Central role of the apical membrane H⁺-ATPase in electrogenesis and epithelial transport in Malpighian tubules

The effects of bafilomycin A₁, a blocker of V-type H⁺-ATPases, were investigated in Malpighian tubules of Aedes aegypti. Bafilomycin A₁ reduced rates of transepithelial fluid secretion and the virtual short-circuit current (vI(sc)) with an IC₅₀ of approximately 5 μmol l^-1. As vI(sc) decreased, the electrical resistance increased across the whole epithelium and across the apical membrane, indicating effects on electroconductive pathways. Bafilomycin A₁ had no effect when applied from the tubule lumen, pointing to the relative impermeability of the apical membrane of bafilomycin A₁. Thus, bafilomycin A₁ must take a cytoplasmic route to its blocking site in the proton channel of the H⁺-ATPase located in the apical membrane of principal cells. The inhibitory effects of bafilomycin A₁ were qualitatively similar to those of dinitrophenol in that voltages across the epithelium (V(t)), the basolateral membrane (V(bl)) and the apical membrane (V(a)) depolarized towards zero in parallel. Moreover, V(bl) always tracked V(a), indicating electrical coupling between the two membranes through the shunt. Electrical coupling allows the H⁺-ATPase to energize not only the apical membrane, but also the basolateral membrane. Furthermore, electrical coupling offers a balance between electroconductive entry of cations across the basolateral membrane and extrusion across the apical membrane to support steady-state conditions during transepithelial transport.