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Stochastic synaptic plasticity underlying compulsion in a model of addiction

Naturevolume 564pages366371 (2018) | Download Citation

Abstract

Activation of the mesolimbic dopamine system reinforces goal-directed behaviours. With repetitive stimulation—for example, by chronic drug abuse—the reinforcement may become compulsive and intake continues even in the face of major negative consequences. Here we gave mice the opportunity to optogenetically self-stimulate dopaminergic neurons and observed that only a fraction of mice persevered if they had to endure an electric shock. Compulsive lever pressing was associated with an activity peak in the projection terminals from the orbitofrontal cortex (OFC) to the dorsal striatum. Although brief inhibition of OFC neurons temporarily relieved compulsive reinforcement, we found that transmission from the OFC to the striatum was permanently potentiated in persevering mice. To establish causality, we potentiated these synapses in vivo in mice that stopped optogenetic self-stimulation of dopamine neurons because of punishment; this led to compulsive lever pressing, whereas depotentiation in persevering mice had the converse effect. In summary, synaptic potentiation of transmission from the OFC to the dorsal striatum drives compulsive reinforcement, a defining symptom of addiction.

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The dataset is available from https://doi.org/10.5281/zenodo.1474531.

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Acknowledgements

We thank E. C. O’Connor for discussion and comments on the manuscript; C. Gerfen for providing Cre-mouse lines through the MMRC repository. This study was financed by a grant from the Swiss National Science Foundation (Ambizone grant to P.V. and core grant to C.L.), the National Center of Competence in Research (NCCR) SYNAPSY-The Synaptic Bases of Mental Diseases, and an advanced grant from the European Research Council (MeSSI).

Reviewer information

Nature thanks J. P. Britt and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Author information

Affiliations

  1. Department of Basic Neurosciences, Faculty of Medicine, University of Geneva, Geneva, Switzerland

    • Vincent Pascoli
    • , Agnès Hiver
    • , Ruud Van Zessen
    • , Michaël Loureiro
    • , Ridouane Achargui
    • , Masaya Harada
    • , Jérôme Flakowski
    •  & Christian Lüscher
  2. Clinic of Neurology, Department of Clinical Neurosciences, Geneva University Hospital, Geneva, Switzerland

    • Christian Lüscher

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Contributions

V.P. conceived the experiments and performed patch recordings and behavioural experiments. A.H. did surgeries for viral infection, behavioural experiments and in vivo recordings. R.A. carried out the retrograde tracing with the rabies strategy. R.V.Z. implanted the photometry and carried out analyses and recordings. M.L. and M.H. carried out patch recordings. J.F. carried out clustering analysis. C.L. conceptualized and supervised the study, and prepared the manuscript with the help of all authors.

Competing interests

C.L. is a member of the following scientific advisory boards: Stalicla SA; Phénix Foundation; International research in paraplegia (IRP) Foundation, Geneva.

Corresponding author

Correspondence to Christian Lüscher.

Extended data figures and tables

  1. Extended Data Fig. 1 No correlation between perseverance and VTA infection.

    a, Serial coronal sections of a renouncing and a persevering mouse centred on the VTA infected with AAV5-EF1a-DIO-ChR2-eYFP and nuclear Hoechst staining (Ho). We infected 109 mice and took coronal images of brains from 71 mice. b, High-magnification images of VTA. c, oDASS perseverance as a function of the infection rate and group data (n = 71 mice, Pearson’s r = 0.075). Infection rate was determined as the number of ChR2–eYFP-positive cells normalized to the total number of cells based on Hoechst staining. Note that mice from which sagittal sections were obtained are not included in this quantification. d, Coronal sections of a renouncing and a persevering mouse that show VTA as above. Sections were additionally stained for tyrosine hydroxylase (TH) using a Cy3-conjugated secondary antibody. Staining was performed in slices from 15 mice. e, High-magnification images of VTA. f, Quantification of TH-positive neurons in the VTA from a subset of renouncing and persevering mice. g, Percentage of TH neurons infected with ChR2. Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics.

