Apical ouabain-sensitive and ouabain-insensitive ATPases in rat colonic epithelium

Active resorption of potassium ions in the colon is mediated by ouabain-sensitive and ouabain-insensitive H,K-ATPases localized in the apical membrane of colonic enterocytes (colonocytes). The present study was performed to investigate distribution patterns of apical ATPases using catalytic histochemistry and immunohistochemistry. Activity of the ATPases was localized both at the apical and basolateral regions of these cells. The basolateral activity was almost completely inhibited by ouabain, but apical ATPase activity was only partially inhibited by ouabain. Ultracytochemically, activity was localized at the cytoplasmic side of the apical and basolateral membrane and thus we assume that the activity represents isoforms of apical H,K-ATPases and basolateral Na,K-ATPase. With the use of a polyclonal antibody raised against H,K-ATPase alpha-subunit, we demonstrated immunostaining only in the apical region of colonic enterocytes whereas positive staining was not observed at the basolateral membrane. On the other hand, when antibodies against alpha1-subunit Na,K-ATPase were applied, immunostaining was localized only in the basolateral membrane domain. Therefore, we conclude that ATPases as demonstrated histochemically in the present study were identified immunohistochemically as colonic alpha-subunit H,K-ATPase (in the apical cell membrane of colonocytes along the entire length of crypts) and as alpha1-subunit Na,K-ATPase (in the basolateral membrane).