pH sensitivity of ammonium transport by Rhbg

Rhbg is a membrane glycoprotein that is involved in N[H.sub.3]/N[H.sup.+.sub.4] transport. Several models have been proposed to describe Rhbg, including an electroneutral N[H.sup.+.sub.4]/[H.sup.+] exchanger, a uniporter, an N[H.sup.+.sub.4] channel, or even a gas channel. In this study, we characterized the pH sensitivity of Rhbg expressed in Xenopus oocytes. We used two-electrode voltage clamp and ion-selective microelectrodes to measure N[H.sup.+.sub.4]-induced [and methyl ammonium ([MA.sup.+])] currents and changes in intracellular pH ([pH.sub.i]), respectively. In oocytes expressing Rhbg, 5 mM N[H.sub.4]Cl (N[H.sub.3]/N[H.sup.+.sub.4]) at extracellular pH ([pH.sub.o]) of 7.5 induced an inward current, decreased [pH.sub.i], and depolarized the cell. Raising [pH.sub.o] to 8.2 significantly enhanced the N[H.sup.+.sub.4]-induced current and [pH.sub.i] changes, whereas decreasing bath pH to 6.5 inhibited these changes. Lowering [pH.sub.i] (decreased by butyrate) also inhibited the N[H.sup.+.sub.4]-induced current and [pH.sub.i] decrease. In oocytes expressing Rhbg, 5 mM methyl amine hydrochloride (MA/[MA.sup.+]), often used as an N[H.sub.4]Cl substitute, induced an inward current, a [pH.sub.i] increase (not a decrease), and depolarization of the cell. Exposing the oocyte to MA/[MA.sup.+] at alkaline bath pH (8.2) enhanced the [MA.sup.+]-induced current, whereas lowering bath pH to 6.5 inhibited the [MA.sup.+] current completely. Exposing the oocyte to MA/[MA.sup.+] at low [pH.sub.i] abolished the [MA.sup.+]-induced current and depolarization; however, [pH.sub.i] still increased. These data indicate that 1) transport of N[H.sup.+.sub.4] and MA/[MA.sup.+] by Rhbg is pH sensitive; 2) electrogenic N[H.sup.+.sub.4] and [MA.sup.+] transport are stimulated by alkaline [pH.sub.o] but inhibited by acidic [pH.sub.i] or [pH.sub.o]; and 3) electroneutral transport of MA by Rhbg is likely but is less sensitive to pH changes. N[H.sup.+.sub.4]- transport; Rh B glycoprotein; methyl ammonium; ammonium/ hydrogen exchange; ammonium channel; gas channel doi: 10.1152/ajpcell.00211.2010.