Aminopeptidase N mediates the utilization of the GSH precursor CysGly by cultured neurons

Neurons in culture rely on the supply of exogenous cysteine for their gluta thione synthesis. After application of cysteine to neuron-rich primary cult ures, the glutathione content was doubled after a 4-hr incubation. The dipe ptide cysteinylglycine (CysGly) was able to substitute for cysteine as exog enous glutathione precursor. In kidneys, the ectopeptidase aminopeptidase N (ApN) has been reported to hydrolyze CysGly. Expression of mRNA of ApN in rat brain and cultured rat neurons was demonstrated by reverse transcriptas e polymerase chain reaction and sequencing of the cDNA fragment obtained. I n addition, the presence of ApN protein in cultured neurons was demonstrate d by its immunocytochemical localization. In the presence of an activity-in hibiting antiserum against ApN the utilization of CysGly as neuronal glutat hione precursor was completely prevented, whereas that of cysteine plus gly cine was not affected. The data presented demonstrates that cultured rat ne urons express ApN and that this ectopeptidase participates in the utilizati on of CysGly as precursor for neuronal glutathione. (C) 2001 Wiley-Liss, In c.