Abstract
Intracellular proteolysis was measured in primary cultures of newborn rat skeletal (gastrocnemius) and cardiac muscle cells by release to the medium of trichloroacetic acid‐soluble label from cells grown in the presence of ^14^C‐labeled phenylalanine. Exposure of the cultured cells to 10^−7^M dexamethasone for 5 days starting at day 0 of culture resulted in an enhancement of proteolysis in skeletal muscle but not in cardiac muscle cells. Dexamethasone did not affect cell viability measured by release of label from cells preloaded with Na~2~ ^51^CrO~4~, release of creatine phosphokinase, and release of lactic dehydrogenase into the culture medium. Incorporation of ^3^H‐thymidine into the cells increased during the first 3 to 4 days of culture and subsequently decreased, indicating that cell proliferation ceases at that time. When the exposure to dexamethasone was started on day 4 of culture, i.e., at a postmitotic stage, and continued for 4 days, proteolysis was again found to increase in skeletal but not cardiac cells, thereby suggesting that the response to the hormone is independent of the proliferative state of the culture.
Ammonium chloride at a concentration of 10 mM produced a 50% reduction of the basal proteolysis in cultures of skeletal muscle cells and did not affect proteolysis in cardiac muscle cells. Exposure to ammonium chloride did not prevent the dexamethasone‐induced increase of proteolysis in skeletal muscle cells. Serum‐deprivation induced enhanced proteolysis which was not affected by NH~4~Cl in both cell types. It is concluded that the differential responses of the two cultures to dexamethasone reflects the sparing of heart proteins and concomitant wasting of skeletal muscle proteins by glucocorticoid hormones in vivo, and that the enhancement of proteolysis by the glucocorticoid hormone or by serum‐deprivation is not sensitive to the lysosomotropic agent NH~4~Cl. Thus, while a lysosomal‐autophagic enzyme system is responsible for almost half of the basal proteolysis, the accelerated proteolysis occurs via ammonium chloride‐insensitive enzymes.