A substantial number of mutations have been identified in voltage-gated sodium channel genes that result in various forms of human epilepsy. SCN1A mutations result in a spectrum of severity ranging from mild febrile seizures to Dravet syndrome, an infant-onset epileptic encephalopathy. Dravet syndrome patients experience multiple seizures types that are often refractory to treatment, developmental delays, and elevated risk for SUDEP. The same sodium channel mutation can produce epilepsy phenotypes of varying clinical severity. This suggests that other factors, including genetic, modify the primary mutation and change disease severity. Mouse models provide a useful tool in studying the genetic basis of epilepsy. The mouse strain background can alter phenotype severity, supporting a contribution of genetic modifiers in epilepsy. The Scn1a+/- mouse model has a strain-dependent epilepsy phenotype. Scn1a+/- mice on the 129S6/SvEvTac (129) strain have a normal phenotype and lifespan, while [129xC57BL/6J]F1-Scn1a+/- mice experience spontaneous seizures, hyperthermia-induced seizures and high rates of premature death. We hypothesize the phenotypic differences are due to strain-specific genetic modifiers that influence expressivity of the Scn1a+/- phenotype. Low resolution mapping of Scn1a+/- identified several Dravet syndrome modifier (Dsm) loci responsible for the strain-dependent difference in survival. One locus of interest, Dsm1 located on chromosome 5, was fine mapped to a 9 Mb region using interval specific congenics. RNA-Seq was then utilized to identify candidate modifier genes within this narrowed region. Three genes with significant total gene expression differences between 129S6/SvEvTac and [129xC57BL/6J]F1 were identified, including the GABAA receptor subunit, Gabra2. Further analysis of Gabra2 demonstrated allele-specific expression. Pharmological manipulation by clobazam, a common anticonvulsant with preferential affinity for the GABRA2 receptor, revealed dose-dependent protection against hyperthermia-induced seizures in Scn1a+/- mice. These findings support Gabra2 as a genetic modifier of the Scn1a+/- mouse model of Dravet syndrome.

The α(7)β(1)-integrin is an adhesion molecule highly expressed in skeletal muscle that can enhance regeneration in response to eccentric exercise. We have demonstrated that mesenchymal stem cells (MSCs), predominantly pericytes, accumulate in muscle (mMSCs) overexpressing the α(7B)-integrin (MCK:α(7B); α(7)Tg) and contribute to new fiber formation following exercise. Since vascularization is a common event that supports tissue remodeling, we hypothesized that the α(7)-integrin and/or mMSCs may stimulate vessel growth following eccentric exercise. Wild-type (WT) and α(7)Tg mice were subjected to single or multiple (3 times/wk, 4 wk) bouts of downhill running exercise. Additionally, 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) -labeled mMSCs were intramuscularly injected into WT recipients. A subset of recipient mice were run downhill before injection to recapitulate the exercised microenvironment. While total number of CD31(+) vessels declined in both WT and α(7)Tg muscle following a single bout of exercise, the number of larger CD31(+) vessels with a visible lumen was preferentially increased in α(7)Tg mice following eccentric exercise training (P < 0.05). mMSC transplantation similarly increased vessel diameter and the total number of neuron-glial antigen-2 (NG2(+)) arterioles postexercise. Secretion of arteriogenic factors from mMSCs in response to mechanical strain, including epidermal growth factor and granulocyte macrophage-colony stimulating factor, may account for vessel remodeling. In conclusion, this study demonstrates that the α(7)-integrin and mMSCs contribute to increased vessel diameter size and arteriolar density in muscle in response to eccentric exercise. The information in this study has implications for the therapeutic treatment of injured muscle and disorders that result in vessel occlusion, including peripheral artery disease.

Summary More than 1200 mutations in neuronal voltage-gated sodium channel genes have been identified in patients with several epilepsy syndromes. A common feature of genetic epilepsies is variable expressivity among individuals with the same mutation. The Scn2aQ54 transgenic mouse model has a mutation in Scn2a that results in spontaneous epilepsy. Scn2aQ54 phenotype severity varies depending on the genetic strain background, making it a useful model for identifying and characterizing epilepsy modifier genes. Scn2aQ54 mice on the [C57BL/6JxSJL/J]F1 background exhibit earlier seizure onset, elevated spontaneous seizure frequency, and decreased survival compared to Scn2aQ54 mice congenic on the C57BL/6J strain. Genetic mapping and RNA-Seq analysis identified Cacna1g as a candidate modifier gene at the Moe1 locus, which influences Scn2aQ54 phenotype severity. In this study, we evaluated the modifier potential of Cacna1g, encoding the Cav3.1 voltage-gated calcium channel, by testing whether transgenic alteration of Cacna1g expression modifies severity of the Scn2aQ54 seizure phenotype. Scn2aQ54 mice exhibited increased spontaneous seizure frequency with elevated Cacna1g expression and decreased seizure frequency with decreased Cacna1g expression. These results provide support for Cacna1g as an epilepsy modifier gene and suggest that modulation of Cav3.1 may be an effective therapeutic strategy.

