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)

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 > School of Cardiovascular & Metabolic Health
• Journal Name: Cold Spring Harbor Protocols
• Publisher: Cold Spring Harbor Laboratory Press
• ISSN: 1940-3402
• ISSN (Online): 1559-6095