Effects of chemical modification of carboxyl groups on the voltage-clamped nerve fiber of the frog

Abstract

Voltage-clamped single nerve fibers of the frog Rana esculenta were treated with the carboxyl group activating reagent N-ethoxy-carbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) in the presence of different primary amines and without added amine. Carboxyl groups form stable amide bonds with primary amines in the presence of EEDQ. EEDQ treatment reduced the sodium current considerably and irreversibly, regardless of the presence of a primary amine in the Ringer's solution. The potassium current was also reduced. After modification the reduced sodium currents inactivated slowly and incompletely. The descending branch of the sodium current-voltage relation, INa(E), was shifted along the voltage axis in the depolarizing direction. The size of the shift was strongly dependent on the amine present during modification with EEDQ. The voltage-dependence of sodium inactivation, h infinity (E), was shifted to more positive values of membrane potential by EEDQ in the presence of ethylenediamine (11 mV) and glucosamine (3 mV). In contrast, a small shift to more negative potentials occurred in the presence of taurine (-3 mV) or without the addition of an amine (-2 mV). A tenfold increase of the calcium concentration still shifted the INa(E) and h infinity (E) curves of the chemically modified fibers. However, these shifts were smaller than those observed on untreated fibers. The currents remaining after the modification were completely blocked by tetrodotoxin; no change of the reversal potential occurred.