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

Xiyuan Lu, Kenneth S Ginsburg, Sarah Kettlewell, Julie B C Bossuyt, Godfrey L. Smith, Donald M Bers

Research output: Contribution to journalArticle

58 Citations (Scopus)

Abstract

RATIONALE:: Mitochondrial [Ca2+] ([Ca2+] mito) regulates mitochondrial energy production, provides transient Ca 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. OBJECTIVE:: To directly measure local [Ca2+]mito during normal SR Ca release in intact myocytes, and evaluate potential subsarcomeric spatial [Ca2+] mito gradients. METHODS AND RESULTS:: Using the mitochondrially targeted inverse pericam indicator Mitycam, calibrated in situ, we directly measured [Ca2+]mito during SR Ca 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 Ca 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 Ca release sites, because the mitochondrial Ca uniporter was homogeneously distributed, and elevated [Ca2+] applied laterally did not produce longitudinal [Ca2+]mito gradients. 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 Ca signaling.

Original languageEnglish (US)
Pages (from-to)424-431
Number of pages8
JournalCirculation Research
Volume112
Issue number3
DOIs
StatePublished - Feb 1 2013

Fingerprint

Sarcoplasmic Reticulum
Cardiac Myocytes
Sarcomeres
Muscle Cells
Mitochondria
Excitation Contraction Coupling
Periodicity
Confocal Microscopy
Cytosol
Cell Death
Rabbits

Keywords

  • calcium
  • cardiac myocytes
  • mitochondria
  • sarcoplasmic reticulum Ca release

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

Cite this

Measuring local gradients of intramitochondrial [Ca2+] in cardiac myocytes during sarcoplasmic reticulum Ca2+ release. / Lu, Xiyuan; Ginsburg, Kenneth S; Kettlewell, Sarah; Bossuyt, Julie B C; Smith, Godfrey L.; Bers, Donald M.

In: Circulation Research, Vol. 112, No. 3, 01.02.2013, p. 424-431.

Research output: Contribution to journalArticle

@article{f9bea9a411914a28a9813ca87850b6f5,
title = "Measuring local gradients of intramitochondrial [Ca2+] in cardiac myocytes during sarcoplasmic reticulum Ca2+ release",
abstract = "RATIONALE:: Mitochondrial [Ca2+] ([Ca2+] mito) regulates mitochondrial energy production, provides transient Ca 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. OBJECTIVE:: To directly measure local [Ca2+]mito during normal SR Ca release in intact myocytes, and evaluate potential subsarcomeric spatial [Ca2+] mito gradients. METHODS AND RESULTS:: Using the mitochondrially targeted inverse pericam indicator Mitycam, calibrated in situ, we directly measured [Ca2+]mito during SR Ca 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 Ca 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 Ca release sites, because the mitochondrial Ca uniporter was homogeneously distributed, and elevated [Ca2+] applied laterally did not produce longitudinal [Ca2+]mito gradients. 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 Ca signaling.",
keywords = "calcium, cardiac myocytes, mitochondria, sarcoplasmic reticulum Ca release",
author = "Xiyuan Lu and Ginsburg, {Kenneth S} and Sarah Kettlewell and Bossuyt, {Julie B C} and Smith, {Godfrey L.} and Bers, {Donald M}",
year = "2013",
month = "2",
day = "1",
doi = "10.1161/CIRCRESAHA.111.300501",
language = "English (US)",
volume = "112",
pages = "424--431",
journal = "Circulation Research",
issn = "0009-7330",
publisher = "Lippincott Williams and Wilkins",
number = "3",

}

TY - JOUR

T1 - Measuring local gradients of intramitochondrial [Ca2+] in cardiac myocytes during sarcoplasmic reticulum Ca2+ release

AU - Lu, Xiyuan

AU - Ginsburg, Kenneth S

AU - Kettlewell, Sarah

AU - Bossuyt, Julie B C

AU - Smith, Godfrey L.

AU - Bers, Donald M

PY - 2013/2/1

Y1 - 2013/2/1

N2 - RATIONALE:: Mitochondrial [Ca2+] ([Ca2+] mito) regulates mitochondrial energy production, provides transient Ca 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. OBJECTIVE:: To directly measure local [Ca2+]mito during normal SR Ca release in intact myocytes, and evaluate potential subsarcomeric spatial [Ca2+] mito gradients. METHODS AND RESULTS:: Using the mitochondrially targeted inverse pericam indicator Mitycam, calibrated in situ, we directly measured [Ca2+]mito during SR Ca 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 Ca 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 Ca release sites, because the mitochondrial Ca uniporter was homogeneously distributed, and elevated [Ca2+] applied laterally did not produce longitudinal [Ca2+]mito gradients. 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 Ca signaling.

AB - RATIONALE:: Mitochondrial [Ca2+] ([Ca2+] mito) regulates mitochondrial energy production, provides transient Ca 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. OBJECTIVE:: To directly measure local [Ca2+]mito during normal SR Ca release in intact myocytes, and evaluate potential subsarcomeric spatial [Ca2+] mito gradients. METHODS AND RESULTS:: Using the mitochondrially targeted inverse pericam indicator Mitycam, calibrated in situ, we directly measured [Ca2+]mito during SR Ca 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 Ca 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 Ca release sites, because the mitochondrial Ca uniporter was homogeneously distributed, and elevated [Ca2+] applied laterally did not produce longitudinal [Ca2+]mito gradients. 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 Ca signaling.

KW - calcium

KW - cardiac myocytes

KW - mitochondria

KW - sarcoplasmic reticulum Ca release

UR - http://www.scopus.com/inward/record.url?scp=84873410873&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84873410873&partnerID=8YFLogxK

U2 - 10.1161/CIRCRESAHA.111.300501

DO - 10.1161/CIRCRESAHA.111.300501

M3 - Article

VL - 112

SP - 424

EP - 431

JO - Circulation Research

JF - Circulation Research

SN - 0009-7330

IS - 3

ER -