The aims of this study were to determine 1) the effects of anoxia and reoxygenation on electrolytes and ATP content of isolated hepatocytes and 2) whether exogenous ATP-MgCl2 has any beneficial effects on the cellular alterations that are produced during the reoxygenation period. After 90 min anoxia, intracellular Na and inorganic phosphate (P(i)) increased, K and ATP decreased, whereas Ca and Mg did not change. After 60 min of reoxygenation, intracellular Mg and Na levels decreased, Ca increased, and P(i) levels returned to normal, but ATP levels remained low. These results suggest that the plasma membrane is relatively impermeable to divalent ions such as Ca2+, Mg2+, and HPO42- during anoxia but becomes permeable to them after reoxygenation. When anoxic cells were treated with ATP-MgCl2 during reoxygenation, intracellular ATP and Mg levels increased, but accumulation of Ca and P(i) was also observed. Thus enhancement of Mg transport and specific stimulation of Ca-P(i) sequestration occurs with ATP-MgCl2 treatment. Na and K levels exhibited biphasic reciprocal dose-response changes to ATP-MgCl2 treatment; i.e., 0.1 mM ATP-MgCl2 (low-dose) treatment increased K and decreased Na, whereas 1 mM ATP-MgCl2 (high-dose) treatment decreased K and increased Na. These results indicate that although addition of a single high-dose ATP-MgCl2 was deleterious over a period of 60 min, low-dose ATP-MgCl2 accelerates the recovery of postanoxic cellular electrolyte homeostasis.