Transepithelial (psi T) and basolateral (psi BL) potential difference was measured in rabbit proximal convoluted tubules perfused in vitro. In control solution without protein, the mean psi BL was -54 +/- 2.2 mV (n = 57). Luminal substitution of K by Na had no effect. Complete luminal substitution of glucose and alanine, 110 mM substitution of Na or NaCl produced transient hyperpolarizations of psi BL of 14, 10, and 13 mV, respectively, with a return close to the control value within 4-8 min in all cases. Returning to control solution produced similar time-course transient depolarizations of psi BL of 17, 11, and 16 mV, respectively, again with a return to the control value in 4-10 min. Omission of glucose and alanine in the perfusate produced a decrease in cell volume of 14% that was maximal in 4 min with a complete recovery in the post-control period. A 110 mM luminal or peritubular substitution of Cl by cyclamate produced no significant effect on psi BL after taking into account the large psi T generated by the diffusion of Cl across the paracellular pathway. On the other hand, complete peritubular substitution of K by Na and 110 mM substitution of Na or NaCl produced sustained but reversible depolarizations of psi BL of 37.5, 10.2, and 20.4 mV, respectively. The transient nature of the hyperpolarization following luminal substitution of glucose, alanine, or Na can be interpreted in terms of changes in the intracellular sodium activity that would affect the Na-K-ATPase pump. Similarly, the sustained depolarization seen after a peritubular substitution of K and Na would also be compatible with a decrease in the basolateral ionic pump activity.