An early effect of tetanus toxin is the blockade of inhibitory synaptic transmission. Potassium-evoked release of glycine has been used to evaluate tetanus toxin uptake and action in neuronal cell cultures. We tested the hypothesis that toxin is internalized into neurons by receptor-mediated endocytosis and that the toxin forms pH-dependent channels in the endosomal membrane, gaining access to the neuronal cytoplasm. Evidence was provided that toxin action is blocked by monensin, that toxin action correlates with the maintenance of a pH gradient in endosomes, that lowering the external pH allows direct toxin entry through the plasma membrane, by-passing the endosomal system, and that toxin collapses the pH gradient in neuronal endosomes. The time course of toxin action could be modified by alterations in the level of endogenous neuronal activity, suggesting an effect on the uptake mechanism. Preliminary data indicate that Brefeldin A, which disrupts membrane flow and exocytosis, reduces neurotransmitter release in neuronal cultures.
