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Genetically identified amygdala–striatal circuits for valence-specific behaviors

Abstract

The basolateral amygdala (BLA) plays essential roles in behaviors motivated by stimuli with either positive or negative valence, but how it processes motivationally opposing information and participates in establishing valence-specific behaviors remains unclear. Here, by targeting Fezf2-expressing neurons in the BLA, we identify and characterize two functionally distinct classes in behaving mice, the negative-valence neurons and positive-valence neurons, which innately represent aversive and rewarding stimuli, respectively, and through learning acquire predictive responses that are essential for punishment avoidance or reward seeking. Notably, these two classes of neurons receive inputs from separate sets of sensory and limbic areas, and convey punishment and reward information through projections to the nucleus accumbens and olfactory tubercle, respectively, to drive negative and positive reinforcement. Thus, valence-specific BLA neurons are wired with distinctive input–output structures, forming a circuit framework that supports the roles of the BLA in encoding, learning and executing valence-specific motivated behaviors.

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Fig. 1: Fezf2 selectively labels a unique pyramidal neuron population in the BLA.
Fig. 2: Characterization of Fezf2+ and Fezf2 neuron response properties.
Fig. 3: Learning induces valence-specific predictive responses in Fezf2BLa neurons.
Fig. 4: Fezf2BLa neuron activity predicts reward seeking and punishment avoidance.
Fig. 5: Predictive valence-specific signals evolve in Fezf2BLa → NAc and Fezf2BLa → OT neurons during learning.
Fig. 6: Fezf2BLa → NAc and Fezf2BLa → OT neurons receive distinct sets of monosynaptic inputs.
Fig. 7: Fezf2BLa → NAc and Fezf2BLa → OT are differentially required for approach or avoidance.
Fig. 8: Fezf2BLa → NAc and Fezf2BLa → OT neurons differentially instruct learning.

Data availability

All data are contained in the main text, extended data or Supplementary Information. Source data can be downloaded at https://figshare.com/articles/dataset/NN-A72265C/15130017.

Code availability

Source code can be downloaded at https://figshare.com/articles/software/code_for_NN-A72265C/15157614.

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Acknowledgements

We thank T. Russo and R. Sharma for technical assistance, and members of the laboratory of B.L. for helpful discussions. This work was supported by grants from NARSAD (28229, to X.Z.; 27820, to K.Y.), EMBO (ALTF 458-2017, to A.F.), the Swedish Research Council (2017-00333, to A.F.), the NIH (R01MH101214, R01MH108924, R01NS104944 and R01DA050374, to B.L.; R01MH101268, to Z.J.H.), the Human Frontier Science Program (RGP0015/2016, to B.L.), the Stanley Family Foundation (to B.L.), the Simons Foundation (344904, to B.L.), the Wodecroft Foundation (to B.L.), the CSHL and Northwell Health Affiliation (to B.L.), Feil Family Neuroscience Endowment (to B.L.) and the Shanghai Rising-Star Program (18QA1400600, to M.H.).

Author information

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Authors

Contributions

X.Z. and B.L. conceived and designed the study. X.Z. conducted the experiments and analyzed data. W. Guan performed the patch-clamp recording experiments and assisted with rabies tracing experiments. T.Y. developed the one-photon wide-field imaging system and methods. A.F. performed the smFISH experiments. X.X. assisted with generating MATLAB codes for controlling behavioral devices. K.Y. assisted with the behavioral setup. X.A. and W. Galbavy assisted with characterization of Fezf2-expressing neurons. C.R and K.D. developed the Flp-off GCaMP6f virus. K.R. and A.H. developed the optimized rabies viral tracing system. M.H. designed and generated the Fezf2-CreER mouse line and characterized the labeling of neurons in the BLA. Z.J.H. conceived and oversaw the generation and characterization of the Fezf2-CreER and Fezf2-Flp mouse lines, and provided critical reagents and advice. X.Z. and B.L. wrote the paper with input from all authors.

