Sensitized signalling between L-type Ca 2+ channels and ryanodine receptors in the absence or inhibition of FKBP12.6 in cardiomyocytes

<span class="paragraphSection"><div class="boxTitle">Aims</div>The heart contraction is controlled by the Ca²⁺-induced Ca²⁺release (CICR) between L-type Ca²⁺channels and ryanodine receptors (RyRs). The FK506-binding protein FKBP12.6 binds to RyR subunits, but its role in stabilizing RyR function has been debated for long. Recent reports of high-resolution RyR structure show that the HD2 domain that binds to the SPRY2 domain of neighbouring subunit in FKBP-bound RyR1 is detached and invisible in FKBP-null RyR2. The present study was to test the consequence of FKBP12.6 absence on the <span style="font-style:italic;">in situ</span> activation of RyR2.<div class="boxTitle">Methods and results</div>Using whole-cell patch-clamp combined with confocal imaging, we applied a near threshold depolarization to activate a very small fraction of LCCs, which in turn activated RyR Ca²⁺sparks stochastically. FKBP12.6-knockout and FK506/rapamycin treatments increased spark frequency and LCC-RyR coupling fidelity without altering LCC open probability. Neither FK506 nor rapamycin further altered LCC-RyR coupling fidelity in FKBP12.6-knockout cells. In loose-seal patch-clamp experiments, the LCC-RyR signalling kinetics, indexed by the delay for a LCC sparklet to trigger a RyR spark, was accelerated after FKBP12.6 knockout and FK506/rapamycin treatments. These results demonstrated that RyRs became more sensitive to Ca²⁺triggers without FKBP12.6. Isoproterenol (1 μM) further accelerated the LCC-RyR signalling in FKBP12.6-knockout cells. The synergistic sensitization of RyRs by catecholaminergic signalling and FKBP12.6 dysfunction destabilized the CICR system, leading to chaotic Ca²⁺waves and ventricular arrhythmias.<div class="boxTitle">Conclusion:</div>FKBP12.6 keeps the RyRs from over-sensitization, stabilizes the potentially regenerative CICR system, and thus may suppress the life-threatening arrhythmogenesis.</span>

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