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Cited by 704Neurotransmitter Receptor Influence on BehaviorMemory and Neural MechanismsNeuroscience and Neuropharmacology Research

Distinct extended amygdala circuits for divergent motivational states

Open access
Jennings, Joshua H., Sparta, Dennis R., Stamatakis, Alice M., Ung, Randall L., Pleil, Kristen E., Kash, Thomas L., Stuber, Garret D.·2013·Nature
DOI 10.1038/nature12041PMC PMC3778934
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Abstract

The co-morbidity of anxiety and dysfunctional reward processing in illnesses such as addiction and depression suggests that common neural circuitry contributes to these disparate neuropsychiatric symptoms. The extended amygdala, including the bed nucleus of the stria terminalis (BNST), modulates fear and anxiety, but also projects to the ventral tegmental area (VTA), a region implicated in reward and aversion, thus providing a candidate neural substrate for integrating diverse emotional states. However, the precise functional connectivity between distinct BNST projection neurons and their postsynaptic targets in the VTA, as well as the role of this circuit in controlling motivational states, have not been described. Here we record and manipulate the activity of genetically and neurochemically identified VTA-projecting BNST neurons in freely behaving mice. Collectively, aversive stimuli exposure produced heterogeneous firing patterns in VTA-projecting BNST neurons. By contrast, in vivo optically identified glutamatergic projection neurons displayed a net enhancement of activity to aversive stimuli, whereas the firing rate of identified GABAergic (γ-aminobutyric acid-containing) projection neurons was suppressed. Channelrhodopsin-2-assisted circuit mapping revealed that both BNST glutamatergic and GABAergic projections preferentially innervate postsynaptic non-dopaminergic VTA neurons, thus providing a mechanistic framework for in vivo circuit perturbations. In vivo photostimulation of BNST glutamatergic projections resulted in aversive and anxiogenic behavioural phenotypes. Conversely, activation of BNST GABAergic projections produced rewarding and anxiolytic phenotypes, which were also recapitulated by direct inhibition of VTA GABAergic neurons. These data demonstrate that functionally opposing BNST to VTA circuits regulate rewarding and aversive motivational states, and may serve as a crucial circuit node for bidirectionally normalizing maladaptive behaviours.

AI snapshot

Written from the paper’s full text.

TL;DR

In mice, VTA-projecting BNST neurons comprise opposing glutamatergic and GABAergic subcircuits: aversive stimuli increase glutamatergic activity and suppress GABAergic activity, and optogenetic activation of these pathways produces aversive/anxiogenic versus rewarding/anxiolytic states, respectively. These findings identify a bidirectional BNST→VTA circuit node for motivational and emotional processing.

Key findings

  • In vivo antidromic identification found 53/137 BNSTv→VTA projection neurons; aversive stimuli produced heterogeneous firing patterns across these neurons. Identified Vglut2BNSTv→VTA neurons (34/145) showed net enhanced firing during aversive events and shock-associated cues, whereas VgatBNSTv→VTA neurons (33/77) were principally suppressed.
  • Circuit mapping showed that Vglut2BNSTv→VTA and VgatBNSTv→VTA terminals formed functional synapses primarily onto non-dopaminergic and medially located dopaminergic VTA neurons.
  • Photostimulation of Vglut2BNSTv→VTA produced avoidance of a stimulation-paired chamber, reduced active reward seeking, and anxiety-like open-field behavior; the aversive phenotype was abolished by intra-VTA ionotropic glutamate receptor antagonists.
  • Photostimulation of VgatBNSTv→VTA produced place preference, supported optical self-stimulation, and increased open-arm time in the elevated plus maze; these effects were prevented by an intra-VTA GABAA antagonist and recapitulated by direct inhibition of VTA GABAergic neurons. Concurrent VgatBNSTv→VTA stimulation during foot shock reduced freezing and increased later open-arm time/entries.

