New aspects of the location of neuronal nitric oxide synthase in the skeletal muscle: a light and electron microscopic study.
The action of nitric oxide (NO) synthesized by NO synthases (NOS) is spatially restricted. Hence, the intracellular location of NOS might play an important role for the functional interactions of NO with its target molecules. In the skeletal muscle the neuronal NOS (nNOS) is considered to be the predominant isoform expressed as a muscle specific elongated splice variant. There are only a few and highly discrepant reports of the subcellular distribution of nNOS, which prompted us to re-examine the distribution of nNOS in the skeletal muscle of rat and mouse applying immunocytochemistry and NADPH-diaphorase (NADPH-d) histochemistry. Light microscopically, the sarcolemma, areas beneath the sarcolemma, areas around the nuclei, and the cross striation were labeled by antibodies and by the NADPH-d reaction as well. Ultrastructurally, nNOS visualized immunocytochemically or by the histochemical BSPT-reaction, was associated discretely with extrajunctional portions of the sarcolemma. Both reaction products were additionally observed in the vicinity of endoplasmic reticulum and mitochondria, or associated with their outer membranes. In the neuromuscular junction (NMJ)-region NOS was localized to the cytoplasm of nerve terminals and terminal Schwann cells. In contrast to the commonly accepted assumption, the enzyme was found in association with the presynaptic, and not with the postsynaptic membrane. Cytosolic NADPH-d was exhibited especially between mitochondria accumulated in the postsynaptic region of the NMJ. Surprisingly, in nNOS-/--mice the skeletal muscle showed patterns of significant nNOS-immunoreactivity and NADPH-d activity possibly due to alternative nNOS-splice isoforms, which might be up-regulated to compensate for decreased NO formation.
Rothe
Fritz
F
Institute of Medical Neurobiology, Otto-von-Guericke University of Magdeburg, Leipziger Strasse 44, 39120 Magdeburg, Germany. fritz.rothe@medizin.uni-magdeburg.de
Langnaese
Kristina
K
Wolf
Gerald
G
D000818:Animals; D005455:Fluorescent Antibody Technique; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008854:Microscopy, Electron; D016253:Microscopy, Immunoelectron; D018485:Muscle Fibers, Skeletal / Q000378:metabolism; D018482:Muscle, Skeletal / Q000201:enzymology / Q000254:growth & development; D009252:NADPH Dehydrogenase / Q000032:analysis; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000235:genetics; D009469:Neuromuscular Junction / Q000201:enzymology / Q000648:ultrastructure; D019001:Nitric Oxide Synthase / Q000032:analysis* / Q000235:genetics; D052248:Nitric Oxide Synthase Type I; D051381:Rats; D017208:Rats, Wistar; D012508:Sarcolemma / Q000378:metabolism
D009419:Nerve Tissue Proteins; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; C496321:Nos1 protein, mouse; C496322:Nos1 protein, rat; D009252:NADPH Dehydrogenase