Corresponding authors

Correspondence to Xian Zhang or Bo Li.

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Competing interests

The authors declare no competing interests.

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Peer review information Nature Neuroscience thanks Anna Beyeler, Sabine Krabbe, and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Fezf2BLa neurons constitute a subset of pyramidal neurons in the BLa.

a, Confocal images of a coronal brain section containing the BLA from a Fezf2-Flp mouse, showing the distribution of Flp and Fezf2 detected by smFISH, and overlay as indicated. In the right-most panel are high magnification images of the boxed area in the overlay image. b, Quantification of the fraction of Flp+ neurons that co-express Fezf2, and vice versa (n = 3 mice). c, Confocal images of coronal brain sections from Fezf2-CreER;Ai14 mice, in which Fezf2+ neurons in the BLA are labeled with tdTomato (red). The BLa has the highest density of Fezf2+ neurons within the BLA complex. This experiment was repeated in 4 mice. d, Representative confocal images showing Fezf2BLa neurons labeled with tdTomato (left), BLa pyramidal neurons labeled by antibodies recognizing CaMKII (middle), and overlay (right). Arrows indicate Fezf2BLa cells expressing CaMKII. e, Representative images showing Fezf2BLa cells labeled with tdTomato (left), BLa inhibitory neurons labeled by antibodies recognizing GABA (middle), and overlay (right). Arrows indicate Fezf2BLa cells that do not express GABA. f, Quantification of the fraction of Fezf2BLa neurons that are pyramidal neurons or GABAergic neurons (n = 4 mice). g, Quantification of the fraction of BLa pyramidal neurons that are Fezf2+ neurons or not (n = 4 mice). Data are presented as mean ± s.e.m.

Extended Data Fig. 2 Fezf2BLa neurons are not PPP1R1B +neurons.

a, Confocal images of coronal brain sections containing the BLA along the antero-posterior axis from a representative Fezf2-CreER;Ai14 mouse, showing the distribution of Fezf2+ neurons labeled with tdTomato, and the distribution of PPP1R1B expression detected with antibodies. In the rightmost panel are high magnification images of the boxed area in the corresponding overlay images. b, Quantification of the distribution of Fezf2+ neurons and PPP1R1B +neurons in the BLa and BLp (data from 4 mice). Note that the vast majority of Fezf2+ neurons are in the BLa, whereas essentially no PPP1R1B +neurons are in the BLa. c, Quantification of the overlap between neurons that express Fezf2 and PPP1R1B in the BLa (left) and BLp (right) in the same mice as in (b). Values in matrix represent the percent labeling with markers in columns among neurons labeled with markers in rows. For example, in the BLa (left), 0% of Fezf2 (tdTomato)-labeled neurons were labeled with PPP1R1B.

Extended Data Fig. 3 Fezf2BLa neurons are predominately Rspo2+ neurons.

a, Confocal images of coronal brain sections containing the BLA along the antero-posterior axis from a representative Fezf2-CreER;H2B-GFP mouse, showing the distribution of Fezf2+ neurons labeled with H2B-GFP, and the distribution of Rspo2 and Ppp1r1b expression detected with smFISH. This experiment was repeated in 2 mice. b, High magnification images of the boxed areas in the corresponding overlay images in (a). Arrows in the left three panels indicate that, in the BLa, Fezf2+ neurons expressing Rspo2. Arrows in the rightmost panel indicate that, in the BLp, many Ppp1r1b+ neurons express Rspo2, but not Fezf2. This experiment was repeated in 2 mice.

