (4 μM) did not alter the amplitudes of waveforms related to sodium and potassium currents of the nerve terminal action potential, while the waveform related to calcium current was decreased. It is concluded that the toxin acts on both presynaptic and postsynaptic sites of the mouse motor endplate, and that the presynaptic effect is apparently more potent than the postsynaptic effect.