Na+ transport and pH in principal cells of frog skin: effect of antidiuretic hormone.

Intracellular pH (pHi), apical membrane potential (Va), and fractional apical membrane resistance (FRa) were measured in principal cells of isolated frog skin (Rana pipiens) with double-barreled microelectrodes under short-circuit conditions. Basolateral exposure to 10 mU/ml arginine vasotocin (AVT) depolarized Va by 30 mV, decreased FRa by 33%, increased short-circuit current (Isc) by 17 microA, and increased pHi by 0.17 pH units. The response of Va, Isc, and pHi occurred concurrently. Forskolin, theophylline, and 8-(4-chlorophenyl-thio)-adenosine 3',5'-cyclic monophosphate caused similar changes in Va, Isc, and pHi. The enhanced response of Isc, Va, and FRa to short pulses of apical amiloride applied during AVT or cAMP exposure suggests an increase in apical Na+ conductance. The presence of cAMP agonists also enhanced the response of pHi to amiloride. We conclude that the AVT- and cAMP-induced increase in Na+ transport across the apical cell membrane is associated with a change in pHi. These data are consistent with the hypothesis that changes in pHi may play a role in the second messenger cascade initiated by the antidiuretic hormone.

Na+ channel blockers inhibit voltage-dependent intracellular pH changes in principal cells of frog (Rana pipiens) skin.

1. The relationship between Va and pHi was studied with double-barrelled microelectrodes in principal cells of frog skin (Rana pipiens) when (i) the transepithelial potential (Vt) was clamped at different values of Vt and (ii) when the pH of the apical solution was altered. 2. Under all conditions examined here, depolarization of Va was associated with an increase in pHi and hyperpolarization of Va was accompanied by a decrease in pHi. However, the changes in the basolateral cell membrane potential occurred, either in the same or opposite direction to that of Va depending on the conditions. 3. The voltage-dependent changes in pHi were not affected by H+ transport inhibitors or the complete removal of Na+, Cl- and HCO3- but were effectively inhibited by the application of amiloride (10(-4) M) or benzamil (10(-6)M) on the apical side. 4. A decrease in pH of the apical solution hyperpolarized Va and decreased pHi, an effect that was significantly attenuated when benzamil was present on the apical side. 5. The results indicate the presence of an H+ and/or OH- conductive pathway in the apical cell membrane of the principal cells. The effect of Na+ channel blockers suggests that this pathway proceeds through the apical Na+ channels.

Effect of changes in extracellular potassium on intracellular pH in principal cells of frog skin.

Intracellular pH (pHi), apical membrane potential (Va), and fractional apical membrane resistance (FRa) were measured in principal cells of isolated frog skin (Rana pipiens) with double-barreled microelectrodes under short-circuit and open-circuit conditions. Basolateral exposure to high K+ concentration or Ba2+ depolarized V(a), decreased short-circuit current, and increased FRa and pHi. However, an increase in K+ subsequent to Ba2+ application did not induce additional changes in these parameters. High basolateral K+, previously shown to increase apical K+ secretion (N. S. Bricker, T. Biber, and H. H. Ussing. J. Clin. Invest. 41: 88-99, 1963), also increased apical Na+ conductance. The depolarization and intracellular alkalinization induced by high K+ were also observed in absence of Na+, Cl-, and HCO3- and in presence of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid. Under all these conditions pHi moved toward electrochemical equilibrium. Reduced basolateral K+ hyperpolarized V(a) and decreased pHi. The data suggest that depolarization and hyperpolarization of the apical and/or basolateral membrane are associated with an increase and decrease, respectively, in pHi without involvement of Na(+)-H+, Cl(-)-HCO3-, or K(+)-H+ exchange and are apparently also independent of an active H+ secretion pathway. This indicates the presence of a potential-dependent H+ and/or OH- conductance in the apical and/or basolateral cell membrane that may play an important role in pHi regulation and signal transduction.