Methods

  • Adult male C57BL/6J, Vgat-ires-cre, and Vglut2-ires-cre mice received stereotactic viral injections (ChR2-eYFP, NpHR) and optical fibers/MEA implants; experiments included anesthetized and behaving in vivo electrophysiology, patch-clamp recordings in VTA slices, unpredictable foot-shock sessions, and optogenetic behavioral assays.
  • In vivo electrophysiology used a 16-channel MEA in BNSTv and optical fibers above VTA for antidromic identification; units were classified using spike latency, waveform correlation, and z-score firing changes during foot-shock sessions.
  • Behavioral assays included real-time place preference, open field, sucrose self-administration, optical self-stimulation/self-inhibition, elevated plus maze, and foot-shock-induced freezing, with intra-VTA antagonist microinfusions in some experiments.

Limitations

  • The paper states that shock artifacts prevented reliable quantification of neuronal firing 500–1000 ms after foot-shock onset, excluding approximately 0.8–1.66% of collected data during the foot-shock session.
  • Evident from the text: all experiments used adult male mice, so sex differences and generalization to females are not addressed.
  • Evident from the text: the paper does not include a dedicated limitations section; circuit specificity relies on viral targeting, optogenetic manipulation, slice physiology, and pharmacological blockade rather than fully selective circuit-silencing approaches.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1038/nature12041

Cites

33 of 33 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 1 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    Exposure to aversive foot shock or shock-associated contextual cues enhances firing of Vglut2BNSTv→VTA neurons.

    “in vivo optically-identified glutamatergic projection neurons displayed a net enhancement of activity to aversive stimuli”
    • Also stated in: § Abstract · plain
      “Collectively, exposure to the aversive event or associated cues alone enhanced the firing of Vglut2BNSTv→VTA neurons”
    Quote in the paperSubstantial claim
  • Findingown data

    BNSTv→VTA neurons display heterogeneous firing patterns in response to aversive stimuli, segregating into three functionally distinct classes based on changes in normalized firing rates during foot-shock.

    “Collectively, aversive stimuli exposure produced heterogeneous firing patterns in VTA-projecting BNST neurons.”
    • Also stated in: § Abstract · boosted
      “Identified BNSTv→VTA neurons segregated into three functionally distinct classes based on changes in their normalized firing rates throughout the foot-shock session (Supplementary Fig. 3), demonstrating that BNSTv→VTA neurons differentially encode information related to aversive stimuli and their associated contextual cues.”
    Quote in the paperSubstantial claim
  • Findingown data

    Exposure to aversive foot shock or shock-associated contextual cues suppresses firing of VgatBNSTv→VTA neurons.

    “whereas the firing rate of identified GABAergic projection neurons was suppressed”
    • Also stated in: § Abstract · plain
      “while simultaneously suppressing the activity of VgatBNSTv→VTA neurons.”
    Quote in the paperSubstantial claim
  • Findingown data

    Photostimulation of VgatBNSTv→VTA projections produces anxiolytic phenotypes (increased open arm time in elevated plus maze).

    “activation of BNST GABAergic projections produced rewarding and anxiolytic phenotypes”
    • Also stated in: § Abstract · hedged
      “Photostimulation of the VgatBNSTv→VTA pathway and direct inhibition of VgatVTA neurons significantly increased time spent in the open arms of an elevated plus maze, indicative of anxiolysis”
    Quote in the paperSubstantial claim
  • Findingown data

    VgatBNSTv→VTA::ChR2 mice spent significantly less time frozen following foot-shock with concurrent activation.

    “VgatBNSTv→VTA::ChR2 mice spent significantly less time frozen (Fig. 5h)”
    Quote in the paperSubstantial claim
  • May combine two claims. 9
  • Classified as probably not a substantial claim. 19

  • May combine two claims.

    Findingown data

    Photostimulation of CaMKIIaBNSTv→VTA pathway produces both glutamatergic and GABAergic currents in VTA neurons, demonstrating neurochemically distinct BNSTv neurons project to VTA.

