Isotonic contractile impairment due to genetic CLC‐1 chloride channel deficiency in myotonic mouse diaphragm muscle

Abstract

The hallmark of genetic CLC-1 chloride channel deficiency in myotonic humans, goats and mice is delayed muscle relaxation resulting from persistent electrical discharges. In addition to the ion channel defect, muscles from myotonic humans and mice also have major changes in fibre type and myosin isoform composition, but the extent to which this affects isometric contractions remains controversial. Many muscles, including the diaphragm, shorten considerably during normal activities, but shortening contractions have never been assessed in myotonic muscle. The present study tested the hypothesis that CLC-1 deficiency leads to an impairment of muscle isotonic contractile performance. This was tested in vitro on diaphragm muscle from SWR/JClcn1 adr-mto /J myotonic mice. The CLC-1-deficient muscle demonstrated delayed relaxation, as expected. During the contractile phase, there were significant reductions in power and work across a number of stimulation frequencies and loads in CLC-1-deficient compared with normal muscle, the magnitude of which in many instances exceeded 50%. Reductions in shortening and velocity of shortening occurred, and were more pronounced when calculated as a function of absolute than relative load. However, the maximal unloaded shortening velocity calculated from Hill's equation was not altered significantly. The impaired isotonic contractile performance of CLC-1-deficient muscle persisted during fatigue-inducing stimulation. These data indicate that genetic CLC-1 chloride channel deficiency in mice not only produces myotonia but also substantially worsens the isotonic contractile performance of diaphragm muscle.

Improvement of diaphragm and limb muscle isotonic contractile performance by K+ channel blockade

“…The choice of using peak force during 20 Hz stimulation rather than tetanic force to define maximum load was based on two considerations. First, it is consistent with the approach used in our previous studies of muscle isotonic contractile properties [30,31]. Second, the present study was performed in the context of functional electrical stimulation, and thus it is more meaningful to base loads on force produced during the frequency at which the muscle will be stimulated.…”

Fatigue-inducing stimulation resolves myotonia in a drug-induced model

“…The latter issue has been examined to a limited extent in a previous study from our lab in which we examined isotonic contractions of diaphragm muscle from genetically myotonic mice [19]. Diaphragm from these mice had lesser degrees of myotonia than seen in the diaphragm of the present study.…”

“…Peak force was defined as the largest value that occurred during the portion of the contraction while the muscle was being electrically stimulated (and thus did not include mechanical myotonia even if mechanical myotonia exceeded force during active electrical stimulation). These force values were normalized relative to twitch force before drug addition to factor out the effects of variability among muscle samples in absolute force related to variability in animal size and muscle sample size, similar to previous studies of a similar nature [11,15,19,22]. Contraction time was defined as the time from the onset of force production to the top of the first peak of the non-fused contraction; values for contraction time are not presented in those instances in which a distinct first peak was not discernable (which was the case for 50 Hz stimulation for the diaphragm).…”

“…Muscle from humans and animals with myotonia congenita has physiological, structural and biochemical alterations which transcend the loss of CLC-1 channels, including hypertrophy (goats, most humans) or atrophy (mice), alterations in fiber type composition (in particular loss of type IIB fibers, seen in most species with myotonia congenita), and altered amounts of parvalbumin [1,5,7,9,11-13,17,18]. These downstream changes may contribute to the complex abnormalities in contractile performance found in CLC-1 deficient muscle, in particular changes that occur during the contractile phase of the contraction-relaxation cycle, such as impaired force production and alterations in rate of fatigue during isometric contractions [19]. …”

Risk of Myopathy in Patients in Therapy with Statins: Identification of Biological Markers in a Pilot Study

“…In addition, toxicity studies showed that statin therapy plus eccentric exercise stress enable muscle instability, triggering stronger activation of intracellular proteolytic cascades (Urso et al, 2005). At this regard it is known that the mechanical stress during exercise may aggravate ClC-1 function (van Lunteren et al, 2007; Camerino et al, 2014b; Cozzoli et al, 2014) and that the activation of Ubiquitin Ligase Complex specifically mediates the poly-ubiquitination and degradation of ClC-1 (Chen et al, 2015). At the light of these findings, it would be also important to verify if strenuous exercise and/or genetic mutations of ClC-1, may potentiate adverse effects of statins, as already observed for metabolic myopathies (Tay et al, 2008).…”