Evaluation of four channelopathy variants on BK channel current under Ca1.2 activation.

Variants in , encoding the voltage- and calcium-activated K (BK) channel, are associated with human neurological disease. The effects of gain-of-function (GOF) and loss-of-function (LOF) variants have been predominantly studied on BK channel currents evoked under steady-state voltage and Ca conditions. However, in their physiological context, BK channels exist in partnership with voltage-gated Ca channels and respond to dynamic changes in intracellular Ca (Ca). In this study, an L-type voltage-gated Ca channel present in the brain, Ca1.2, was co-expressed with wild type and mutant BK channels containing GOF (D434G, N999S) and LOF (H444Q, D965V) patient-associated variants in HEK-293T cells. Whole-cell BK currents were recorded under Ca1.2 activation using buffering conditions that restrict Ca to nano- or micro-domains. Both conditions permitted wild type BK current activation in response to Ca1.2 Ca influx, but differences in behavior between wild type and mutant BK channels were reduced compared to prior studies in clamped Ca. Only the N999S mutation produced an increase in BK current in both micro- and nano-domains using square voltage commands and was also detectable in BK current evoked by a neuronal action potential within a microdomain. These data corroborate the GOF effect of N999S on BK channel activity under dynamic voltage and Ca stimuli, consistent with its pathogenicity in neurological disease. However, the patient-associated mutations D434G, H444Q, and D965V did not exhibit significant effects on BK current under Ca1.2-mediated Ca influx, in contrast with prior steady-state protocols. These results demonstrate a differential potential for variant pathogenicity compared under diverse voltage and Ca conditions.