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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 1. CaMKII increases Ca2+ spark frequency in WT mouse ventricular myocytes. A, Longitudinal line scan images of Ca2+ sparks in a permeabilized W...Show More

Figure 1. CaMKII increases Ca2+ spark frequency in WT mouse ventricular myocytes. A, Longitudinal line scan images of Ca2+ sparks in a permeabilized WT myocyte under control conditions (left), 3 minutes after exposure to preactivated CaMKII (middle) and 3 minutes after washout of CaM and CaMKII (all at 50 nmol/L [Ca2+]i). B, Pooled data for CaSpF (left) and SR Ca2+ content assessed by the Δ[Ca2+]i on rapid caffeine application (n=5 myocytes; *P<0.05).Show Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 2. CaMKII increases Ca2+ spark frequency in PLB-KO myocytes. A, Ca2+ sparks in a permeabilized PLB-KO myocyte under control conditions (left),...Show More

Figure 2. CaMKII increases Ca2+ spark frequency in PLB-KO myocytes. A, Ca2+ sparks in a permeabilized PLB-KO myocyte under control conditions (left), 3 minutes after exposure to preactivated CaMKII (middle), and 3 minutes after washout (all at 25 nmol/L [Ca2+]i). Pooled data for (B) CaSpF and (C) SR Ca2+ content assessed by caffeine-induced Δ[Ca2+]i (right; n=6 PLB-KO myocytes; *P<0.05). D, Pooled results, n=5 experiments where 1 μmol/L AIP was included throughout.Show Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

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Table 1.
By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Time-Dependent CaMKII Phosphorylation of RyRShow More

Time-Dependent CaMKII Phosphorylation of RyRShow Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 3. CaM decreases Ca2+ sparks in WT and PLB-KO myocytes. A, Dose-dependent CaM effect on CaSpF in WT myocytes (no CaMKII or CaMKII inhibitor, n=...Show More

Figure 3. CaM decreases Ca2+ sparks in WT and PLB-KO myocytes. A, Dose-dependent CaM effect on CaSpF in WT myocytes (no CaMKII or CaMKII inhibitor, n=5). B, Dose-dependent CaM effect on CaSpF in PLB-KO myocytes with 1 μmol/L AIP present in bath (n=5).Show Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 4. RyR Phosphorylation by exogenous CaMKIIα, PKA effects, and FKBP association. A, Time course of 32P incorporation into immunoprecipitated RyR...Show More

Figure 4. RyR Phosphorylation by exogenous CaMKIIα, PKA effects, and FKBP association. A, Time course of 32P incorporation into immunoprecipitated RyR (±OA). Center row shows RyR immunoblot. B, CaSpF after PKA catalytic subunit (1000 U/mL) was incubated in permeabilized cells (5 minutes WT n=5; PLB-KO n=5). C, FKBP12/12.6 expression in ventricular homogenate (Western blots, normalized to WT). D, FKBP12.6 coimmunoprecipitated with RyR antibody, from WT and PLB-KO ventricular homogenates (untreated or treated with PKA catalytic subunit or PKI; n=3 for each).Show Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 5. Endogenous CaMKII activation increases Ca spark frequency. A, Representative line scans in PLB-KO at 10 nmol/L [Ca2+]i, on addition of 1.2 μ...Show More

Figure 5. Endogenous CaMKII activation increases Ca spark frequency. A, Representative line scans in PLB-KO at 10 nmol/L [Ca2+]i, on addition of 1.2 μmol/L CaM with 500 nmol/L [Ca2+]i, and at 5 and 8 minutes after washout of CaM and return of [Ca2+]i to 50 nmol/L. B, Pooled data from this protocol in 6 WT myocytes ([Ca2+]i=50 nmol/L) and (C) 9 PLB-KO myocytes (Bar indicates time of Ca/CaM elevation). D, SR Ca2+ content (caffeine-induced Δ[Ca]i) in WT (n=5, P<0.05) and PLB-KO in both control (Ctl) and after CaMKII activation/washout (w/o; n=5).Show Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 6. Ca2+ sparks duration after endogenous CaMKII activation in PLB-KO. A, Histograms of Ca spark duration (FDHM), normalized to total number of...Show More

