Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Cocaine exposure reorganizes cell type– and input-specific connectivity in the nucleus accumbens

Nature Neuroscience volume 17, pages 1198–1207 (2014)Cite this article

Subjects

This article has been updated

Abstract

Repeated exposure to cocaine alters the structural and functional properties of medium spiny neurons (MSNs) in the nucleus accumbens (NAc). These changes suggest a rewiring of the NAc circuit, with an enhancement of excitatory synaptic connections onto MSNs. However, it is unknown how drug exposure alters the balance of long-range afferents onto different cell types in the NAc. Here we used whole-cell recordings, two-photon microscopy, optogenetics and pharmacogenetics to show how repeated cocaine exposure alters connectivity in the mouse NAc medial shell. Cocaine selectively enhanced amygdala innervation of MSNs expressing D1 dopamine receptors (D1-MSNs) relative to D2-MSNs. We also found that amygdala activity was required for cocaine-induced changes to behavior and connectivity. Finally, we established how heightened amygdala innervation can explain the structural and functional changes evoked by cocaine. Our findings reveal how exposure to drugs of abuse fundamentally reorganizes cell type– and input-specific connectivity in the NAc.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Repeated cocaine exposure enhances excitatory connectivity.
Figure 2: Synaptic plasticity is both cell type– and input-specific.
Figure 3: Changes to the number and strength of connections.
Figure 4: Alterations to structural connections at dendritic spines.
Figure 5: Synaptic reorganization depends on NMDARs.
Figure 6: Enhancement of BLA inputs depends on BLA activity.
Figure 7: Enhanced connectivity reflects BLA innervation.

Similar content being viewed by others

Change history

  • 19 August 2014

    In the version of this article initially published online, the top of the y axis in the right subpanel of Figure 1e was labeled 100. The correct value is 10. The error has been corrected for the print, PDF and HTML versions of this article.

References

  1. Humphries, M.D. & Prescott, T.J. The ventral basal ganglia, a selection mechanism at the crossroads of space, strategy, and reward. Prog. Neurobiol. 90, 385–417 (2010).

    PubMed  Google Scholar 

  2. Sesack, S.R. & Grace, A.A. Cortico-Basal Ganglia reward network: microcircuitry. Neuropsychopharmacology 35, 27–47 (2010).

    PubMed  Google Scholar 

  3. Wilson, C.J. Basal Ganglia in The Synaptic Organization of the Brain (ed., G.M. Shepherd) 361–414 (Oxford University Press, 2004).

  4. Hyman, S.E., Malenka, R.C. & Nestler, E.J. Neural mechanisms of addiction: the role of reward-related learning and memory. Annu. Rev. Neurosci. 29, 565–598 (2006).

    CAS  PubMed  Google Scholar 

  5. Kalivas, P.W. The glutamate homeostasis hypothesis of addiction. Nat. Rev. Neurosci. 10, 561–572 (2009).

    CAS  PubMed  Google Scholar 

  6. Luscher, C. & Malenka, R.C. Drug-evoked synaptic plasticity in addiction: from molecular changes to circuit remodeling. Neuron 69, 650–663 (2011).

    PubMed  PubMed Central  Google Scholar 

  7. Robinson, T.E. & Kolb, B. Structural plasticity associated with exposure to drugs of abuse. Neuropharmacology 47 (suppl. 1): 33–46 (2004).

    CAS  PubMed  Google Scholar 

  8. Russo, S.J. et al. The addicted synapse: mechanisms of synaptic and structural plasticity in nucleus accumbens. Trends Neurosci. 33, 267–276 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Schmidt, H.D. & Pierce, R.C. Cocaine-induced neuroadaptations in glutamate transmission: potential therapeutic targets for craving and addiction. Ann. NY Acad. Sci. 1187, 35–75 (2010).

