Cytosolic free magnesium, ATP and blebbing during chemical hypoxia in cultured rat hepatocytes

Abstract

Cytosolic free Mg2+ concentration was determined in 1-day cultured rat hepatocytes using Multiparameter Digitized Video Microscopy (MDVM) of the fluorescent probe, mag-fura-2. Chemical hypoxia with KCN (5 mM) and iodoacetate (1 mM), a model which mimics the ATP depletion and reductive stress of hypoxia, caused a rapid increase of free Mg2+ from 1.1 +/- 0.2 to 1.6 +/- 0.2 mM within 4 min. Concurrently, numerous small plasma membrane blebs formed and ATP levels dropped from 13.24 to 1.32 nmol/10(6) cells. Removal of KCN and iodoacetate resulted in recovery of ATP to 60-70% of pre-exposure levels, a concomitant decrease in cytosolic free Mg2+ back toward basal levels, and reversal of blebbing (bleb resorption). These results indicate that changes of cytosolic free Mg2+ inversely reflect changes of ATP in a model of hypoxia and reoxygenation. Bleb formation and resorption were dependent on the fall and rise of ATP.