Measuring local gradients of intramitochondrial [Ca²⁺] in cardiac myocytes during sarcoplasmic reticulum Ca²⁺ release

Lu, X., Ginsburg, K.S., Kettlewell, S., Bossuyt, J., Smith, G.L. and Bers, D.M. (2013) Measuring local gradients of intramitochondrial [Ca²⁺] in cardiac myocytes during sarcoplasmic reticulum Ca²⁺ release. Circulation Research, 112(3), pp. 424-431. (doi: 10.1161/CIRCRESAHA.111.300501)

Abstract

<p>Rationale: Mitochondrial [Ca²⁺] ([Ca²⁺]mito) regulates mitochondrial energy production, provides transient Ca²⁺ buffering under stress, and can be involved in cell death. Mitochondria are near the sarcoplasmic reticulum (SR) in cardiac myocytes, and evidence for crosstalk exists. However, quantitative measurements of [Ca²⁺]mito are limited, and spatial [Ca²⁺]mito gradients have not been directly measured.</p> <p>Objective: To directly measure local [Ca²⁺]mito during normal SR Ca release in intact myocytes, and evaluate potential subsarcomeric spatial [Ca²⁺]mito gradients.</p> <p>Methods and Results: Using the mitochondrially targeted inverse pericam indicator Mitycam, calibrated in situ, we directly measured [Ca²⁺]mito during SR Ca²⁺ release in intact rabbit ventricular myocytes by confocal microscopy. During steady state pacing, Δ[Ca²⁺]mito amplitude was 29±3 nmol/L, rising rapidly (similar to cytosolic free [Ca²⁺]) but declining much more slowly. Taking advantage of the structural periodicity of cardiac sarcomeres, we found that [Ca²⁺]mito near SR Ca²⁺ release sites (Z-line) versus mid-sarcomere (M-line) reached a high peak amplitude (37±4 versus 26±4 nmol/L, respectively P<0.05) which occurred earlier in time. This difference was attributed to ends of mitochondria being physically closer to SR Ca²⁺ release sites, because the mitochondrial Ca²⁺ uniporter was homogeneously distributed, and elevated [Ca²⁺] applied laterally did not produce longitudinal [Ca²⁺]mito gradients.</p> <p>Conclusions: We developed methods to measure spatiotemporal [Ca²⁺]mito gradients quantitatively during excitation–contraction coupling. The amplitude and kinetics of [Ca²⁺]mito transients differ significantly from those in the cytosol and are respectively higher and faster near the Z-line versus M-line. This approach will help clarify SR-mitochondrial Ca²⁺ signaling.</p>

• Item Type: Articles
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Smith, Professor Godfrey and Kettlewell, Dr Sarah
• Authors: Lu, X., Ginsburg, K.S., Kettlewell, S., Bossuyt, J., Smith, G.L., and Bers, D.M.
• College/School: College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health
• Journal Name: Circulation Research
• Publisher: American Heart Association
• ISSN: 0009-7330
• ISSN (Online): 1524-4571
• Published Online: 14 December 2012