A new combined triple staining method for neuromusculer junctions and its application to intrinsic laryngeal muscles in man
human intrinsic laryngeal muscle
anti-skeletal fast myosin antibody
A new combined stain for the simultaneous demonstration of motor nerve terminals, cholinesterase and fast myosin at neuromuscular junction (NMJ) s was devised. This method employed dye-staining for cholinesterase, silver-gold impregnation for nerve terminals, and immunostaining used anti-skeletel fast myosin antibody for type 2 muscle fiber. The type 2 muscle fiber stained dark brown with a diaminobenzidine-H2O2 substrate, and the cholinesterase-containing motor endplate stained indigo blue, against which the nerve terminals, from coarse axons to fine terminal branches, stained black and standed out cleary.
One hundred and sixty-five intrinsic laryngeal muscles were obtained from 13 autopsy specimens (including 3 patients with amyotrophic lateral sclerosis (ALS)) and 20 surgical spesimens (half of these were irradiated previously). Using this triple stain, morphological changes of NMJs were studied on each muscle fiber type in human intrinsic laryngeal muscles.
In human intrinsic laryngeal muscles, most of NMJs consisted of one motor endplate and more than one terminal axons. Some muscle fibers possessed more than one NMJ, but this didn't relate to multiple innervation of laryngeal muscles immediately. The NMJ of type 2 muscle fiber had greater motor endplate and much terminal branches than type 1 muscle fiber. Among specialized intrinsic laryngeal muscles, motor endplate length and terminal branching points did not differ. Neuromuscular junction morphology could not account of functional differentiation in intrinsic laryngeal muscles. No significant differences were found between irradiated and nonirradiated groups. This suggested that NMJs were resistant to irradiation. Terminal branching points and segmented motor endplates increased significantly in the ALS specimens, whereas mortor endplate length did not differ. This indicated that nerve sprouting occured in intrinsic laryngeal muscles.
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