Abstract Title

Investigating Cys-loop Cysteine Residues in α3β2 Nicotinic Acetylcholine Receptors

Disciplines

Biology

Abstract

Parkinson’s disease is caused by insufficient release of dopamine from the neurons in the substantia nigra. The release of dopamine is stimulated by the activation of nicotinic acetylcholine receptors (nAchR). The α3β2 isoform of this receptor was the focus of this experiment. We hypothesize that cysteine residues in the cys-loop of the α subunit play an inhibitory role in the binding of acetylcholine. The aim of this study was to characterize the function of the cysteine residues by utilizing site-directed mutagenesis. The native cysteine residues in the α subunit were mutated to alanine and homologous aspartate residues in the β subunit were mutated to cysteine. The mutant mRNA was synthesized and expressed in Xenopus laevis oocytes. Two-electrode voltage clamping was used to compare the efficacy of these mutations. Applications of acetylcholine led to an influx of extracellular sodium and the ionic current was recorded. A comparison of the dose response curves to the wild type will provide insight into the binding affinity of acetylcholine, which is a potential drug target for the treatment of neurodegenerative disorders.

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Investigating Cys-loop Cysteine Residues in α3β2 Nicotinic Acetylcholine Receptors

Parkinson’s disease is caused by insufficient release of dopamine from the neurons in the substantia nigra. The release of dopamine is stimulated by the activation of nicotinic acetylcholine receptors (nAchR). The α3β2 isoform of this receptor was the focus of this experiment. We hypothesize that cysteine residues in the cys-loop of the α subunit play an inhibitory role in the binding of acetylcholine. The aim of this study was to characterize the function of the cysteine residues by utilizing site-directed mutagenesis. The native cysteine residues in the α subunit were mutated to alanine and homologous aspartate residues in the β subunit were mutated to cysteine. The mutant mRNA was synthesized and expressed in Xenopus laevis oocytes. Two-electrode voltage clamping was used to compare the efficacy of these mutations. Applications of acetylcholine led to an influx of extracellular sodium and the ionic current was recorded. A comparison of the dose response curves to the wild type will provide insight into the binding affinity of acetylcholine, which is a potential drug target for the treatment of neurodegenerative disorders.