Summary Dravet syndrome, an early onset epileptic encephalopathy, is most often caused by de novo mutation of the neuronal voltage-gated sodium channel gene SCN1A. Mouse models with deletion of Scn1a recapitulate Dravet syndrome phenotypes, including spontaneous generalized tonic-clonic seizures, susceptibility to seizures induced by elevated body temperature, and elevated risk of sudden unexpected death due to epilepsy. Importantly, the epilepsy phenotype of Dravet mouse models is highly strain-dependent, suggesting a strong influence of genetic modifiers. We previously identified Cacna1g, encoding the Cav3.1 subunit of the T-type calcium channel family, as an epilepsy modifier in the Scn2aQ54 transgenic epilepsy mouse model. In this study, we asked whether transgenic alteration of Cacna1g expression modifies severity of the Scn1a+/− Dravet phenotype. Scn1a+/− mice with decreased Cacna1g expression showed partial amelioration of disease phenotypes with improved survival and reduced spontaneous seizure frequency. However, reduced Cacna1g expression did not alter susceptibility to hyperthermia-induced seizures. Transgenic elevation of Cacna1g expression had no effect on the Scn1a+/− epilepsy phenotype. These results provide support for Cacna1g as a genetic modifier in a mouse model of Dravet syndrome and suggest that Cav3.1 may be a potential molecular target for therapeutic intervention in patients.

-The ␣ 7␤1-integrin is an adhesion molecule highly expressed in skeletal muscle that can enhance regeneration in response to eccentric exercise. We have demonstrated that mesenchymal stem cells (MSCs), predominantly pericytes, accumulate in muscle (mMSCs) overexpressing the ␣ 7B-integrin (MCK:␣7B; ␣7Tg) and contribute to new fiber formation following exercise. Since vascularization is a common event that supports tissue remodeling, we hypothesized that the ␣ 7-integrin and/or mMSCs may stimulate vessel growth following eccentric exercise. Wild-type (WT) and ␣ 7Tg mice were subjected to single or multiple (3 times/wk, 4 wk) bouts of downhill running exercise. Additionally, 1,1=-dioctadecyl-3,3,3=,3=-tetramethylindocarbocyanine perchlorate (DiI) -labeled mMSCs were intramuscularly injected into WT recipients. A subset of recipient mice were run downhill before injection to recapitulate the exercised microenvironment. While total number of CD31 ϩ vessels declined in both WT and ␣7Tg muscle following a single bout of exercise, the number of larger CD31 ϩ vessels with a visible lumen was preferentially increased in ␣7Tg mice following eccentric exercise training (P Ͻ 0.05). mMSC transplantation similarly increased vessel diameter and the total number of neuron-glial antigen-2 (NG2 ϩ ) arterioles postexercise. Secretion of arteriogenic factors from mMSCs in response to mechanical strain, including epidermal growth factor and granulocyte macrophage-colony stimulating factor, may account for vessel remodeling. In conclusion, this study demonstrates that the ␣ 7-integrin and mMSCs contribute to increased vessel diameter size and arteriolar density in muscle in response to eccentric exercise. The information in this study has implications for the therapeutic treatment of injured muscle and disorders that result in vessel occlusion, including peripheral artery disease.integrin; pericyte; remodeling; angiogenesis; arteriole INTEGRINS ARE TRANSMEMBRANE adhesion proteins comprised of noncovalently bound ␣-and ␤-subunits. Integrins form clusters and recruit cytoskeletal and cytoplasmic proteins to the cell membrane to provide a mechanism of communication between the outside and inside of the cell (10). In skeletal muscle, the ␣ 7 ␤ 1 -integrin is the predominant heterodimer based on high levels of protein expression, and its primary function is to ensure adhesion by providing a stabilizing link between the actin cytoskeleton and the external environment, specifically, laminin (6). Transgenic expression of the integrin in mouse muscle can suppress macrophage infiltration, sarcolemmal damage, and reductions in force following eccentric exercise (3, 4, 13). Our recently published work also suggests ␣ 7BX2 -integrin overexpression (MCK:␣ 7BX2 ; ␣ 7 Tg) can increase myogenesis, hypertrophic signaling, myofibrillar content, fiber cross-sectional area, muscle cross-sectional area, and maximal isometric force following single or multiple bouts of eccentric exercise (13, 34), a type of exercise that results in muscle lengthening during...