The Temperature Dependence of Respiration and ATPase in Rat Liver Mitochondria is Altered by Ethanol

Membranes and phospholipids of liver mitochondria from chronic alcoholic rats are resistant to membrane disordering by alcohol.

“…Two-dimensional thin-layer chromatography of lipid fractions and lipids reextracted from liposomes after ethanol treatment did not reveal hydrolysis products. The in the presence and in the absence ofethanol, were significantly higher for membranes and phospholipid vesicles derived from ethanol-treated animals (not shown), a finding that presumably reflects a more rigid membrane structure for the alcoholic rats (5). Similar experiments with 12-doxylstearic acid gave almost identical results, except that the calculated order parameter was considerably lower with this probe, a finding that indicates increased acyl-chain disorder in the membrane's hydrophobic core.…”

“…Although these changes are commonly ascribed to a damaging effect of chronic alcoholism on hepatic metabolism, we have recently suggested (5) that they might reflect an adaptation of the mitochondrial membranes to the presence of ethanol. In mitochondria from rats treated chronically with ethanol, respiration and ATPase activities are lower and more resistant to the stimulating effect of ethanol at physiological temperature (5). This latter effect is probably the result of the "fluidizing" effect ofethanol on phospholipid membranes (6).…”

“…The resistance to the effect of ethanol in mitochondria from chronically intoxicated animals might result from a change in the membrane structure, as suggested by the shifts of the breaks in Arrhenius plots of both respiration and ATPase to higher temperatures, and by the lowering of the energy of activation of these membrane enzymes. We, therefore, sought to detect such structural changes by direct physical measurements in intact membranes and in their isolated lipids to discover whether the membranes become more "rigid" or more resistant to the fluidizing effect ofethanol, as suggested by our previous study (5). We employed the nitroxide spin labels 5-doxyl-and 12-doxylstearic acid as probes ofthe membrane lipid environment (7).…”

“…Liver mitochondria were isolated as described (5). Mitochondrial membrane lipids were extracted according to Keith et al (10).…”

Kinetics and control of oxidative phosphorylation in rat liver mitochondria after chronic ethanol feeding

“…Fractured mitochondria were made by rapidly freeze-thawing liver mitochondria (repeated three times). ATPase activity was measured at 30 mC by recording the pH changes in the incubation chamber during ATP hydrolysis reaction in the presence of 250 mM sucrose, 5 mM MgCl # , 2.5 mM Tris (pH 8) and 1 mM ATP [21]. The buffering capacity of the reaction mixture was determined experimentally by adding a known amount of HCl (5 µl of 50 mM HCl 250 nmol ions H + ) to the reaction mixture after each measurement, and recording the pH change induced by this addition.…”