Methods of blocker-induced noise analysis were used to investigate the way in which forskolin and vasopressin stimulate Na transport at apical membranes of short-circuited frog skin transporting Na at spontaneous rates of transport. Experiments were done under conditions where the apical Ringer solution contained either 100 mM Na or a reduced Na concentration of 5 or 10 mM Na and buffered with either HCO3 or HEPES. Reduction of apical solution Na concentration caused a large autoregulatory increase of Na channel density (NT) similar in magnitude to that observed previously in response to blocker (amiloride) inhibition of apical membrane Na entry. Forskolin at 2.5 microM caused maximal and reversible large increases of NT, which were larger than could be elicited by 30 mU/ml vasopressin. In both the absence and presence of the autoregulatory increase of NT (caused by reduction of apical Na concentration), forskolin caused large increases of NT. Although the fractional increases of NT in response to forskolin were roughly similar, the absolute increases of NT were considerably larger in those tissues studied at reduced Na concentration and where baseline values of NT were markedly elevated by reduction of apical Na concentration. Because the effects on NT were additive, it is likely that the cAMP-dependent and autoregulatory mechanism that lead to changes of NT are distinct. We speculate that autoregulation of NT may involve change of the size of a cytosolic pool of Na-containing vesicles that are in dynamic balance with the apical membranes. cAMP-dependent regulation of NT may involve change of the dynamic balance between vesicles and the apical membranes of these epithelial cells. Alternative hypotheses cannot at present be ruled out, but will require incorporation of the idea that regulation of NT can occur both by hormonal and nonhormonal (autoregulatory) mechanisms of action.

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