Elevated lysosomal pH in neuronal ceroid lipofuscinoses (NCLs)

We report here the intracellular (pH(i)) and lysosomal pH in fibroblasts of six forms of neuronal ceroid lipofuscinoses (NCLs). Acid extrusion rate an d pH(i) values were measured by the membrane-permeant acetoxymethyl ester o f the indicator dye, 2',7'-bis(carboxyethyl)-5-(and-6)-carboxyfluorescein ( BCECF) and lysosomal pH by a spectrofluorometric assay utilizing a novel ac idotropic probe, Lysosensor yellow/blue. Intracellular pH was normal in all NCLs. Elevated lysosomal pH was detected in all NCL forms except CLN2 and CLN8. Elevated pH most probably disturbs the catalytic activity of lysosome s and is one important factor in explaining accumulation of ceroid and lipo fuscin-like autofluorescent lipopigments characteristic of NCLs. Using the novel spectrofluorometric assay introduced in this study provides a fast an d repeatable technique to measure intralysosomal pH from cell suspensions.