    CAS  PubMed  Google Scholar 

  10. Wolf, M.E. The Bermuda Triangle of cocaine-induced neuroadaptations. Trends Neurosci. 33, 391–398 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Brown, T.E. et al. A silent synapse-based mechanism for cocaine-induced locomotor sensitization. J. Neurosci. 31, 8163–8174 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Dietz, D.M. et al. Rac1 is essential in cocaine-induced structural plasticity of nucleus accumbens neurons. Nat. Neurosci. 15, 891–896 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Dumitriu, D. et al. Subregional, dendritic compartment, and spine subtype specificity in cocaine regulation of dendritic spines in the nucleus accumbens. J. Neurosci. 32, 6957–6966 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Kim, J., Park, B.H., Lee, J.H., Park, S.K. & Kim, J.H. Cell type-specific alterations in the nucleus accumbens by repeated exposures to cocaine. Biol. Psychiatry 69, 1026–1034 (2011).

    CAS  PubMed  Google Scholar 

  15. LaPlant, Q. et al. Dnmt3a regulates emotional behavior and spine plasticity in the nucleus accumbens. Nat. Neurosci. 13, 1137–1143 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Maze, I. et al. Essential role of the histone methyltransferase G9a in cocaine-induced plasticity. Science 327, 213–216 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Gerfen, C.R. et al. D1 and D2 dopamine receptor-regulated gene expression of striatonigral and striatopallidal neurons. Science 250, 1429–1432 (1990).

    CAS  PubMed  Google Scholar 

  18. Lobo, M.K. & Nestler, E.J. The striatal balancing act in drug addiction: distinct roles of direct and indirect pathway medium spiny neurons. Front. Neuroanatomy 5, 41 (2011).

    Google Scholar 

  19. Smith, Y., Bevan, M.D., Shink, E. & Bolam, J.P. Microcircuitry of the direct and indirect pathways of the basal ganglia. Neuroscience 86, 353–387 (1998).

    CAS  PubMed  Google Scholar 

  20. Kravitz, A.V., Tye, L.D. & Kreitzer, A.C. Distinct roles for direct and indirect pathway striatal neurons in reinforcement. Nat. Neurosci. 15, 816–818 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Tai, L.H., Lee, A.M., Benavidez, N., Bonci, A. & Wilbrecht, L. Transient stimulation of distinct subpopulations of striatal neurons mimics changes in action value. Nat. Neurosci. 15, 1281–1289 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Yawata, S., Yamaguchi, T., Danjo, T., Hikida, T. & Nakanishi, S. Pathway-specific control of reward learning and its flexibility via selective dopamine receptors in the nucleus accumbens. Proc. Natl. Acad. Sci. USA 109, 12764–12769 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Lee, K.W. et al. Cocaine-induced dendritic spine formation in D1 and D2 dopamine receptor-containing medium spiny neurons in nucleus accumbens. Proc. Natl. Acad. Sci. USA 103, 3399–3404 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Ferguson, S.M. et al. Transient neuronal inhibition reveals opposing roles of indirect and direct pathways in sensitization. Nat. Neurosci. 14, 22–24 (2011).

    CAS  PubMed  Google Scholar 

  25. Hikida, T., Kimura, K., Wada, N., Funabiki, K. & Nakanishi, S. Distinct roles of synaptic transmission in direct and indirect striatal pathways to reward and aversive behavior. Neuron 66, 896–907 (2010).

    CAS  PubMed  Google Scholar 

  26. Lobo, M.K. et al. Cell type-specific loss of BDNF signaling mimics optogenetic control of cocaine reward. Science 330, 385–390 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Ambroggi, F., Ishikawa, A., Fields, H.L. & Nicola, S.M. Basolateral amygdala neurons facilitate reward-seeking behavior by exciting nucleus accumbens neurons. Neuron 59, 648–661 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Britt, J.P. et al. Synaptic and behavioral profile of multiple glutamatergic inputs to the nucleus accumbens. Neuron 76, 790–803 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Pascoli, V. et al. Contrasting forms of cocaine-evoked plasticity control components of relapse. Nature 509, 459–464 (2014).

    CAS  PubMed  Google Scholar 

  30. Pascoli, V., Turiault, M. & Luscher, C. Reversal of cocaine-evoked synaptic potentiation resets drug-induced adaptive behaviour. Nature 481, 71–75 (2012).

    CAS  Google Scholar 

  31. Stuber, G.D. et al. Excitatory transmission from the amygdala to nucleus accumbens facilitates reward seeking. Nature 475, 377–380 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Voorn, P., Vanderschuren, L.J., Groenewegen, H.J., Robbins, T.W. & Pennartz, C.M. Putting a spin on the dorsal-ventral divide of the striatum. Trends Neurosci. 27, 468–474 (2004).