Figure 6. Ca2+ sparks duration after endogenous CaMKII activation in PLB-KO. A, Histograms of Ca spark duration (FDHM), normalized to total number of events in each case (703 for Control, 2408 for CaMKII). B, Integration of events from longest to shortest duration. Half of events were longer than 32 ms in CaMKII and 26 ms in control. C, Frequency of macrosparks (FWHM >5 μm) and miniwaves (example in inset), where the latter are counted as 2 macrosparks. Note that fully propagating waves are greatly limited by the 1 mmol/L EGTA in permeabilized fiber solution.Show Less
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Ca²⁺/Calmodulin-Dependent Protein Kinase II Phosphorylation of Ryanodine Receptor Does Affect Calcium Sparks in Mouse Ventricular Myocytes
Tao Guo, Tong Zhang, Ruben Mestril, Donald M. Bers

Circulation Research August 2006, 99 (4) 398-406; DOI: https://doi.org/10.1161/01.RES.0000236756.06252.13

By Tao Guo, Tong Zhang, Ruben Mestril and Donald M. Bers

Figure 7. Time course of phosphorylation by endogenous CaMKII. A, RyR back-phosphorylation of immunoprecipitated RyR from cell lysates pretreated as i...Show More

Figure 7. Time course of phosphorylation by endogenous CaMKII. A, RyR back-phosphorylation of immunoprecipitated RyR from cell lysates pretreated as indicated (top). Lower panels show the time course of PLB-Thr17 phosphorylation (with total PLB) from same samples as in upper panel. Myocytes pretreated with AIP and alkaline phosphatase had the least PLB phosphorylation, but most availability for RyR back phosphorylation (and vice versa for pretreatment with exogenous activated CaMKIIα). B, Pooled RyR back-phosphorylation data normalized to the control (n=4, P<0.05), calculated as the reciprocal of CaMKII- dependent 32P incorporated signals. C, Pooled data for PLB Thr-17 phosphorylation (normalized to the CaMKIIα treatment).Show Less
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Increased Sarcoplasmic Reticulum Calcium Leak but Unaltered Contractility by Acute CaMKII Overexpression in Isolated Rabbit Cardiac Myocytes
Michael Kohlhaas, Tong Zhang, Tim Seidler, Darya Zibrova, Nataliya Dybkova, Astrid Steen, Stefan Wagner, Lu Chen, Joan Heller Brown, Donald M. Bers and Lars S. Maier

Circulation Research. 2006;98:235-244, originally published February 2, 2006
https://doi.org/10.1161/01.RES.0000200739.90811.9f

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Increased Sarcoplasmic Reticulum Calcium Leak but Unaltered Contractility by Acute CaMKII Overexpression in Isolated Rabbit Cardiac Myocytes
Michael Kohlhaas, Tong Zhang, Tim Seidler, Darya Zibrova, Nataliya Dybkova, Astrid Steen, Stefan Wagner, Lu Chen, Joan Heller Brown, Donald M. Bers, Lars S. Maier

Circulation Research February 2006, 98 (2) 235-244; DOI: https://doi.org/10.1161/01.RES.0000200739.90811.9f

By Michael Kohlhaas, Tong Zhang, Tim Seidler, Darya Zibrova, Nataliya Dybkova, Astrid Steen, Stefan Wagner, Lu Chen, Joan Heller Brown, Donald M. Bers and Lars S. Maier

Figure 1. CaMKIIδC overexpression in rabbit ventricular myocytes. A, Western blots (MOI100, 24 hours) showing expression levels of CaMKIIδ (n=5) vs La...Show More

Figure 1. CaMKIIδC overexpression in rabbit ventricular myocytes. A, Western blots (MOI100, 24 hours) showing expression levels of CaMKIIδ (n=5) vs LacZ control (n=5) and MOI-dependent increase in phosphorylation status. B, Immunohistochemical staining and confocal images showing cytosolic (but not nuclear) expression of CaMKIIδC vs no staining in LacZ. C, Myocyte volume in CaMKIIδC (n=66) vs LacZ (n=154). *P<0.05 vs LacZ.Show Less