Extended Data Fig. 4 Characterization of behavior and Fezf2BLa neuron activity in the AAA task.

a, Top: licking (left) and running (right) events, sorted according to trial types, for a representative mouse in the early stage of training in the AAA task. Bottom: average licking rate (left) or running velocity (right) of this mouse in different types of trials as indicated. b, Average lick rate (left) and running velocity (right) of all the mice in the different types of trials in the AAA task during early stage of training. c, d, Same as (a, b), respectively, except that mice were in the late stage of training in the AAA task. e, Heatmaps of the activities (z-scores) in individual Fezf2BLa neurons from a representative mouse, obtained from different types of trials in the early stage of training in the AAA task. Dashed lines indicate the onset of CS, decision window and US, as indicated. Each row in each type of trials (reward, neutral and punishment) represents the temporal activities of one neuron. Neurons are sorted according to their average z-scores during the presentation of different CSs. f, Left: heatmaps of the activities (z-scores) in individual Fezf2BLa neurons from a representative mouse, obtained from different types of trials in the late stage of training in the AAA task. Dashed lines indicate the onset of CS, decision window and US as indicated. Each row represents the temporal activities of the same neuron in reward (left), neutral (middle) and punishment (right) trials. Neurons are sorted according to their average z-scores during the presentation of CSR in the reward trials. Right: same dataset as that in the left, except that neurons are sorted according to their average z-scores during the presentation of CSP in the punishment trials. g, Left: heat-maps of the responses of Fezf2BLa neurons to CSR (left) and CSP (right) in the late training stage in the AAA task. Responses from all Fezf2BLa neurons in all the mice (n = 10) are shown. Each row represents the responses of the same neuron to CSR and CSP. Right: PCA was applied to the CS responses in the left, followed by hierarchical clustering using the first three principal components (PCs) to sort Fezf2BLa neurons into three clusters. h, The average responses of all neurons in each of the three clusters to different CSs. Data are presented as mean ± s.e.m. Shaded areas represent s.e.m.

Extended Data Fig. 5 Anterograde and retrograde tracing of Fezf2BLa neurons.

a-i, Characterization of axonal projections from Fezf2BLa neurons in target areas. a, A schematic of the approach and the sections used to quantify GFP fluorescence in the major target areas (mPFC, medial prefrontal cortex; VS, ventral striatum; LS, lateral septum). b, Upper: diagrams of the four major target areas. Lower: representative confocal images of coronal brain sections containing the corresponding target areas. Areas selected for analysis are marked by red boxes. c, Quantification of axonal fluorescence signals as the fluorescence intensity in target areas normalized by the intensity in the NAc for each animal (n = 4 mice; Friedman test (F-stat 11.1) with Dunn’s post-hoc test: P = 0.0009; n.s., P > 0.05; *P< 0.05; **P < 0.01). d, Left: a schematic of the approach to selectively label Fezf2BLa → NAc neurons. Right: a confocal image showing injection location of AAVrg-fDIO-Cre in the NAc, as indicated by fluorescent beads (arrow). e, Representative confocal images showing axons of Fezf2BLa → NAc neurons in the mPFC (left), VS (middle) and LS (right). f, Quantification of Fezf2BLa → NAc neuron axonal fluorescence signals. Data are represented as the fluorescence intensity in target areas normalized by the intensity in the NAc for each animal (n = 4 mice; Friedman test (F-stat 11.15) with Dunn’s post-hoc test: P = 0.0006; n.s., P > 0.05; *P< 0.05). g, Left: a schematic of the approach to selectively label Fezf2BLa → OT neurons. Right: a confocal image showing injection location of AAVrg-fDIO-Cre in the OT, as indicated by fluorescent beads (arrow). h, Representative confocal images showing axons of Fezf2BLa → OT neurons in the mPFC (left), VS (middle) and LS (right). i, Quantification of Fezf2BLa → OT neuron axonal fluorescence signals. Data are represented as the fluorescence intensity in target areas normalized by the intensity in the OT for each animal (n = 4 mice; Friedman test (F-stat 11.1) with Dunn’s post-hoc test: P = 0.0009; n.s., P > 0.05; *P< 0.05; **P < 0.01). j-l, Retrograde tracing of Fezf2BLa → NAc and Fezf2BLa → OT neurons with CTB. j, A schematic of the approach (left) and a histology image (right) showing CTB-647 and CTB-555 injected into the NAc and OT of Fezf2-CreER;LSL-H2B-GFP mice, respectively. k, Representative confocal images of the BLa in the same mouse as that in (j), showing Fezf2BLa neurons labelled by H2B-GFP, CTB-555 and CTB-647. In the lower panel are high magnification images of the boxed area in the image in the upper panel. Arrowheads and arrows indicate the Fezf2BLa → OT and Fezf2BLa → NAc neurons, respectively; on the upper right corner (indicated by a vertical arrow) is a Fezf2BLa neuron projecting to both the NAc and the OT (triple positive for H2B-GFP, CTB-555 and CTB-647). l, Quantification of the overlap between Fezf2BLa → OT and Fezf2BLa → NAc neurons. Data are presented as mean ± s.e.m.