    “photostimulation of the CaMKIIaBNSTv→VTA pathway produced both glutamatergic and γ-aminobutyric acid (GABAergic) currents in VTA neurons (Supplementary Fig. 4), demonstrating that neurochemically distinct BNSTv neurons project to the VTA.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Photostimulation of ChR2-containing fibers from Vglut2 or Vgat expressing BNSTv neurons produces excitatory or inhibitory postsynaptic currents respectively.

    “photostimulation of ChR2-containing fibers originating from Vglut2 (Vglut2BNSTv→VTA::ChR2) or Vgat (VgatBNSTv→VTA::ChR2) expressing BNSTv neurons produced excitatory or inhibitory postsynaptic currents respectively”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    Data collected during the time epochs surrounding individual delivered foot shocks were excluded from analysis.

    “Thus, data collected during these time epochs were excluded from analysis.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    Drug injections into the VTA were given in a counterbalanced design.

    “into the VTA in a counter balanced design (all drugs from Tocris).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    Reward-related responses to VgatBNSTv→VTA activation are recapitulated by directly inhibiting VgatVTA neurons, providing a circuit mechanism for the VgatBNSTv→VTA pathway to regulate motivated behavior.

    “Together, these results show that reward-related responses to VgatBNSTv→VTA activation are recapitulated by directly inhibiting VgatVTA neurons, thus providing a circuit mechanism for the VgatBNSTv→VTA pathway to regulate motivated behavior.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim. May combine two claims.

    Study design and conductown data

    Individual units were discriminated offline using principal component analysis, and auto- and cross-correlograms, firing characteristics, and inter-spike interval distributions were examined to ensure units were well-isolated.

    “Briefly, discrimination of individual units was performed offline using principal component analysis to separate individual units from the same electrode. In addition, auto- and cross-correlograms, firing characteristics, and inter-spike interval distributions were examined to ensure units were well-isolated.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    Placements of recording electrode tips within the BNSTv and optical fibers within the VTA were verified with histological examination.

    “Placements of recording electrode tips within the BNSTv and optical fibers within the VTA were verified with histological examination of brain tissue following the experiments.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    The exclusion of data during the time epochs surrounding individual delivered foot shocks represents an approximate loss of 0.8-1.66% of the collected data during the foot-shock session.

    “This represents an approximate loss of 0.8 – 1.66% of the collected data during the foot-shock session.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    Large shock artifacts were readily apparent during the time epoch surrounding individual delivered foot shocks.

    “During the time epoch surrounding the individual delivered foot shocks, large shock artifact were readily apparent.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    Resulting data were binned in 30 s epochs to minimize skewing of results due to data lost by shock artifacts.

    “Resulting data was binned in 30 s epochs in order to minimize the skewing of our results due to data lost by the shock artifacts.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Electrical stimulation of the BNST produces both excitatory and inhibitory responses in VTA neurons in vivo, implying distinct subcircuits may exist.

    “Electrical stimulation of the BNST produces both excitatory and inhibitory responses in VTA neurons in vivo6 implying that distinct subcircuits may exist.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [6] Georges, F. & Aston-Jones, G. Potent regulation of midbrain dopamine neurons by the bed nucleus of the stria terminalis. J. Neurosci. 21, RC160 (2001) source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    BNSTv neurons were classified as antidromic-responsive if they met three criteria: stable antidromic spike latency, ability to respond reliably to high frequency photostimulation, and collision between orthodromic- and antidromic-evoked spikes.

    “BNSTv neurons were classified as antidromic-responsive, if the following 3 criteria were met: 1) stable antidromic spike latency (< 0.2 ms), 2) ability to respond reliably to high frequency photostimulation, 3) collision between orthodromic- and antidromic-evoked spikes.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim. May combine two claims.

    Study design and conductnone given

    Neuronal units were included in the data only if the signal-to-noise ratio was high and the mean firing rate was between 0.5 and 25 Hz during baseline recording periods.

    “Neuronal units were included in the data if the signal-to-noise ratio was high, and the mean firing rate was between 0.5 – 25 Hz during baseline recording periods.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Mice had 2 days off between each VTA microinjection.

    “Mice had 2 days off between each VTA microinjection.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Recording sites were verified histologically using electrolytic lesions at 200 μA for 5 s.