    CAS  PubMed  Google Scholar 

  33. MacAskill, A.F., Little, J.P., Cassel, J.M. & Carter, A.G. Subcellular connectivity underlies pathway-specific signaling in the nucleus accumbens. Nat. Neurosci. 15, 1624–1626 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Li, Y. et al. Both glutamate receptor antagonists and prefrontal cortex lesions prevent induction of cocaine sensitization and associated neuroadaptations. Synapse 34, 169–180 (1999).

    CAS  PubMed  Google Scholar 

  35. Gong, S. et al. A gene expression atlas of the central nervous system based on bacterial artificial chromosomes. Nature 425, 917–925 (2003).

    CAS  PubMed  Google Scholar 

  36. Shuen, J.A., Chen, M., Gloss, B. & Calakos, N. Drd1a-tdTomato BAC transgenic mice for simultaneous visualization of medium spiny neurons in the direct and indirect pathways of the basal ganglia. J. Neurosci. 28, 2681–2685 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Little, J.P. & Carter, A.G. Subcellular synaptic connectivity of layer 2 pyramidal neurons in the medial prefrontal cortex. J. Neurosci. 32, 12808–12819 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Little, J.P. & Carter, A.G. Synaptic mechanisms underlying strong reciprocal connectivity between the medial prefrontal cortex and basolateral amygdala. J. Neurosci. 33, 15333–15342 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Matsuzaki, M. et al. Dendritic spine geometry is critical for AMPA receptor expression in hippocampal CA1 pyramidal neurons. Nat. Neurosci. 4, 1086–1092 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Wolf, M.E. The role of excitatory amino acids in behavioral sensitization to psychomotor stimulants. Prog. Neurobiol. 54, 679–720 (1998).

    CAS  PubMed  Google Scholar 

  41. Wolf, M.E. & Jeziorski, M. Coadministration of MK-801 with amphetamine, cocaine or morphine prevents rather than transiently masks the development of behavioral sensitization. Brain Res. 613, 291–294 (1993).

    CAS  PubMed  Google Scholar 

  42. Armbruster, B.N., Li, X., Pausch, M.H., Herlitze, S. & Roth, B.L. Evolving the lock to fit the key to create a family of G protein-coupled receptors potently activated by an inert ligand. Proc. Natl. Acad. Sci. USA 104, 5163–5168 (2007).

    PubMed  PubMed Central  Google Scholar 

  43. Kourrich, S. & Thomas, M.J. Similar neurons, opposite adaptations: psychostimulant experience differentially alters firing properties in accumbens core versus shell. J. Neurosci. 29, 12275–12283 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Lee, B.R. et al. Maturation of silent synapses in amygdala-accumbens projection contributes to incubation of cocaine craving. Nat. Neurosci. 16, 1644–1651 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Huang, Y.H. et al. In vivo cocaine experience generates silent synapses. Neuron 63, 40–47 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Kourrich, S., Rothwell, P.E., Klug, J.R. & Thomas, M.J. Cocaine experience controls bidirectional synaptic plasticity in the nucleus accumbens. J. Neurosci. 27, 7921–7928 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Conrad, K.L. et al. Formation of accumbens GluR2-lacking AMPA receptors mediates incubation of cocaine craving. Nature 454, 118–121 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. McCutcheon, J.E., Wang, X., Tseng, K.Y., Wolf, M.E. & Marinelli, M. Calcium-permeable AMPA receptors are present in nucleus accumbens synapses after prolonged withdrawal from cocaine self-administration but not experimenter-administered cocaine. J. Neurosci. 31, 5737–5743 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Dobi, A., Seabold, G.K., Christensen, C.H., Bock, R. & Alvarez, V.A. Cocaine-induced plasticity in the nucleus accumbens is cell specific and develops without prolonged withdrawal. J. Neurosci. 31, 1895–1904 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  50. Penzes, P., Cahill, M.E., Jones, K.A., VanLeeuwen, J.E. & Woolfrey, K.M. Dendritic spine pathology in neuropsychiatric disorders. Nat. Neurosci. 14, 285–293 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Petreanu, L., Huber, D., Sobczyk, A. & Svoboda, K. Channelrhodopsin-2-assisted circuit mapping of long-range callosal projections. Nat. Neurosci. 10, 663–668 (2007).