Extended Data Fig. 6 Characterization of Fezf2BLa → OT and Fezf2BLa → NAc neurons.

a, Confocal images of coronal brain sections containing the BLa or BLp along the antero-posterior axis from a representative Fezf2-CreER mouse, showing the distribution of NAc- or OT-projecting Fezf2+ neurons. In the rightmost panel are high magnification images of the boxed area in the corresponding overlay images. b, A schematic of the approach to target OT- or NAc-projecting Fezf2+ neurons in the BLA. c, Quantification of the proportion of overlapping neurons among Fezf2BLA → NAc neurons (13 ± 3.1%) or Fezf2BLA → OT neurons (11.8 ± 3.2%; 4 mice). d, Quantification of the OT- or NAc-projecting Fezf2+ neurons in the BLa and BLp. e, Quantification of the expression of PPP1R1B in NAc- or OT-projecting Fezf2+ neurons, in the BLa and BLp (n = 4; Friedman test (F-stat 10.1) with Dunn’s post-hoc test: P = 0.003; n.s., P > 0.05). Data are presented as mean ± s.e.m.

Extended Data Fig. 7 Functional connectivity between Fezf2BLa neurons and ventral striatum neurons.

a, A schematic of the intersectional approach to selectively target Fezf2BLa → NAc neurons for optogenetic activation. b, A schematic of the approach to record the synaptic responses of NAc and OT neurons to optogenetic activation of the axons originating from Fezf2BLa → NAc neurons. c, Traces of excitatory postsynaptic currents (EPSCs) recorded from two representative neurons, with one in the NAc (top) and the other in the OT (bottom), in the presence of TTX and 4-AP. The EPSCs were evoked by optogenetic activation of axon fibers originating from Fezf2BLa → NAc neurons. The upward square pulses (1 ms duration) in the blue traces (top) indicate the timing of photo-stimulation. d. Quantification of connection probability between Fezf2BLa → NAc neurons and neurons in the NAc or OT (36 out of 39 cells in the NAc, and 30 out of 36 cells in the OT had evoked EPSCs). e, Quantification of peak amplitudes of the EPSCs (NAc, 36 cells, OT, 30 cells; 6 mice; **P = 0.0018, unpaired t test). f-h, Same as a-c, respectively, except that Fezf2BLa → OT neurons were selectively targeted for optogenetic activation. i. Quantification of connection probability between Fezf2BLa → OT neurons and neurons in the NAc or OT (18 out of 20 cells in the NAc, and 17 out of 19 cells in the OT had evoked EPSCs). j, Quantification of peak amplitudes of the EPSCs (NAc, 18 cells, OT, 17 cells; 4 mice; n.s., P = 0.78, unpaired t test). Data are presented as mean ± s.e.m.

