Perturbation of cellular energy state in complete ischemia: Relationship to dissipative ion fluxes

Publisher Website
Google Scholar
Abstract
Loss of cellular ion homeostasis during anoxia, with rapid downhill fluxes of K+, Ca2+, Na+ and Cl-, is preceded by a slow rise in extracellular K+ concentration (K e + ), probably reflecting early activation of a K+ conductance. It has been proposed that this conductance is activated by either a rise in intracellular calcium concentration (Ca i 2+ ), or by a fall in ATP concentration. In a previous study from this laboratory (Folbergrová et al. 1990) we explored whether the early activation of a K+ conductance could be triggered by a rise in Ca i 2+ . To that end, labile metabolites and phosphorylase a, a calcium sensitive enzyme, were measured after 15, 30, 60 and 120 s of complete ischemia (“anoxia”). In the present study, we investigated whether brief anoxia is accompanied by changes in ATP/ADP ratio, or in the phosphate potential, which could cause activation of a K+ conductance. To provide information on this issue, we added a group with 45 s of anoxia to the previously reported groups, and derived changes in intracellular pH (pHi). This allowed calculations of the free concentrations of ADP (ADPf) and AMP (AMPf) from the creatine kinase and adenylate kinase equilibria, and hence the derivation of ATP/ADPf ratios. In performing these calculations we initially assumed that the free intracellular Mg2+ concentration remained unchanged at 1 mM. However we also explored how a change in Mg i 2+ of the type described by Brooks and Bachelard (1989) influenced the calculation. The results showed that ADPf must have risen to 150–200% of control within 15 s, and to 330–350% of control within 45 s of anoxia. The concentration of AMPf should have increased 2–4 fold in 15 s and 10–20 fold in 45 s. Thus although tissue ATP concentration usually remains >90% of control within the first 30s of anoxia, and >80% of control within the first 45 s, the ATP/ADPf ratios change markedly at a time when alterations in ion homeostasis are dominated by a moderate rise in K e + , and long before massive ion fluxes occur and the cells depolarise (after about 60–70 s). Such early changes in ATP/ADPf ratio, or in phosphate potential, could well influence reactions which are coupled to ATP hydrolysis, and perhaps lead to activation of ATP-dependent K+ conductances.