This protocol provides a detailed method for studying voltage- and ligand-dependent activation of ion channels expressed in Xenopus oocytes using patch-clamp techniques.
Enables precise patch-clamp control of voltage and intracellular milieu in oocytes; leaves open adoption in cardiovascular ion channel studies pending validation.
The Xenopus oocyte expression system is ideal for electrophysiological characterization of voltage-dependent and ligand-dependent ion channels because of its relatively low background of endogenous channels and the large size of the cell. Here, we present a protocol to study voltage- and ligand-dependent activation of ion channels expressed in Xenopus oocytes using patch-clamp techniques designed to control both the membrane voltage and the intracellular solution. In this protocol, the large conductance voltage- and Ca 2+ -activated K + (BK) channel is studied as an example. After injection of BK channel mRNA, oocytes are incubated for 2–7 d at 18°C. Inside-out membrane patches containing single or multiple BK channels are excised with perfusion of different solutions during recording. The protocol can be used to study structure–function relations for ion channels and neurotransmitter receptors.
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Zhang et al. (2018) studied this question.
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