  2. Extended Data Fig. 2 Examples from two mice of lever presses and outcomes during oDASS.

    Raster plots for two mice, one keeping a high stimulation rate during punished sessions (example 2) and one falling to a low stimulation rate during punished sessions (example 1). Every action and the associated outcome as a function of time is shown for three baseline (B) and four punished (P) sessions.

  3. Extended Data Fig. 3 Emergence of two clusters of mice with punished oDASS.

    a, Hierarchical clustering of the entire dataset (time-event for eight parameters during three baseline and four punished sessions). Each column corresponds to a mouse. Heat maps for delays between the active lever presses, the number of laser stimulation events, the time of the last laser stimulation, the time remaining in a trial and the number of inactive lever press in one mouse are plotted. Two clusters are found in the resulting dendrogram (green, renouncer; pink, perseverer). Vertical dashed lines separate renouncing from persevering mice. Heat map (greyscale) represents the perseverance (measured as the oDASS rate during punished sessions 3 and 4, normalized to the baseline sessions) of each mouse as a function of the clustering. b, Before clustering, we applied a nonlinear dimension reduction to project the high-dimensional dataset into a two-dimensional representation (left). Mapping the perseverance onto this map (right) shows that this variable can be used to categorize the mice as renouncing and persevering mice.

  4. Extended Data Fig. 4 Stability of perseverance and oDASS acquisition parameters.

    a, Three blocks of two baseline and four punished oDASS sessions were performed during a two-month period. Left, oDASS rate as a function of advancing sessions and average for the two groups. Middle, perseverance measured during punishment sessions 3–4 was compared to sessions 11–12. Perseverance was calculated as the average oDASS during the two punished sessions normalized to the corresponding baseline rate. No effect of punishment block was detected. Right, correlation between perseverance between first and last block (sessions 3–4 versus session 11–12), Pearson’s r = 0.88 (n = 26). b, Left, perseverance and baseline rate for male and female mice of the two clusters (for perseverance, ANOVA followed two-sided t-test: *P < 0.0001, t60 = 23.02 for persevering versus renouncing female mice (n = 39 and 23 mice, respectively); *P < 0.0001, t45 = 22.30 for persevering versus renouncing male mice (n = 20 and 27 mice, respectively). Right, no difference in baseline rate was detected between the two clusters, nor between male and female mice (n = 109 mice). c, Left, cumulative active presses as a function of the progressive ratio. Middle, perseverance as a function of the breakpoint during the progressive ratio schedule for male (42, squares) and female (56, circles). Note that in Fig. 1e, data are presented with cumulative active presses, which avoids the steps that are observed inherent to the breakpoint plot. No difference in breakpoint was detected between the two clusters, nor between male and female mice (n = 98 mice). d, Temporal structure of an oDASS trial showing the delays between active press during baseline and punished sessions for cluster 1 and cluster 2 mice (renouncing and persevering mice, respectively). Delays were increased in renouncing mice (ANOVA followed by two-sided t-test: *P < 0.05 in punished sessions for persevering versus renouncing mice for every delay (n = 66 and 43 mice, respectively); #P < 0.05; baseline/punished for renouncing mice for every delay). Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics.

  5. Extended Data Fig. 5 Characterization of the connection between OFC and striatum.

    a, Retrograde tracing from the striatum with cholera toxin subunit B coupled to a red dye (CTB-555). Scale bar, 50 μm. Experiment was repeated in n = 8 mice. b, OFC–striatum optogenetic stimulation and recordings of EPSCs, blocked by the AMPAR antagonist (NBQX) and IPSCs blocked by GABAA antagonist (picrotoxin). Rise time and time to peak for IPSCs and EPSCs indicate a feed-forward circuit between principal neurons of the OFC and striatum (two-sided paired t-test for IPSCs versus EPSCs: t25 = 9.55, *P < 0.0001 and t25 = 7.57, P < 0.0001 for rise time and time to peak, respectively, n = 26 cells from five mice). Scale bars, 20 ms, 100 pA. c, Schematic of retrograde tracing from specific cell types of the striatum using a rabies virus that was injected in transgenic Cre-mouse lines encoding the dopamine D1 or D2 receptor and parvalbumin. A first injection (red) leads to cell-type-specific expression of the EnvA receptor TVA and the RG protein. After two weeks an EnvA-pseudotyped and glycoprotein (ΔG)-deleted rabies virus (EnvA and RVΔG–GFP) is injected (green) and taken up by the cells that express TVA and thus turn yellow (starters). Trans-complemented with glycoprotein by infection of the AAV8-CA-flex-RG and RVΔG–GFP transsynaptically caused a spread to upstream neurons (inputs). The injection site in the striatum (left) and high-magnification images show starter cells. Retrogradely infected neurons in the OFC at low and higher magnification (right). See Supplementary Table 1 for complete statistics. a, c, Images reproduced from Paxinos and Franklin44, copyright © 2001.

