BK channel activation by L-type Ca²⁺ channels Ca_V1.2 and Ca_V1.3 during the subthreshold phase of an action potential

Abstract

Mammalian circadian (24-hour) rhythms are timed by the pattern of spontaneous action potential firing in the suprachiasmatic nucleus (SCN). This oscillation in firing is produced through circadian regulation of several membrane currents, including large-conductance Ca2+- and voltage-activated K+ (BK) and L-type Ca2+ channel (LTCC) currents. During the day, steady-state BK currents depend mostly on LTCCs for activation, while at night, they depend predominantly on RyRs. However, the contribution of these Ca2+ channels to BK channel activation during action potential firing has not been thoroughly investigated. In this study, we used a pharmacological approach to determine that both LTCCs and RyRs contribute to the baseline membrane potential of SCN action potential waveforms, as well as action potential-evoked BK current, during the day and night, respectively. Since the baseline membrane potential is a major determinant of circadian firing rate, we focused on the LTCCs contributing to low voltage activation of BK channels during the subthreshold phase. For these experiments, two LTCC subtypes found in SCN (CaV1.2 and CaV1.3) were co-expressed with BK channels in heterologous cells, where their differential contributions could be separately measured. CaV1.3 channels produced currents that were shifted to more hyperpolarized potentials compared to CaV1.2, resulting in increased subthreshold Ca2+ and BK currents during an action potential command. These results show that while multiple Ca2+ sources in SCN can contribute to the activation of BK current during an action potential, specific BK-CaV1.3 partnerships may optimize the subthreshold BK current activation that is critical for firing rate regulation.

Contributions of CaV1.3 Channels to Ca2+ Current and Ca2+-Activated BK Current in the Suprachiasmatic Nucleus

“…To examine this idea, C57BL6 WT BK currents were recorded in two Ca 2+ buffering conditions. As a control, BK currents were first recorded under standard whole-cell voltage-clamp conditions with 0.9 mM EGTA in the internal solution, conditions which permit the endogenous Ca 2+ sources to contribute to BK current activation (Fakler and Adelman, 2008 ; Whitt et al, 2018 ; Plante et al, 2021 ). These BK currents recorded with 0.9 mM EGTA were then compared to BK currents recorded with internal solution containing 5 mM BAPTA, a Ca 2+ chelator with fast kinetics that disrupts the functional coupling of BK channels located >10–20 nm from their Ca 2+ source (Berkefeld et al, 2006 ; Fakler and Adelman, 2008 ; Cox, 2014 ).…”

“…Correlated with the change in current magnitude, inhibition of LTCC current with nimodipine decreases SCN firing selectively during the day (Pennartz et al, 2002 ; Whitt et al, 2018 ; McNally et al, 2020 ). One mechanism for the effect on SCN firing is via LTCC-dependent activation of Ca 2+ -activated BK K + channels (Plante et al, 2021 ). During the day, LTCCs are responsible for the majority of BK current activation (Jackson et al, 2004 ; Whitt et al, 2018 ; Plante et al, 2021 ).…”

“…One mechanism for the effect on SCN firing is via LTCC-dependent activation of Ca 2+ -activated BK K + channels (Plante et al, 2021 ). During the day, LTCCs are responsible for the majority of BK current activation (Jackson et al, 2004 ; Whitt et al, 2018 ; Plante et al, 2021 ). However, at night, their role is reduced, and BK current activation relies primarily on release of Ca 2+ from intracellular stores (Whitt et al, 2018 ; Plante et al, 2021 ).…”

“…During the day, LTCCs are responsible for the majority of BK current activation (Jackson et al, 2004 ; Whitt et al, 2018 ; Plante et al, 2021 ). However, at night, their role is reduced, and BK current activation relies primarily on release of Ca 2+ from intracellular stores (Whitt et al, 2018 ; Plante et al, 2021 ). Either Ca V 1.2 or Ca V 1.3 could contribute to the LTCC current that drives daytime activation of the BK current, as both channel subtypes are known to activate BK currents produced by BK channel variants from SCN (Plante et al, 2021 ) or within other excitable cells (Roberts et al, 1990 ; Wisgirda and Dryer, 1994 ; Prakriya and Lingle, 1999 ; Sun et al, 2003 ; Grunnet and Kaufmann, 2004 ; Berkefeld et al, 2006 ; Marcantoni et al, 2010 ; Hou et al, 2016 ; Bellono et al, 2017 ; Vivas et al, 2017 ; Plante et al, 2021 ).…”

“…However, at night, their role is reduced, and BK current activation relies primarily on release of Ca 2+ from intracellular stores (Whitt et al, 2018 ; Plante et al, 2021 ). Either Ca V 1.2 or Ca V 1.3 could contribute to the LTCC current that drives daytime activation of the BK current, as both channel subtypes are known to activate BK currents produced by BK channel variants from SCN (Plante et al, 2021 ) or within other excitable cells (Roberts et al, 1990 ; Wisgirda and Dryer, 1994 ; Prakriya and Lingle, 1999 ; Sun et al, 2003 ; Grunnet and Kaufmann, 2004 ; Berkefeld et al, 2006 ; Marcantoni et al, 2010 ; Hou et al, 2016 ; Bellono et al, 2017 ; Vivas et al, 2017 ; Plante et al, 2021 ). Little is known about the specific role of Ca V 1.2 on SCN firing, but Ca V 1.2 expression is higher at night and mouse knockouts display altered nighttime phase-shifting behavior (Schmutz et al, 2014 ).…”