Overview
Epilepsy is characterized by recurrent brain seizures and ranks as the most common neurological affliction in the human population. Our studies focus on absence epilepsy, which is characterized by brief losses of consciousness accompanied by distinctive electroencephalographic tracings (EEGs). Several mouse models with this form of spontaneous generalized epilepsy have been characterized and the mutated genes have been identified. Significantly, mutations in the voltage-dependent calcium channel subunits represent four of these mouse models, including tottering, lethargic, ducky and stargazer. We have primarily focused on stargazer and the two related waggler and stargazer3J mutants. These three mouse mutants all have defects in the expression of the gamma2 gene, Cacng2. As we discover more about the proteins that are associated with absence seizures, we can address why mutations lead to disturbances of the neuronal circuits. Greater understanding will not only give us insight into specific disturbances like absence seizures, but potentially yield a better understanding of the basic mechanisms underlying related human neurological disorders, including convulsive epilepsies, migraines and episodic ataxias.
Scientific report
The Stargazer mouse and absence epilepsy
Epilepsy is the most common neurological affliction in the human population. It is caused by recurrent brain seizures, with variable age of onset and severity of disease. Our studies are focused on absence epilepsy, a generalized form of epilepsy that is most frequently diagnosed in children and is characterized by brief losses of consciousness. These episodes are accompanied by distinctive electroencephalographic recordings (EEGs) showing repetitive spike-wave discharges (SWDs).
Several mouse models with this form of spontaneous generalized epilepsy have been characterized and the mutated genes have been identified. Significantly, mutations in the voltage-dependent calcium channel (VDCC) subunits represent four of these models, including tottering, lethargic, ducky, and stargazer. We have primarily focused on the mouse mutant, stargazer, and two related mutants, waggler and stargazer 3 Jackson. All have mutations in the same voltage-dependent calcium channel gamma 2 subunit gene, Cacng2. To understand how mutations in VDCC gamma regulatory subunits contribute to absence seizures, we are exploring the seizure profile in this stargazer allelic series and in engineered targeted disruptions of closely related gamma genes. To further delineate the connection between the loss of calcium channel subunit expression and absence seizures, we are collaborating to study the calcium channel activity in brains from the single and double mutant combinations.
The stargazer mouse is noted for its long and frequently recurring seizure episodes. The three alleles of stargazer show heterogeneous phenotypes; in particular, the mildest allele, stargazer 3 Jackson, has no seizure activity. We have further generated a targeted mutation in the closely related gamma 4 gene, Cacng4. This targeted mutant also has no seizures, but when coupled with the Cacng2 mutants, the double mutants display significant SWD activity. These results indicate that the loss of gamma4 expression can result in an increase in seizure activity but only when gamma2 protein expression is also compromised. Our preliminary results of the double mutant combinations between all three alleles of stargazer, the targeted mutation in Cacng4, and the mutations in lethargic and tottering mutant strains, confirm that the severity of the seizures is influenced by the particular combination of mutated subunits. We are investigating this more thoroughly by testing genetic combinations to assess gene dosage effects on the incidence and duration of SWD episodes. From these results we will be able to determine the minimum threshold of in vivo gamma expression sufficient to suppress seizures, and how this threshold is altered in the presence of other VDCC subunit mutations.
Investigators have determined, using electrophysiological recordings from neurons within the thalamus, that the activity of both the high- and low-voltage calcium channels is significantly increased in stargazer mice. Furthermore, the gamma subunits are also required for the transport of the neurotransmitter AMPA receptors to their active sites at the post-synaptic membrane. We are determining whether VDCC calcium channel and AMPA receptor activity are modified in our double mutant combinations that lack functional gamma proteins. This will allow us to evaluate the pathways linking the gamma subunits with VDCC calcium channel activity and AMPA receptor localization. Ultimately, we are seeking to establish an understanding of how gamma subunits contribute to normal brain function, and why, by deleting their expression, we observe absence seizures in the mouse.
As we discover more about the proteins that are associated with absence seizures, we can address why mutations lead to the perturbations of the neuronal circuits observed as recurrent spike-wave discharge (SWD) bursts. In our studies, we have focused on absence seizures in mice, but mutations in the same calcium channel subunits in humans reveal a greater diversity of phenotypes, including absence and convulsive seizures, episodic ataxia and hemiplegic migraines. Thus, understanding the role of the gamma subunits in the mouse models described here will not only give us insight into absence seizures, but potentially yield a better understanding of the basic mechanisms underlying other human neurological disorders.
Lab staff
Principal Investigator: Verity A. Letts, Ph.D.Research Assistant III: Connie L. Mahaffey, M.S.
Research Administrative Assistant: Heidi Stanton-Drew, B.A.
Publication listings
Letts VA, Kang MG, Mahaffey CL, Beyer B, Tenbrink H, Campbell KP, Frankel WN. 2003. Phenotypic heterogeneity in the stargazin allelic series. Mamm Genome 14:506-513.
Yang Y, Beyer BJ, Otto JF, O'Brien TP, Letts VA, White HS, Frankel WN. 2003. Spontaneous deletion of epilepsy gene orthologs in a mutant mouse with a low electroconvulsive threshold. Hum Mol Genet 12:975-984.
Frankel WN, Beyer B, Maxwell CR, Pretel S, Letts VA, Siegel SJ. 2005. Development of a new genetic model for absence epilepsy: Spike-wave seizures in C3H/He and backcross mice. J Neurosci 25:3452-3458.
Letts VA, Mahaffey CL, Beyer B, Frankel WN. 2005. A targeted mutation in Cacng4 exacerbates spike-wave seizures in stargazer (Cacng2) mice. Proc Natl Acad Sci USA 102:2123-2128.
Letts VA. 2005. A Mouse to Seize! Epilepsy Currents 5:161-165.
Beyer B, Deleuze C, Letts VA, Mahaffey CL, Boumil RM, Lew TA, Huguenard JR, Frankel WN. 2008. Absence seizures in C3H/HeJ and knockout mice caused by mutation of the AMPA receptor subunit Gria4. Hum Mol Genet. 17:1738-1749.
Miki, T. Zwingman, TA, Wakamori, M, Lutz, CM, Cook, SA, Hosford, DA, Herrup, K, Fletcher, CF, Mori, Y, Frankel, WN, Letts, VA. 2008. Two novel alleles of tottering with distinct Ca(v)2.1 calcium channel neuropathologies. Neuroscience 155:31-44.