Skeletal muscle Na currents in mice heterozygous for Six5 deficiency

Abstract

Myotonic dystrophy results from a trinucleotide repeat expansion between the myotonic dystrophy protein kinase gene (Dmpk), which encodes a serine-threonine protein kinase, and the Six5 gene, which encodes a homeodomain protein. The disease is characterized by late bursts of skeletal muscle Na channel openings, and this is recapitulated in Dmpk -/- and Dmpk +/- murine skeletal muscle. To test whether deficiency of the nearby Six5 gene also affected Na channel gating in murine skeletal muscle, we measured Na currents from cell-attached patches in Six5 +/- mice and age-matched wild-type and Dmpk +/- mice. Late bursts of Na channel activity were defined as an opening probability >10% measured from 10 to 110 ms after depolarization. There was no significant difference in the occurrence of late Na channel bursts in wild-type and Six5 +/- muscle, whereas in Dmpk +/- muscle there was greater than fivefold increase in late bursts (P < 0.001). Compared with wild-type mice, Na current amplitude was unchanged in Six5 +/- muscle, whereas in Dmpk +/- muscle it was 36% reduced (P < 0.05). Thus, since Six5 +/- mice do not exhibit the Na channel gating abnormality of Dmpk deficiency, we conclude that Six5 deficiency does not contribute to the Na channel gating abnormality seen in dystrophia myotonica patients.

Abnormal Na channel gating in murine cardiac myocytes deficient in myotonic dystrophy protein kinase

“…The DMPKϪ/Ϫ 129SV mouse model that we studied has been described previously (5,20,25,(32)(33)(34). Homozygous DmpkϪ/Ϫ and heterozygous Dmpkϩ/Ϫ mice were studied at Ͼ60 wk of age, when the Na channel phenotype in skeletal muscle is most profound.…”

“…Our techniques for recording membrane potentials and Na currents from isolated mammalian muscle cells have been reported (20,25). Briefly, recordings were made at room temperature (23 Ϯ 0.1°C) using an amplifier (Axopatch model 200A; Axon Instruments, Foster City, CA) and pCLAMP (Axon) hardware and software.…”

“…If this were the case, then both heterozygous and homozygous mutant animals would show a similar phenotype if the Dmpk threshold level required for normal function is more than 50% of the wild-type levels. Interestingly, mice partially deficient in Six5, the gene upstream of Dmpk, exhibit a reduction of Dmpk mRNA levels to 75% of normal (35), and these mice do not exhibit the skeletal muscle Na channel defect characteristic of Dmpk deficiency (20). The effects of partial deficiency of Six5 on cardiac Na channels have not been reported, but if effects of Six5 deficiency on cardiac and skeletal muscle Na channels are similar, this observation suggests that the threshold of Dmpk deficiency required to produce the Na channel defect is between 75% and 50% of normal.…”

Transgenic mouse models for myotonic dystrophy type 1 (DM1)

“…Remarkably, also in Dmpk +/-mice a first degree heart block was observed, which is surprisingly similar to what is observed in DM1 patients. Furthermore, in skeletal muscle cells isolated from Dmpk -/-and Dmpk +/-mice an abnormal Na + channel opening was observed (Mounsey et al, 2000;Mistry et al, 2001;Reddy et al, 2002). This feature was reminiscent of abnormal Na + channel gating found in muscle biopsies from DM1 patients (Franke et al, 1990) and may contribute to muscle hyperexcitability.…”

“…Although Six5 is expressed in (developing) skeletal muscle and is involved in muscle development in Drosophila (see above), no abnormal skeletal muscle function in Six5 -/-mice was observed. In addition, Six5 +/-mice did not show the Na + channel gating abnormality of Dmpk +/-mice (Mistry et al, 2001). Six5 being a transcription factor, both knock-out mouse lines were screened for down-stream effects of Six5 loss, in particular on the transcript levels of Atp1a1, encoding the Na + /K + ATPase ·1 subunit.…”