Evidence for acidity of prelysosomal autophagic/endocytic vacuoles (amphisomes)

[¹⁴C]Lactose electroinjected into isolated rat hepatocytes is normally autophagocytosed, transferred to lysosomes and degraded by lysosomal β-galactosidase, but at high concentrations of asparagine the transfer is inhibited and lactose accumulates in prelysosomal autophagic/endocytic vacuoles (amphisomes). The accumulation can be prevented by addition of yeast, β-galactosidase, which is transferred to the lactose-containing vacuoles by endocytosis. Propylamine, a weak base capable of neutralizing acidic vacuoles, protects autophagocytosed lactose against both endogenous and exogenous, β-galactosidase, suggesting that amphisomes, like lysosomes, have an acidic internal environment.