Characterization and Localization of Acid Hydrolase Activity in the Synaptosomal Fraction From Rat Cerebral Cortex

Synaptosomes were prepared from the cerebral cortex of adult rats by a rapid technique of centrifugation in a Ficoll‐sucrose discontinuous gradient. The synaptosomal fraction contained 40 per cent of the total gradient activity of acid α‐naphthyl phosphatase (EC 3.1.3.2). Quantitative electron microscopy of this fraction revealed rare, typical, extrasynaptosomal dense body lysosomes. pH‐activity profiles of free and Triton X‐100 (total) activities were prepared for α‐naphthyl phosphatase, β‐glucuronidase (EC 3.2.1.31), β‐galactosidase (EC 3.2.1.23), arylsulfatase (EC 3.1.6.1) and N‐acetylglucosaminidase (EC 3.2.1.30). The ratios of total to free activity varied in the order: arylsulfatase > β‐galactosidase > β‐glucuronidase > N‐acetylglucosaminidase > acid phosphohydrolase. Incubation of synaptosomal fractions at pH 5 and 37°C produced significant activation of β‐galactosidase and N‐acetylglucosaminidase but no activation of cryptic lactate dehydrogenase (EC 1.1.1.27). Hyposmotic suspension and subfractionation of the synaptosomal fraction produced considerable solubilization of lactate dehydrogenase, arylsulfatase and β‐galactosidase but only partial liberation of α‐naphthyl phosphatase, the remainder being associated with synaptosomal membrane fragments. Incomplete equilibrium sedimentation of synaptosomes in a continuous sucrose gradient (0·55‐1·5 M) provided a broad lactate dehydrogenase and Na + K ATPase (EC 3.6.1.4) peak (peak I) at low sucrose densities. β‐Glucuronidase, β‐glucosidase and α‐naphthyl phosphatase were significantly present in peak I. Conversely, N‐acetylglucosaminidase, arylsulphatase and β‐galactosidase were predominantly located in denser particles sedimenting through 1·2 M sucrose (peak II). Electron microscopy confirmed the heterogeneity of this second peak and the presence of numerous extrasynapto‐somal dense body lysosomes.