    CAS  PubMed  Google Scholar 

  52. Yasuda, R. et al. Imaging calcium concentration dynamics in small neuronal compartments. Sci. STKE 2004, pl5 (2004).

    PubMed  Google Scholar 

  53. Carter, A.G. & Sabatini, B.L. State-dependent calcium signaling in dendritic spines of striatal medium spiny neurons. Neuron 44, 483–493 (2004).

    CAS  PubMed  Google Scholar 

  54. Rodriguez, A., Ehlenberger, D.B., Hof, P.R. & Wearne, S.L. Rayburst sampling, an algorithm for automated three-dimensional shape analysis from laser scanning microscopy images. Nat. Protoc. 1, 2152–2161 (2006).

    CAS  PubMed  Google Scholar 

  55. Zhu, H. et al. Chemogenetic inactivation of ventral hippocampal glutamatergic neurons disrupts consolidation of contextual fear memory. Neuropsychopharmacology 39, 1880–1892 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank members of the Carter laboratory, M. Farrant and R. Froemke for helpful discussions and comments on the manuscript, P. Kennedy for advice on cocaine administration, and C. Farb, R. Sears and H. Seong for help with anatomy. This work was supported by the Wellcome Trust (A.F.M.), and the Dana Foundation, McKnight Foundation and US National Institutes of Health R01DA038138 (A.G.C.).

Author information

Author notes

  1. Andrew F MacAskill

    Present address: Present address: Department of Neuroscience, Physiology and Pharmacology, University College London, London, UK.,

Authors and Affiliations

  1. Center for Neural Science, New York University, New York, New York, USA

    Andrew F MacAskill, John M Cassel & Adam G Carter

Authors
  1. Andrew F MacAskill
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. John M Cassel
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Adam G Carter
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

A.F.M. and A.G.C. designed the experiments. A.F.M. performed the experiments and analyzed the data. J.M.C. performed the stereotaxic injections. A.F.M. and A.G.C. wrote the paper.

Corresponding author

Correspondence to Adam G Carter.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Development of behavioral sensitization and viral injection sites.

a. Locomotion of saline (grey, top) or cocaine (black, bottom) mice, measured for 10 min immediately after injection on day 1 (left) and day 5 (right). Scale bar = 5 cm.

b. Summary of time course of behavioral sensitization. On day 0, all mice received a saline injection; on days 1 – 5, mice received either an injection of saline (grey) or 15 mg / kg cocaine (black); on day 8, all mice received a challenge injection of 15 mg / kg cocaine. Note that the effect of cocaine increases after each injection (cocaine day 1 vs. day 5: t(10) = 5.0, p = 0.001, paired t-test; saline vs. cocaine day 8: U = 14, p = 0.006, Mann-Whitney; n = 9 (s), 11 (c) mice). Summary shows mean ± SEM. * denotes p < 0.05.

c. Confocal images showing ChR2 expression at viral injection site in BLA (left), VH (center) and PFC (right). Scale bar = 100 μm.

Supplementary Figure 2 BLA and VH EPSCs at multiple stimulus intensities.

a. Average light-evoked BLA EPSCs recorded at neighboring D1‑MSNs (black) and D2‑MSNs (red) on day 8 from animals injected with saline on days 1 to 5. The duration of light pulse was varied between 0.5 ms and 2 ms. Darker colors show longer stimulus durations. Scale bar = 50 pA, 20 ms.

b. Summary of BLA EPCSs at D1-MSNs (black) and D2-MSNs (red). In saline conditions, there is no difference between the two cell types at any light intensity.

c, d. Similar to (a, b) but for mice injected with cocaine on days 1 to 5. After repeated cocaine exposure, D1‑MSNs have larger BLA EPSCs than neighboring D2-MSNs.

e. Summary of D1/D2 ratio for BLA EPSCs recorded in response to 1 ms light pulse in animals treated with either saline (s) or cocaine (c) (D1/D2 ratio: U = 5, p = 0.01, Mann-Whitney; n = 8 (s), 7 (c) pairs).

f - j. Similar to (a - e), but for VH EPSCs. Note that VH input is larger onto D1-MSNs in saline conditions, but this difference is no longer apparent after cocaine treatment (D1/D2 ratio: U = 11, p = 0.01, Mann-Whitney; n = 8 (s), 10 (c) pairs).