Extended Data Fig. 8 Chemogenetic inhibition of Fezf2BLa neurons impairs both reward seeking and punishment avoidance.

a, A schematic of the approach. b, Representative confocal images showing the BLa neurons expressing KORD. c, A schematic of the AAA task. d, Top: licking events sorted according to trial types (which were randomly interleaved) for a representative mouse before (left), during (middle) and after (right) chemogenetic inhibition of Fezf2BLa neurons. Bottom: the average rate of the licking responses in the corresponding top panels. e, Top: running events sorted according to trial types (which were randomly interleaved) for a representative mouse before (left), during (middle) and after (right) chemogenetic inhibition of Fezf2BLa neurons. Bottom: the average velocity of the running responses in the corresponding top panels. f, Chemogenetic inhibition of Fezf2BLa neurons reduced the licking responses during the decision window (n = 7 mice; F(2, 18) = 5.13, P = 0.01; Pre-SALB and SALB, *P = 0.0211; SALB and DMSO, *P = 0.0299; Pre-SALB and DMSO, n.s. (nonsignificant), P = 0.97; one-way ANOVA followed by Tukey’s multiple comparisons test). g, Chemogenetic inhibition of Fezf2BLa neurons did not affect the licking responses evoked by water (US) delivery to the mouth (n = 7; t(6) = 1.4, P = 0.22; paired t-test). h, Treatment with SALB does not change the licking responses during the decision window in control mice in which the Fezf2BLa neurons expressed GFP (n = 6; t(5) = -0.33, P = 0.75; paired t-test). i, Chemogenetic inhibition of Fezf2BLa neurons reduced the running responses during the decision window (n = 7 mice; F(2, 18) = 3.95, P = 0.03; Pre-SALB and SALB, *P = 0.0386; SALB and DMSO, *P = 0.0048; Pre-SALB and DMSO, n.s. (nonsignificant), P = 0.6742; one-way ANOVA followed by Tukey’s multiple comparisons test). j, Chemogenetic inhibition of Fezf2BLa neurons did not affect the running responses evoked by air-puff (US) blowing to the face (n = 7; t(6) = 0.95, P = 0.38; paired t-test). k, Treatment with SALB does not change the running responses during the decision window in control mice in which the Fezf2BLa neurons expressed GFP (n = 6; t(5) = -0.02, P = 0.98; paired t-test). Data are presented as mean ± s.e.m.

Extended Data Fig. 9 Optogenetic manipulation of Fezf2BLa → NAc and Fezf2BLa → OT neurons.

a-f, Optogenetic inhibition of Fezf2BLa → NAc and Fezf2BLa → OT neurons. a, A schematic of the approach to selectively inhibit Fezf2BLa → NAc neurons with optogenetics. b, Left: a confocal image showing Fezf2BLa → NAc neurons expressing stGtACR2. Locations of optical fibers for optogenetics are indicated. Right: a confocal image showing injection location of AAVrg-fDIO-Cre in the NAc, as indicated by fluorescent beads (arrow). c, A schematic of the approach to selectively inhibit Fezf2BLa → OT neurons with optogenetics. d, Left: a confocal image showing Fezf2BLa → OT neurons expressing stGtACR2. Locations of optical fibers for optogenetics are indicated. Right: a confocal image showing injection location of AAVrg-fDIO-Cre in the OT, as indicated by fluorescent beads (arrow). e, f, Schematics of the AAA task (e; blue bar on time axis indicates laser delivery) and experimental procedure (f). g-p, Optogenetic activation of Fezf2BLa → NAc and Fezf2BLa → OT neurons increases pupil size. g, A schematic of the approach to selectively activate Fezf2BLa → NAc neurons with optogenetics. h, Left: a confocal image showing Fezf2BLa → NAc neurons expressing ChR2. Locations of optical fibers for optogenetics are indicated. Right: a confocal image showing injection location of AAVrg-fDIO-Cre in the NAc, as indicated by fluorescent beads (arrow). i, A schematic of the approach to selectively activate Fezf2BLa → OT neurons with optogenetics. j, Left: a confocal image showing Fezf2BLa → OT neurons expressing ChR2. Locations of optical fibers for optogenetics are indicated. Right: confocal image showing injection location of AAVrg-fDIO-Cre in the OT, as indicated by fluorescent beads (arrow). k, Images of the pupil in a representative mouse, before (left) and after (right) photoactivation of Fezf2BLa → NAc neurons. l, Trial-by-trial pupil size changes in response to photoactivation (blue bar, 2 s) of Fezf2BLa → NAc neurons in an example mouse. m, Quantification of pupil size change in response to laser stimulation in the BLa in all the mice in which ChR2 (n = 9) or GFP (n = 7) was expressed in Fezf2BLa → NAc neurons (Kruskal-Wallis test (K-stat 18.88) with Dunn’s post-hoc test: P = 0.0003; ChR2: **P = 0.002; GFP: P = 0.99). n, o, same as (k, l), respectively, except that Fezf2BLa → OT neurons were photoactivated. p, Quantification of pupil size change in response to laser stimulation in the BLa in all the mice in which ChR2 (n = 10) or GFP (n = 7) was expressed in Fezf2BLa → OT neurons (Kruskal-Wallis test (K-stat 21.9) with Dunn’s post-hoc test: P < 0.0001; ChR2: ****P < 0.0001; GFP: P = 0.99).

