Measuring sarcoplasmic reticulum Ca2+ content, fractional release, and Ca2+ buffering in cardiac myocytes

Macquaide, N. , Bito, V. and Sipido, K. R. (2015) Measuring sarcoplasmic reticulum Ca2+ content, fractional release, and Ca2+ buffering in cardiac myocytes. Cold Spring Harbor Protocols, 2015(4), pdb.prot07. (doi: 10.1101/pdb.prot076976)

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Abstract

Here, we describe a protocol for the reliable measurement of the amount of Ca2+ in the sarcoplasmic reticulum (SR) Ca2+ store of cardiac myocytes. The whole-cell patch-clamp method is used to provide controlled loading of the SR during conditioning depolarizing pulses, followed by rapid application of a high dose of caffeine to release all SR Ca2+ and to prevent Ca2+ reuptake by the SR. Simultaneous measurement of membrane currents records Ca2+ extruded through the Na+–Ca2+ exchanger. The integral of the caffeine-induced Na+–Ca2+ exchange current is then used as a measure of the SR Ca2+. Derived measurements include the Ca2+ buffering capacity and measurement of fractional release as an indicator of coupling gain. Caveats, advantages, and disadvantages of this method and alternative methods are discussed.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:MacQuaide, Dr Niall
Authors: Macquaide, N., Bito, V., and Sipido, K. R.
College/School:College of Medical Veterinary and Life Sciences > Institute of Cardiovascular and Medical Sciences
Journal Name:Cold Spring Harbor Protocols
Publisher:Cold Spring Harbor Laboratory Press
ISSN:1940-3402
ISSN (Online):1559-6095

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