title: Cardiac ryanodine receptor activation by a high Ca²⁺ store load is reversed in a reducing cytoplasmic redox environment.
abstract: Here, we report the impact of redox potential on isolated cardiac ryanodine receptor (RyR2) channel activity and its response to physiological changes in luminal [Ca(2+)]. Basal leak from the sarcoplasmic reticulum is required for normal Ca(2+) handling, but excess diastolic Ca(2+) leak attributed to oxidative stress is thought to lower the threshold of RyR2 for spontaneous sarcoplasmic reticulum Ca(2+) release, thus inducing arrhythmia in pathological situations. Therefore, we examined the RyR2 response to luminal [Ca(2+)] under reducing or oxidising cytoplasmic redox conditions. Unexpectedly, as luminal [Ca(2+)] increased from 0.1 to 1.5 mM, RyR2 activity declined when pretreated with cytoplasmic 1 mM DTT or buffered with GSH∶GSSG to a normal reduced cytoplasmic redox potential (-220 mV). Conversely, with 20 µM cytoplasmic 4,4'-DTDP or buffering of the redox potential to an oxidising value (-180 mV), RyR2 activity increased with increasing luminal [Ca(2+)]. The luminal redox potential was constant at -180 mV in each case. These responses to luminal [Ca(2+)] were maintained with cytoplasmic 2 mM Na2ATP or 5 mM MgATP (1 mM free Mg(2+)). Overall, the results suggest that the redox potential in the RyR2 junctional microdomain is normally more oxidised than that of the bulk cytoplasm.
authors.affiliation: John Curtin School of Medical Research, Australian National University, Canberra, ACT 6200, Australia.
authors.forename: Amy D
authors.initials: AD
authors.lastname: Hanna
authors.affiliation: John Curtin School of Medical Research, Australian National University, Canberra, ACT 6200, Australia.
authors.forename: Alex
authors.initials: A
authors.lastname: Lam
authors.affiliation: John Curtin School of Medical Research, Australian National University, Canberra, ACT 6200, Australia.
authors.forename: Chris
authors.initials: C
authors.lastname: Thekkedam
authors.affiliation: John Curtin School of Medical Research, Australian National University, Canberra, ACT 6200, Australia.
authors.forename: Esther M
authors.initials: EM
authors.lastname: Gallant
authors.affiliation: Centre for Research in Therapeutic Solutions, University of Canberra, Bruce, ACT 2617, Australia.
authors.forename: Nicole A
authors.initials: NA
authors.lastname: Beard
authors.affiliation: John Curtin School of Medical Research, Australian National University, Canberra, ACT 6200, Australia angela.dulhunty@anu.edu.au.
authors.forename: Angela F
authors.initials: AF
authors.lastname: Dulhunty
mesh_terms: D000818:Animals; D001145:Arrhythmias, Cardiac / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling; D002478:Cells, Cultured; D060833:Cellular Microenvironment; D003593:Cytoplasm / Q000378:metabolism*; D004285:Dogs; D021962:Membrane Microdomains / Q000378:metabolism*; D008564:Membrane Potentials; D032383:Myocytes, Cardiac / Q000502:physiology*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D012756:Sheep
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium
keywords: Cardiac ryanodine receptor; Luminal Ca2+ sensitivity; Redox potential; Sarcoplasmic reticulum