Application of alpha-latrotoxin (alpha-LT), the active component of black widow spider venom (BWSV), to a vertebrate neuromuscular junction, in the presence of millimolar bath concentrations of Ca2+ or Mg2+, greatly increases the frequency of miniature end-plate potentials (Fmepp). We have further characterized the cation dependence of alpha-LT action at the frog cutaneous pectoris neuromuscular junction. The divalent cations, Ca, Sr, Ba at less than or equal to 50 microM, Zn, Mn, Cd at greater than or equal to 50-100 microM, and Mg at greater than or equal to 1.0 mM, as well as the trivalent cation La at greater than or equal to 15 microM, all increase Fmepp exponentially to greater than or equal to 100-200 s-1 over several minutes time. The exponential rate of rise is graded with extracellular cation concentration and can be reduced by increasing [K+] of the bath from 2 to 25-40 mM. Long-term exposure to alpha-LT in the presence of Sr2+ or Mn2+ results in the exhaustion of the releasable quantal store of transmitter, which in the case of Mn2+ correlates well with depletion of synaptic vesicles. These data support the hypothesis that BWSV promotes an increase in Fmepp by increasing nerve terminal permeability to multivalent cations that enter the nerve terminal down their electrochemical gradients and then may bind to quantal release activating sites or displace Ca2+ from intracellular stores.