The effect of tetanus toxin on neuropeptide hormone release from isolated nerve endings of the neural lobe of rat pituitaries (neurosecretosomes) was measured in a perfusion system. Tetanus toxin inhibited depolarization-evoked release of oxytocin and vasopressin in a time- and dose-dependent manner. At 1 microgram/ml, tetanus toxin blocked stimulated release by 85%. Tetanus toxin that was preincubated with a neutralizing monoclonal antibody or heated to 100 degrees C had no effect on hormone release. The ionophores A23187 and ionomycin were potent stimulators of hormone release in control nerve endings, but were not able to overcome the effect of tetanus toxin in intoxicated nerve endings. 8-Bromo-cyclic GMP, which has been reported to reverse the action of tetanus toxin in PC12 cells, had no effect on the action of tetanus toxin in neurosecretosomes. Neurosecretosomes are the first system in which tetanus toxin has been shown to block release from peptidergic nerve terminals. They appear to be a valuable in vitro system for studying the biochemical mechanism of tetanus toxin action.