    “In addition, recording sites was verified histologically using electrolytic lesions at 200 µA for 5 s.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    Neuronal firing responses in the 500-1000 ms following foot shock onset could not be reliably quantified.

    “neuronal firing responses in the 500 – 1000 ms following foot shock onset could not be reliably quantified”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given

    All mice were infused at a rate of 0.1 μl per minute, and the injector remained in place for approximately 2 min following infusion to ensure proper diffusion of drug into the VTA.

    “All mice were infused at a rate of 0.1 µl per minute. The injector remained in place for approximately 2 min following infusion to ensure proper diffusion of drug into the VTA.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim. May combine two claims.

    Study design and conductown data

    Recorded BNSTv units were classified as light-responsive, and thus VTA-projecting, if they met both criteria: latency of the first spike after light stimulation onset was less than 20 ms for ≥20% of trials, and light-evoked and spontaneous waveform shapes had a correlation coefficient >0.90.

    “Recorded BNSTv units were classified as light-responsive, and thus VTA-projecting, if they met both of the following two criteria. 1) The latency of the first spike after light stimulation onset was less than 20 ms for ≥ 20% of the trials. 2) Light-evoked and spontaneous waveform shapes had a correlation coefficient of > 0.90.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductnone given

    Neural activity was recorded from the same mice during cue exposure, foot shock, and cue re-exposure sessions, allowing reliable recording from the same population of neurons during each of the three sessions.

    “Neural activity was recorded from the same mice during the cue exposure, foot shock, and cue re-exposure sessions, therefore we were able to reliably record activity from the same population of neurons during each of the three sessions.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Timestamp data for foot-shock session start/end and light pulse delivery were synchronized with electrophysiological data.

    “In addition, timestamp data to signify the start and end of foot-shock sessions, and the delivery of light pulses to optical fibers was synchronized with electrophysiological data.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Mice were run daily at each photostimulation frequency (1, 5, 10, 20, 40 Hz) in a counterbalanced design.

    “Following the 1 day 20 Hz training session, mice were run daily at each photostimulation frequency (1, 5, 10, 20, 40 Hz) in a counter-balanced design.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    Shock artifacts were easily isolated and excluded from the analyzed waveform data using offline spike sorting.

    “While these were easily isolated and excluded from the analyzed waveform data using offline spike sorting”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    The injector needle extended approximately 1 mm past the cannula to ensure drug delivery 0.5 mm below the optical fiber.

    “The injector needle (33 gauge steel tube, McMasters-Carr) extended approximately 1 mm past the cannula to ensure drug delivery 0.5 mm below the optical fiber.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Mice received constant 20 Hz photostimulation during the entire 30 min sucrose session only after stable nose-poking behavior for 15% sucrose was observed (approximately 100 active nose pokes on at least 2 consecutive days).

    “Once stable nose-poking behavior for 15% sucrose was observed (approx. 100 active nose pokes on at least 2 consecutive days), all mice received constant 20 Hz photostimulation during the entire 30 min sucrose session.”
    Quote in the paperSubstantial claim
24 of 24

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Optogenetic studies show projection-specific effects of BNST neurons on behavior.

    “This is supported by optogenetic studies showing the projection-specific effects of BNST neurons on behavior (87–90).”

    From Sexual differentiation of neural mechanisms of stress sensitivity during puberty (2023) · cites this paper as [89]

  • Individual subregions of the BNST differentially regulate separable features of the anxiety phenotype.

    “Moreover, individual subregions of the BNST have been shown to differentially regulate separable features of the anxiety phenotype31,85 (Fig. 3).”

    From Resolving the neural circuits of anxiety (2015) · cites this paper as [85]

  • Activity in glutamatergic ventral BNST neurons projecting to the VTA produces anxiety-like behaviors, whereas GABAergic neurons in the ventral BNST exert anxiolysis via a parallel pathway.

    “activity in glutamatergic ventral BNST neurons projecting to the VTA produces anxiety-like behaviors, whereas GABAergic neurons in the ventral BNST exert anxiolysis via a parallel pathway85.”

