Expression Of The Embryonic Cav1.1 Splice Variant In Adult Mice Alters Excitation-Contraction Coupling But Does Not Cause Dystrophic Myotonia

CaV1.1e is the calcium channel splice variant of embryonic skeletal muscle. It functions as voltage sensor in EC coupling, but in contrast to the adult CaV1.1a variant it also supports sizeable calcium currents activating at the same membrane potential as SR calcium release. Mis-splicing of CaV1.1 results in elevated levels of the CaV1.1e variant in mature muscle that correlate with the severity of muscle weakness in patients suffering from dystrophic myotonia. Therefore we hypothesize that exclusive expression of CaV1.1e in exon 29 knockout mice will cause muscle weakness. In muscles of wildtype mice expression of CaV1.1e declines after birth. However, two weeks after sciatic nerve resection CaV1.1e transcript levels are upregulated in paralyzed muscles, indicative of an activity-dependent regulation of CaV1.1e splicing. Quantitative RT-PCR analysis demonstrates that exon 29 knockouts express exclusively the CaV1.1e splice variant at levels comparable to total CaV1.1 in wildtype muscle. Combined voltage clamp and fluorescence calcium recordings of isolated muscle fibers from exon 29 knockout mice revealed a substantial influx-dependent component of the calcium transient. This not only alters myoplasmic calcium handling, but is further expected to prolong the muscle action potential and consequently reduce the tetanic fusion frequency. Surprisingly, these severe alterations of the muscle fiber properties were not accompanied by a dystrophic myotonia phenotype in knockout mice. A battery of behavioral tests (home-cage activity, voluntary and forced running, grip strength and rotarod tests) revealed no signs of muscle weakness or altered motor activity compared to wildtype siblings. Thus, development of dystrophic myotonia may require the combinatorial action of several mis-spliced genes. Alternatively, strong compensatory mechanisms may cause the absence of the disease phenotype in exon 29 knockout mice. Support: FWF P23479, W1101, OTKA-NN107765, TAMOP-4.2.2./B.-10/1-2010-0024.