Transforming growth factor beta 1 (TGFβ1) is involved in promoting cell death by apoptosis in the liver, whereas the activation of Na+/H+ exchanger has been related to cell proliferation. The aim of this study was to gain information on the effects of TGFβ1 on intracellular pH and Na+H+ exchange activity in isolated periportal (PP) and perivenular (PV) rat hepatocytes using the pH-sensitive dye BCECF in a perfused subconfluent hepatocyte monolayer. Steady-state intracellular pH (pHi) in a bicarbonate-free solution (HEPES) were 7.17 ± 0.031 in PP and 7.15 ± 0.041 in PV cells. Treatment with TGFβ1 (120 pmol/L) for 7 hours increased the number of apoptotic bodies by 25% and 38%, and decreased steady-state pHi to 7.11 ± 0.018 (P = .05) and to 7.07 ± 0.021 (P ± .02), respectively, in PP and PV hepatocytes. In HEPES, cells recovered from an acid load, extruding protons at a rate (JH) of 4.85 ± 1.01 mmol/L/min in PP cells and of 4.91 ± 0.99 mmol/L/min in PV hepatocytes. This recovery appeared amiloride inhibitable (1 mmol/L). Culture with TGFβ1 for 7 hours induced (in HEPES) a decrease of pHi recovery rate from an acid load more in PV (by 46%) than in PP hepatocytes (by 35%, P ± .05). Acute administration of epidermal growth factor (EGF) (10 to 100 ng/mL) induced an increase in Na+/H+ exchange activity by 32% and 27%, respectively, in PP and PV cells compared with controls. In contrast, in cells cultured for 7 hours with 120 pmol/L TGFβ1, the acute administration of EGF slightly increased Na+/H+ exchange activity (by 18%, P ± .05) only in PP cells. This study demonstrates that pHi and Na+/H+ exchange activity are decreased by TGFβ1, which increases the number of apoptotic bodies in periportal and perivenular rat hepatocyte primary cultures. (Hepatology 1995; 22:1488–1498).