Basolateral membrane potential and conductance in frog skin exposed to high serosal potassium

Abstract

In studies of apical membrane current-voltage relationships, in order to avoid laborious intracellular microelectrode techniques, tight epithelia are commonly exposed to high serosal K concentrations. This approach depends on the assumptions that high serosal K reduces the basolateral membrane resistance and potential to insignificantly low levels, so that transepithelial values can be attributed to the apical membrane. We have here examined the validity of these assumptions in frog skins (Rana pipiens pipiens). The skins were equilibrated in NaCl Ringer's solutions, with transepithelial voltage Vt clamped (except for brief perturbations delta Vt) at zero. The skins were impaled from the outer surface with 1.5 M KCl-filled microelectrodes (Rel greater than 30 M omega). The transepithelial (short-circuit) current It and conductance gt = -delta It/delta Vt, the outer membrane voltage Vo (apical reference) and voltage-divider ratio (Fo = delta Vo/delta Vt), and the microelectrode resistance Rel were recorded continuously. Intermittent brief apical exposure to 20 microM amiloride permitted estimation of cellular (c) and paracellular (p) currents and conductances. The basolateral (inner) membrane conductance was estimated by two independent means: either from values of gt and Fo before and after amiloride or as the ratio of changes (-delta Ic/delta Vi) induced by amiloride. On serosal substitution of Na by K, within about 10 min, Ic declined and gt increased markedly, mainly as a consequence of increase in gp. The basolateral membrane voltage Vi (= -Vo) was depolarized from 75 +/- 4 to 2 +/- 1 mV [mean +/- SEM (n = 6)], and was partially repolarized following amiloride to 5 +/- 2 mV.(ABSTRACT TRUNCATED AT 250 WORDS)