  6. Extended Data Fig. 6 Activity and manipulation of OFC–striatum projection during oDASS.

    a, Calcium signal (ΔF/F0, mean ± s.e.m.) around active press 1 and 2 of the FR3 schedule (numbers 1, 4, 7 and 2, 5, 8) during a baseline session for renouncing and persevering mice (green diamonds and pink bars indicate a significant deviation from baseline, n = 4 and 6 mice). b, Averaged calcium signal (ΔF/F0, mean ± s.e.m.) around active press 1 and 2 of the FR3 schedule (all but press number 8), around the active press number 8 (leading to the shock-associated cue) and around the lever press that terminates the non-shock FRs in punished sessions for renouncing and persevering mice (green diamonds and pink bars indicate a significant deviation from baseline, n = 4 and 6 mice). c, Example of trial activity map of the calcium signal (ΔF/F0, mean ± s.e.m.) around an unpredictable foot shock (500 ms, 0.25 mA, repeated in 10 times in one mouse). For each animal, 10 unpredictable foot shocks were delivered during a separate recording. Grouped data for the calcium signal (ΔF/F0, mean ± s.e.m.) around an unpredictable foot shock for renouncing and persevering mice (green diamonds and pink bars indicate a significant deviation from baseline, n = 4 and 6 mice). See Supplementary Table 1 for statistics. d, Scheme of a mouse brain infected with eArchT3.0–eYFP in the OFC and with ChR2–eYFP in the VTA (left). For persevering mice, OFC inhibition with eArchT3.0 between oDASS and the next FR initiation (or for a maximum of 90 s) delayed the next press of persevering mice (ANOVA followed by two-sided t-test: *P < 0.05 when comparing control versus eArchT3.0 delays during punished sessions, n = 13 mice). Perseverance changed (from 73% to 46%) as a consequence of eArchT3.0 stimulation (two-sided paired t-test: t12 = 9.13, *P < 0.0001, n = 13 mice for control versus eArchT3.0 before each FR initiation). e, For renouncing mice, OFC inhibition with eArchT3.0, after the punishment-predictive cue, between punished oDASS and the next FR initiation (or for a maximum of 90 s) or between each oDASS and the next FR initiation slightly delayed the next press (ANOVA followed two-sided paired t-test: *P < 0.05 when comparing control and eArchT3.0 delays during punished sessions). Perseverance was reduced as a consequence of eArchT3.0 stimulation between each oDASS and the next FR initiation (two-sided paired t-test: t7 = 2.62, *P = 0.034, n = 8 mice for control versus eArchT3.0 before each FR initiation). The oDASS rate during a baseline session was not significantly changed between punished oDASS and the next FR initiation (or for a maximum of 90 s) by inhibition with eArchT3.0. Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics. d, Line drawing modified from Paxinos and Franklin44, copyright © 2007.

