METHAMPHETAMINE NEUROTOXICITY INVOLVES VACUOLATION OF ENDOCYTIC ORGANELLES AND DOPAMINE-DEPENDENT INTRACELLULAR OXIDATIVE STRESS

Methamphetamine (MA) produces selective degeneration of dopamine (DA) neuron terminals without cell body loss. While excitatory amino acids (EAAs) contribute to MA toxicity, terminal loss is not characteristic of excitotoxic lesions nor is excitotoxicity selective for DA fibers; rather, EAAs may modulate MA-induced DA turnover, suggesting that DA-d ependent events play a key role in MA neurotoxicity. To examine this p ossibility, we used postnatal ventral midbrain DA neuron cultures main tained under continuous EAA blockade. As in vivo, MA caused neurite de generation but minimal cell death. We found that MA is a vacuologenic weak base that induces swelling of endocytic compartments; MA also ind uces blebbing of the plasma membrane. However, these morphological cha nges occurred in MA-treated cultures lacking DA neurons. Therefore, wh ile collapse of endosomal and lysosomal pH gradients and vacuolation m ay contribute to MA neurotoxicity, this does not explain selective DA terminal degeneration. Alternatively, MA could exert its neurotoxic ef fects by collapsing synaptic vesicle proton gradients and redistributi ng DA from synaptic vesicles to the cytoplasm. This could cause the fo rmation of DA-derived free radicals and reactive metabolites. To test whether MA induces oxidative stress within living DA neurons, we used 2,7-dichlorofluorescin diacetate (DCF), an indicator of intracellular hydroperoxide production. MA dramatically increased the number of DCF- labeled cells in ventral midbrain cultures, which contain about 30% DA neurons, but not in nucleus accumbens cultures, which do not contain DA neurons. In the DA neuron cultures, intracellular DDF labeling was localized to axonal varicosities, blebs, and endocytic organelles. The se results suggest that MA redistributes DA from the reducing environm ent within synaptic vesicles to extravesicular oxidizing environments, thus generating oxygen radicals and reactive metabolites within DA ne urons that may trigger selective DA terminal loss.