Encoding of conditioned fear in central amygdala inhibitory circuits

1. NatureVolume:468,Pages:277–282 Date published:(11 November 2010) Encoding of conditioned fear in centralamygdala inhibitory circuits Stephane Ciocchi1*, Cyril Herry1*{, François Grenier1, Steffen B. E.Wolff1, Johannes J. Letzkus1, Ioannis Vlachos2, Ingrid Ehrlich1{, Rolf Sprengel3, Karl Deisseroth4, Michael B. Stadler1, Christian Mu¨ller1 & Andreas Lu¨thi1

2. BACKGROUND • The central amygdala (CEA)has been considered to beprimarily involved in the behavioural expression of conditioned fearresponses • CEAoutput neurons(CEm) project to downstream targets in the brainstemand in the hypothalamus where theyorchestrate conditioned autonomicand motor responses • CEmoutput neurons are under tight inhibitory control from the lateral andcapsular subdivisions (together referred to as CEl)

3. How the acquisition and expression of conditioned fear are encoded within CEA inhibitory circuits ?

4. WHAT WE WILL KNOW IN THIS ARTICLE • Neuronal activity in the lateral subdivision of the central amygdala (CEl) is required for fear acquisition. • Conditioned fear responses are driven by output neurons in the medial subdivision (CEm). • Inhibitory CEA microcircuits are highly organized • Cell-type-specific plasticity of phasic and tonic activity in the CEl to CEm pathway may gate fear expression and regulate fear generalization.

5. TECHNOLOGY

6. Optogenetics TECHNOLOGY

7. TECHNOLOGY For more information, go to www.stanford.edu/group/dlab/optogenetics/

8. TECHNOLOGY Microiontophoresis is the technique whereby ions and charged molecules canbe ejected in very small amounts from solutions contained inmicropipettes OFTEN USED FOR: (1) deposition of dyes and neural transport tracers for histological examination (2) for administration of neuroactive compounds (e.g. transmitters, modulators, drugs or hormones) by microiontophoresis to examine their effects on firing parameters of single neurons in vivo. For more information, go to www.kationscientific.com/basics.html

9. RESULTS Differential role of CEl andCEm 1.The impact of neuronal activity in CEm on freezing behavior OPTOGENETICS (Activation) Virus expressingChR2 in neurons

10. RESULTS • Bilateral activation ofCEm induced strong and reversible freezing responses

11. RESULTS MICROIONTOPHORESIS (Inhibition) Fluorescently labelledGABAA receptor agonist (muscimol-bodipy (BPY)) • Bilateral inactivation of CEm, or of the entire CEA (CEm and CEl), did not elicit freezing behaviour. • Inactivation of CEl alone induced unconditioned freezing.

12. RESULTS 2. The contribution of distinct CEA subnuclei to theacquisition of conditioned freezing

13. RESULTS 3. The role of CEl and CEm for memory retrieval or expression Local application of muscimol-BPY 24 h after conditioning Inactivation of the entire CEA or CEm resulted in a retrieval/ expression deficit. Inactivation of CEl did not reduce conditioned freezing levels

14. CONCLUSION 1 Conditioned and unconditioned freezingbehaviour is driven by CEm output neurons which are under tonicinhibitory control originating in CEl. CEl as an essential component of the neuronal circuitry underlying theacquisition of conditioned fear.

15. RESULTS Organization of CEA inhibitory networks 1. Fearconditioning-induced changes in CSevokedneuronal firing in the CEl 30% of units acquiredan excitatory response (CElon neurons) (Fig.a) 25% of CEl neuronsdisplayed a strong inhibitory response to the CS1 after fear conditioning (CEloff neurons) (Fig.b)

16. RESULTS

17. RESULTS The inverse direction of fear-conditioning-induced plasticity in CEIon and CEIoff neurons indicated that ...

18. RESULTS CElon to CEloff, 9 of 35 pairs; CEloff to CElon, 3 of 35 pairs;

19. RESULTS 2.Anatomical and functional connectivitybetween CEl and CEm. Injections into CEm resulted in intense retrogradelabelling of neurons in CEl CEm remained largely devoid of GFPafter injections into CEl

20. RESULTS 2.1 Whether identified CElon or CEloff neurons project to CEm • The axons of bothsubtypesarborize locally within CEl, and sendcollaterals to CEm

21. RESULTS 2.2Whether CElon and CEloff neurons functionallyinhibit CEm neurons Two distinctsubclasses of CEl neurons inhibit CEm neurons in vivo

22. 3.Considering that CElon and CEloff neurons exhibited oppositechanges in CS-evoked firing during fear conditioning, this raises thequestion of whether at the level of CEm output neurons fear conditioningresults in CS-evoked inhibition or disinhibition.

23. RESULTS

24. RESULTS Conditioned CS responses ofCEm output neurons reflect the integration of both excitatory and disinhibitory inputs.

25. CONCLUSION 2 CEl contains two functionallydistinct subpopulations of neurons forming highly organizedlocal inhibitory circuits which inhibit CEm output neurons. Conditioned fear responses are drivenby CS-evoked disinhibition of CEm output neurons

26. RESULTS Tonic inhibition and fear generalization 1.Whether spontaneous activity in CEl and CEm is subject to regulation ? 2.How plasticity of spontaneous activity might contribute to the encoding of conditioned fear responses ?

27. RESULTS • Spontaneous activity of CEm output neurons was markedly • decreased after fear conditioning. • CEloff neurons exhibited increased spontaneous activity • after fear conditioning • On average CElon neurons showed a slight decrease

28. RESULTS

29. RESULTS 3.What might be the behavioural relevance of plasticity of tonic activity in CEA inhibitory circuits? After fear conditioning, absoluteand z-scored levels of tonic activitywere only poorly correlated withfreezing

30. RESULTS Fear-conditioninginducedchanges in tonic activity were not limited to periods ofCS+ exposure, but were also manifest during CS-stimulation

31. RESULTS A decrease in tonic activity of CEm output neurons was associated with generalization, CEloff neuronsand CElon neurons exhibited the inverse correlation.

32. SUMMARY SECTION 1、2 SECTION 3

33. Thank you !