  7. Extended Data Fig. 7 Plasticity at OFC–striatum synapses correlates with compulsive oDASS.

    a, Average of 10 sweeps for AMPAR EPSCs recorded at −70 mV and for EPSCs recorded at +40 mV in slices from renouncing and persevering mice, with or without SPN identification with tdTomato. NMDA amplitude was analysed 20 ms after the peak of the EPSC recorded at +40 mV. The AMPAR/NMDAR ratio for every recorded neuron as a function of perseverance, data are mean ± s.e.m. per animal and correlation (Pearson’s r = 0.84, P < 0.0001; n = 127 cells from 16 mice). Data are mean AMPAR/NMDAR ratio for renouncing and persevering mice (ANOVA followed by two-sided t-test: t17 = 3.13, *P = 0.007 and t19 = 3.53; *P = 0.002 for tdTomato+ and tdTomato, respectively (n = 8 and 11 mice; 9 and 12 cells); t85 = 6.68, *P < 0.0001, renouncing versus persevering for non-identified neurons (42 and 45 cells, respectively)). Data are mean AMPAR/NMDAR ratio for naive mice and for mice yoked to renouncing or persevering mice, with Drd1a–tdTomato identification (naive mice, n = 14 tdTomato+ cells from 5 mice and n = 12 tdTomato cells from 7 mice; mice yoked to renouncing mice, n = 6 tdTomato+ cells from 2 mice and n = 9 tdTomato cells from 2 mice; mice yoked to persevering mice, n = 6 tdTomato+ cells from 2 mice and n = 8 tdTomato cells from 2 mice). b, Average of 10 sweeps for AMPAR EPSCs recorded at −70 mV, with two short light pulses spaced with a 76-ms interval in slices from renouncing and persevering mice, with or without SPN identification using tdTomato. PPR is the ratio of the amplitudes of the second EPSC over the first. PPR for every recorded neuron as a function of perseverance, mean ± s.e.m. per animal and correlation (Pearson’s r = −0.77, P < 0.0001; n = 137 cells from 16 mice). Data are mean PPR for renouncing and persevering mice (ANOVA followed by two-sided t-test: t23 = 3.97, *P = 0.0005 and t22 = 1.72, P = 0.183 for tdTomato+ and tdTomato, respectively (n = 9 and 15 mice; 9 and 16 cells); t86 = 5.66, *P < 0.0001, renouncing versus persevering for non-identified neurons (n = 38 and 50 cells, respectively)). Data are mean PPR for naive mice and for mice yoked to renouncing or persevering mice, with Drd1a–tdTomato identification (naive mice, n = 13 tdTomato+ cells from 5 mice and n = 11 tdTomato cells from 7 mice; mice yoked to renouncing mice, n = 6 tdTomato+ cells from 2 mice and n = 10 tdTomato cells from 2 mice; mice yoked to persevering mice, n = 7 tdTomato+ cells from 2 mice and n = 8 tdTomato cells from 2 mice). c, Left, average of 10 sweeps for AMPAR EPSCs recorded at −70, 0 and +40 mV in slices from renouncing and persevering mice, with or without SPN identification using tdTomato. The rectification of the AMPAR EPSCs was calculated as the ratio of the chord conductance calculated at negative potential divided by chord conductance at positive potential. Middle, rectification index for every recorded neuron as a function of perseverance, mean ± s.e.m. per animal and correlation (Pearson’s r = 0.008, P = 0.971; n = 121 cells from 16 mice). Right, rectification index for renouncing and persevering in tdTomato+ or tdTomato cells (8 and 11 mice; 9 and 11 cells) and in unidentified SPNs (42 and 40 cells, respectively). Data are mean rectification index for naive mice and for mice yoked to renouncing or persevering mice with Drd1a–tdTomato identification (naive mice, n = 13 tdTomato+ cells from 5 mice and n = 11 tdTomato cells from 7 mice; mice yoked to renouncing mice, n = 6 tdTomato+ cells from 2 mice and n = 9 tdTomato cells from 2 mice; mice yoked to persevering mice, n = 7 tdTomato+ cells from 2 mice and n = 8 tdTomato cells from 2 mice). d, Average of 10 sweeps for EPSCs recorded at −70 mV and IPSCs recorded at 0 mV in slices from renouncing and persevering mice and in slices from naive mice. Five pulses were given at different frequencies (5, 10, 20 and 40 Hz) and the charge transfer was measured (area under the curve). The excitatory/inhibitory ratio (E/I) was calculated as the ratio charge transfer for EPSCs over IPSCs. The charge transfer of EPSCs was higher in slices from persevering mice at low frequencies (ANOVA followed by two-sided t-test: t27 = 4.75, *P < 0.0001; t27 = 3.75, *P = 0.0007; t27 = 2.37, P = 0.057; t27 = 1.64, P = 0.306 for 5, 10, 20 and 40 Hz, respectively (17 and 12 cells)). The charge transfer of IPSCs was not different between persevering and renouncing mice (17 and 12 cells, respectively). The ratio of charge transfer for EPSCs over IPSCs was higher in slices from persevering mice (ANOVA followed by two-sided t-test: t27 = 6.22, *P < 0.0001; t27 = 4.39, *P < 0.0001; t27 = 4.07, P = 0.0002; t27 = 3.67, P = 0.001 for 5, 10, 20 and 40 Hz, respectively (17 and 12 cells)). Measurements were obtained from four renouncing mice, three persevering mice and three naive mice. e, Average of 10 sweeps for AMPAR EPSCs in the presence of D-AP5 (50 μM) and NMDAR EPSCs isolated by subtraction for oDASS mice and for naive mice. Mean AMPAR/NMDAR ratio, PPR and rectification index for naive and oDASS mice (*P < 0.05 for t-test comparing naive/oDASS). Each dot represents the mean ± s.e.m. for all cells obtained in a given mouse. Recordings were obtained from six naive mice and seven oDASS mice (PPR: 48 cells from oDASS mice compared to 29 cells from naive mice; rectification index: 52 cells from oDASS mice compared to 24 cells from naive mice; AMPAR/NMDAR ratio with pharmacological isolation: 42 cells from oDASS mice compared to 23 cells from naive mice; AMPAR/NMDAR ratio without pharmacological isolation: 52 cells from oDASS mice compared to 26 cells from naive mice). Scale bars, 200 pA, 50 ms. Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics.

