Cytolysin produced by Vibrio vulnificus has been incriminated as one of the important virulence determinants in V. vulnificus infection. Ion selectivity of cytolysin-induced pores was examined in a CPAE cell, a cell line of pulmonary endothelial cell, using inside-out patch clamp techniques. In symmetrical NaCl concentration (140 mM), intracellular or extracellular application of cytolysin formed ion-permeable pores with a single channel conductance of 37.5⁑4.0 pS. The pore currents were consistently maintained after washout of cytolysin. Replacement of Na in bath solution with monovalent ions (K, Cs or TEA) or with divalent ions (Mg2, Ca2) did not affect the pore currents. When the NaCl concentration in bath solution was lowered from 140 to 60 and 20 mM, the reversal potential shifted from 0 to 11.8 and 28.2 mV, respectively. The relative permeability of the cytolysin pores to anions measured at 40 mV was Cl = NO2 ≥ Br = I ＞ SCN ＞ acetate ＞ isethionate ＞ ascorbic acid ＞ EDTA2, in descending order. The cytolysin-induced pore current was blocked by Cl channel blockers or nucleotides. These results indicate that V. vulnificus cytolysin forms anion-selective pores in CPAE cells.
