Differential contribution of Ca2+ sources to day and night BK current activation in the circadian clock

Abstract

Large conductance K+ (BK) channels are expressed widely in neurons, where their activation is regulated by membrane depolarization and intracellular Ca2+ (Ca2+ i). To enable this regulation, BK channels functionally couple to both voltage-gated Ca2+ channels (VGCCs) and channels mediating Ca2+ release from intracellular stores. However, the relationship between BK channels and their specific Ca2+ source for particular patterns of excitability is not well understood. In neurons within the suprachiasmatic nucleus (SCN)-the brain's circadian clock-BK current, VGCC current, and Ca2+ i are diurnally regulated, but paradoxically, BK current is greatest at night when VGCC current and Ca2+ i are reduced. Here, to determine whether diurnal regulation of Ca2+ is relevant for BK channel activation, we combine pharmacology with day and night patch-clamp recordings in acute slices of SCN. We find that activation of BK current depends primarily on three types of channels but that the relative contribution changes between day and night. BK current can be abrogated with nimodipine during the day but not at night, establishing that L-type Ca2+ channels (LTCCs) are the primary daytime Ca2+ source for BK activation. In contrast, dantrolene causes a significant decrease in BK current at night, suggesting that nighttime BK activation is driven by ryanodine receptor (RyR)-mediated Ca2+ i release. The N- and P/Q-type Ca2+ channel blocker ?-conotoxin MVI IC causes a smaller reduction of BK current that does not differ between day and night. Finally, inhibition of LTCCs, but not RyRs, eliminates BK inactivation, but the BK ?2 subunit was not required for activation of BK current by LTCCs. These data reveal a dynamic coupling strategy between BK channels and their Ca2+ sources in the SCN, contributing to diurnal regulation of SCN excitability. Copyright 2018 Whitt et al.