Inhibition of bioenergetics alters intracellular calcium, membrane composition, and fluidity in a neuronal cell line.
The effect of inhibited bioenergetics and ATP depletion on membrane composition and fluidity was examined in cultured neuroblastoma-glioma hybrid NG108-15 cells. Sodium cyanide (CN) and 2-deoxyglucose (2-DG) were used to block, oxidative phosphorylation and anaerobic glycolysis, respectively. Endoplasmic reticulum (ER) Ca(2+)-pump activity measured by 45Ca2+ uptake was > 92% inhibited in intact cells incubated with CN (1 mM) and 2-DG (20 mM) for 30 min. In addition, exposure of cells to CN and 2-DG caused a 134% increased release of isotopically labeled arachidonic acid (3H-AA) or arachidonate-derived metabolites from membranes. Removal of Ca2+ from the incubation medium ablated the CN/2-DG induced release of 3H-AA or its metabolites. Membrane fluidity of intact cells was measured by electron spin resonance spectroscopy using the spin label 12-doxyl stearic acid. The mean rotational correlation time (tau c) of the spin label increased 49% in CN/2-DG exposed cells compared to controls, indicating a decrease in membrane fluidity. These results show that depletion of cellular ATP results in inhibition of the ER Ca(2+)-pump, loss of AA from membranes, and decreased membrane fluidity. We propose that impaired bioenergetics can increase intracellular Ca2+ as a result of Ca(2+)-pump inhibition and thereby activate Ca(2+)-dependent phospholipases causing membrane effects. Since neurons derive energy predominantly from oxidative metabolism, ATP depletion during brain hypoxia may initiate a similar cytotoxic mechanism.
Ray
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P
Department of Biology, Walter Reed Army Institute of Research, Washington, DC 20307-5100.
Ray
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R
Broomfield
C A
CA
Berman
J D
JD
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D003847:Deoxyglucose / Q000494:pharmacology*; D004734:Energy Metabolism / Q000187:drug effects*; D005910:Glioma; D006019:Glycolysis / Q000187:drug effects; D006822:Hybrid Cells; D007424:Intracellular Fluid / Q000378:metabolism; D007700:Kinetics; D008560:Membrane Fluidity / Q000187:drug effects / Q000502:physiology*; D009447:Neuroblastoma; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010085:Oxidative Phosphorylation / Q000187:drug effects; D012966:Sodium Cyanide / Q000494:pharmacology*
D000255:Adenosine Triphosphate; D003847:Deoxyglucose; D012966:Sodium Cyanide; D002118:Calcium