Elsevier

Methods in Enzymology

Volume 207, 1992, Pages 699-707
Methods in Enzymology

[49] Planar bilayer recording of ryanodine receptors of sarcoplasmic reticulum

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This chapter describes methods to incorporate sarcoplasmic reticulum (SR) Ca2+ channels, also called “ryanodine receptors,” into planar bilayers. Recordings of ryanodine receptors are best made using CsCI as current carrier, instead of the Ca-HEPES and Tris-HEPES solutions. The use of CsCI instead of Ca-HEPES and Tris-HEPES eliminate the need for perfusion and the need for large and unphysiological gradients of Ca2+, which severely inactivate the channel. SR Cl- channels can be separated from ryanodine receptors based on reversal potential. The polarity of channels incorporated into the bilayer is constant, in the majority of cases. The myoplasmic end of the receptor faces into the cis solution, and the intravesicular end faces into the trans solution. Polarity can be easily confirmed by the cis-activation of channels by adenosine triphosphate (ATP) and micromolar Ca2+, which are myoplasmic activators of ryanodine receptors.

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