    From Resolving the neural circuits of anxiety (2015) · cites this paper as [85]

  • The BNST contains distinct anxiogenic and anxiolytic GABAergic neuronal subsets with unique patterns of long-range connections.

    “It has recently been shown that the BNST contains distinct anxiogenic and anxiolytic GABAergic neuronal subsets with unique patterns of long-range connections (Jennings et al., 2013; Kim et al., 2013).”

    From Control of Stress-Induced Persistent Anxiety by an Extra-Amygdala Septohypothalamic Circuit (2014) · cites this paper as Jennings et al., 2013

  • Our results provide further evidence of the importance of long-range GABAergic projection neurons in the control of anxiety.

    “Our results provide further evidence of the importance of long-range GABAergic projection neurons in the control of anxiety (Jennings et al., 2013; Kim et al., 2013).”

    From Control of Stress-Induced Persistent Anxiety by an Extra-Amygdala Septohypothalamic Circuit (2014) · cites this paper as Jennings et al., 2013

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Recording electrode differs from the stated requirement for all in vivo experiments

    Minor

    The methods say mice in all in vivo electrophysiology experiments received a 16-wire array, whereas the anesthetized in vivo electrophysiology procedure describes recording with a glass electrode.

  • Injection laterality differs for an in vivo electrophysiology experiment

    Minor

    The methods say all in vivo electrophysiology mice received unilateral BNSTv injections, but the anesthetized in vivo electrophysiology procedure specifies bilateral injections.

2 of 2

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Related

Authors
Jennings, J. H., Sparta, D. R., Stamatakis, A. M., Ung, R. L., Pleil, K. E., Kash, T. L., & Stuber, G. D.
Affiliations
University of North Carolina at Chapel Hill (US)
Corresponding author
Stuber, Garret D.
ORCID
Venue
Nature
Publisher
Springer Science and Business Media LLC
Published
2013-03-20
Volume
496
Issue
7444
Pages
224-228
ISSN
  • 0028-0836 (ISSN-L)
  • 1476-4687
DOI
10.1038/nature12041
PMC
PMC3778934
OpenAlex
W2067203693
Semantic Scholar
b094f8c7bc24b675130d787985a6c38dc2555363
Cited by
704
References
33
Citations by year
2026: 34 · 2025: 53 · 2024: 30 · 2023: 44 · 2022: 35 · 2021: 65 · 2020: 43 · 2019: 55 · 2018: 57 · 2017: 55 · 2016: 78 · 2015: 66 · 2014: 58 · 2013: 29
FWCI
37.77
Type
journal article
Language
en
Open access
Licence
www.springer.com/tdm (tdm)
Locations
  • Nature (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
Funders
  • Foundation for the National Institutes of Health
  • Whitehall Foundation
  • National Institute on Alcohol Abuse and Alcoholism (F32AA018610, AA011605, AA 007573, F32AA021043, AA018610, AA021043, P60AA011605, T32AA007573)
  • National Institute on Drug Abuse (F31DA034472, R01DA032750, DA034472, DA029325, R21DA029325, DA032750, R37DA032750)
  • National Institute of Neurological Disorders and Stroke (P30NS045892, NS 007431, T32NS007431)
Topics
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neurotransmitter Receptor Influence on Behavior
  • Life Sciences › Neuroscience › Cognitive Neuroscience › Memory and Neural Mechanisms
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neuropharmacology Research
Keywords
bed nucleus of the stria terminalis, ventral tegmental area, extended amygdala, glutamatergic projections, GABAergic projections, anxiety, reward, aversion, motivational states, depression, channelrhodopsin-2, aversive stimuli
MeSH terms
Channelrhodopsins, Amygdala, Animals, Anxiety, Avoidance Learning, Behavior, Animal, Cues, Electroshock, Glutamine, Male, Mice, Inbred C57BL, Motivation, Phenotype, Reward, Septal Nuclei, Ventral Tegmental Area, Mice, GABAergic Neurons, Optogenetics

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