Gene expression profiling with cre-conditional pseudorabies virus reveals a subset of midbrain neurons that participate in reward circuitry

TY - JOUR

T1 - Gene expression profiling with cre-conditional pseudorabies virus reveals a subset of midbrain neurons that participate in reward circuitry

AU - Pomeranz, Lisa E.

AU - Ekstrand, Mats I.

AU - Latcha, Kaamashri N.

AU - Smith, Gregory A.

AU - Enquist, Lynn W.

AU - Friedman, Jeffrey M.

N1 - Publisher Copyright: © 2017 the authors.

PY - 2017/4/12

Y1 - 2017/4/12

N2 - The mesolimbic dopamine pathway receives inputs from numerous regions of the brain as part of a neural system that detects rewarding stimuli and coordinates a behavioral response. The capacity to simultaneously map and molecularly define the components of this complex multisynaptic circuit would thus advance our understanding of the determinants of motivated behavior. To accomplish this, we have constructed pseudorabies virus (PRV) strains in which viral propagation and fluorophore expression are activated only after exposure to Cre recombinase. Once activated in Cre-expressing neurons, the virus serially labels chains of presynaptic neurons. Dual injection of GFP and mCherry tracing viruses simultaneously illuminates nigrostriatal and mesolimbic circuitry and shows no overlap, demonstrating that PRV transmission is confined to synaptically connected neurons. To molecularly profile mesolimbic dopamine neurons and their presynaptic inputs, we injected Cre-conditional GFP virus into the NAc of (anti-GFP) nanobody-L10 transgenic mice and immunoprecipitated translating ribosomes from neurons infected after retrograde tracing. Analysis of purified RNA revealed an enrichment of transcripts expressed in neurons of the dorsal raphe nuclei and lateral hypothalamus that project to the mesolimbic dopamine circuit. These studies identify important inputs to the mesolimbic dopamine pathway and further show that PRV circuitdirected translating ribosome affinity purification can be broadly applied to identify molecularly defined neurons comprising complex, multisynaptic circuits.

AB - The mesolimbic dopamine pathway receives inputs from numerous regions of the brain as part of a neural system that detects rewarding stimuli and coordinates a behavioral response. The capacity to simultaneously map and molecularly define the components of this complex multisynaptic circuit would thus advance our understanding of the determinants of motivated behavior. To accomplish this, we have constructed pseudorabies virus (PRV) strains in which viral propagation and fluorophore expression are activated only after exposure to Cre recombinase. Once activated in Cre-expressing neurons, the virus serially labels chains of presynaptic neurons. Dual injection of GFP and mCherry tracing viruses simultaneously illuminates nigrostriatal and mesolimbic circuitry and shows no overlap, demonstrating that PRV transmission is confined to synaptically connected neurons. To molecularly profile mesolimbic dopamine neurons and their presynaptic inputs, we injected Cre-conditional GFP virus into the NAc of (anti-GFP) nanobody-L10 transgenic mice and immunoprecipitated translating ribosomes from neurons infected after retrograde tracing. Analysis of purified RNA revealed an enrichment of transcripts expressed in neurons of the dorsal raphe nuclei and lateral hypothalamus that project to the mesolimbic dopamine circuit. These studies identify important inputs to the mesolimbic dopamine pathway and further show that PRV circuitdirected translating ribosome affinity purification can be broadly applied to identify molecularly defined neurons comprising complex, multisynaptic circuits.

KW - Mesolimbic

KW - Pseudorabies virus

KW - RNASeq

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U2 - 10.1523/JNEUROSCI.3193-16.2017

DO - 10.1523/JNEUROSCI.3193-16.2017

M3 - Article

C2 - 28283558

SN - 0270-6474

VL - 37

SP - 4128

EP - 4144

JO - Journal of Neuroscience

JF - Journal of Neuroscience

IS - 15

ER -