CA2-PERFUSED RAT HEPATOCYTES FROM ANOXIC INJURY( ANTAGONISTS DO NOT PROTECT ISOLATED)

Ca2+ antagonists were studied during anoxia in perfused isolated rat h epatocytes. Cytosolic free calcium (Ca(i)2+) was measured with aequori n. Anoxia was induced for 2 h by saturating the perfusate with 95% N2/ 5% CO2. Anoxia increased Ca(i)2+ in two distinct phases reaching a max imum of 1.5 muM. The increase in Ca(i)2+ was caused by Ca2+ influx fro m the extracellular fluids because the main Ca(i)2+ surge was totally abolished in Ca2+-free media. LDH release increased 6-fold during the second hour of anoxia, but when Ca2+ was removed from the perfusate du ring the anoxic period, LDH rose only 2.7-fold. Ca2+ antagonists (10(- 7) to 10(-5) M) did not prevent the increase in Ca(i)2+ and the rise i n LDH release. On the contrary, high concentrations (10(-6) and 10(-5) M) of the blockers nifedipine and diltiazem significantly increased a noxic cell injury. The observation that the increase in LDH and the ri se in Ca(i)2+ were not suppressed by Ca2+ antagonists suggests that (i ) Ca2+ antagonists protect the whole liver from anoxic injury by actin g on cells other than parenchymal cells; (ii) the influx of Ca2+ respo nsible for the massive increase in hepatocyte Ca(i)2+ evoked by anoxia did not take place through voltage-sensitive Ca2+ channels but must h ave occurred via the Na+-Ca2+ antiporter operating in the reverse mode (Ca2+ influx vs. Na+ efflux), and (iii) high concentrations of Ca2+ a ntagonists may be deleterious to the parenchymal cells of the liver.