Investigation of mitochondrial inner membrane ion conductance by planar lipid bilayer electrophysiology

Mitochondrial ion channels are proteins of the inner and outer mitochondrial membranes that regulate ion flux and control various cellular processes, including calcium signaling, bioenergetic and metabolic functions, and cell death. Their precise regulation is essential to maintaining normal mitochondrial function and preventing pathological processes. Patch-clamp and planar lipid bilayer electrophysiology techniques have been used to measure ion flow directly across the membrane, thereby revealing the gating kinetics and pharmacological profile of ion channels in real time. Here, we describe a planar lipid bilayer electrophysiology approach for assessing mitochondrial ion channel conductance using mitochondrial inner membrane vesicles (IMVs). The comparative electrophysiology analysis between IMVs and purified mitochondrial proteins, ATP synthase, and the adenine nucleotide translocator (ANT), demonstrates that planar lipid bilayer electrophysiology is a robust tool for biophysical characterization of mitochondrial ion channels using IMVs. This approach is particularly valuable for investigating ion channel properties under controlled yet physiologically relevant conditions and for evaluating the direct modulatory effects of different pharmacological agents.