Pathogenic point mutations in a transmembrane domain of the ε subunit increase the Ca²⁺ permeability of the human endplate ACh receptor

The ε subunit of the human endplate ACh receptor (AChR) is a key determinant of the large fraction of the ACh-evoked current carried by Ca²⁺ ions (P_f). Consequently, missense mutations in the ε subunit are potential targets for altering the P_f of human AChR. In this paper we investigate the effects of two pathogenic point mutations in the M2 transmembrane segment AChR ε subunit, εT264P and εV259F, that cause slow-channel syndromes (SCS). When expressed in GH4C1 cells, the mutant receptors subunits raise Ca²⁺ permeability of the receptors ∼1.5 and ∼2-fold above that of wild-type, to attain P f values of 11.8% (εT264P) and 15.4% (εV259F). The latter value exceeds most P_f values reported to date for ligand-gated ion channels. Consistent with these findings, the biionic Ca²⁺ permeability ratio (P_Ca/P_Cs) of the mutant AChRs is also increased. Upon repetitive stimulation with ACh, the mutant receptors show an enhanced current run-down compared with wild-type, leading to a strong reduction of their function. We propose that the enhanced Ca²⁺ permeability of the mutant receptors overrides the protective effect of desensitization and, together with the prolonged opening events of the AChR channel, is an important determinant of the excitotoxic endplate damage in the SCS.