Combined Chronic Oral Methylphenidate and Fluoxetine Decreases D2R Levels in the Caudate Putamen and Nucleus Accumbens

Abstract

Methylphenidate (MP) is a commonly prescribed psychostimulant for treating Attention-Deficit/Hyperactive Disorder (ADHD). Many patients with ADHD also experience anxiety and depression, often leading to co-dosing with selective serotonin reuptake inhibitors (SSRIs), such as fluoxetine (FLX), commonly used for ADHD-related and adolescent depression. Our laboratory and others have shown that MP increases striatal dopamine (DA) transporters and DA type 1 receptor binding (D1R) in rats, and FLX has been shown to affect the DA reward pathway through the effect DA receptors play on increased cellular serotonin (5-HT). However, the effects of combined MP and FLX on DA receptor binding remain unclear. This study investigated how MP, FLX, and their combination affect D1R and DA type 2 (D2R) binding. At three weeks of age, adolescent rats received four weeks of oral drug treatments via a previously established dosing paradigm that replicates human pharmacokinetics. Rats were separated into four groups, receiving water, MP, FLX, or MP + FLX. Following treatment, autoradiography binding was conducted on coronal brain sections and showed chronic combined treatment with MP + FLX resulted in significant decreases in D2R levels relative to controls in the: Dorsal Caudate Putamen (DCPU) (51.5%), Dorsolateral Caudate Putamen (DLCPU) (50.4%), Nucleus Accumbens Core (Nac Core) (44.8%), Ventral Caudate Putamen (VCPU) (47.7%), and Ventromedial Caudate Putamen (VMCPU) (49.1%). No significant effects were reported for D1R binding. Thus, the combined treatment of MP + FLX in attenuating D2R levels may be involved in the mechanism that prior literature has described an increased risk for substance use disorder, cognitive deficits and motor dysregulation.

Keywords: Autoradiography; Brain Mapping; Dopamine; Psychostimulants; Reward; Serotonin Reuptake Inhibitors.

Fig. 1

Diagram detailing dosing schedule for animals from 9:00–17:00. Rats in the MP group received a dosage of 30 mg/kg, rats in the FLX group briefly received a dosage of 20 mg/kg FLX, and rats in the MP + FLX group briefly received a dosage of 30 mg/kg of MP and 20 mg/kg FLX for one hour (9:00–10:00). For the following 7 h, rats in the MP group received a dosage of 60 mg/kg, rats in the FLX group received a dosage of 20 mg/kg, and rats in the MP + FLX group received a dosage of 30 mg/kg of MP and 20 mg/kg of FLX

Fig. 2

Representative figures showing D1R binding using [³H] SCH23390 to assess binding levels, with attached atlas figure showing: Dorsomedial Caudate Putamen (DMCPU), Dorsolateral Caudate Putamen (DLCPU), Ventromedial Caudate Putamen (VMCPU), Ventromedial Caudate Putamen (VMCPU), Ventrolateral Caudate Putamen (VLCPU), Nucleus Accumbens Core (NacC), Nucleus Accumbens Shell (NacS), and Olfactory Tubercle (OT)

Fig. 3

Mean [³H] SCH 23,390 binding (µCi/g) ± SEM in the Basal Ganglia following 4 weeks of treatment, with n = 8/9 per group. Quantitative autoradiography of [³H] 23,390 (D1R-like) binding levels within the Dorsal Caudate putamen (D CPU), Dorsolateral Caudate Putamen (DL CPU), Dorsomedial Caudate Putamen (DM CPU), Nucleus Accumbens Core (Nac Core), Nucleus Accumbens Shell (Nac Sh Shell), Olfactory Tubercle (OT), Substantia Nigra (SNR), Ventral Caudate Putamen (V CPU), Ventrolateral Caudate Putamen (VL CPU), and the Ventromedial Caudate Putamen (VM CPU) across all drug treatment groups. Measurements of the regions of interest were carried out at the bregma coordinates taken from the Paxinos & Watson rat brain atlas. No significant difference was observed (p > 0.05) across any of the groups. Each bar represents the group mean for dopamine D1 receptor binding

Fig. 4

Representative figures showing D2R binding using [³H] spiperone to assess binding levels with attached atlas figure showing: Dorsomedial Caudate Putamen (DMCPU), Dorsolateral Caudate Putamen (DLCPU), Ventromedial Caudate Putamen (VMCPU), Ventrolateral Caudate Putamen (VLCPU), Nucleus Accumbens Core (NacC), Nucleus Accumbens Shell (NacS), and Olfactory Tubercle (OT)

Fig. 5

Mean [³H] spiperone binding (µCi/g) ± SEM in the Basal Ganglia following 4 weeks of treatment, with n = 8/9 per group. * Denotes a significant difference (p < 0.05) between MP + FLX and Water. $ denotes a significant difference (p< 0.05) between MP + FLX and FLX. ROIs consisted of the Dorsal Caudate putamen (D CPU), Dorsolateral Caudate Putamen (DL CPU), Dorsomedial Caudate Putamen (DM CPU), Nucleus Accumbens Core (Nac Core), Nucleus Accumbens Shell (Nac Shell), Olfactory Tubercle (OT), Ventral Caudate Putamen (V CPU), Ventrolateral Caudate Putamen (VL CPU), and the Ventromedial Caudate Putamen (VM CPU)