Permeation of divalent cations through alpha-latrotoxin channels in lipid bilayers: steady-state current-voltage relationships.

alpha-Latrotoxin, a polypeptide neurotoxin known to cause massive release of transmitter from vertebrate nerve terminals, is thought to act by forming cation-selective channels in plasma membranes. This paper describes the steady-state current carried by Ca2+, Sr2+ and Ba2+ through pores of alpha-LaTx molecules incorporated in artificial bilayer membranes made of neutral lipids. Even when the solutions separated by the membrane are identical, the I-V relations rectify strongly, the current being higher when the side to which the toxin is added is positive. The polarity of the rectification is consistent with the hypothesis that the mechanism of action of the toxin is, at least in part, that of promoting inwardly directed flow of cations, and thus, accumulation of Ca2+ and other ions in the intracellular spaces. The dependence of the I-V characteristics on voltage and Ca2+ concentration is well described by a one-site, one-ion model for a channel. Three parameters of the model are deduced: the binding constant of the site for Ca2+, K = 1.5 M-1 (or K = 7 M-1 when activities are used instead of concentrations); the "electrical" distance of the site from the toxin-containing solution, alpha = 0.3; the free energy difference between the two barrier peaks, delta F = 0.26 kT. The values of the parameters deduced by studying the channel in the presence of Ca2+ give theoretical curves that also fit the data with Sr2+ and Ba2+, indicating a low level of discrimination among these three cations.