title: Protein phosphatases but not reactive oxygen species play functional role in acute amphetamine-mediated dopamine release.
abstract: Drug abuse-induced neurodegeneration can be triggered by elevated production of reactive oxygen species (ROS). Involvement of oxidative stress in acute amphetamine (AMPH)-mediated dopamine (DA) release, however, has not been completely understood yet. In order to elucidate the dopaminergic response of PC12 cells to a single dose of 10 μM AMPH, ROS production was measured as related to the extracellular DA level. Due to the spontaneous oxidation of peroxide-sensitive fluorophore 2',7'-dichlorofluorescin diacetate (DCFH-DA) to 2',7'-dichlorofluorescein (DCF), the increase in fluorescence could not be unambiguously attributed to AMPH-triggered ROS production. Based on Amplex Red fluorescence, no ROS production was detected after acute AMPH application. Our data strongly suggest that ROS development was not the main triggering factor for immediate DA release after acute AMPH treatment. On the other hand, AMPH-induced elevation of DA levels in rat brain striatal slices was quenched by the water soluble antioxidant, N-acetylcysteine (NAC) at 10 mM. In this study, we also investigated the contribution of protein phosphatases to the AMPH-induced rat brain striatal dopaminergic response. The experimental protocol, double AMPH challenge was applied for screening the effect of NAC and cantharidin on AMPH-mediated DA release. Here we show that AMPH-mediated DA release increased nearly twofold in striatal rat brain slices pretreated for 30 min with 1000 μM cantharidin, a selective PP1 and PP2A inhibitor. These findings prove the lack of ROS inhibitory action on protein phosphatase activity in acute AMPH-mediated DA efflux.
authors.affiliation: Department of Pharmacology and Toxicology, Faculty of Veterinary Science, Szent István University, Budapest, Hungary. gere.erzsebet@aotk.szie.hu
authors.forename: Erzsebet
authors.initials: E
authors.lastname: Paszti-Gere
authors.forename: Judit
authors.initials: J
authors.lastname: Jakus
mesh_terms: D000661:Amphetamine / Q000494:pharmacology*; D000818:Animals; D002193:Cantharidin / Q000494:pharmacology; D004298:Dopamine / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000737:chemistry; D006861:Hydrogen Peroxide / Q000378:metabolism; D017072:Neostriatum / Q000187:drug effects / Q000378:metabolism; D010078:Oxazines / Q000737:chemistry; D018384:Oxidative Stress / Q000187:drug effects; D016716:PC12 Cells; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D005456:Fluorescent Dyes; D010078:Oxazines; D017382:Reactive Oxygen Species; C470430:Amplex Red; D006861:Hydrogen Peroxide; D000661:Amphetamine; D010749:Phosphoprotein Phosphatases; D002193:Cantharidin; D004298:Dopamine