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Acute High Glucose Exposure Impairs Synaptosomal Vesicle Release Activity
Alshakhshir, Nadine Hani Sa'da
Alshakhshir, Nadine Hani Sa'da
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Abstract
An increasing volume of evidence has demonstrated a clear association between Alzheimer’s disease (AD) and diabetes. AD brains have recently been shown to exhibit hyperglycemia, a phenotype shared with diabetic brains that can possibly serve as a mechanistic pathological link between AD and diabetes. The vacuolar-type ATPase (V-ATPase) is a proton pump widely distributed in eukaryotic cells that serves many essential functions. Reversible disassembly of V-ATPase is a mechanism by which cells regulate V-ATPase function where disassembled V-ATPase is inactive, however, this process is reversible and reassembly restores the activity of V-ATPase. There is available evidence demonstrating an association of brain lysosomal V-ATPase dysregulation with AD. At synapses, V-ATPase is present on the membranes of synaptic vesicles and is essential for neurotransmitter concentration in synaptic vesicles and subsequent neuronal transmission, however, its role in AD pathology is unclear.This thesis was carried out to investigate whether acute hyperglycemia could modify neuronal synaptic V-ATPase function and synaptic vesicular activity in a model of functional synaptosomes freshly isolated from wild type mice brains. The release of the fluorescent dye Acridine Orange and the neurotransmitter glutamate from synaptosomes were used to study the effect of hyperglycemia on synaptic vesicular activity. Neuronal vesicular V-ATPase activity under hyperglycemia was tested through measuring V-ATPase assembly using Co-Immunoprecipitation (Co-IP) and specific V-ATPase released phosphates. The results demonstrate that hyperglycemia reduces synaptic vesicular activity as indicated by attenuated Acridine Orange and glutamate release. Results from V-ATPase activity experiments indicate that hyperglycemia reduces V-ATPase assembly but not phosphate release. Overall, these data suggest that hyperglycemia can potentially impair neuronal synaptic vesicular exocytosis in part by reduced V- iv ATPase assembly, contributing to the increased risk of cognitive decline associated with AD and diabetes. A pitfall of this study is that the steps for the V-ATPase phosphate release experiment did not allow for the total sample incubation time under hyperglycemia to be consistent with the other experiments. This might have contributed to the insignificant results obtained for the V-ATPase activity assay. The work presented in this thesis studied the impact of acute hyperglycemic exposure on synaptosomes from wild type brains. Further studies are needed to investigate the effects of chronic hyperglycemia on synaptic vesicular activity and V-ATPase function, for example, through comparisons between wild type and diabetic animals.
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2021-05-31
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University of Kansas
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Pharmacology, Alzheimer's, brain hyperglycemia, diabetes, synaptic impairment, V-ATPase function, V-ATPase reversible disassembly