Identification and Localization of H+-ATPase, NHE2 and NHE3 in the gills of the southern Appalachian brook trout, Salvelinus fontinalis

Southern brook trout (Salvelinus fontinalis) native to the Great Smoky Mountains are found in headwater streams. These habitats experience increased instances of stream acidification, lowering stream pH on average ~1.0 pH unit. It is well documented that lowering environmental pH results in fish experiencing systemic acidosis. We hypothesize that during acidification events, brook trout suffer acid/base perturbation. Previous studies on other species of freshwater fishes have demonstrated an alteration in the relative expression of H+-ATPase, NHE2 and NHE3 in gill tissue during pH disturbances. Due to a lack of physiological information available, the mechanism utilized by S. fontinalis to regulate systemic pH is unknown. We hypothesized that S. fontinalis will utilize H+-ATPase, NHE2 and NHE3 to excrete excess protons when experiencing episodic stream acidification. To date, we cloned ORFs for H+-ATPase, NHE2, and NHE3 which are homologous to rainbow trout (87, 94 and 92% identical respectively). Immunohistochemical analysis demonstrated H+-ATPase, NHE2 and NHE3 expression is localized to the apical membrane and sub-apical regions of the MRCs in gill epithelia. Also, the relative expression of H+-ATPase decreased along an elevation gradient, whereas NHE3 does not.