Forskolin increases apical sodium conductance in cultured toad kidney cells (A6) by stimulating membrane insertion

The role of membrane traffic in the stimulation of apical Na+ permeability caused by increases in cytoplasmic cyclic AMP was assessed by measuring the effects of forskolin on transepithelial capacitance (C-T), transepithelial conductance (G(T)), and short-circuit current (I-sc) in A6 cultured toad k idney cells. Apical water per meability was probed by recording cell volume changes after reducing the osmolality of the apical bath. We found that fo rskolin does not increase the osmotic water permeability of the apical memb rane of A6 cells, and thus does not stimulate the insertion of water channe ls. Comparison of the effects of forskolin and insulin on Na+ transport dem onstrated that both agents produce reversible increases in C-T, G(T) and I- sc. G(T) and C-T increased proportionally during the rising phase of the in sulin response. However, a non-linear relationship between both parameters was recorded when forskolin was given in NaCl Ringer's solution. The relati onship between C-T and G(T) became linear after the effects of forskolin on Cl- conductances were eliminated by substituting Cl- by an impermeant anio n. In contrast, in Cl--containing Na+-free solutions, the nonlinearity beca me more pronounced. Successive additions of insulin and forskolin caused ad ditive increases in C-T. Because increases in C-T and Na+ transport occurre d in the absence of stimulation of water permeability and increases of C-T and G(T) were directly proportional when Na+ was the major permeating ion a cross the apical membrane, we suggest that the increase in apical Na+ perme ability in the presence of either forskolin or insulin is due to the insert ion of channels residing in intracellular pools. In contrast, the increased Cl- permeability caused by forskolin may be related to the activation of c hannels already present in the membrane.