Article

Adherent Primary Cultures of Mouse Intercostal Muscle Fibers for Isolated Fiber Studies

Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, 108 N. Greene Street, Baltimore, MD 21201, USA.
BioMed Research International (Impact Factor: 2.71). 08/2011; 2011:393740. DOI: 10.1155/2011/393740
Source: PubMed

ABSTRACT Primary culture models of single adult skeletal muscle fibers dissociated from locomotor muscles adhered to glass coverslips are routine and allow monitoring of functional processes in living cultured fibers. To date, such isolated fiber cultures have not been established for respiratory muscles, despite the fact that dysfunction of core respiratory muscles leading to respiratory arrest is the most common cause of death in many muscular diseases. Here we present the first description of an adherent culture system for single adult intercostal muscle fibers from the adult mouse. This system allows for monitoring functional properties of these living muscle fibers in culture with or without electrical field stimulation to drive muscle fiber contraction at physiological or pathological respiratory firing patterns. We also provide initial characterization of these fibers, demonstrating several common techniques in this new model system in the context of the established Flexor Digitorum Brevis muscle primary culture model.

Download full-text

Full-text

Available from: Erick O Hernández-Ochoa, Jul 07, 2015
0 Followers
 · 
152 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The NFATc transcription factor family is responsible for coupling cytoplasmic calcium signals to transcription programs in a wide variety of cell types. In skeletal muscle, these transcription factors control the fiber type in response to muscle activity. This excitation-transcription (E-T) coupling permits functional adaptation of muscle according to use. The activity dependence of these transcription programs is sensitive to the firing patterns of the muscle, not merely the period of activity, enabling a nuanced adaptation to various functional tasks. Isolated skeletal muscle fibers expressing exogenous fluorescent NFATc1 were studied by confocal microscopy under stimulation both with and without pharmacological inhibitors. Western blots of whole muscle lysates were also used. This study investigates the activity dependent response of NFATc1 skeletal muscle fibers cultured from mice, comparing fibers of respiratory origin to muscles responsible for limb locomotion. Using patterns of stimulation known to strongly activate NFATc1 in the commonly cultured flexor digitorum brevis and soleus muscles, we have observed significant deactivation of NFATc1 in cultured intercostal muscle fibers. This effect is at least partially dependent on the action of JNK and CaMKII in intercostal fibers. Our findings highlight the role of lineage in the NFAT pathway, showing that the respiratory intercostal muscle fibers decode similar E-T coupling signals into NFAT transcriptional programs in a different manner from the more commonly studied locomotor muscles of the limbs.
    01/2014; 4(1):1. DOI:10.1186/2044-5040-4-1