Regulation by PKC isoforms of Na+/H+exchanger in luminal membrane vesicles isolated from cortical tubules

Abstract

The present study was designed to determine the Na/H exchanger isoforms present in luminal membrane vesicles (LMV) isolated from rat kidney cortical tubule suspensions, as well as the effects of acute phorbol ester (phorbol myristate acetate, PMA) and angiotensin II (ANG II) pretreatment of suspensions on NHE activity and protein kinase C (PKC) isoform abundance. In LMV, both NHE3 and NHE2 proteins were found by Western blot analysis, but only ethylisopropylamiloride-sensitive and almost completely Hoe-694-resistant Na/H exchange activity was observed from22Na uptake and thus attributed to NHE3. PMA pretreatment increased Na/H exchange activity and PKC isoforms α, δ, and ε abundance in LMV, and these effects were prevented by PKC inhibition. Low-dose ANG II (10−11 M) pretreatment increased Na/H exchange activity and only PKC-ζ abundance in LMV, and these effects were also prevented by PKC inhibition. After high-dose ANG II (10−7 M), Na/H exchange activity was decreased in LMV. PKC inhibition did not prevent this effect. In conclusion, the stimulating effects of PMA and low-dose ANG II are explained by the translocation of different isoforms of PKC in LMV, whereas the inhibitory effect of high-dose ANG II is not PKC dependent.