Effects of amiloride analogues on adult Notophthalmus viridescens limb stump currents.

We have previously investigated the relevance to limb regeneration of epidermally driven, Na+-dependent limb stump currents by blocking epidermal Na+ channels with amiloride, 3,5-diamino-6-chloro-N-(diaminomethylene)pyrazinecarboxamide. In order to reduce Notophthalmus viridescens stump currents more effectively than with amiloride, we have examined six amiloride analogues. Of these, only benzamil, 3,5-diamino-6-chloro-N-[(benzylamino)aminomethylene]pyrazinecarboxamide, was more effective than amiloride. The concentration of benzamil that reduced stump currents to half their initial value was 0.034 microM, while this concentration for amiloride was 0.42 microM. We also found a contribution of calcium ions to these currents. When immersed in water whose Ca2+ concentration decreased stepwise from 1 to 0 mM, stump currents decreased significantly, although to a variable extent, depending on the batch of newts. With 30 microM benzamil and 0.5 mM calcium (in water that also contained 1.5 mM NaCl and 0.06 mM KCl) stump currents could be reduced to very low levels and, in many cases, changed to incurrents.