Mechanism of verapamil action on wild‐type and slow‐channel mutant human muscle acetylcholine receptor

Abstract

Verapamil, a Ca 2+ channel blocker widely used in clinical practice, also affects the properties of frog and mouse muscle acetylcholine receptor (AChR). Here, we examine the mechanism of action of verapamil on human wild-type and slowchannel mutant muscle AChRs harboring in any subunit a valine-to-alanine mutation of 13¢ residue of the pore-lining M2 transmembrane segment. Verapamil, after a pre-treatment of 0.5-10 s, accelerated the decay of whole-cell or macroscopic outside-out currents within milliseconds of ACh application even at clinically attainable doses. Recordings of unitary events in the cell-attached and outside-out configurations showed that verapamil does not alter single-channel conductance, but reduces channel open probability, by prolonging the dwell time into the closed state for wild-type and all mutant AChR. The duration of channel openings decreased only for the eV265A-AChR, by shortening the longest exponential component of the open-time distribution. These results provide a rationale for the therapeutic use of verapamil in the slow-channel syndrome and emphasize the major role played by e subunit in controlling the functional properties of human muscle AChR, as revealed by the peculiar alterations imparted by mutations in this subunit.

Physiological characterization of human muscle acetylcholine receptors from ALS patients

“…The larger EC 50 observed for ALS AChRs in injected oocytes could be due to a variable proportion of γ-and ε-AChRs in the muscle membranes derived from ALS patients, reflecting the cycles of denervation/reinnervation typical of ALS. Values reported for human γ-or ε-AChR (29,30) are in the same range, as the EC 50 we report for AChR from denervated patients. Whereas it is clear that human myotubes from ALS or denervated patients express the fetal form of the AChR, our biopsied muscle tissues from ALS patients transplanted in oocytes may express mixed γ/ε-containing receptors.…”

Nicotinic acetylcholine receptor and the structural basis of neuromuscular transmission: insights fromTorpedopostsynaptic membranes

“…For example, in oocyte expression studies, mutation of the leucine (position 9′) to serine or threonine, in any of the subunits, increases the opening sensitivity of the channel by roughly equal amounts (Filatov & White, 1995; Labarca et al 1995). Mutation of the valine (position 13′) to alanine, in any of the subunits, gives similar results (Moriconi et al 2010). Likewise, a small molecule that binds in the interstitial space behind α γ M2 should be able open the channel using contraction around the bound ligand, rather than tilting of a subunit, to destabilize the gate.…”

Fluoxetine prevents acetylcholine-induced excitotoxicity blocking human endplate acetylcholine receptor

“…However, the drop in channel opening frequency, together with (and possibly more than) the reduction of channel open duration, causes the marked decrease in ACh evoked charge influx, which in turn likely underlies prevention of cell death. Neither verapamil nor salbutamol protects cells expressing ε WT ‐AChR against ACh‐induced death, but at clinically relevant concentrations salbutamol has little effect on the function of ε WT ‐AChR, and verapamil depresses ACh evoked charge movement less than fluoxetine (compare results in this study and in previous work) …”