Extended Data Fig. 10 Fezf2BLa → NAc and Fezf2BLa → OT projections differentially instruct valence-specific learning.

a, A schematic of the approach to stimulate the Fezf2BLa → NAc pathway. b, Confocal images of the BLa (left) and ventral striatum (right) from a representative mouse, showing Fezf2BLa neurons expressing ChR2 and ChR2+ axon fibers originating from Fezf2BLa neurons, respectively. The placement of optical fiber in the NAc is also shown. Antibodies recognizing SP were used to label the ventral pallidum, which lies in between the NAc and OT. c, d, Same as a, b, respectively, except that the Fezf2BLa → OT pathway was the target for optogenetic stimulation (c), and the optical fiber was placed over the OT (d). e, Images of the pupil in a representative mouse, before (left) and after (right) photoactivation of the Fezf2BLa → NAc pathway. f, Trial-by-trial pupil size changes in response to photoactivation (blue bar, 2 s) of the Fezf2BLa → NAc pathway in an example mouse. g, Quantification of pupil size change in response to laser stimulation in the NAc in all the mice in which ChR2 (n = 8) or GFP (n = 7) was expressed in Fezf2BLa → NAc neurons (F(1, 26) = 32.52, P < 0.0001; ****P < 0.0001, n.s., P = 0.9999; two-way ANOVA followed by Bonferroni’s multiple comparisons test). h, i, same as e, f, respectively, except that the Fezf2BLa → OT pathway was photoactivated. j, Quantification of pupil size change in response to laser stimulation in the OT in all the mice in which ChR2 (n = 8) or GFP (n = 7) was expressed in Fezf2BLa → OT neurons (F(1, 26) = 30.34, P < 0.0001; ***P < 0.0001, n.s., P = 0.9999; two-way ANOVA followed by Bonferroni’s multiple comparisons test). k, Movement trajectory of a representative mouse at baseline (left), or in a situation whereby entering the left (middle) or right (right) side of the chamber triggered photoactivation of the Fezf2BLa → NAc pathway. l, Quantification of mouse activity as shown in (k), for mice in which ChR2 (n = 8) or GFP (n = 7) was expressed in Fezf2BLa neurons. The ChR2 mice, but not the GFP mice, avoided the side associated with the photo-stimulation (F(1, 26) = 89.75, P < 0.0001; ****P < 0.0001, n.s., P > 0.05; two-way ANOVA followed by Bonferroni’s multiple comparisons test). m, n, Same as k, l, respectively, except that the Fezf2BLa → OT pathway was targeted. The ChR2 mice (n = 8), but not the GFP mice (n = 7), preferred the side associated with the photo-stimulation (F(1, 26) = 63.28, P < 0.0001; ****P < 0.0001, n.s., P > 0.05; two-way ANOVA followed by Bonferroni’s multiple comparisons test). o, A schematic of the approach. p, Raster plot of nose-poking events at active or inactive port, for a mouse in which ChR2 was expressed in Fezf2BLa neurons and photo-stimulation was delivered to the NAc at the active port. q, Quantification of nose-poking events in a 60-min session for mice in which ChR2 (n = 8) or GFP (n = 7) was expressed in Fezf2BLa neurons and photo-stimulation was delivered to the NAc at the active port (F(1, 26) = 1.4, P = 0.25; n.s., P > 0.05; two-way ANOVA). r, Quantification of nose-poke events of the ChR2 mice in q, in the first 2 mins of the reversal test, in which the active and inactive ports were switched (P = 0.57; Mann-Whitney test, U = 24). s, Same as p, except that photo-stimulation was delivered to the OT at the active port. t, Same as q, except that the Fezf2BLa → OT pathway was targeted. The ChR2 mice (n = 8), but not the GFP mice (n = 7), vigorously poked the active port (F(1, 26) = 35.43, P < 0.0001; ***P < 0.0001, n.s., P > 0.05; two-way ANOVA followed by Bonferroni’s multiple comparisons test). u, Quantification of nose-poke events of the ChR2 mice in t, in the first 2 mins of the reversal test, in which the active and inactive ports were switched (***P = 0.0003; Mann-Whitney test, U = 0.5). v, A schematic of the approach. w, Top: trial-by-trial heat-maps of running velocity for an example mouse in the active avoidance task; bottom: average running velocity of the mouse in early (first 10 trials) and late (last 10 trials) trials. x, Quantification of running velocity during the decision window in early (first 10) and late (last 10) trials (ChR2 mice, n = 8, GFP mice, n = 7; (F(1, 26) = 29.66, P < 0.0001; ****P < 0.0001, n.s., P > 0.05; two-way ANOVA followed by Bonferroni’s multiple comparisons test). y, z, Same as w, x, respectively, except that the Fezf2BLa → OT pathway was targeted (ChR2 mice, n = 8, GFP mice, n = 7; (F(1, 26) = 2.976, P = 0.096; n.s., P > 0.05; two-way ANOVA followed by Bonferroni’s multiple comparisons test). Data are presented as mean ± s.e.m.

