Zinc controls RyR2 activity during excitation-contraction coupling

Cardiac excitation-contraction (EC) coupling is a process which governs contractility of the heart through the controlled release of Ca2C from the sarcoplasmic reticulum (SR). The type-2 ryanodine receptor (RyR2) is the route through which Ca2C is released from the SR providing the necessary driving force for cellular contraction. In heart failure, RyR2-channels become abnormally active, or ‘leaky’, and are unable to remain closed during diastole resulting in unwanted irregular contractile and electrical activity. Defective Zn2C handling has been shown to contribute to the cellular pathology of certain cardiomyopathies which give rise to impaired contractility including heart failure. This is likely a consequence of altered EC coupling as a result of modified RyR2 function. How zinc impacts upon the contractile force and the release of calcium from intracellular stores in heart is not fully understood. In the recent study by Woodier and coworkers it was shown that cytosolic Zn2C can act as a high affinity activator of RyR2. In the aforementioned study, single RyR2 channels were incorporated into phospholipid bilayers under voltageclamp conditions and the direct action of Zn2C at the cytosolic face of the channel studied. This approach enabled the study of RyR2 function under tight control of the chemical environment. Concentrations of free Zn2C 1 nM potentiated RyR2 activity but the presence of activating levels of cytosolic Ca2C was a requirement for channel activation. At concentrations of free Zn2C > 1 nM, the main activating ligand became Zn2C and the requirement of Ca2C for channel activation was removed. Under these conditions channel gating was altered and RyR2 gated in exceptionally long-lived open states. The ability of Zn2C at a concentration of 1 nM to directly activate RyR2 reveals that RyR2 has a much higher affinity for Zn2C than Ca2C (by »3-orders of magnitude). These data suggest that RyR2-mediated Ca2C-homeostasis is intimately related to intracellular Zn2C levels. Woodier et al. also showed that Zn2C modulated both the frequency and amplitude of Ca2C-waves in cardiomyocytes in a concentration-dependent manner. Reduction of the concentration of intracellular Ca2C to sub-activating concentrations did not abolish Ca2Cwaves in the presence of 1 nM Zn2C. This suggests that RyR2 gating is altered under these conditions whereby RyR2 gates in a Ca2C-independent manner with Zn2C the sole activating ligand. These data indicate that channel dysregulation, through aberrant Zn2C homeostasis, may play a fundamental role in the generation of heart failure and other arrhythmic diseases. Cardiomyocytes contain a small but measurable pool of free Zn2C in the cytosol reported to be »100 pM. Since small changes in the Zn2C level will have a marked effect on RyR2 activity, this becomes highly relevant when we consider that concentrations of Zn2C have recently been reported to be transiently elevated to »50 nM during Zn2C-signaling events. Live-cell detection of intracellular Zn2C and Ca2C using selective fluorophores reveal that intracellular Zn2C concentrations are altered during cardiac EC coupling and that spatio-temporal fluctuations in free Zn2C levels are comparable to those of Ca2C.6 Extracellular Zn2C can also enter cardiomyocytes through the L-type Ca2C channel in a similar manner to Ca2C.7 These Zn2C fluctuations may serve to modulate RyR2 activity highlighting a potential role for Zn2C in fine-tuning graded Ca2C-release events to control the force and duration of © Alan J Stewart and Samantha J Pitt *Correspondence to: Samantha J Pitt; Email: [email protected]