Abstract
M-channel inhibitors, especially XE991, are increasingly being used in animal experiments. However, insufficient characterization of XE991 at times confounds the interpretation of results when using this compound. Here, we demonstrate that XE991 and linopirdine are state-dependent inhibitors that favor the activated-subunit of neuronal Kv7/KCNQ channels. We performed patch clamp experiments on homomeric Kv7.2 or heteromeric Kv7.2/3 channels expressed in Chinese hamster ovary cells in order to characterize XE991 and linopirdine. Both inhibitors were not efficacious around the resting membrane potential of cells in physiological conditions. Inhibition of Kv7.2 and Kv7.2/3 channels by XE991 was closely related with channel activation. When voltage dependence of activation was left-shifted by retigabine or right-shifted by the mutation, Kv7.2(R214D), the shift in half activation voltage proportionally coincided with the shift in the half-effective potential for XE991 inhibition. Inhibition kinetics during XE991 wash-in were facilitated at depolarized potentials. Ten-minute washout of XE991 resulted in ~30% current recovery, most of which was attributed to surface transport of Kv7.2 channels. Linopirdine also exhibited similar inhibition characteristics with the exception of near complete current recovery after washout at depolarized potentials. Inhibition kinetics of both XE991 and linopirdine were not as sensitive to voltages as would be predicted by open channel inhibition. Instead, they were well explained by binding to a single activated subunit. The characteristics of XE991 and linopirdine should be taken into account when these M-channel inhibitors are used in experiments.
- The American Society for Pharmacology and Experimental Therapeutics