  8. Extended Data Fig. 8 Synaptic properties in persevering mice after 20-Hz stimulation in vivo.

    a, Average traces for EPSCs recorded to determine PPR, rectification index (RI) and AMPAR/NMDAR ratio without pharmacological isolation. Ex vivo measurement of the PPR, rectification index and AMPAR/NMDAR ratio after in vivo stimulation of OFC–striatum terminals at 20 Hz for 1 min in renouncing mice (for PPR: control versus 20 Hz, two-sided t-test: t45 = 0.48, P = 0.63, n = 29 and 18 cells, respectively; for rectification index: control versus 20 Hz: t53 = 0.90, P = 0.37, n = 32 and 23 cells respectively; for AMPAR/NMDAR ratio: control versus 20 Hz: t58 = 6.79, *P < 0.0001, n = 32 and 28 cells, respectively). b, Effect of 20-Hz stimulation of OFC–striatum prior to a baseline session (n = 8 mice). Delay to engage the next action was not changed (ANOVA followed by two-sided t-test: *P < 0.05 when comparing control and 20 Hz). oDASS rate was not modified by 20 Hz stimulation before a baseline session (t7 = 0.97, P = 0.36, n = 8 mice for control versus 20 Hz before a baseline session). Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics.

  9. Extended Data Fig. 9 Absence of an effect on compulsive behaviour by normalization of release probability in persevering mice.

    a, Average traces for EPSCs recorded immediately before and 30 min after the LTD protocol (10 Hz for 5 min) and grouped data. In slices from persevering mice, LTD is unmasked by bath application of mGluR5 PAM (CDPPB, 100 μM) and MK801 (NMDAR blocker, 10 μM) (two-sided t-test: t15 = 2.89, *P = 0.037, n = 8 and 9 cells, respectively). b, AMPAR/NMDAR ratio was left unchanged by in vivo stimulation of OFC–striatum terminals with 10 Hz in presence of MK801 and CDPPB (0.3 and 30 mg kg−1, respectively) in persevering mice (two-sided t-test: t53 = 1.86, P = 0.069 for control versus 10 Hz treated with MK801 and CDPPB (n = 29 and 26 cells, respectively)). c, PPR was normalized by in vivo stimulation of OFC–striatum terminals with 10 Hz in the presence of MK801 and CDPPB in persevering mice (two-sided t-test: t61 = 4.94, P < 0.0001 for control versus 10 Hz with MK801 and CDPPB (n = 38 and 25 cells, respectively)). The rectification index was different between controls and mice treated in vivo with the stimulation of OFC–striatum terminals at 10 Hz in the presence of MK801 and CDPPB (two-sided t-test: t53 = 2.25, *P = 0.029 for control versus 10 Hz with MK801 and CDPPB (n = 29 and 26 cells, respectively)). AMPAR/NMDAR ratio without pharmacological isolation was not different between controls and mice treated in vivo with the stimulation of OFC–striatum terminals at 10 Hz in the presence of MK801 and CDPPB (two-sided t-test: t54 = 1.57, P = 0.12 for control versus 10 Hz with MK801 and CDPPB (n = 30 and 26 cells, respectively)). d, Plots for delay between lever presses in punished sessions 12 h after 10 Hz, MK801 and CDPPB or 10 Hz with MK801and CDPPB (ANOVA followed by two-sided paired t-test: *P < 0.05 when comparing control/treatment delays during punished sessions). Perseverance is not modified by any treatment (two-sided t-test: t12 = 2.12, P = 0.056, n = 13 mice for control versus 10 Hz; t4 = 0.73, P = 0.51, n = 5 mice for control versus MK801 and CDPPB; t7 = 1.31, P = 0.231, n = 8 mice for control versus 10 Hz with MK801 and CDPPB). e, During additional punished sessions without renewal of the intervention, perseverance remained unchanged (n = 8 mice). Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics.

