The work proposed will examine the effects of ethanol, in vitro, on gamma-aminobutyric acid (GABA) binding, uptake and release by isolated rat forebrain synaptosomes and synaptic plasma membranes; and the effects of ethanol, in vitro, on calcium release by isolated sarcoplasmic reticulum vesicles. Cholesterol depletion of synaptic plasma membranes, with consequent membrane fluidization, results in decreased binding and uptake of GABA, but not choline. This work will use ethanol, a pharmacological membrane fluidizing agent, to examine the basis of this effect. Binding, uptake and release of GABA by isolated synaptosomes or synaptic plasma membranes will be measured isotopically in the presence and absence of ethanol. Similar measurements will be made of synaptosomal choline uptake and acetylcholine release, to examine the specificity of ethanol effects. A nonspecific lipid transfer protein will be used to load the plasma membranes of isolated synaptosomes with cholesterol to greater-than-normal levels (with consequent membrane rigidification). GABA binding, uptake and release will be measured using these cholesterol-loaded preparations, in the presence and absence of ethanol. These experiments will determine if the effects of cholesterol depletion on GABA binding and uptake are attributable to a biophysical change in the synaptosomal plasma membrane (a decrease in microviscosity) or to specific requirement for cholesterol. They will also determine if membrane rigidification (by means of cholesterol loading) confers a protective effect against ethanol effects. Ethanol produces both acute and chronic muscle disease. Physiological studies have indicated that ethanol, in vitro, alters muscle excitation-contraction coupling. This work will use isolated sarcoplasmic reticulum vesicles to investigate the effects of ethanol, in vitro, on calcium release from the sarcoplasmic reticulum. Vesicles will be loaded with calcium phosphate, and calcium release measured isotopically or spectrophotometrically using a calcium-sensitive dye. Spontaneous calcium release, calcium-induced calcium release, caffeine-induced calcium release and passive vesicular calcium permeability will be determined in the presence and absence of ethanol. These experiments will determine if an ethanol-induced alteration in calcium release from sarcoplasmic reticulum exists which might underlie ethanol-induced muscle dysfunction.
| Mrak, R E (1992) Opposite effects of dimethyl sulfoxide and ethanol on synaptic membrane fluidity. Alcohol 9:513-7 |
| Mrak, R E (1990) Ethanol inhibition of spontaneous calcium release from isolated sarcoplasmic reticulum of alcoholic rats. Neurotoxicology 11:367-73 |
| Mrak, R E; North, P E (1989) Triphasic effects of short chain n-alcohols on synaptic membrane transport of choline and of gamma-aminobutyric acid. Biochim Biophys Acta 984:97-103 |
| North, P E; Mrak, R E (1989) Synaptosomal uptake of choline and of gamma-aminobutyric acid: effects of ethanol and of dimethylsulfoxide. Neurotoxicology 10:569-76 |
| Mrak, R E (1989) Ethanol inhibition of spontaneous calcium release from isolated sarcoplasmic reticulum. Neurotoxicology 10:445-54 |
| Mrak, R E; North, P E (1988) Ethanol inhibition of synaptosomal high-affinity choline uptake. Eur J Pharmacol 151:51-8 |
