3-Methyladenine (3-MeA) is a specific inhibitor of autophagic/lysosomal protein degradation in isolated rat hepatocytes. At a concentration of 5 mM, 3-MeA inhibits endogenous protein degradation by about 60% without affecting the degradation of an exogenous protein (asialofetuin), protein synthesis, or intracellular ATP levels. The inhibitory effect of 3-MeA is specific to the autophagic/lysosomal pathway, as it is not influenced by amino acids or the lysosomotropic amine propylamine. The depression of intracellular amino acid levels, which occurs with 3-MeA, further supports its specificity. Additionally, 3-MeA suppresses the formation of electron microscopically visible autophagosomes, suggesting that it acts as a specific inhibitor of autophagy. The study highlights the unique selectivity of 3-MeA among purines and its potential utility in understanding the role of autophagy under various physiological conditions.3-Methyladenine (3-MeA) is a specific inhibitor of autophagic/lysosomal protein degradation in isolated rat hepatocytes. At a concentration of 5 mM, 3-MeA inhibits endogenous protein degradation by about 60% without affecting the degradation of an exogenous protein (asialofetuin), protein synthesis, or intracellular ATP levels. The inhibitory effect of 3-MeA is specific to the autophagic/lysosomal pathway, as it is not influenced by amino acids or the lysosomotropic amine propylamine. The depression of intracellular amino acid levels, which occurs with 3-MeA, further supports its specificity. Additionally, 3-MeA suppresses the formation of electron microscopically visible autophagosomes, suggesting that it acts as a specific inhibitor of autophagy. The study highlights the unique selectivity of 3-MeA among purines and its potential utility in understanding the role of autophagy under various physiological conditions.