The use and effects of vital fluorescent dyes: observation of motor nerve terminals and satellite cells in living frog muscles

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Abstract
Several different fluorescent mitochondrial dyes were tested as vital stains for motor nerve terminals and other cells in frog skeletal muscles. It was found that 3,3′diethyloxadicarbocyanine iodide and 4-(4-diethylaminostyryl)-N-methylpyridinium iodide were most useful. Both dyes labelled motor nerve terminals with high reliability. Electrophysiological and morphological control experiments showed that these dyes could be used to repeatedly observe neuromuscular junctions in living animals without affecting synaptic growth or remodelling. The importance of appropriate controls was emphasized by the finding that illumination, if excessively intense or prolonged, can cause physiological and structural damage to nerve terminals. Additional observations indicated that these dyes may be useful for determining the mitochondrial content, and therefore oxidative capacity, of living muscle fibres. It was also found that the fluorescent dyes labelled cells identified as muscle satellite cells, and that these myoblast precursors could be visualized in fixed whole mounts with a nitroblue tetrazolium stain. Both methods were used to study reactive cells that were closely associated with muscle fibres in lesioned muscles. Mitochondrial dyes also labelled the microvasculature, associated axons and other cells.