Evidence for low affinity of GABA at the vesicular monoamine transporter VMAT2. Implications for transmitter co-release from dopamine neurons

Abstract Background and Purpose Midbrain dopamine (DA) neurons comprise a heterogeneous population of cells. For instance, some DA neurons express the vesicular glutamate transporter VGLUT2 allowing these cells to co-release DA and glutamate. Additionally, GABA may be co-released from DA neurons. However, most cells do not express the canonical machinery to synthesize GABA or the vesicular GABA transporter VGAT. Instead, GABA seems to be taken up into DA neurons by a plasmalemmal GABA transporter (GAT1) and stored in synaptic vesicles via the vesicular monoamine transporter VMAT2. Yet, it remains unclear whether GABA indeed interacts with VMAT2, or whether another transmitter could be responsible for the observed inhibitory effects attributed to GABA. Experimental Approach We used radiotracer flux measurements in VMAT2 expressing HEK-293 cells and synaptic vesicles from rodents to determine whether GABA qualifies as substrate at VMAT2. mRNA in situ hybridization was employed to determine expression of VMAT2 and GAT1 transcripts in DA neurons of mouse and in human midbrains. Key Results We found that GABA reduced uptake of VMAT2 substrates in rodent synaptic vesicle preparations from striatum and cerebellum at millimolar concentrations but had no effect in VMAT2-expressing cells indicating that key components are missing in a non-neuronal system. Roughly 60 % of murine and human DA neurons in the substantia nigra express VMAT2 and GAT1 suggesting that many may be capable of co-releasing DA and GABA. Conclusion and Implication Our experiments suggest that GABA is a low-affinity substrate at VMAT2 with potential implications for basal ganglia physiology and disease. Bullet point summary What is already known Subpopulations of dopamine neurons co-release glutamate and/or GABA. While glutamate is loaded into vesicles by VGLUT2, GABA co-release depends on GAT1 and VMAT2. What this study adds The relative affinity of GABA at VMAT2 was found to be in the millimolar range. Human midbrain dopamine neurons express GAT1. Clinical significance GABA co-release from midbrain dopamine neurons may also occur in humans. GABA co-release from dopamine neurons may play a role in neuropsychiatric diseases