Stimulants cocktail: Methylphenidate plus caffeine impairs memory and cognition and alters mitochondrial and oxidative status

Highlights

• Methylphenidate used in an abusive context elicits antisocial behavior.

• The association of methylphenidate and caffeine increases anxiety.

• The association of methylphenidate and caffeine impairs memory.

Abstract

Methylphenidate (MPH) is a psychostimulant widely misused to increase wakefulness by drivers and students. Also, MPH can be found in dietary supplements in a clandestine manner aiming to burst performance of physical exercise practitioners. The abusive use of high doses of caffeine (CAF) in these contexts is equally already known. Here, we demonstrate the behavioral, oxidative and mitochondrial effects after acute exposure to high doses of MPH (80 mg/L) and CAF (150 mg/L), alone or associated (80 mg/L + 150 mg/L, respectively). We used zebrafish as animal model due to its high translational relevance. We evaluated the behavioral effects using the Novel Tank Test (NTT), Social Preference Test (SPT) and Y-maze Task and analyzed biomarkers of oxidative stress and activity of mitochondrial respiratory chain complexes. MPH alone induced antisocial behavior. MPH inhibited lipid peroxidation. The association of MPH + CAF presented memory impairment and anxiogenic behavior. In oxidative status, it inhibited lipid peroxidation, increased protein carbonylation and mitochondrial complex II, III and IV activity. Our results demonstrate that MPH and CAF alone negatively impact the typical behavioral of zebrafish. When associated, changes in cognition, memory, oxidative and mitochondrial status are more relevant.

Introduction

Methylphenidate (MPH) is a psychostimulant that improves attention, concentration and reduces impulsive behavior in Attention Deficit Hyperactivity Disorder (ADHD) (Challman and Lipsky, 2000; Leonard et al., 2004). Due to these effects, its nonmedical use is explored in order to promote increased cognitive function, productivity and performance (Faraone et al., 2020). It is also used as a drug of abuse due to its amphetamine-like effects with easier access (Maier et al., 2013).

MPH is one of the most frequently detected drug in the doping test according to the World Anti-Doping Agency (WADA) (Wada, 2018). The fact that MPH appears as one of the most widely used stimulants by athletes may be due to its addition in some dietary supplements, instead of being consumed as medication. The irregular presence of MPH is frequently seen in samples of seized dietary supplements. Generally when they present positive results for MPH, they also present other stimulants, the most prevalent being high-dose caffeine (CAF) (Santos et al., 2018).

CAF (1,3,7-trimethylxanthine) is a stimulant of the Central Nervous System (CNS) (Tunnicliffe et al., 2008) with high consumption mainly by practitioners of high-performance physical activities and in order to improve wakefulness by students and workers (Gurley et al., 2014; Neves and Caldas, 2017). However, it can be toxic when consumed in high doses and particular in combination with other stimulants, which is very common in dietary supplements (Bloomer et al., 2013; Venhuis et al., 2014).

Chronic use of high doses of MPH can cause oxidative stress in rats, evidenced by the increase in lipoperoxidation and oxidized glutathione (Motaghinejad et al., 2015). In addition, the chronic use of pharmacological doses of MPH increases mitochondrial enzymes involved in brain metabolism (Fagundes et al., 2010). Studies show that chronic exposure to different doses of CAF can damage and significantly increase mitochondrial function, in a dose dependent manner (Abdelkader et al., 2013; Rah et al., 2017; Saiki et al., 2011), increase apoptosis (Rah et al., 2017) and alters oxidative status in different species (Baldissera et al., 2019; De Carvalho et al., 2019; Silva et al., 2018). However, effects of MPH + CAF abusive use remains unknown.

In this context, here we tested the hypotheses that MPH, especially when associated with high doses of CAF, cause toxic effects in humans and that oxidative stress and mitochondrial impairment might be related to its mechanisms. The Danio rerio fish were chosen as a model organism, considering the relationship between this animal model and the human species since zebrafish genome shows 70% homology to the human genome (Howe et al., 2013). Additionally, the basic structure of the central nervous system in zebrafish, and other teleosts has all the major domains found in the mammalian brain with the same modulatory neurotransmitters (Maximino and Herculano, 2010; Panula et al., 2010).