Methylphenidate alters Akt‐mTOR signaling in rat pheochromocytoma cells

Abstract

The exponential increase in methylphenidate (MPH) prescriptions in recent years has worried researchers about its misuse among individuals who do not meet the full diagnostic criteria for attention‐deficit/hyperactivity disorder (ADHD) such as young children and students in search of cognitive improvement or for recreational reasons. The action of MPH is based mainly on inhibition of dopamine transporter, but the complete cellular effects are still unknown. Based upon prior studies, we attempted to determine whether the treatment with MPH (1μM) influences protein kinase B‐mammalian target of rapamycin complex 1 signaling pathways (Akt‐mTOR), including translation repressor protein (4E‐BP1) and mitogen activated protein kinase (S6K), in rat pheochromocytoma cells (PC12), a well characterized cellular model, in a long or short term. MPH effects on the Akt substrates [cAMP response element‐binding protein (CREB), forkhead box protein O1 (FoxO1), and glycogen synthase kinase 3 beta (GSK‐3β)] were also evaluated. Whereas short term MPH treatment decreased the pAkt/Akt, pmTOR/mTOR and pS6K/S6K ratios, as well as pFoxO1 immunocontent in PC12 cells, long term treatment increased pAkt/Akt, pmTOR/mTOR and pGSK‐3β/GSK‐3β ratio. Phosphorylation levels of 4E‐BP1 were decreased at 15 and 30 min and increased at 1 and 6 h by MPH. pCREB/CREB ratio was decreased. This study shows that the Akt‐mTOR pathway, as well as other important Akt substrates which have been described as important regulators of protein synthesis, as well as being implicated in cellular survival, synaptic plasticity and memory consolidation, was affected by MPH in PC12 cells, representing an important step in exploring the MPH effects.

Attention-Deficit and Hyperactivity Disorder: A Disorder or a Fraud?

“…Moreover, phosphorylation levels of 4E-BP1 were decreased at 15 and 30 min (short term treatment) and increased at 1 and 6 h (long term treatment). These findings prove that methylphenidate® alters cell signaling in PC12 cells and its responses differ according to the time of exposure to this psychostimulant (Schmitz et al, 2019).…”

“…This is probably because of D2R activation promoted by a further increase in DA levels triggered by methylphenidate® treatment in a short term treatment. However, phosphorylated Akt level was increased after a long methylphenidate® treatment (1 h) in PC12 cells (Schmitz et al, 2019); most likely due to an increase in the extracellular DA metabolites DOPAC and HVA levels which break DA after methylphenidate® treatment (Bartl et al, 2010). Based on this, it was deduced that the decrease in Akt phosphorylation in the first minutes followed by its increase in PC12 cells after long term methylphenidate® treatment, may be associated with increased DA levels and D2R activation in the first place, followed by its increased degradation decrease and the eventual D2R inactivation (Schmitz et al, 2019).…”

“…Results show that methylphenidate® decreases phosphorylated mTOR ratio, as well as phosphorylation of S6K in PC12 cells after a short term treatment. Moreover 4E-BP1 hyperphosphorylated levels were decreased, whereas 4E-BP1 hypophosphorylated levels increased in PC12 cells after a short term treatment (Schmitz et al, 2019). Hence, the decreased Akt phosphorylation observed after short treatment with methylphenidate®, as well as the decreased phosphorylation of mTOR induced translation impairment, affecting processes which require protein synthesis as synaptic plasticity, axonal growth, and regeneration.…”

“…Thus, children are wrongly diagnosed with ADHD every time they demonstrate an "active" behavior while 90% of these 4.5 million kids do not have any abnormal DA metabolism or biological basis (Kagan, 2009). This exponential increase in methylphenidate® prescriptions in recent years has worried researchers about its misuse among individuals who do not meet the full diagnostic criteria for ADHD such as young children and students in search of cognitive improvement (Schmitz, Chao, & Wyse, 2019). Hence, the answer for "ADHD: a disorder?"…”

“…These cells are also used to study the effects of methylphenidate® (Bartl et al, 2010;Grünblatt et al, 2013), since it is a dopaminergic neuronal cell that can synthesize, store, secrete, and take up DA (Pan, Zhu, Hwu, & Jankovic, 2012). Based on that, the PC12 cells' response to methylphenidate® treatment was studied by analyzing Akt-mTOR pathway, as well as the mTOR substrates, 4E-BP1 and S6K kinase (Schmitz et al, 2019). Results show that short term methylphenidate® treatment decreased phosphorylated Akt, mTOR, and S6K ratio in PC12 cells.…”