  10. Extended Data Fig. 10 Effects of SCH23390 and 1 Hz in vivo in persevering mice.

    a, Average traces of EPSCs recorded to determine PPR, rectification index and AMPAR/NMDAR ratio without pharmacological isolation. Ex vivo measurement of the PPR, rectification index and AMPAR/NMDAR ratio after in vivo stimulation of OFC–striatum terminals at 1 Hz for 5 min in persevering mice, in the presence or absence of SCH23390 (ANOVA followed by two-sided t-test: for PPR: t32 = 1.10, P = 0.83; t39 = 0.64, P > 0.99; and t43 = 0.57, P > 0.99 for control versus 1 Hz (n = 19 and 15 cells, respectively); 1 Hz versus 1 Hz with SCH23390 (n = 15 and 26 cells, respectively); and control versus 1 Hz with SCH23390 (n = 19 and 26 cells, respectively); for rectification index: t27 = 0.74, P > 0.99; t39 = 0.17, P > 0.99; and t38 = 1.00, P = 0.97 for control versus 1 Hz (n = 14 and 15 cells, respectively); 1 Hz versus 1 Hz with SCH23390 (n = 15 and 26 cells, respectively); and control versus 1 Hz with SCH23390 (n = 14 and 26 cells, respectively); for AMPAR/NMDAR ratio: t27 = 1.64, P = 0.32; t39 = 5.93, P < 0.0001; and t38 = 3.96, P = 0.0007 for control versus 1 Hz (n = 14 and 15 cells, respectively); 1 Hz versus 1 Hz with SCH23390 (n = 15 and 26 cells, respectively); and control versus 1 Hz with SCH23390 (n = 14 and 26 cells, respectively)). b, Delay to engage the next action was not changed (ANOVA followed by two-sided paired t-test: *P < 0.05 when comparing control versus 1 Hz or control versus 1 Hz with SCH23390, n = 10 mice). oDASS rate was not modified by 1 Hz or 1 Hz with SCH23390 prior to a baseline session (t5 = 0.22, P = 0.84, n = 6 mice for control versus 1 Hz before a baseline session and t9 = 1.48, P = 0.17, n = 10 mice for control versus 1 Hz with SCH23390 before a baseline session). Data are mean ± s.e.m. See Supplementary Table 1 for complete statistics.

Supplementary information

  1. Supplementary Table

    This file contains a Statistical Table: summary of statistical analysis referring to individual figures.

  2. Reporting Summary

  3. Video 1: Anterograde labeling of the OFC to DS projection.

    AAV8-hSyn-chrimson-tdTomato injected in the OFC anterogradely labeled fiber terminals in the centro-ventral part of the dorsal striatum. 3D reconstruction from lightsheet image stack after tissue clarification (see methods).

  4. Video 2: Retrograde tracing of the OFC to DS projection.

    Rabies virus strategy injected in transgenic D2 cre-mouse line. 3D reconstruction from lightsheet image stack after tissue clarification (see methods).

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https://doi.org/10.1038/s41586-018-0789-4

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