Graduate Thesis Or Dissertation

 

A Mutation in Cav2.1 Linked to a Severe Neurodevelopmental Disorder Impairs Channel Gating Public Deposited

https://scholar.colorado.edu/concern/graduate_thesis_or_dissertations/9c67wn12v
Abstract
  • Ca2+ flux via voltage-gated Ca2+ channels is essential to the regulation of membrane excitability, neurotransmission, and a variety of intracellular signaling processes. Cav2.1 is the predominant Ca2+ channel present in the presynaptic terminals of both neuromuscular junctions and many central synapses. Point mutations in Cav2.1 is the predominant can drastically alter channel gating and expression, and indeed have been linked to two paroxysmal disorders – Episodic Ataxia Type 2, and Familial Hemiplegic Migraine Type 1. With the novel whole-exome sequencing technique, mutations linked to a new class of more severe disorders have been found with phenotypes like Episodic Ataxia Type 2 with an additional developmental component. One of these mutations is an arginine to proline substitution in the S4 voltage-sensing region of the fourth membrane-bound Repeat of Cav2.1 (R1673P). This mutation was proposed to cause a gain-of-function in Cav2.1 based on the ability of the mutant channel to rescue the photoreceptor response in Cav2.1-deficient Drosophila cacophony larvae. Here, I show that the R1673P mutation actually results in a profound loss-of-channel-function. Voltage-clamp analysis of tsA-201 cells expressing the mutant channel revealed a ~25 mV depolarizing shift in the voltage-dependence of activation coupled with delayed activation kinetics. These alterations in activation implies that a significant fraction of Cav2.1 channels resident in presynaptic terminals are unlikely to open in response to an action potential, thereby increasing the probability of synaptic failure at both NMJs and central synapses. Indeed, the mutant channel supported only minimal Ca2+ flux in response to an action potential-like waveform. Application of the Cav2.1 agonist GV-58 shifted mutant activation to more hyperpolarizing potentials and slowed deactivation. Consequently, GV-58 was able to rescue some Ca2+ flux in response to an action potential-like stimulus. My thesis suggests that therapeutic agents like GV-58 that increase channel open probability may be effective in combatting this and other severe neurodevelopmental disorders caused by loss-of-function mutations in Cav2.1.

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  • 2019
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  • 2020-01-27
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