Abstracts

RATIONALE: Nine mutations that cause generalized epilepsy with febrile seizures plus (GEFS+) have been identified in the [italic]SCN1A[/italic] gene encoding the [alpha] subunit of the Na[sub]v[/sub]1.1 voltage-gated sodium channel. We have previously described the functional properties of two of these mutations (T875M and R1648H). T875M was shown to enhance slow inactivation, while R1648H dramatically accelerated recovery from inactivation. A third mutation, W1204R, changes a residue in the DII-DIII cytoplasmic linker region located approximately 9 amino acids from the start of the DIIIS1 transmembrane segment that is evolutionarily conserved in vertebrate and invertebrate channels. The mutation was identified in all six affected members of a four generation family. The clinical course was variable, including individuals with isolated febrile seizures, severe myoclonic seizures, and severe epilepsy with mental retardation (Escayg et. al., Am J Hum Genet 2001; 68(4): 866-73). The objective of this study is to determine the effects of the W1204R mutation on sodium channel function while attempting to define a possible common molecular mechanism by which mutations in [italic]SCN1A[/italic] result in the clinical disorder GEFS+2.
METHODS: The mutation was cloned into the orthologous rat channel, rNa[sub]v[/sub]1.1, and the electrophysiological properties of the mutant channels were determined in the absence and presence of the [beta]1 subunit in [italic]Xenopus[/italic] oocytes using the cut-open oocyte and two-electrode voltage clamp techniques.
RESULTS: The W1204R mutation resulted in approximately 11 mV hyperpolarized shifts in the voltage-dependence of activation and steady-state inactivation when expressed as an [alpha] subunit alone. When the channels were coexpressed with the [beta]1 subunit, the hyperpolarized shifts were still present but smaller, approximately 5 mV in magnitude. All other fast and slow-gated properties that we examined were comparable for the mutant and wild-type channels.
CONCLUSIONS: The negative shift in activation would increase channel excitability, whereas the negative shift in inactivation would decrease excitability. The negative shifts in both properties also shifted the window current, which is the voltage region in which sodium channels can continue to open because some percentage of channels are activated and not all of the channels are inactivated. The shift in window current for the W1204R mutation could result in hyperexcitability because the neuron[ssquote]s potential is more likely to reach the more negative range. These results demonstrate that a third [italic]SCN1A[/italic] mutation that causes GEFS+2 alters the properties of the sodium channel in a different manner than the previous two mutations that were studied. The diversity in functional effects for these three mutations indicates that a similar clinical phenotype can result from very different underlying sodium channel abnormalities.
[Supported by: National Institutes of Health Grant NS26729]