Summary shows mean ± SEM. * denotes p < 0.05.

Supplementary Figure 3 Repeated exposure to cocaine does not change PFC D1/D2 ratio.

a. (Top) Schematic of AAV9-CAG-hChR2-mCherry injection in medial prefrontal cortex (PFC, grey) and projections to the NAc. (Bottom) Coronal section of the NAc medial shell (dotted outline), showing distribution of PFC afferents. Scale bar = 100 μm.

b. Light-evoked PFC EPSCs at D1‑MSNs (mean in black, error in grey) and D2‑MSNs (mean in red, error in pink) on day 8 from animals injected with saline (left) or cocaine (right) on days 1 to 5. EPSCs at D1‑MSNs are normalized to those at neighboring D2‑MSNs. Scale bar = unity, 20 ms.

c. (Left) Summary of PFC EPSCs at D1-MSNs and D2-MSNs in saline (top) and cocaine (bottom) mice. (Right) Summary of D1/D2 ratio of PFC EPSCs. Note that cocaine does not alter the D1/D2 ratio of PFC EPSCs (D1/D2 ratio: U = 15, p = 0.7, Mann-Whitney; n = 6 pairs).

Summary shows mean ± SEM. * denotes p < 0.05.

Supplementary Figure 4 Repeated exposure to cocaine does not alter BLA or VH PPR.

a. Schematic of recording configuration showing neighboring D1‑MSNs (black) and D2‑MSNs (red) receiving BLA input. Afferent fibers were stimulated at distal locations in order to avoid confounds due to direct excitation of ChR2 in the presynaptic terminal.

b. Light-evoked BLA EPSCs at D1‑MSNs (black) and D2‑MSNs (red) on day 8 from animals injected with saline (left) or cocaine (right) on days 1 to 5. Inter-stimulus interval = 50 ms.

c. Summary of paired-pulse ratio (PPR) for BLA EPSCs at D1-MSNs and D2-MSNs in saline and cocaine mice (no significant effects, two-way ANOVA).

d. Summary of D1/D2 ratio of BLA PPR. Note that cocaine has no effect on BLA PPR (D1/D2 ratio: U = 27, p = 0.96, Mann-Whitney; n = 8 (s), 7 (c) pairs).

e - h. Similar to (a - d) but for VH input. Note that cocaine has no effect on VH PPR (no significant effects, two-way ANOVA; D1/D2 ratio: U = 21, p = 1, Mann-Whitney; n = 6 (s), 7 (c) pairs).

Summary shows mean ± SEM. * denotes p < 0.05.

Supplementary Figure 5 Synaptic calcium signals at dendritic spines.

a. Two-photon image of dendrites and spines of MSN filled with Alexa Fluor 594 and Fluo-4FF via the patch pipette, before (left) and after (right) two-photon glutamate uncaging. Orange circles show uncaging location. Note easily detectable calcium signal. Scale bar = 2 μm.

b. Summary of amplitudes of uncaging-evoked calcium signals at D1-MSNs (black) and D2-MSNs (red) with increasing distance from the soma (left) or with increasing spine volume (right). Note that all spines have detectable calcium signals, showing no change in amplitude with either distance along the dendrite or spine volume.

c. Summary of sampled dendrite locations for BLA (purple) and VH (blue) two-photon mapping experiments in saline (s) and cocaine (c) mice (D1/D2 ratio: BLA: U = 11, p = 0.54, n = 6 (s), 5 (c) pairs; VH: U = 11, p = 0.84, n = 5 (s), 5 (c) pairs; Mann-Whitney). Original data is shown in Fig. 4.

Summary shows mean ± SEM. * denotes p < 0.05.

