Functional Characterization and Neuronal Modeling of the Effects of Childhood Absence Epilepsy Variants ofCACNA1H, a T-Type Calcium Channel

Abstract

Sequencing of the T-type Ca2ϩ channel gene CACNA1H revealed 12 nonsynonymous single nucleotide polymorphisms (SNPs) that were found only in childhood absence epilepsy (CAE) patients. One SNP, G773D, was found in two patients. The present study reports the finding of a third patient with this SNP, as well as analysis of their parents. Because of the role of T-channels in determining the intrinsic firing patterns of neurons involved in absence seizures, it was suggested that these SNPs might alter channel function. The goal of the present study was to test this hypothesis by introducing these polymorphisms into a human Ca v 3.2a cDNA and then study alterations in channel behavior using whole-cell patch-clamp recording. Eleven SNPs altered some aspect of channel gating. Computer simulations predict that seven of the SNPs would increase firing of neurons, with three of them inducing oscillations at similar frequencies, as observed during absence seizures. Three SNPs were predicted to decrease firing. Some CAE-specific SNPs (e.g., G773D) coexist with SNPs also found in controls (R788C); therefore, the effect of these polymorphisms were studied. The R788C SNP altered activity in a manner that would also lead to enhanced burst firing of neurons. The G773D-R788C combination displayed different behavior than either single SNP. Therefore, common polymorphisms can alter the effect of CAE-specific SNPs, highlighting the importance of sequence background. These results suggest that CACNA1H is a susceptibility gene that contributes to the development of polygenic disorders characterized by thalamocortical dysrhythmia, such as CAE.

δ Subunit Susceptibility Variants E177A and R220H Associated with Complex Epilepsy Alter Channel Gating and Surface Expression of α4β2δ GABAAReceptors

“…Twin studies have also suggested a strong genetic basis for the complex epilepsies (Berkovic et al, 1998); however, description and validation of the underlying genetic variation for these more common epilepsies has only just begun. Changes in channel properties of CACNA1H are consistent with complex epilepsy (Khosravani et al, 2004(Khosravani et al, , 2005Vitko et al, 2005). The present study reinforces and extends the previously reported role of GABRD as another susceptibility gene underlying complex epilepsy.…”

The I–II Loop Controls Plasma Membrane Expression and Gating of Cav3.2 T-Type Ca2+Channels: A Paradigm for Childhood Absence Epilepsy Mutations

“…Notably, deletion of multiple regions produced nearly (100% in A). The various CAE SNPs are presented as by Vitko et al (2005). One construct harboring two SNPs, G773D and R788C (Chen et al, 2003b;Vitko et al, 2005), is labeled Double.…”

“…Altogether, these results define two roles for this intracellular loop: (1) as an intracellular gating factor that controls the voltage dependence of channel gating and (2) as a regulator of channel surface expression. Previous studies have shown that the polymorphisms associated with CAE are capable of altering channel function (Khosravani et al, 2004;Vitko et al, 2005). The effects of the polymorphisms were modest, and the only common finding was that they led to an increase in persistent currents.…”

Molecular Targets for Antiepileptic Drug Development

“…85,86 Functional expression studies with several of these mutations have revealed a gain of function. 87,88 At least 30 mutations in CACNA1H, some involving splicing defects, are associated with CAE and other idiopathic generalized epilepsies. 89 Five mutations in EFHC1, a gene encoding a protein with an EF-hand motif, have been found in families with juvenile myoclonic epilepsy (JME).…”