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Functional coupling of calcineurin and protein kinase A in mouse ventricular myocytes
Author(s) -
Santana L. F.,
Chase E. G.,
Votaw V. S.,
Nelson Mark T.,
Greven R.
Publication year - 2002
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2002.020552
Subject(s) - calcineurin , myocyte , calcium , patch clamp , ryanodine receptor , voltage dependent calcium channel , endocrinology , medicine , chemistry , phosphatase , biophysics , okadaic acid , protein kinase a , voltage clamp , biology , kinase , membrane potential , phosphorylation , biochemistry , receptor , transplantation
We examined the role of the Ca 2+ ‐regulated protein phosphatase calcineurin in controlling Ca 2+ signalling in mouse ventricular myocytes. Membrane currents and voltage were measured in single myocytes using the patch‐clamp technique. Cytoplasmic Ca 2+ concentration ([Ca 2+ ] i ) was measured in cells loaded with the fluorescent Ca 2+ indicators fluo‐4 or fura‐2 using a confocal or epifluorescence microscope. Inhibition of calcineurin with cyclosporin A (CsA, 100 n m ) or the calcineurin auto‐inhibitory peptide (CiP, 100 μM), increased the amplitude and rate of decay of the evoked [Ca 2+ ] i transient and also prolonged the action potential (AP) of ventricular myocytes to a similar extent. The effects of CsA (100 n m ) and 100 μM CiP on the [Ca 2+ ] i transient and AP were not additive. Calcineurin inhibition did not modify the K + currents responsible for repolarisation of the mouse ventricle. Instead, inhibition of calcineurin increased the amplitude of the Ca 2+ current ( I Ca ) and the evoked calcium transient normalized to the I Ca . Calcium sparks, which underlie the [Ca 2+ ] i transient, had a higher frequency and amplitude, suggesting an elevation of SR calcium load. Inhibition of protein kinase A (PKA) prevented the effects of calcineurin inhibition, indicating that calcineurin opposes the actions of PKA. Finally, immunofluorescence images suggest that calcineurin and PKA co‐localize near the T‐tubules of ventricular myocytes. We propose that calcineurin and PKA are co‐localized to control Ca 2+ influx through calcium channels and calcium release through ryanodine receptors.