Supplementary Figure 6 Additional MK-801 behavior experiments.

a. (Left) Schematic of experimental protocol. Mice were pretreated with MK-801 or saline 30 min before each saline injection on days 1 to 5, followed by a challenge injection of cocaine on day 8 to monitor behavior. (Right) Summary of distance traveled by saline- (s) and MK-801- (m) pretreated mice after the challenge injection of cocaine on day 8 (U = 14, p = 0.93, Mann-Whitney; n = 5 (s), 6 (m) mice).

b. (Left) Schematic of experimental protocol. Mice were pretreated with MK-801 or saline 30 min before a cocaine injection on days 1 to 5, followed by a baseline locomotion measurement on day 8. (Right) Summary of distance traveled by saline- (s) and MK‑801- (m) pretreated mice before any cocaine challenge on day 8 (U = 29, p = 0.54, Mann-Whitney; n = 8 (s), 9 (m) mice).

c. (Left) Schematic of experimental protocol. Mice were pretreated with MK-801 or saline 30 min before a challenge injection of cocaine on day 8. (Right) Summary of distance traveled by saline- (s) and MK-801- (m) pretreated mice after the challenge injection of cocaine on day 8 (U = 5, p = 0.1, Mann-Whitney; n = 5 (s), 6 (m) mice).

Summary shows mean ± SEM. * denotes p < 0.05.

Supplementary Figure 7 Characterization of DREADDs in the BLA.

a. Wide-field fluorescence image showing expression of mCitrine after viral injection of AAV8-CaMKIIa-HA-hM4D-IRES-mCitrine into the BLA. Scale bar = 100 μm.

b. Wide-field fluorescence image showing lack of expression of mCitrine in the VH after viral injection into the BLA. Scale bar = 100 μm.

c. Confocal image showing expression of hM4D-positive axons in the NAc after viral injection into the BLA. Scale bar = 10 μm.

d. Current-clamp recording from an hM4D-expressing BLA neuron, with current injection eliciting action potential firing before but not after bath application of 10 μM CNO. Scale bar = 20 mV, 400 ms.

e. Summary of effect of CNO on action potential firing at hM4D-expressing BLA neurons (t(4) = 4.5, p = 0.01, paired t-test; n = 5 cells).

f. Wide-field fluorescence image showing expression of mCitrine (green) and ChR2-mCherry (red) after viral injections into the BLA. Scale bar = 300 μm.

g. Enlargement of boxed region in (f), showing overlapping expression of mCitrine and ChR2-mCherry. Scale bar = 100 μm.

h. Confocal images showing co-expression of hM4D (green) and ChR2-mCherry (red) in BLA neurons. Scale bar = 20 μm.

I, j. Quantification of neurons in the BLA infected with ChR2 alone (red), hM4D alone (green), and both ChR2 and hM4D (orange).

Supplementary Figure 8 Additional BLA DREADD behavior experiments.

a. (Left) Schematic of experimental protocol. Mice were injected with AAVs to express hM4D bilaterally in the basolateral amygdala (BLA). Two weeks later, mice were pretreated with saline 30 min before each cocaine injection on days 1 to 5, followed by a challenge injection of cocaine on day 8 to monitor behavior. (Right) Summary of distance traveled by control or hM4D-expressing mice after cocaine challenge injection on day 8 (U = 20, p = 0.62, Mann-Whitney; n = 7 (Ctrl), 7 (hM4D) mice).

b. (Left) Schematic of experimental protocol. Mice were injected with AAVs to express hM4D bilaterally in the basolateral amygdala (BLA). Two weeks later, mice were pretreated with CNO 30 min before each saline injection on days 1 to 5, followed by a challenge injection of cocaine on day 8 to monitor behavior. (Right) Summary of distance traveled by control or hM4D-expressing mice after cocaine challenge injection on day 8 (U = 10, p = 0.69, Mann-Whitney; n = 5 (Ctrl), 5 (hM4D) mice).

c. (Left) Schematic of experimental protocol. Mice were injected with AAVs to express hM4D bilaterally in the basolateral amygdala (BLA). Two weeks later, mice were pretreated with CNO 30 min before each cocaine injection on days 1 to 5, followed by a baseline locomotion measurement on day 8. (Right) Summary of distance traveled by control or hM4D-expressing mice before any cocaine challenge on day 8 (U = 15, p = 0.28, Mann-Whitney; n = 6 (Ctrl), 8 (hM4D) mice).

