Objective—To investigate whether protein kinase C
(PKC) isoforms are expressed in equine skeletal muscle
and determine their distribution in various types of
fibers by use of immunofluorescence microscopy.
Animals—5 healthy adult Dutch Warmblood horses.
Procedure—In each horse, 2 biopsy specimens were
obtained from the vastus lateralis muscle.
Cryosections of equine muscle were stained with
PKC isoform (α, β1, β2, δ, ξ, or ζ)-specific polyclonal
antibodies and examined by use of a fluorescence
microscope. Homogenized muscle samples were
evaluated via western blot analysis.
Results—The PKC α, β1, β2, δ, ξ, and ζ isoforms
were localized within the fibers of equine skeletal
muscle. In addition, PKC α and β2 were detected near
or in the plasma membrane of muscle cells. For some
PKC isoforms, distribution was specific for fiber type.
Staining of cell membranes for PKC α was observed
predominantly in fibers that reacted positively with
myosin heavy chain (MHC)-IIa; PKC δ and ξ staining
were more pronounced in MHC-I-positive fibers. In
contrast, MHC-I negative fibers contained more PKC
ζ than MHC-I-positive fibers. Distribution of PKC β1
was equal among the different fiber types.
Conclusions and Clinical Relevance—Results indicated
that PKC isoforms are expressed in equine
skeletal muscle in a fiber type-specific manner.
Therefore, the involvement of PKC isoforms in signal
transduction in equine skeletal muscle might be
dependent on fiber type. ( Am J Vet Res 2004;
Objective—To investigate the effects of exercise on activation of mitogen-activated protein kinase (MAPK) signaling proteins in horses.
Animals—6 young trained Standardbred geldings.
Procedure—Horses performed a 20-minute bout of exercise on a treadmill at 80% of maximal heart rate. Muscle biopsy specimens were obtained from the vastus lateralis and pectoralis descendens muscles before and after exercise. Amount of expression and intracellular location of phosphospecific MAPK pathway intermediates were determined by use of western blotting and immunofluorescence staining.
Results—Exercise resulted in a significant increase in phosphorylation of p38 pathway intermediates, c-Jun NH2 terminal kinase (JNK), and heat shock protein 27 (HSP27) in the vastus lateralis muscle, whereas no significant changes were found in phosphorylation of extracellular regulated kinase. In the pectoralis descendens muscle, phosphorylation of p38 and HSP27 was significantly increased after exercise. Immunohistochemical analysis revealed fiber-type– specific locations of phosphorylated JNK in type 2a/b intermediate and 2b fibers and phosphorylated p38 in type 1 fibers. Phosphorylated HSP27 was strongly increased after exercise in type 1 and 2a fibers.
Conclusions and Clinical Relevance—The p38 pathway and JNK are activated in the vastus lateralis muscle after a single 20-minute bout of submaximal exercise in trained horses. Phosphorylation of HSP27 as detected in the study reported here is most likely induced through the p38 signaling pathway.
Procedures—Percutaneous biopsy specimens were
obtained from the vastus lateralis, pectoralis descendens,
and triceps brachii muscles. Cryosections were
stained with combinations of GLUT4 and myosin
heavy chain (MHC) specific antibodies or FAT/CD36
and MHC antibodies to assess the fiber specific
expression of GLUT4 and FAT/CD36 in equine skeletal
muscle via indirect immunofluorescent
Results—Immunofluorescent staining revealed that
GLUT4 was predominantly expressed in the cytosol
of fast type 2B fibers of equine skeletal muscle,
although several type 1 fibers in the vastus lateralis
muscle were positive for GLUT4. In all muscle fibers
examined microscopically, FAT/CD36 was strongly
expressed in the sarcolemma and capillaries. Type 1
muscle fibers also expressed small intracellular
amounts of FAT/CD36, but no intracellular FAT/CD36
expression was detected in type 2 fibers.
Conclusions and Clinical Relevance—In equine
skeletal muscle, GLUT4 and FAT/CD36 are expressed
in a fiber type selective manner. ( Am J Vet Res 2004;65:951–956)