Ventral tegmental area GABA, glutamate, and glutamate-GABA neurons are heterogeneous in their electrophysiological and pharmacological properties.

The ventral tegmental area (VTA) contains dopamine neurons intermixed with GABA-releasing (expressing vesicular GABA transporter, VGaT), glutamate-releasing (expressing vesicular glutamate transporter 2, VGluT2), and glutamate-GABA co-releasing (co-expressing VGluT2 and VGaT) neurons. By delivering INTRSECT viral vectors into the VTA of double vglut2-Cre/vgat-Flp transgenic mice, we targeted specific VTA cell populations for ex vivo recordings. We found that VGluT2+ VGaT- and VGluT2+ VGaT+ neurons on average had relatively hyperpolarized resting membrane potential, greater rheobase, and lower spontaneous firing frequency compared to VGluT2- VGaT+ neurons, suggesting that VTA glutamate-releasing and glutamate-GABA co-releasing neurons require stronger excitatory drive to fire than GABA-releasing neurons. In addition, we detected expression of Oprm1mRNA (encoding µ opioid receptors, MOR) in VGluT2+ VGaT- and VGluT2- VGaT+ neurons, and that the MOR agonist DAMGO hyperpolarized neurons with these phenotypes. Collectively, we demonstrate the utility of the double transgenic mouse to access VTA glutamate, glutamate-GABA, and GABA neurons to determine their electrophysiological properties. SIGNIFICANT STATEMENT: Some physiological properties of VTA glutamate-releasing and glutamate-GABA co-releasing neurons are distinct from those of VTA GABA-releasing neurons. µ-opioid receptor activation hyperpolarizes some VTA glutamate-releasing and some GABA-releasing neurons.

The Ventral Tegmental Area has calbindin neurons with the capability to co-release glutamate and dopamine into the nucleus accumbens.

The ventral tegmental area (VTA) has three major classes of neurons: dopaminergic (expressing tyrosine hydroxylase; TH), GABAergic (expressing vesicular GABA transporter; VGaT) and glutamatergic (expressing vesicular glutamate transporter 2; VGluT2). While VTA dopaminergic and GABAergic neurons have been further characterized by expression of calcium-binding proteins (calbindin, CB; calretinin, CR or parvalbumin, PV), it is unclear whether these proteins are expressed in rat VTA glutamatergic neurons. Here, by a combination of in situ hybridization (for VGluT2 mRNA detection) and immunohistochemistry (for CB-, CR- or PV-detection), we found that among the total population of VGluT2 neurons, 30% coexpressed CB, 3% coexpressed PV and <1% coexpressed CR. Given that some VGluT2 neurons coexpress TH or VGaT, we examined whether these neurons coexpress CB, and found that about 20% of VGluT2-CB neurons coexpressed TH and about 13% coexpressed VGaT. Because VTA TH-CB neurons are known to target the nucleus accumbens (nAcc), we determined whether VGluT2-CB-TH neurons innervate nAcc, and found that about 80% of VGluT2-CB neurons innervating the nAcc shell coexpressed TH. In summary, (a) CB, PV and CR are detected in subpopulations of VTA-VGluT2 neurons; (b) CB is the main calcium-binding protein present in VTA-VGluT2 neurons; (c) one-third of VTA-VGluT2 neurons coexpress CB; (d) some VTA-VGluT2-CB neurons have the capability to co-release dopamine or GABA, and (e) a subpopulation of VTA glutamatergic-dopaminergic neurons innervates nAcc shell. These findings further provide evidence for molecular diversity among VTA-VGluT2 neurons, neurons that may play a role in specific circuitry and behaviours.