Abstracts

Rationale: KCNQ3, encoding the voltage-gated potassium channel subunit Kv7.3, was one of the first identified human epilepsy genes. Inherited loss-of-function missense variants in KCNQ3 cause self-limited familial neonatal epilepsy. In contrast, de novo variants that neutralize arginine 230 (R230) within the voltage sensor result in gain-of-function (GoF) effects including increased current and loss of voltage-dependence. In patients, R230 variants cause intellectual disability, autistic symptoms and abundant sleep-activated multifocal spikes on electroencephalography (EEG). Our objective is to identify changes in neuronal physiology, EEG, and behavior resulting from increased Kv7.3 current in a mouse model of the KCNQ3 GoF disorder. We hypothesize that severe behavioral and electroclinical phenotypes will arise as a result of dampened neuronal excitability.

Methods: We generated mice harboring a R231H mutation in Kcnq3 (C57BL6/J), orthologous to the human pathogenic R230H GoF variant. RNA and protein were analyzed to determine expression levels of Kcnq3 and Kv7.3 respectively. EEG was performed and spike-wave discharges (SWD) identified using interactive Assyst 3.1 software (Kaoskey, Inc). Growth and ultrasonic vocalizations were measured from postnatal day 4 to 10, while adult behaviors, electroconvulsive threshold (ECT), and local field potentials (LFP) recordings were examined in Kcnq3^R231H/+ mice on the (FVB X C57BL6/J)F₁ hybrid background at 7 to 12 weeks.

Results: We observed no significant differences in RNA expression between Kcnq3 (p= 0.89) or its paralogue Kcnq2 (p= 0.85) in Kcnq3^R231H/+ mice (n= 9) compared to wildtype (Wt) (n= 9). Similarly, no differences in total Kv7.3 protein expression (p= 0.65) was observed between the genotypes (n= 5 each). Interestingly, Kv7.3 was increased significantly (p= 0.01) in crude neuronal membranes in Kcnq3^R231H/+ mice (n= 7) compared to Wt (n= 7). While Kcnq3^R231H mice did not show convulsive seizure behavior, but did show frequent SWD (p= 0.02; n= 6), and decreased maximal ECT (p= 0.0024). An anxiety-like phenotype was observed in adult mice: on open field (p= 0.04) and elevated plus maze (EPM) assays (p= 0.0023; n= 13-15 per group). Kcnq3^R231H/+ mice also had decreased ambulatory activity (p= 0.0038), and heightened cued fear response (p= 0.02). Preliminary LFP recordings of electrically stimulated hippocampal responses in Kcnq3^R231H/+ mice (n=3; n=4) showed alterations in both pre- and postsynaptic components in CA3. Paradoxically, the magnitude of a low-latency response corresponding to presynaptic activity was reduced while the magnitude of a longer-latency postsynaptic response was increased, suggesting a possible impairment in axon function but an increase in network excitability overall.

Conclusions: Our preliminary data suggest that the Kcnq3 R231H GoF mutation causes molecular, behavioral, and electroclinical alterations in mice, suggesting the relevance of this model for study of the KCNQ3 GoF disorder. Experiments are ongoing to further delineate the mechanisms by which altered Kv7.3 function results in human neurodevelopmental disease and to lay the preclinical groundwork for targeted treatment of the KCNQ3 GoF disorder.

Funding: Please list any funding that was received in support of this abstract.: None.