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In situ calibration of nucleoplasmic versus cytoplasmic Ca²+ concentration in adult cardiomyocytes.

ABSTRACT: Quantification of subcellularly resolved Ca²? signals in cardiomyocytes is essential for understanding Ca²? fluxes in excitation-contraction and excitation-transcription coupling. The properties of fluorescent indicators in intracellular compartments may differ, thus affecting the translation of Ca²?-dependent fluorescence changes into [Ca²?] changes. Therefore, we determined the in situ characteristics of a frequently used Ca²? indicator, Fluo-4, and a ratiometric Ca²? indicator, Asante Calcium Red, and evaluated their use for reporting and quantifying cytoplasmic and nucleoplasmic Ca²? signals in isolated cardiomyocytes. Ca²? calibration curves revealed significant differences in the apparent Ca²? dissociation constants of Fluo-4 and Asante Calcium Red between cytoplasm and nucleoplasm. These parameters were used for transformation of fluorescence into nucleoplasmic and cytoplasmic [Ca²?]. Resting and diastolic [Ca²?] were always higher in the nucleoplasm. Systolic [Ca²?] was usually higher in the cytoplasm, but some cells (15%) exhibited higher systolic [Ca²?] in the nucleoplasm. Ca²? store depletion or blockade of Ca²? leak pathways eliminated the resting [Ca²?] gradient between nucleoplasm and cytoplasm, whereas inhibition of inositol 1,4,5-trisphosphate receptors by 2-APB reversed it. The results suggest the presence of significant nucleoplasmic-to-cytoplasmic [Ca²?] gradients in resting myocytes and during the cardiac cycle. Nucleoplasmic [Ca²?] in cardiomyocytes may be regulated via two mechanisms: diffusion from the cytoplasm and active Ca²? release via inositol 1,4,5-trisphosphate receptors from perinuclear Ca²? stores.

SUBMITTER: Ljubojevic S

PROVIDER: S-EPMC3093572 | biostudies-other | 2011 May

REPOSITORIES: biostudies-other

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In situ calibration of nucleoplasmic versus cytoplasmic Ca²+ concentration in adult cardiomyocytes.

Ljubojević Senka S Walther Stefanie S Asgarzoei Mojib M Sedej Simon S Pieske Burkert B Kockskämper Jens J

Biophysical journal 20110501 10

Quantification of subcellularly resolved Ca²⁺ signals in cardiomyocytes is essential for understanding Ca²⁺ fluxes in excitation-contraction and excitation-transcription coupling. The properties of fluorescent indicators in intracellular compartments may differ, thus affecting the translation of Ca²⁺-dependent fluorescence changes into [Ca²⁺] changes. Therefore, we determined the in situ characteristics of a frequently used Ca²⁺ indicator, Fluo-4, and a ratiometric Ca²⁺ indicator, Asante Calcium ...[more]

PMID: 21575569

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Dataset Information

In situ calibration of nucleoplasmic versus cytoplasmic Ca²+ concentration in adult cardiomyocytes.

Publications

In situ calibration of nucleoplasmic versus cytoplasmic Ca²+ concentration in adult cardiomyocytes.

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