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Receptor‐dependent translocation of diphtheria toxin across the surface membrane of Vero cells was studied using patch clamp techniques. Translocation was induced by exposing cells with surface‐bound toxin to low pH. Whole cell current and voltage clamp recordings showed that toxin translocation was associated with membrane depolarization and increased membrane conductance. The conductance increase was voltage independent, with a reversal potential of approximately 15 mV. This value was unaffected by changing the Cl‐ gradient across the membrane and microfluorometric measurements showed that the cytosolic Ca2+ concentration was only marginally elevated by the translocation. The conductance increase is thus mainly due to monovalent cations. Exposing outside‐out and cell‐attached patches with bound toxin to low pH induced a new type of ion channel in the membrane. The channel current was inward at negative membrane potentials and the single channel conductance was approximately 30 pS. This value is about three times larger than for receptor‐independent channels induced by diphtheria toxin or toxin fragments in artificial lipid membranes.

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The marine flagellate Prymnesium patelliferum produces toxins lethal to fish. The toxin extracted from the alga has haemolytic, cytotoxic and neurotoxic effects, but the action mechanisms of the toxin are not known in detail. We have examined the toxin effects on the voltage sensitive Ca(2+)-currents, the cytosolic Ca(2+)-level ([Ca2+]i) and the prolactin release in clonal rat anterior pituitary GH4C1 cells, which possess T- and L-type Ca(2+)-channels. The trans-membrane Ca(2+)-current was recorded using whole-cell voltage clamp. After 5-15 min exposure to the algal toxin at a final concentration of 50,000-100,000 cells mL-1, the Ca(2+)-currents through both the T- and L-channels showed a 2-3-fold enhancement. The voltage sensitivity of the Ca(2+)-currents was not affected by the algal toxin, and the toxin-induced currents were inhibited by 100 microM of the Ca(2+)-channel blocker D-600. In toxin-exposed cells microfluorometric measurements based on fura-2 revealed an increase of [Ca2+]i from 100-150 to 300-500 nM. This elevation was delayed and partially inhibited by 100 microM D-600. The algal toxin induced prolactin release in a dose-dependent manner, and this effect was inhibited by the Ca(2+)-channel blocker verapamil. We therefore conclude that the toxin of P. patelliferum affects the Ca2+ homeostasis of the pituitary cells by increasing the leak through voltage sensitive Ca(2+)-channels, resulting in increased [Ca2+]i and secretion of prolactin.

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