1. The mechanism of a long-term potentiation of transmitter release (pre-LTP) induced by a tetanic stimulation (33 Hz for 1-30 s) applied to the preganglionic nerve was examined by intracellularly recording the fast excitatory postsynaptic potentials (fast EPSPs) in bull-frog sympathetic ganglia. 2. Short-term facilitation induced by paired pulses was decreased during the course of pre-LTP; the extent of reduction paralleled with the magnitude of pre-LTP. 3. The frequency of miniature EPSPs increased after tetanic stimulation that produced the pre-LTP. 4. The Ca2+ ionophore, A23187, increased both the amplitude and quantal content of fast EPSPs and frequency of miniature EPSPs while it decreased short-term facilitation. 5. A Ca2+ chelating agent, Quin-2, loaded as acetoxymethyl ester, reduced the amplitude and quantal content of fast EPSPs and short-term facilitation, and blocked the generation of pre-LTP. 6. Activators of protein kinase C, phorbol 12,13-dibutyrate and 1-oleoyl-2-acetyl-rac-glycerol, and its inhibitors, H-7 and staurosporine, did not block the generation of pre-LTP, while the activators enhanced transmitter release. 7. Inhibitors of calmodulin, trifluoperazine and W-7, blocked the generation of pre-LTP, whereas the amplitude and quantal content of fast EPSPs were not influenced. 8. These results suggest that the pre-LTP results from a sustained rise in the basal level of intraterminal Ca2+ and an activation of the Ca(2+)-calmodulin-dependent process in the preganglionic nerve terminals.