Challenges and Insights in Patch-Clamp Studies: From Cell-Attached to Whole-Cell Configurations

The patch-clamp technique is widely regarded as the gold standard in cellular electrophysiology and can be applied in several configurations. In the cell-attached (C-A) mode, it enables the recording of single-channel currents, whereas the whole-cell (W-C) mode allows for the measurement of macroscopic currents, representing the collective activity of many channels. When the recording configuration was switched from C-A to W-C on the same cell, the current amplitude increased dramatically, while action currents (ACs) were completely abolished, indicating a profound alteration in the cell’s electrophysiological response under the new setup. In excitable cells, the occurrence of ACs, representing propagated action potentials, can interfere with C-A single-channel recordings. To address this, a high-K+ solution is typically applied to the bath to suppress the ACs. The inwardly rectifying K+ (Kir), ATP-sensitive K+ (KATP) and large-conductance Ca2+-activated K+ (BKCa) channels are crucial members of the K+ channel family that facilitate the efflux of K+ ions, driven by the K+ electrochemical gradient. These channels are primarily distinguished by their rectification properties and gating kinetics. For instance, KATP channels exhibit a bursting kinetic pattern with inward rectifying property, while BKCa channels display strong outward rectification. Mitoxantrone, which belongs to a class of drugs called anthracenediones, can suppress the activity of Kir channels in differentiated RAW 264.7 cells, with no change in single-channel conductance. The respiratory stimulator GAL-021 acts as a BKCa channel inhibitor, and it suppresses channel activity and shifts the activation curve to the right, suggesting a voltage-dependent blockade that stabilizes the channel in a closed state. GAL-021 does not change the single-channel conductance, indicating it is a gating modifier rather than an open-pore blocker. The functional roles of ion channels are fundamentally important. Correspondingly, the field is transitioning to artificial intelligence for automated single-cell patch-clamp experiments, though brain slice recordings still require manual techniques.