Supplementary information

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Supplementary Figs. 1–8

Reporting Summary

Supplementary Video 1

Extraction of the spatial and temporal components of GCaMP6 signals from individual neurons with CNMF-E. Shown are videos of GCaMP6 signals from Fezf2+ BLA neurons imaged through a GRIN lens in a representative mouse (the same mouse as that in Fig. 2a) receiving water and air-puff presentations. These videos are short clips of movies stitched together, with each clip representing a trial (200 frames). These videos are played in synchrony, and represent the following contents: ‘raw data’, GCaMP6 signals without any processing; ‘denoised’, the remaining signals after subtraction of the background from the raw data; ‘demixed’, the deconvolved spatiotemporal activity for each neuron using CNMF-E. In the ‘demixed’ video, the contours of four representative neurons are traced and numbered. The temporal activities of these four neurons are displayed in the bottom, in which the moving line indicates the passage of time synchronized with all the videos, and the dashed green and red lines denote the onset of water and air puff, respectively. Neurons 1 and 2 are water-responsive neurons, whereas neurons 3 and 4 are air-puff-responsive neurons. The scale bar beside each of the videos or response traces denotes ΔF values. These four neurons are also identified and labeled in Fig. 2a. See Methods for more information.

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Zhang, X., Guan, W., Yang, T. et al. Genetically identified amygdala–striatal circuits for valence-specific behaviors. Nat Neurosci 24, 1586–1600 (2021). https://doi.org/10.1038/s41593-021-00927-0

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