Single tottering mutations responsible for the neuropathic phenotype of the P-type calcium channel

Recent genetic and molecular biological analyses have revealed many forms o f inherited channelopathies. Homozygous ataxic mice, tottering (tg) and lea ner (tg(1a)) mice, have mutations in the P/Q-type Ca2+ channel alpha(1A) su bunit gene. Although their clinical phenotypes, histological changes, and l ocations of gene mutations are known, it remains unclear what phenotypes th e mutant Ca2+ channels manifest, or whether the altered channel properties are the primary consequence of the mutations. To address these questions, w e have characterized the electrophysiological properties of Ca2+ channels i n cerebellar Purkinje cells, where the P-type is the dominant Ca2+ channel, dissociated from the normal, tg, and tg(1a) mice, and compared them with t he properties of the wild-type and mutant alpha(1A) channels recombinantly expressed with the alpha(2) and beta subunits in baby hamster kidney cells. The most striking feature of Ca2+ channel currents of mutant Purkinje cell s was a marked reduction in current density, being reduced to similar to 60 and similar to 40% of control in tg and tg(1a) mice, respectively, without changes of cell size. The Ca2+ channel currents in the tg Purkinje cells s howed a relative increase in non-inactivating component in voltage-dependen t inactivation. Besides the same change, those of the tg(1a) mice showed a more distinct change in voltage dependence of activation and inactivation, being shifted in the depolarizing direction by similar to 10 mV, with a bro ader voltage dependence of inactivation. In the recombinant expression syst em, the tg channel with a missense mutation (P601L) and one form of the two possible tg(1a) aberrant splicing products, tg(1a) (short) channel, showed a significant reduction in current density, while the other form of the tg (1a) channels, tg(1a) (long), had a current density comparable to the norma l control. On the other hand, the shift in voltage dependence of activation and inactivation was observed only for the tg(1a) (long) channel. Comparis on of properties of the native and recombinant mutant channels suggests tha t single tottering mutations are directly responsible for the neuropathic p henotypes of reduction in current density and deviations in gating behavior , which lead to neuronal death and cerebellar atrophy.