Tetanus toxin (TeTX) inhibits the evoked release of acetylcholine (ACh) at rat soleus end-plates. The effects of various procedures which evoke ACh release by raising the level of free intracellular calcium have been investigated at various stages of tetanus intoxication. At all stages studied TeTX has little or no effect on either the frequency or the amplitude of spontaneous miniature end-plate potentials (m.e.p.p.s). After TeTX poisoning, e.p.p. latency is more variable than normal and the slope of the relationship between ln m (quantal content) and ln [Ca]o is reduced from the control value of about 4. Plots of m-1/n against 1/[Ca]o, for n = 1-4, suggest that mmax, the maximum number of quanta releasable by nerve stimulation, is reduced at intoxicated end-plates. Blocking delayed rectification with 3-aminopyridine (1-5 mM) increases m, but has little or no effect on either the slope of ln m-ln [Ca]o plots or estimates of mmax. Several treatments which raise m.e.p.p. rate (high [Ca]o, hyperosmotic medium, addition of lanthanum) are less effective after TeTX poisoning. Some of the tested agents increase m.e.p.p. frequency by a mechanism which is thought to involve a mobilization of calcium from intracellular stores. The decline in m.e.p.p. rate after a period of high-frequency nerve stimulation is different at normal and TeTX-treated end-plates. At tetanus-intoxicated end-plates, the decline differs from that expected if TeTX acted simply to block calcium entry into the terminal. In addition, the increase in m.e.p.p. frequency observed with a high rate of nerve stimulation suggests that considerable amounts of calcium can enter the terminal with each action potential. It is concluded that TeTX blocks transmitter release by acting at a step between calcium influx to the terminal and transmitter release such that the mechanism for ACh release shows a reduced sensitivity to intracellular calcium. The possibility of an additional effect on the presynaptic calcium conductance cannot be excluded. Some differences between the properties of end-plates poisoned with TeTX and botulinum toxin are discussed.