Nonuniform and variable arrangements of ryanodine receptors within mammalian ventricular couplons.
BACKGROUND
Single-tilt tomograms of the dyads in rat ventricular myocytes indicated that type 2 ryanodine receptors (RYR2s) were not positioned in a well-ordered array. Furthermore, the orientation and packing strategy of purified type 1 ryanodine receptors in lipid bilayers is determined by the free Mg2+ concentration. These observations led us to test the hypothesis that RYR2s within the mammalian dyad have multiple and complex arrangements.


OBJECTIVE
To determine the arrangement of RYR2 tetramers in the dyads of mammalian cardiomyocytes and the effects of physiologically and pathologically relevant factors on this arrangement.


RESULTS
We used dual-tilt electron tomography to produce en-face views of dyads, enabling a direct examination of RYR2 distribution and arrangement. Rat hearts fixed in situ; isolated rat cardiomyocytes permeabilized, incubated with 1 mmol/L Mg2+, and then fixed; and sections of human ventricle, all showed that the tetramer packing within a dyad was nonuniform containing a mix of checkerboard and side-by-side arrangements, as well as isolated tetramers. Both phosphorylation and 0.1 mmol/L Mg2+ moved the tetramers into a predominantly checkerboard configuration, whereas the 4 mmol/L Mg2+ induced a dense side-by-side arrangement. These changes occurred within 10 minutes of application of the stimuli.


CONCLUSIONS
The arrangement of RYR2 tetramers within the mammalian dyad is neither uniform nor static. We hypothesize that this is characteristic of the dyad in vivo and may provide a mechanism for modulating the open probabilities of the individual tetramers.
From the Department of Cellular and Physiological Sciences (P.A., D.R.L.S., E.D.W.M.), Department of Pediatrics (S.S.), and Department of Surgery (S.K.G., A.I.M.C.), University of British Columbia, Vancouver, British Columbia, Canada.
Parisa
P
Asghari
From the Department of Cellular and Physiological Sciences (P.A., D.R.L.S., E.D.W.M.), Department of Pediatrics (S.S.), and Department of Surgery (S.K.G., A.I.M.C.), University of British Columbia, Vancouver, British Columbia, Canada.
David R L
DR
Scriven
From the Department of Cellular and Physiological Sciences (P.A., D.R.L.S., E.D.W.M.), Department of Pediatrics (S.S.), and Department of Surgery (S.K.G., A.I.M.C.), University of British Columbia, Vancouver, British Columbia, Canada.
Shubhayan
S
Sanatani
From the Department of Cellular and Physiological Sciences (P.A., D.R.L.S., E.D.W.M.), Department of Pediatrics (S.S.), and Department of Surgery (S.K.G., A.I.M.C.), University of British Columbia, Vancouver, British Columbia, Canada.
Sanjiv K
SK
Gandhi
From the Department of Cellular and Physiological Sciences (P.A., D.R.L.S., E.D.W.M.), Department of Pediatrics (S.S.), and Department of Surgery (S.K.G., A.I.M.C.), University of British Columbia, Vancouver, British Columbia, Canada.
Andrew I M
AI
Campbell
From the Department of Cellular and Physiological Sciences (P.A., D.R.L.S., E.D.W.M.), Department of Pediatrics (S.S.), and Department of Surgery (S.K.G., A.I.M.C.), University of British Columbia, Vancouver, British Columbia, Canada. edmoore@mail.ubc.ca.
Edwin D W
ED
Moore
D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D055032:Electron Microscope Tomography; D004789:Enzyme Activation / Q000187:drug effects; D056966:Excitation Contraction Coupling; D006352:Heart Ventricles / Q000737:chemistry* / Q000166:cytology / Q000648:ultrastructure; D006801:Humans; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D032383:Myocytes, Cardiac / Q000737:chemistry* / Q000187:drug effects / Q000648:ultrastructure; D010766:Phosphorylation; D011494:Protein Kinases / Q000502:physiology; D011499:Protein Processing, Post-Translational; D051381:Rats; D017208:Rats, Wistar; D019837:Ryanodine Receptor Calcium Release Channel / Q000032:analysis* / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000502:physiology
D019837:Ryanodine Receptor Calcium Release Channel; D011494:Protein Kinases; D008274:Magnesium
calcium; electron microscope tomography; excitation contraction coupling; phosphorylation; ryanodine receptor calcium release channel; sarcoplasmic reticulum