Experiments were carried out on the isolated rat phrenic nerve diaphragm preparations and chick biventer cervicis preparations. Cu2+ at cumulative concentrations inhibited muscle contractile response to indirect stimulation without affecting the conductivity of the nerve. The inhibiting action of Cu2+ on the muscle contractile response to indirect stimulation was stronger than that to direct stimulation. Cu2+ did not affect the Ach sensitivity of the chick biventer cervicis muscle. The effects of Cu2+ on neuromuscular transmission were antagonized by Ca2+ and L-cysteine. It is concluded that Cu2+ produces neuromuscular blockade by acting mainly on the presynaptic nerve terminals, binding with the -SH groups of the membrane and thus altering the transmembrane movement of Ca2+ and reducing the release of Ach from the nerve terminals.