Activation of Na+ /H+ exchange on rat preadipocyte plasma membrane and its role in cell proliferation and differentiation

An in vitro cultured rat perirenal preadipocyte (PA) was established as a model system to investigate the role of the intracellular pH (pHi) and of the Na /H exchanger during PA proliferation and differentiation. pH sensitive probe, 2' ,7'-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein(BCECF) , was employed to measure the pHi of PA and to determine the Na /H exchange activity. The results showed that there was Na /H exchange activity in the plasma membrane of PA, FCS stimulated DNA synthesis measured by H-TdR incorporation, and the activation of Na /H exchanger resulted in pHi increase (nearly 0.2 pH unit) within 2 min. Ethyl-isopropyl-amiloride (EIPA) , a specific Na /H exchange inhibitor, inhibited Na /H exchange activity and DNA synthesis. In the absence of serum insulin did not stimulate DNA synthesis but did induce PA differentiation characterized by the appearance of adiposome in the cell and the enhancement of glycerol-3-phosphate dehydrogenase (G3 PDHase) activity. Meantime, insulin was also found to stimulate Na /H exchange activity and pHi increase. EIPA inhibited Na /H exchanger activation induced by insulin and also partially inhibited the enhancement of G3PDHase activity. These results demonstrated that the activation of Na /H exchange and the resulting pHi increase are the early events related to both proliferation and differentiation of PA.