Altered Cross Bridge Kinetics in Skeletal Myofibrils from NEBDex55, a Novel Mouse Model of Nebulin-Based Nemaline Myopathy

Nemaline Myopathy is the most common non-dystrophic congenital myopathy, clinically characterized by muscle weakness. The disease is associated with mutations in the nebulin gene and the nebulin-based disease is referred to as NEM2. Recent work on skinned muscle fibres from NEM2 patients revealed remarkable phenotypic similarities to fibres from nebulin KO mice (Ottenheijm et al, 2012). Here we investigated mechanics and kinetics of single myofibrils from a novel NEM2 mouse model (NEB Δex55) that mimics a deletion in the nebulin gene found in a large group of NEM2 patients. We used rapid solution switching (Tesi et al.,2002) to compare maximal tension and kinetics of contraction and relaxation of myofibrils isolated from frozen skeletal muscles (tibialis cranialis of neonatal mice) of WT and NEB Δex55 mice. Myofibrils, mounted in a force recording apparatus (15 °C), were maximally Ca2+-activated (pCa 4.5) and fully relaxed (pCa 9.0). Maximal isometric tension was markedly reduced in NEB Δex55 mouse myofibrils (49.7±10.6 mN mm−2n=11) compared to WT (135.3±16.9mN mm−2n=9).The rate constant of active tension generation following maximal Ca2+ activation (kACT) was significantly reduced inNEB Δex55 mouse myofibrils (1.46±0.07s−1) compared to WT (2.75±0.27 s−1). Force relaxation kinetics was remarkably faster in NEB Δex55 mouse myofibrils than in WT, evidence that the apparent rate with which cross-bridges leave the force generating states is accelerated in the NEB Δex55sarcomeres. Reduction of the rate with which cross-bridges enter force generating states and of cross bridge dissociation can markedly contribute to reducing maximal tension. This is expected to increase the energetic cost of tension generation of the NEB Δex55sarcomeres. Results suggest that nebulin plays a significant role in contraction regulation and that altered cross bridge kinetics contribute to NEM2 pathogenesis