Blood acid/base regulation by specialized epithelia such as gills and kidney requires the ability to sense blood acid/base status. Here, we developed primary cultures of ray (Urolophus halleri) gill cells to study mechanisms for acid/base sensing without the interference of whole-animal hormonal regulation. Ray gills had abundant base-secreting cells, identified by being rich in V-H⁺-ATPase (VHA), and also expressed the evolutionarily conserved acid/base sensor soluble adenylyl cyclase (sAC). Exposing cultured cells to extracellular alkalosis (pH 8.0; 40 mM HCO₃^-) triggered VHA translocation to the cell membrane, similar to previous reports in live animals experiencing blood alkalosis. VHA translocation was dependent on sAC, as it was blocked by the sAC-specific inhibitor KH7. Ray gill base-secreting cells also express transmembrane adenylyl cyclases (tmACs); however, tmAC inhibition by 2'5'-dideoxyadenosine did not prevent alkalosis dependent VHA translocation, and tmAC activation by forskolin reduced the abundance of VHA at the cell membrane. This study demonstrates that sAC is a necessary and sufficient sensor of extracellular alkalosis in ray gill base-secreting cells. In addition, this study indicates that different sources of cAMP differentially modulate cell biology.