Abstracts

RATIONALE: The stargazer and waggler mice have mutations in the voltage-dependent calcium channel gamma subunit gene, Cacng2. Noticeably these mice have absence seizures throughout their lives. There are many other related gamma subunits genes, several of which are predominantly expressed in the brain. If these genes are disrupted, do they also cause absence epilepsy in mice? To pursue this question we have made a targeted disruption of the Cacng4 gene.
METHODS: The Cacng4 gene was disrupted by replacing exon 4 encoding the carboxy terminus of the protein with a DNA cassette including the lacZ gene. This disruption was introduced into mice using standard knockout technology and homozygous mice for the targeted disruption were studied. EEGs were measured from bipolar electrodes implanted into each cortical hemisphere. Recordings were taken daily over a three hour interval.
RESULTS: The homozygous mice carrying both targeted alleles of Cacng4 were overtly normal. EEG recordings from these mice reveal that they have no significant absence seizure activity. We have now set up crosses between these mice and wagglers to introduce the Cacng4 targeted disruption onto the Cacng2 mutant background. On measuring the absence seizure activity in the double homozygotes, we noticed that the activity was markedly enhanced in the double mutant compared to the waggler mutation alone. We have further crossed the gamma4 targeted mutant to other mutants associated with voltage-dependent calcium channel mutations, including stargazer-3J (a new allele of stargazer), lethargic and tottering mouse mutants. The same seizure phenotype was also observed in stargazer-3J gamma4 double homozygotes although the stargazer 3J mutant itself shows no absence seizure activity
CONCLUSIONS: Both waggler and stargazer-3J have residual Cacng2 expression and neither shows the marked absence seizure activity observed in the original stargazer mutant. By introducing the mutation of the Cacng4 allele onto these backgrounds, the absence phentoype becomes more pronounced. These results indicate that these two closely-related gamma subunits both have a role in absence seizure activity. The expression pattern of these two molecules shows some overlap, for instance in the granule cell layer of the cerebellum. However Cacng2 is predominantly expressed in dorsal regions of the brain, including the cortex, hippocampus and cerebellum, whereas Cacng4 shows higher expression levels in more ventral structures, the habenulae and caudate putamen. Thus it is probable that these results are not due to an interaction between the Cacng2 and 4 molecules but rather, that an overall depletion of these two gamma subunits confers a more severe absence phenotype.
[Supported by: NIH NS 32801.]