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Journal of Neuroscience, Vol 11, 1231-1242, Copyright © 1991 by Society for Neuroscience
Metabolic and contractile protein expression in developing rat diaphragm muscle
AM Kelly, BW Rosser, R Hoffman, RA Panettieri, S Schiaffino, NA Rubinstein and PM Nemeth
Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia 19104.
Progressive changes in myosin isozyme expression and in energy- generating
enzyme activities were followed in the diaphragm and, for comparison, in
axial and appendicular muscles of rats from 18 d gestation to maturity.
Native myosins were characterized by pyrophosphate gel electrophoresis.
Myosin heavy-chain (MHC) isozymes were measured with ELISA using monoclonal
antibodies and were localized by immunocytochemistry. RNA transcripts for
the MHCs were demonstrated on Northern blots and by RNase protection
assays. Quantitative activities of malate dehydrogenase (MDH),
beta-hydroxyacyl CoA dehydrogenase (beta OAC), 1-phosphofructokinase (PFK),
lactate dehydrogenase (LDH), creatine kinase (CK), and adenylokinase (AK)
were measured in muscle homogenates and in individual fibers by
fluorometric pyridine nucleotide-dependent assays. Compared to limb
muscles, expression of neonatal myosin in the diaphragm is precocious.
Neonatal MHC mRNA is prominent in the diaphragm at 19 d gestation, and
neonatal myosin is the major MHC isoform present at birth. Slow and fast
IIa MHCs are also present at birth. Transcripts for IIa MHC are detectable
in the diaphragm at 21 d gestation and are upregulated at birth. Comparable
signal for IIa MHC mRNA is not found in the gastrocnemius until 10 d
postpartum. Adult fast IIb MHC mRNA was detected only as a faint signal at
30-40 d in the diaphragm and then disappeared. Results indicate that a
separate phenotype, the IIx type, matures late in diaphragmatic
development. The activities of enzymes representing all of the major energy
pathways are higher in the fetal diaphragm than in the fetal hindlimb
muscles. For example, beta OAC had sixfold higher activity in the diaphragm
than in the extensor digitorum longus (EDL) muscle at birth, activity in
the diaphragm than in the extensor digitorum longus (EDL) muscle at birth.
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