THE MAMMALIAN NA+/H+ ANTIPORTERS NHE-1, NHE-2, AND NHE-3 ARE ELECTRONEUTRAL AND VOLTAGE INDEPENDENT, BUT CAN COUPLE TO AN H+ CONDUCTANCE

Na+/H+ exchange in vertebrates is thought to be electroneutral and insensitive to the membrane voltage. This basic concept has been challenged by recent reports of antiport-associated currents in the turtle colon epithelium (Post and Dawson, 1992, 1994). To determine the electrogenicity of mammalian antiporters, we used the whole-cell patch clamp technique combined with microfluorimetric measurements of intracellular pH (pH(i)). In murine macrophages, which were found by RT-PCR to express the NHE-1 isoform of the antiporter, reverse (intracellular Na+-driven) Na+/H+ exchange caused a cytosolic acidification and activated an outward current, whereas forward (extracellular Na+-driven) exchange produced a cytosolic alkalinization and reduced a basal outward current. The currents mirrored the changes in pH(i), were strictly dependent on the presence of a Na+ gradient and were reversibly blocked by amiloride. However, the currents were seemingly not carried by the Na+/H+ exchanger itself, but were instead due to a shift in the voltage dependence of a preexisting H+ conductance. This was supported by measurements of the reversal potential (E(rev)) of tail currents, which identified Ht (equivalents) as the charge carrier. During Na+/H+ exchange, E(rev) changed along with the measured changes in pH(i) (by 60-69 mV/pH). Moreover, the current and Na+/H+ exchange could be dissociated. Zn2+, which inhibits the Hf conductance, reversibly blocked the currents without altering Na+/H+ exchange. In Chinese hamster ovary (CHO) cells, which lack the H+ conductance, Na+/H+ exchange produced pH(i) changes that were not accompanied by transmembrane currents. Similar results were obtained in CHO cells transfected with either the NHE-1, NHE-2, or NHE-3 isoforms of the antiporter, indicating that exchange through these isoforms is electroneutral. In all the isoforms tested, the amplitude and time-course of the antiport-induced pH(i) changes were independent of the holding voltage. We conclude that mammalian NHE-1, NHE-2, and NHE-3 are electroneutral and voltage independent. In cells endowed with a pH-sensitive H+ conductance, such as macrophages, activation of Na+-H+ exchange can modulate a transmembrane H+ current. The currents reported in turtle colon might be due to a similar ''cross-talk'' between the antiporter and a H+ conductance.