Date of Award
1994
Degree Type
Thesis
Degree Name
Master of Science in Biochemistry
Department
Biochemistry, Microbiology, and Molecular Genetics
First Advisor
Dennis E. Rhoads
Abstract
The present study is composed of two relatively independent analyses of ethanol and fatty acids as potential effectors of neuronal Ca²⁺ homeostasis. Never ending preparations (synaptosomes) from rat brains were used to examine the effects of ethanol and fatty acids on intrasynaptosomal free Ca²⁺ ([Ca²⁺]i). [Ca²⁺]i was measured using Fura2, a fluorescent Ca²⁺ indicator dye.
Ca²⁺has been proposed as a possible mediator of some of the effects of ethanol on the mammalian brain. The present study evaluates the action of ethanol on [Ca²⁺]i and the relationship between the change in [Ca²⁺]i caused by ethanol and the activation of a calnaktin-like inhibitor of a Na⁺, K⁺-ATPase. A small increase in synaptosomal [Ca²⁺]i was observed in the presence of ethanol. However, the concentrations of ethanol required for significant effects on [Ca²⁺]i were not relevant to alcohol intoxication and greater effects on [Ca²⁺]i would be needed to increase calnaktin activity. Therefore, these results suggest pharmacologically relevant concentrations of ethanol (<200mM) do not elevate [Ca²⁺]i to levels necessary for the inhibition of Na⁺, K⁺-ATPase.
Two mechanisms have been proposed for the inhibition of neurotransmitter amino acid uptake by unsaturated fatty acids: the uncoupling of Na⁺/amino acid symport and the depolarization of the plasma membrane. The preset study demonstrates a potent decrease in [Ca²⁺]i by two unsaturated fatty acids, but not by a saturated fatty acid. This result is not consistent with the depolarization mechanism and supports the proposed uncoupling mechanism. The target site for the decease in [Ca²⁺]i by unsaturated fatty acids appears to be distinct from the site for inhibition of amino acid uptake. Protein kinase C (PKC) does not seem to be required since a PKC inhibitor , H-7, did not alter the decrease in [Ca²⁺]i caused by unsaturated fatty acids. Unsaturated fatty acids also increased Ca²⁺ uptake into microsomes. The increased microsomal Ca²⁺ uptake was observed even in the presence of the Ca²⁺-ATPase inhibitor, orthovanadate, but was partially inhibited in the presence of a physiological concentration of ATP. The reduction in synaptosomal [Ca²⁺]i by unsaturated fatty acids is most likely due to enhanced mechanisms for extrustions of Ca²⁺ across the plasma membrane.
Recommended Citation
Adachi, Megumi, "The Effects of Ethanol and Fatty Acids on Intrasynptosomal Calcium Concentration" (1994). Open Access Master's Theses. Paper 821.
https://digitalcommons.uri.edu/theses/821
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