Behavioral Phenotype Analysis
Objective: Characterization of ataxia, dyskinesia, and absence epilepsy phenotypes in adult purky mice with delayed postnatal loss of P/Q-type calcium channels
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Materials1
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Protocol Steps
Genetic construct creation
Created a novel conditional Cacna1a knock-in mouse by inserting the floxed green fluorescent protein derivative Citrine into the first exon of Cacna1a
Note: This is a genetic engineering step to create the conditional knockout mouse line
View evidence from paper
“created a novel conditional Cacna1a knock-in mouse by inserting the floxed green fluorescent protein derivative Citrine into the first exon of Cacna1a”
Breeding for Cre-mediated deletion
Crossed the conditional Cacna1a knock-in mouse with a postnatally expressing PCP2-Cre line to achieve delayed Purkinje cell gene deletion within the cerebellum and sparsely in forebrain
Note: PCP2-Cre line provides postnatal, cell-type specific expression in Purkinje cells
View evidence from paper
“crossed it with a postnatally expressing PCP2-Cre line for delayed Purkinje cell (PC) gene deletion within the cerebellum and sparsely in forebrain”
Phenotypic characterization in adult mice
Examined adult purky mice for neurological deficits including ataxia, dyskinesia, and absence epilepsy phenotypes
Note: Adult purky mice exhibited the full spectrum of neurological deficits seen in mice with genomic Cacna1a ablation
View evidence from paper
“adult purky mice exhibited the full spectrum of neurological deficits seen in mice with genomic Cacna1a ablation”
Cellular analysis of Purkinje cells
Analyzed Purkinje cells in purky mice for lack of P/Q-type calcium channel protein and currents, altered spontaneous firing, and impaired neurotransmission
Note: PCs in purky mice lacked P/Q-type calcium channel protein and currents within the first month after birth
View evidence from paper
“PCs in purky mice lacked P/Q-type calcium channel protein and currents within the first month after birth, displayed altered spontaneous firing, and showed impaired neurotransmission”