Variants in the KCNT1 gene, which encodes a Na+-activated K+ channel, cause several severe
childhood epilepsy disorders that are largely refractory to treatment. Previously, pathogenic
KCNT1 variants were shown to increase channel current when assayed in non-neuronal cells,
leading to the fundamental question of how increasing a current that is usually associated with
dampening neuronal excitability leads to disorders characterized by excessive, synchronous
neuronal activity. We have created two mouse models with orthologous pathogenic human
KCNT1 variants that have frequent seizures. In both models, cortical inhibitory neurons (cINs)
show strong impairments in membrane excitability and action potential generation, whereas
excitatory neurons (cENs) do not. These data suggest that KCNT1 gain-of-function (GOF)
variants cause childhood epilepsy by impairing the function of cINs, and that KCNT1 channels
play cell-type-specific roles in regulating neuronal excitability. This proposal will test these
hypotheses by (1) determining how KCNT1 block and GOF affect membrane excitability of cIN
subtypes, (2) using newly-developed KCNT1-selective inhibitors to measure the current and
how it is altered by disease-causing variants in cIN subtypes, and (3) testing how impaired
membrane excitability in subpopulations of cINs affects their activity in vivo, and how this relates
to seizures or cortical hyperexcitability. Determining these vulnerable cell types, the underlying
mechanisms, and the in vivo effects of their dysfunction will allow us to relate cellular deficits to
epileptiform activity and seizures. The results will advance our understanding of the regulation
of cIN excitability, the physiological roles of the KCNT1-mediated current, and the disease
mechanisms of KCNT1 GOF in an in vivo model of epilepsy. This will narrow the knowledge gap
between the biophysical effects of ion channel variants and the resulting dysfunction of
networks, and has the potential to improve the targeting of precision therapies for severe
childhood epilepsies.