Summary shows mean ± SEM. * denotes p < 0.05.

Supplementary Figure 9 Characterization of DREADDs in the VH.

a. Wide-field fluorescence image showing expression of mCitrine after viral injection of AAV8-CaMKIIa-HA-hM4D-IRES-mCitrine into the VH. Scale bar = 100 μm.

b. Wide-field fluorescence image showing lack of expression of mCitrine in the BLA after viral injection into the VH. Scale bar = 100 μm.

c. Confocal image showing expression of hM4D-positive axons in the NAc after viral injection into the VH. Scale bar = 10 μm.

d. Current-clamp recording from an hM4D-expressing VH neuron, with current injection eliciting action potential firing before but not after bath application of 10 μM CNO. Scale bar = 20 mV, 200 ms.

e. Summary of effect of CNO on action potential firing at hM4D-expressing VH neurons (t(4) = 3.8, p = 0.02, paired t-test; n = 5 cells).

f. Wide-field fluorescence image showing expression of mCitrine (green) and ChR2-mCherry (red) after viral injections into the VH. Scale bar = 300 μm.

g. Enlargement of boxed region in (f), showing overlapping expression of mCitrine and ChR2-mCherry. Scale bar = 100 μm.

h. Confocal images showing co-expression of hM4D (green) and ChR2-mCherry (red) in VH neurons. Scale bar = 20 μm.

i, j. Quantification of neurons in the VH infected with ChR2 alone (red), hM4D alone (green), and both ChR2 and hM4D (orange).

Supplementary Figure 10 Enhancement of VH inputs depends on VH activity.

a. Schematic of experimental protocol. Mice were injected with AAVs to express hM4D bilaterally in the ventral hippocampus (VH). Two weeks later, mice were pretreated with CNO 30 min before each cocaine injection on days 1 to 5, followed by a cocaine challenge on day 8 to monitor behavioral sensitization (b), or electrophysiology on day 8 without a cocaine challenge to assess synaptic connectivity (c - f).

b. (Left) Locomotion of control (top) or hM4D-expressing (bottom) mice, measured for 10 min immediately after a challenge injection of cocaine on day 8. Scale bar = 5 cm. (Right) Summary of distance traveled by control or hM4D-expressing mice after the cocaine challenge. Dashed line indicates saline control. Note that inhibiting VH activity does not prevent behavioral sensitization (U = 17, p = 0.23, Mann-Whitney; n = 8 (Ctrl), 7 (hM4D) mice).

c. (Top) Schematic of AAV injection to express hM4D or control GFP bilaterally in the VH, and ChR2 unilaterally in the BLA. (Bottom) Light-evoked BLA EPSCs at D1‑MSNs (black) and D2‑MSNs (red) from control (left) and hM4D-expressing (right) mice. EPSCs at D1‑MSNs are normalized to those at neighboring D2‑MSNs (see Figure 2). Scale bar = unity, 20 ms.

d. (Left) Summary of BLA EPSCs at D1-MSNs (top) and D2-MSNs (bottom) in control (top) and hM4D-expressing (bottom) mice. (Right) Summary of D1/D2 ratio of BLA EPSCs. Note that inhibiting VH activity does not prevent the biasing of BLA EPSCs onto D1-MSNs (D1/D2 ratio: U = 23, p = 0.38, Mann-Whitney; n = 8 (Ctrl), 8 (hM4D) pairs).

e-f. Similar to (c - d), with hM4D or control GFP expressed bilaterally in the VH, and ChR2 expressed unilaterally in the VH. Note that inhibiting VH activity reverses the normalization of VH ESPCs at D1- and D2-MSNs (D1/D2 ratio: U = 23, p = 0.001, Mann-Whitney; n = 11 (Ctrl), 15 (hM4D) pairs).

Bar graphs show mean ± SEM. * denotes p < 0.05.

Supplementary information

Source data

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

MacAskill, A., Cassel, J. & Carter, A. Cocaine exposure reorganizes cell type– and input-specific connectivity in the nucleus accumbens. Nat Neurosci 17, 1198–1207 (2014). https://doi.org/10.1038/nn.3783

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nn.3783

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing