title: Functional Connectome Analysis of Dopamine Neuron Glutamatergic Connections in Forebrain Regions.
abstract: In the ventral tegmental area (VTA), a subpopulation of dopamine neurons express vesicular glutamate transporter 2 and make glutamatergic connections to nucleus accumbens (NAc) and olfactory tubercle (OT) neurons. However, their glutamatergic connections across the forebrain have not been explored systematically. To visualize dopamine neuron forebrain projections and to enable photostimulation of their axons independent of transmitter status, we virally transfected VTA neurons with channelrhodopsin-2 fused to enhanced yellow fluorescent protein (ChR2-EYFP) and used DAT(IREScre) mice to restrict expression to dopamine neurons. ChR2-EYFP-expressing neurons almost invariably stained for tyrosine hydroxylase, identifying them as dopaminergic. Dopamine neuron axons visualized by ChR2-EYFP fluorescence projected most densely to the striatum, moderately to the amygdala and entorhinal cortex (ERC), sparsely to prefrontal and cingulate cortices, and rarely to the hippocampus. Guided by ChR2-EYFP fluorescence, we recorded systematically from putative principal neurons in target areas and determined the incidence and strength of glutamatergic connections by activating all dopamine neuron terminals impinging on recorded neurons with wide-field photostimulation. This revealed strong glutamatergic connections in the NAc, OT, and ERC; moderate strength connections in the central amygdala; and weak connections in the cingulate cortex. No glutamatergic connections were found in the dorsal striatum, hippocampus, basolateral amygdala, or prefrontal cortex. These results indicate that VTA dopamine neurons elicit widespread, but regionally distinct, glutamatergic signals in the forebrain and begin to define the dopamine neuron excitatory functional connectome.
  
  
  CONCLUSIONS
  Dopamine neurons are important for the control of motivated behavior and are involved in the pathophysiology of several major neuropsychiatric disorders. Recent studies have shown that some ventral midbrain dopamine neurons are capable of glutamate cotransmission. With conditional expression of channelrhodopsin in dopamine neurons, we systematically explored dopamine neuron connections in the forebrain and identified regionally specific dopamine neuron excitatory connections. Establishing that only a subset of forebrain regions receive excitatory connections from dopamine neurons will help to determine the function of dopamine neuron glutamate cotransmission, which likely involves transmission of precise temporal signals and enhancement of the dynamic range of dopamine neuron signals.
authors.affiliation: Department of Psychiatry, Columbia University, and Department of Molecular Therapeutics, NYS Psychiatric Institute, New York, New York 10032, and.
authors.forename: Susana
authors.identifier: http://orcid.org/0000-0002-0401-4317
authors.initials: S
authors.lastname: Mingote
authors.affiliation: Department of Psychiatry, Columbia University, and Department of Molecular Therapeutics, NYS Psychiatric Institute, New York, New York 10032, and.
authors.forename: Nao
authors.initials: N
authors.lastname: Chuhma
authors.affiliation: Departments of Psychiatry and Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232.
authors.forename: Sheila V
authors.initials: SV
authors.lastname: Kusnoor
authors.affiliation: Department of Psychiatry, Columbia University, and Department of Molecular Therapeutics, NYS Psychiatric Institute, New York, New York 10032, and.
authors.forename: Bianca
authors.identifier: http://orcid.org/0000-0002-0297-949X
authors.initials: B
authors.lastname: Field
authors.affiliation: Departments of Psychiatry and Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232.
authors.forename: Ariel Y
authors.initials: AY
authors.lastname: Deutch
authors.affiliation: Department of Psychiatry, Columbia University, and Department of Molecular Therapeutics, NYS Psychiatric Institute, New York, New York 10032, and stephen.rayport@columbia.edu.
authors.forename: Stephen
authors.identifier: http://orcid.org/0000-0001-9755-7486
authors.initials: S
authors.lastname: Rayport
mesh_terms: D000818:Animals; D000075402:Channelrhodopsins; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D050483:Dopamine Plasma Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D059290:Dopaminergic Neurons / Q000502:physiology*; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects / Q000502:physiology; D005260:Female; D018698:Glutamic Acid / Q000378:metabolism*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009415:Nerve Net / Q000502:physiology*; D009434:Neural Pathways / Q000502:physiology*; D018377:Neurotransmitter Agents / Q000494:pharmacology; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D016548:Prosencephalon / Q000166:cytology* / Q000502:physiology; D014161:Transduction, Genetic; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D017557:Ventral Tegmental Area / Q000166:cytology
chemical_list: D000075402:Channelrhodopsins; D004268:DNA-Binding Proteins; D050483:Dopamine Plasma Membrane Transport Proteins; D018377:Neurotransmitter Agents; D018698:Glutamic Acid; D014446:Tyrosine 3-Monooxygenase; D010751:Phosphopyruvate Hydratase
keywords: cotransmission; entorhinal cortex; glutamate; nucleus accumbens; optogenetics