Transepithelial Na+ transport in tight epithelia involves the movement of ions across two separate barriers: the apical and basolateral membranes. We are characterizing the involvement of these membranes in the response to regulatory influences using the frog skin as a model epithelium. An important part of our experiments involves the use of intracellular microelectrodes to determine the current-voltage relationships (IV) of the two barriers. With respect to the apical membrane we are testing whether the relationship between the Na current and the membrane potential is described by the Goldman constant-field equation over a wide variety of circumstances. Conformity with this equation allows the calculation of the conductance, permeability and electromotive force of this barrier. We are using the IV relationship of the basolateral border to attempt to characterize the changes of this border during the action of regulators. Ion selective microelectrodes for Na+ and K+ will be used to examine the dependence of the transport parameters of the two barriers on cytoplasmic ion activity. The measurements of intracellular ion activities will also be used to verify the conclusions reached from measurements of the IV relationship. In addition, the microelectrode data will allow us to determine the conditions in which to use noise analysis to examine the relationship between the electrochemical driving force and the single channel current of the apical border. In addition to studying the better characterized modifiers of ion transport, special attention will be given to the regulation exerted by cell volume and cell Ca2+ activity. We will particularly examine the role of prostaglandin release in the responses to volume changes. Finally, in a different group of experiments we will examine whether Cl- moves across the epithelium through the same cells that transport Na+ or through a different pathway.
