Vacuolar H+ -ATPases and Voltage-Gated Proton Channels: Two Electrogenic, Proton-Selective Membrane Transport Mechanisms Co-Existed in Osteoclasts

The vacuolar-type H+-ATPase (V-ATPase) is an electrogenic H+ pump that is distributed widely in living organisms. Voltage-gated proton channels (H+ channels) are found in many types of cells including phagocytes. Although their transport mechanisms are distinct, both depend on pH and voltage and share the outcomes, intracellular alkalinization (extracellular acidification) and hyperpolarization. So far, V-ATPases and H+ channels have been studied separately in different cells, using the different approaches under different conditions. V-ATPases and H+ channels are expressed in the plasma membrane of osteoclasts, multinuclear bone-resorbing cells, which allowed us to investigate the two electrogenic H+ transfer mechanisms quantitatively under the same conditions. The current-voltage relationships of the V-ATPases are linear and outward currents were maintained even when the extracellular space was more acidic than the cell inside. Protons flowed through the V-ATPase continuously. In contrast, H+ channels were silent at voltages lower than the threshold. However, once it opened, the H+ channel current surpassed the V-ATPase current. Consequently, the profile of H+ efflux was composed of two phases, one dominated by the V-ATPase and the other, by the H+ channel. Inhibition of V-ATPases by bafilomycin was often accompanied by decreases in the H+ channel current. These results suggested that the V-ATPase and the H+ channel work over different ranges of pH- and voltage-gradients across the membrane in single cells and co-regulate the pH environments of osteoclasts.