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. Previous research has shown that long-term exposure to MPH causes dopamine-releasing neurons within the nigrostriatal pathway to die when exposed to the Parkinsonian toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MPH acts by blocking dopamine transporters and norepinephrine transporters, preventing the reuptake and removal of these neurotransmitters following release and increasing the time outside of the protective environment of the neuron’s vesicles. We hypothesize that spontaneous oxidation of excess dopamine to a quinone metabolite is rendering these neurons within this particular pathway to be more sensitive to MPTP. The dopamine quinone may be bound by the antioxidant glutathione (GSH) in an effort to protect the cell against oxidative stress. However, as the finite amount of GSH is depleted, the quinone may lead to the production of highly reactive molecules, leading to mitochondrial damage and cell death which may be accelerated by MPTP. In order to examine this hypothesis, we chose to study adolescent male Swiss-Webster mice, which have been shown to be resistant to MPTP’s toxic effects. They were divided into 3 cohorts and administered either saline (control), 1 mg/kg MPH (therapeutic 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 20 mg/kg, every 2 hours), while the other half was administered 4 injections of sterile saline. Either 3 or 7 days after MPTP or saline treatment, the mice were sacrificed, brains were removed, and the substantia nigra (SN) and striatum (STR) were collected. These areas of the brain make up the nigrostriatal pathway and are affected by Parkinson’s disease. Oxidative stress related to increased dopamine levels was determined using the glutathione assay to measure GSH content, near-infrared fluorescence dot blots to measure free and protein-bound ortho-quinones, and an ATP luciferase assay to measure mitochondrial function. Interestingly, there was a significant decrease in GSH as the dose of MPH was increased with both saline and MPTP samples. Furthermore, a significant increase in quinones was observed as the dose of MPH increased. We also expect to see a decrease in ATP inversely proportional to the dose of MPH indicating increased oxidative stress. 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.
| Identifer | oai:union.ndltd.org:ETSU/oai:dc.etsu.edu:asrf-1440 |
| Date | 12 April 2019 |
| Creators | McWethy, David, Oakes, Hannah, Ketchem, Shannon, Ensley, Tucker, Dema, Blerim, Pond, Brooks B |
| Publisher | Digital Commons @ East Tennessee State University |
| Source Sets | East Tennessee State University |
| Detected Language | English |
| Type | text |
| Source | Appalachian Student Research Forum |
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