Abstract
Small Ankyrins (sAnk1) are muscle-specific isoforms generated by the Ank1 gene that participate in the organization of the sarcoplasmic reticulum (SR) of striated muscles. Accordingly, the volume of SR tubules localized around the myofibrils is strongly reduced in skeletal muscle fibers of 4- and 10-month-old sAnk1 knockout (KO) mice, while additional structural alterations only develop with aging. To verify whether the lack of sAnk1 also alters intracellular Ca2+ handling, cytosolic Ca2+ levels were analyzed in stimulated skeletal muscle fibers from 4- and 10-month-old sAnk1 KO mice. The SR Ca2+ content was reduced in sAnk1 KO mice regardless of age. The amplitude of the Ca2+ transients induced by depolarizing pulses was decreased in myofibers of sAnk1 KO with respect to wild type (WT) fibers, while their voltage dependence was not affected. Furthermore, analysis of spontaneous Ca2+ release events (sparks) on saponin-permeabilized muscle fibers indicated that the frequency of sparks was significantly lower in fibers from 4-month-old KO mice compared to WT. Furthermore, both the amplitude and spatial spread of sparks were significantly smaller in muscle fibers from both 4- and 10-month-old KO mice compared to WT. These data suggest that the absence of sAnk1 results in an impairment of SR Ca2+ release, likely as a consequence of a decreased Ca2+ store due to the reduction of the SR volume in sAnk1 KO muscle fibers.
Highlights
The sarcoplasmic reticulum (SR) of skeletal muscle fibers is a convoluted organelle consisting of a network of membrane-limited tubules where Ca2+ is stored
We measured the resting [Ca2+]i in unstimulated Fura-2 loaded flexor digitorum brevis (FDB) fibers from sAnk1 KO and wild type (WT) mice. These measurements indicated that the resting Ca2+ levels were not significantly different in both 4- and 10-month-old FDB fibers from sAnk1 KO mice compared to WT mice (Figure 1C)
Membrane, binds to the sarcomeric giant protein obscurin. This protein–protein interaction is necessary for the accurate positioning of the SR tubules, where Ca2+ is stored, around the myofibrils, in order to regulate muscle contraction
Summary
The sarcoplasmic reticulum (SR) of skeletal muscle fibers is a convoluted organelle consisting of a network of membrane-limited tubules where Ca2+ is stored. The membrane of the terminal cisternae facing the T-tubule accommodates the ryanodine receptor (RyR1) Ca2+ release channels, and additional SR proteins, including triadin, junctin, and calsequestrin, which, together with RyR1 channels, participate in the mechanisms of excitation–contraction (e–c) coupling. This mechanism transduces depolarization-induced activation of the dihydropyridine receptor (DHPR), the voltage sensor on T-tubules, into a release of Ca2+ from the SR to allow muscle contraction [1]. Alterations of this mechanism are associated with different skeletal muscle diseases in humans [2,3,4,5], including Central and Multi Mini Core Disease, and centronuclear myopathies associated with mutations in RyR and myotubularin, respectively [6,7,8]
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