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Cellular and Temporal Dissection of KCNQ3 Gain-of-Function Disorder

KCNQ3 功能获得障碍的细胞和颞叶解剖

基本信息

批准号：
10591921
负责人：
TRISTAN T SANDS
金额：
$ 16.45万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-15 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10591921
关键词：
Action Potentials Alleles Arginine Axon Biophysics Cells Cerebral cortex Clinical Cre driver DNA cassette Data Development Disease Dissection Electroencephalography Electrophysiology (science)Enterobacteria phage P1 Cre recombinase Epilepsy Excision Fire - disasters Functional disorder Genes Genetic Genetic Predisposition to Disease Genetic Recombination Goals Grant Human In Vitro Intellectual functioning disability Internal Ribosome Entry Site Investigation Knock-in Knock-in Mouse Knowledge Lead LoxP-flanked allele Mediating Mission Modeling Mus Mutant Strains Mice Mutation Neurodevelopmental Disability Neurons Pathogenicity Pathologic Patient Care Patients Phenotype Physiology Population Potassium Potassium Channel Public Health Recurrence Reporter Reporting Research Seizures Site Sleep Subgroup Symptoms System Tamoxifen Testing Thalamic structure Therapeutic Translating United States National Institutes of Health Variant Voltage-Gated Potassium Channel Work autism spectrum disorder autistic beta Actin biophysical properties cell type critical period design disability disease phenotype excitatory neuron experimental study gain of function gain of function mutation improved inhibitory neuron innovation mouse model mutant neuronal excitability novel prevent programs recombinase targeted treatment tool translational approach

项目摘要

ABSTRACT Intellectual disability and autism are caused – in up to 1 in 1,000 cases – by recurrent de novo missense variants that alter a single arginine residue (R230) in the KCNQ3 gene. Patients with R230 variants have non-verbal intellectual disability, autistic symptoms, and develop near-continuous multifocal spikes on electroencephalography (EEG). KCNQ3 encodes a voltage-gated potassium channel that is active at subthreshold potentials and concentrated at the axon initial segment, where it opposes depolarizing currents that drive neurons to fire action potentials. We recently reported that R230 variants impart gain-of-function (GoF)) biophysical properties to the channel in vitro, but how these alterations lead to disease phenotypes is unknown. Moreover, identification of the genetic etiology has yet to translate into targeted therapy. We have developed a mouse model of the KCNQ3 GoF Disorder with a robust electroclinical phenotypes (frequent spike-wave discharges and lowered electroconvulsive threshold) with relevance to the human condition. We now propose the development of an innovative and versatile conditional version of this model to grant us control over expression of the GoF mutant allele thereby permitting cellular and temporal dissection of the KCNQ3 GoF Disorder. This mouse includes a floxed stop cassette for Cre recombinase mediated activation. In addition, the GoF allele, (with an IRES-eGFP reporter) will be flanked by frt sites to permit Flpo recombinase mediated deactivation. Using this tool to sequentially turn on and then off the GoF mutant channel with spatial and temporal control, we will begin to dissect out (Aim 1. Cellular dissection) which neuronal populations are responsible for the electroclinical phenotypes and (Aim 2. Temporal dissection) whether there are limited temporal windows for preventing or reversing these phenotypes. We will begin these investigations with a pair of experiments that test different aspects of the mouse design. In Aim 1, we will selectively activate the mutant allele in 1) inhibitory neurons using a Gad2-IRES-Cre knock-in driver line, or in 2) excitatory neurons using an Emx-IRES-Cre knock- in driver line, to ask if expression of the GoF allele in either broad neuronal subgroup is sufficient to reproduce the robust electroclinical phenotypes observed in the constitutive GoF mutant mouse. In Aim 2, we will use an inducible Flpo driver line, R26FlpoER, to examine the reversibility of the SWD phenotype. These experiments represent only the introductory forays into the kind of genetic interrogations this conditional model will afford. This genetic work is part of a larger program investigating the KCNQ3 GoF disorder, including extensive electrophysiological investigations of the underlying physiology of impacted circuits and neurons. Pairing these tools will advance our understanding and inform translational approaches to treatment.

摘要