The grants which preceded this application have resulted in significant contributions to understanding the cellular basis and interrelation between adaptive and pathological changes in sensitivity. the long-term goals of the laboratory have been, and continue to be, the identification of the cellular and molecular mechanisms responsible for altered sensitivity and their biological significance. The proposed work is directly in line with those goals and contains three specific aims. (1) Determine if the decrease in Na+, K+ pump sites associated with adaptive supersensitivity of the guinea-pig vas deferens is the result of a reduction in the concentrations of subunits of Na+, K+ ATPase and/or in the in the appropriate mRNAs. (2) Investigate the cause of the selective supersensitivity of the mesenteric vasculature of Dahl salt-sensitive and spontaneously hypertensive rats to norepinephrine at the level of alpha1 adrenoceptors, the inositol phosphate system and appropriate mRNAs. (3) Extend to the cerebral cortex and hippocampus the investigation of selective subsensitivity of cerebellar neurons of genetically epilepsy prone rats to GABA. The cellular and molecular basis of the subsensitivity will be investigated at the level of GABAA receptors, subunits of the GABA receptor and chloride channels. Methods to be used include intra- and extracellular electrical recording, single ion channel patch clamping, radioligand receptor binding, quantification of inositol phosphates by column chromatography and ligand scintillation spectrometry, quantification of specific proteins by Western analysis and of corresponding messenger RNAs by Northern analysis. As appropriate, G protein analysis will continue to be done in collaboration with our consultants, Drs. Torphy and Stadel. The results will increase existing knowledge of an adaptive characteristic common to many excitable cells and, in particular, will expand the understanding of the pathological basis of genetic forms of epilepsy and hypertension. Experiments are designed to progress from the functional level of nerve and muscle activity to the cellular transduction processes and hence to the level of specific proteins and their regulation. Each step will be determined by the outcome of experiments at the preceding level.
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