Increased iNOS and Nitrosative Stress in Dopaminergic Neurons of MDMA-Exposed Rats

MDMA (ecstasy) neurotoxicity may involve nitrosative stress driven by the enzyme iNOS rather than by NOX enzymes that produce reactive oxygen species. In rats given MDMA, iNOS expression increased, and cells positive for 3-nitrotyrosine (a marker of nitrosative damage) rose in the frontal cortex. No changes occurred in NOX2, NOX1, or NOX4 immunoreactivity or in the oxidative stress marker 8OHdG. MDMA and nitrosative markers colocalized with dopamine-transporter-positive cells, which were neurons, not microglia or astrocytes. The findings indicate that iNOS-derived nitrosative stress, not NOX enzymes, likely contributes to MDMA-induced neurotoxicity.

Study at a glance

Abstract

Several mechanisms underlying 3,4-Methylenedioxy-N-methylamphetamine (MDMA) neurotoxicity have been proposed, including neurochemical alterations and excitotoxicity mediated by reactive oxygen species (ROS), nitric oxide (NO), and reactive nitrogen species (RNS). However, ROS, NO, and RNS sources in the brain are not fully known. We aimed to investigate possible alterations in the expression of the ROS producer NOX enzymes (NOX2, NOX1, and NOX4), NO generators (iNOS, eNOS, and nNOS), markers of oxidative (8-hydroxy-2′-deoxyguanosine, 8OHdG), and nitrosative (3-nitrotyrosine, NT) stress, as well as the colocalization between cells positive for the dopamine transporter (DT1) and cells expressing the neuronal nuclei (NeuN) marker, in the frontal cortex of rats receiving saline or MDMA, sacrificed 6 h, 16 h, or 24 h after its administration. MDMA did not affect NOX2, NOX1, and NOX4 immunoreactivity, whereas iNOS expression was enhanced. The number of NT-positive cells was increased in MDMA-exposed animals, whereas no differences were detected in 8OHdG expression among experimental groups. MDMA and NT markers colocalized with DT1 positive cells. DT1 immunostaining was found in NeuN-positive stained cells. Virtually no colocalization was observed with microglia and astrocytes. Moreover, MDMA immunostaining was not found in NOX2-positive cells. Our results suggest that iNOS-derived nitrosative stress, but not NOX enzymes, may have a crucial role in the pathogenesis of MDMA-induced neurotoxicity, highlighting the specificity of different enzymatic systems in the development of neuropathological alterations induced by the abuse of this psychoactive compound.