ULTRASTRUCTURAL OBSERVATIONS OF ISOLATED INTACT AND FRAGMENTED JUNCTIONS OF SKELETAL-MUSCLE BY USE OF TANNIC-ACID MORDANTING

Tannic acid mordanting during fixation of isolated vesicles from [rabbit] skeletal muscle enhanced the resolution of the images. Isolated triadic junctions displayed 2 characteristic features not previously described: a clear gap separated terminal cisternae from transverse tubules; this gap was bridged by a separating array of structures which resembled the feet of intact muscle. When the triad was broken in a French press and subsequently reassembled by joining the 2 organelles, a similar gap was seen but the structure of the feet was less well defined. When the membrane of the triad was extracted by Triton X-100, the junctional region was retained and a similar gap between the 2 organelles could be discerned. The terminal cisternae characteristically displayed a thickening of the cytoplasmic leaflet of the membrane in select areas in which electron-dense material was apposed on the luminal leaflet. This thickened membrane was not observed in longitudinal reticulum or in terminal cisternae regions distal to the electron-dense matter. This thickened leaflet was not invariably associated with the junction; some junctional regions did not display discernible thickening of the membrane. When the triad was treated with KCl, the electron-dense aggregate was dispersed and the thickened leaflet of the terminal cisternae dissipated, whereas the triadic junctional region with its feet remained unchanged. KCl treatment caused dissolution of 3 proteins of MW = 77,000, 43,000 and 38,000. Treatment of Triton-resistant vesicles with KCl caused the loss of electron-dense aggregate but did not otherwise influence the appearance of the junction. A good degree of correlation both qualitatively and in quantitative parameters between the isolated vesicles and the intact muscle was observed.