Acid incubation causes exocytic insertion of NHE3 in OKP cells

Incubation of opossum kidney clone P (OKP) cells in acid media (pH 6.8) cau ses activation of Na+/H+ exchanger 3 (NHE3) at 6, 12, and 24 h. OKP cell NH E3 protein abundance was increased by 45% at 24 h of acid incubation but wa s unaffected at 3-12 h. By contrast, apical membrane NHE3, measured by surf ace biotinylation, increased approximately twofold at 6, 12, and 24 h, mirr oring the increase in activity. Acid incubation caused a 76% increase in ex ocytic insertion of NHE3 into the apical membrane but had no effect on endo cytic internalization at 6 h. Latrunculin B, an inhibitor of microfilament organization, inhibited the acid-induced increases in apical membrane NHE3, exocytic insertion of NHE3, and NHE3 activity at 6 h. These studies demons trate two mechanisms for acid-induced increases in NHE3 activity. Beginning at 6 h, there is an increase in apical membrane NHE3 that is due to stimul ated exocytic insertion and is required for increased NHE3 activity. At 24 h, there is an additional increase in total cellular NHE3.