IMMUNOGOLD ULTRASTRUCTURAL-LOCALIZATION OF CALPASTATIN, THE CALPAIN INHIBITOR, IN RABBIT SKELETAL-MUSCLE

The ultrastructural localization of calpastatin, the endogenous inhibi tor of the neutral calcium-dependent proteases (calpains), was investi gated in rabbit skeletal muscle fibers using a polyclonal antibody aga inst the 34 kDa form of the inhibitor isolated from rabbit. Quantitati ve studies by pre- and postembedding immunogold techniques revealed th at the distribution pattern of the specific immunoreactivity included: 1) the sarcolemma with the adjacent cytoplasm (about 1 mum wide); 2) the myofibrils; 3) the mitochondria and 4) the nuclei (condensed as we ll as extended chromatin). Other cell substructures, such as lysosomes and the intermyofibrillar cytoplasm, were substantially devoid of imm unoreactivity. Furthermore, in accordance to previous light microscope immunohistochemical experiments, an extracellular (endomysial) locali zation of specific immunoreactivity was confirmed. These results favou r the view, which is also supported by a series of biochemical evidenc es, that calpastatin in rabbit skeletal muscle is present in cell stru ctures also containing calpains and/or their putative substrates. The above multiple patterns of distribution also suggest that the muscular calpain-calpastatin system in skeletal muscle fibers may play differe nt physiological roles in the various subcellular compartments.