Dopamine Cell Loss within the Nigrostriatal Pathway Due to Oxidative Stress from Chronic Methylphenidate

• Main Authors: Ketchem, Shannon, Ensley, Tucker, Oakes, Hannah, Pond, Brooks B.
• Format: Others
• Published: Digital Commons @ East Tennessee State University 2018
• Subjects: Methylphenidate; Oxidative Stress; MPTP; Parkinsons; Other Pharmacy and Pharmaceutical Sciences
• Online Access: https://dc.etsu.edu/asrf/2018/schedule/104; https://dc.etsu.edu/cgi/viewcontent.cgi?article=1166&context=asrf

Attention deficit hyperactivity disorder (ADHD) is a neurobehavioral disorder that affects 11% of children in the US alo­ne. Methylphenidate (MPH) is the most commonly prescribed drug for the treatment of ADHD. Given the fact that ADHD symptoms persist in up to 50% of patients, many children receive MPH from childhood to early adulthood. Unfortunately, most of the scientific literature focuses on the short-term consequences of MPH, even though individuals are taking MPH for many years. MPH acts by blocking dopamine (DA) transporters and norepinephrine transporters, preventing the reuptake of these catecholamines following release. Previous research has shown that long-term exposure to MPH causes dopaminergic neurons within the nigrostriatal pathway to be more sensitive to the Parkinsonian toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We hypothesize that oxidative stress caused by the spontaneous oxidation of the excess DA in the synaptic cleft is what’s rendering dopaminergic neurons within the nigrostriatal pathway to be more sensitive to MPTP. Adolescent male Swiss-Webster mice were divided into three cohorts and administered either saline (control), 1 mg/kg MPH (normal dose) or 10 mg/kg (abusive dose) via intraperitoneal (IP) injections for 12 weeks. Mice were injected twice daily, Monday through Friday, mimicking a school-week dosing schedule. After 12 weeks, all animals received a drug washout period of 7 days. Then, half of each cohort was treated with MPTP (4 x 20mg/kg, every 2 hours), while the other half was administered 4 injections of sterile saline. Seven days after MPTP or saline treatment, the mice were sacrificed, brains were removed, and the substantia nigra (SN) and striatum (STR) were collected. Oxidative stress related to increased DA levels was determined using the glutathione assay to measure glutathione (GSH) content and near-infrared fluorescence dot blots to measure free and protein-bound ortho-quinones. GSH is an important antioxidant and thus its depletion would be indicative of oxidative stress. Additionally, since DA may be oxidized to a quinone, increases in free and protein-bound ortho-quinones also indicate oxidative stress. Interestingly, we observed a significant decrease in GSH as the dose of MPH increased with both saline and MPTP samples. Furthermore, there was a significant increase in quinones as the dose of MPH increased. In conclusion, it appears that long-term exposure to MPH sensitizes dopaminergic neurons within the nigrostriatal pathway to oxidative stress, rendering them vulnerable to further insults, such as MPTP exposure. As such, these studies provide insight into the risks of long-term psychostimulant exposure.