Cooper E. Roache

KCNMA1 encodes the voltage- and calcium-activated K⁺ (BK) channel, which regulates suprachiasmatic nucleus (SCN) neuronal firing and circadian behavioral rhythms. Gain-of-function (GOF) and loss-of-function (LOF) alterations in BK channel activity disrupt circadian behavior, but the effect of human disease-associated KCNMA1 channelopathy variants has not been studied on clock function. Here, we assess circadian behavior in two GOF and one LOF mouse lines. Heterozygous Kcnma1^N999S/WT and homozygous Kcnma1^D434G/D434G mice are validated as GOF models of paroxysmal dyskinesia (PNKD3), but whether circadian rhythm is affected in this hypokinetic locomotor disorder is unknown. Conversely, homozygous LOF Kcnma1^H444Q/H444Q mice do not demonstrate PNKD3. We assessed circadian behavior by locomotor wheel running activity. All three mouse models were rhythmic, but Kcnma1^N999S/WT and Kcnma1^D434G/D434G showed reduced circadian amplitude and decreased wheel activity, corroborating prior studies focused on acute motor coordination. In addition, Kcnma1^D434G/D434G mice had a small decrease in period. However, the phase-shifting sensitivity for both GOF mouse lines was abnormal. Both Kcnma1^N999S/WT and Kcnma1^D434G/D434G mice displayed increased responses to light pulses and took fewer days to re-entrain to a new light:dark cycle. In contrast, the LOF Kcnma1^H444Q/H444Q mice showed no difference in any of the circadian parameters tested. The enhanced sensitivity to phase-shifting stimuli in Kcnma1^N999S/WT and Kcnma1^D434G/D434G mice was similar to other Kcnma1 GOF mice. Together with previous studies, these results suggest that increasing BK channel activity decreases circadian clock robustness, without rhythm ablation.

KCNMA1 forms the pore of BK K ⁺ channels, which regulate neuronal and muscle excitability. Recently, genetic screening identified heterozygous KCNMA1 variants in a subset of patients with debilitating paroxysmal non-kinesigenic dyskinesia, presenting with or without epilepsy (PNKD3). However, the relevance of KCNMA1 mutations and the basis for clinical heterogeneity in PNKD3 has not been established. Here we evaluate the relative severity of three KCNMA1 patient variants in BK channels, neurons, and mice. In heterologous cells, BK ^N999S and BK ^D434G channels displayed gain-of-function (GOF) properties, whereas BK ^H444Q channels showed loss-of-function (LOF) properties. The relative degree of channel activity was BK ^N999S > BK ^D434G > WT > BK ^H444Q . BK currents and action potential firing were increased, and seizure thresholds decreased, in Kcnma1 ^N999S/WT and Kcnma1 ^D434G/WT transgenic mice but not Kcnma1 ^H444Q/WT mice. In a novel behavioral test for paroxysmal dyskinesia, the more severely affected Kcnma1 ^N999S/WT mice became immobile after stress, consistent with stress-induced immobility episodes observed in PNKD3-affected individuals. Homozygous Kcnma1 ^D434G/D434G mice showed similar immobility, but in contrast, homozygous Kcnma1 ^H444Q/H444Q mice displayed hyperkinetic behavior. These data establish the relative pathogenic potential of patient alleles as N999S > D434G > H444Q and validate Kcnma1 ^N999S/WT mice as a model for PNKD3 with increased seizure propensity.