Introduction

The ventral tegmental area (VTA) is a deep brain nucleus that contains dopaminergic, GABAergic and glutamatergic neuronal projections. Functional neuroimaging and behavioral studies have shown that electrical stimulation in the VTA results in fMRI signal activation along mesocorticolimbic pathways, and plays a key role in reinforcement learning¹. Cyclic voltammetry measurements have demonstrated that VTA stimulation leads to dopamine release in vivo^2,3. Yet, the contributions of dopamine receptor subtypes in downstream signaling or the potential involvement of other neurotransmitters in to VTA are still largely unknown. Previous results have shown that changes in D2 receptor-specific due to acute VTA stimulation can be challenging to detect with [¹¹C]raclopride-PET⁴. The purpose of this study was to determine the underlying receptor-specific mechanisms of VTA stimulation by determining dopaminergic receptor subtype contributions and their whole-brain circuit modulation using fMRI in combination with pharmacological blocking and receptor-specific PET imaging in non-human primates (NHP).

Methods

A combined PET/MRI scanner with an NHP-specific custom coil (8-ch) was used to acquire fMRI and PET data in an anesthetized baboon (1.5% isoflurane) with a chronic microstimulation implant in the right VTA. Gradient-echo EPI data were acquired continuously during acute stimulation and ferumoxytol was used as a contrast agent. In PET/fMRI sessions (n=8), [¹¹C]raclopride dynamic scans were acquired to determine D2 receptor (D2R) binding. In order to distinguish receptor-specific contributions, separate fMRI sessions (n=5) were designed to stimulate before and after a blocking dose of the D1 receptor (D1R) antagonist SCH23390 (0.03-0.09 mg/kg i.v.) or the D2R antagonist prochlorperazine (0.1 mg/kg i.v.). Stimulation parameters consisted of a pulse trains of 200 ms at 100 Hz and a current of 1 mA, which lasted for 15 s and alternated with 93 s of rest. This pattern was repeated with a total of 5 blocks for 10 min (Figure 1). Block-design fMRI data were analyzed with a GLM and converted into cerebral blood volume (CBV) changes. PET data were analyzed with the simplified reference tissue model⁵.

Results

fMRI data due to stimulation without any pharmacological blocking showed robust positive CBV signals in the right (implanted) hemisphere, with the largest signals up to 6% CBV observed in right caudate, nucleus accumbens and putamen. Comparisons of the stimulation-induced fMRI signal before and after pharmacological blocking of D2R resulted in a small but significant signal increase in all previously activated regions (Figure 2). After D2R blocking, the %CBV signal was on average 1.3±0.2 times larger than in the pre-drug stimulation condition in putamen, caudate and nucleus accumbens. Additional activation was observed in the prefrontal cortex and the contralateral side of the striatum. Pharmacological blocking of D1R showed a signal decrease by a factor of 0.5±0.1 on average in the previously activated regions (Figure 3). Baseline PET binding potential (BP_ND) values across sessions were 3.8±0.3 (putamen), 2.8±0.3 (caudate) and 2.1±0.1 (nucleus accumbens), with no significant lateral differences (Figure 4).

Discussion

In this study, we showed that microstimulation in the VTA resulted in overall robust positive CBV changes, consistent with previous findings of activation due to VTA stimulation¹. The altered CBV changes we observed after selective pharmacological blocking of receptor subtypes suggest that VTA stimulation is partially driven by dopamine at D1 and D2 receptors: The large CBV decrease after D1R blocking is consistent with blocking of an excitatory D1R contribution and the CBV increase after D2R blocking is consistent with suppression of a smaller inhibitory D2R contribution. Our findings indicate that the magnitude of endogenous dopamine due to acute stimulation-induced changes may be close to the limit of detection for [¹¹C]raclopride-PET competition experiments. This sheds light on limits in dynamic sensitivity with PET, and that fMRI can have superior sensitivity, especially in study designs with fast stimuli. Similar lateral binding potential values demonstrate that D2R concentrations did not change or drive the CBV signal. Finally, the fact that the overall CBV signal remains positive after D1R or D2R blocking suggests that a part of the remaining signal is driven by non-dopaminergic excitatory neurotransmitters/receptors. Overall, the results from this study provide novel insight into the receptor mechanisms of VTA stimulation, providing important insights for deep brain stimulation of mesocorticolimbic pathway modulation.

Acknowledgements

This work was supported by NIH grants 1K99DA043629, P41EB015896, S10RR026666, S10RR022976, S10RR019933 and S10RR017208, and FWO-Flanders, FG0D5817N, G090714N, and G0007.12.