Abstract

‘Sundowning’ in dementia and Alzheimer’s disease is characterized by early-evening agitation and aggression. While such periodicity suggests a circadian origin, whether the circadian clock directly regulates aggressive behavior is unknown. We demonstrate that a daily rhythm in aggression propensity in male mice is gated by GABAergic subparaventricular zone (SPZGABA) neurons, the major postsynaptic targets of the central circadian clock, the suprachiasmatic nucleus. Optogenetic mapping revealed that SPZGABA neurons receive input from vasoactive intestinal polypeptide suprachiasmatic nucleus neurons and innervate neurons in the ventrolateral part of the ventromedial hypothalamus (VMH), which is known to regulate aggression. Additionally, VMH-projecting dorsal SPZ neurons are more active during early day than early night, and acute chemogenetic inhibition of SPZGABA transmission phase-dependently increases aggression. Finally, SPZGABA-recipient central VMH neurons directly innervate ventrolateral VMH neurons, and activation of this intra-VMH circuit drove attack behavior. Altogether, we reveal a functional polysynaptic circuit by which the suprachiasmatic nucleus clock regulates aggression.

  • Subscribe to Nature Neuroscience for full access:

    $59

    Subscribe

Additional access options:

Already a subscriber?  Log in  now or  Register  for online access.

Additional information

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

References

  1. 1.

    Manfredini, R. et al. Day-night variation in aggressive behavior among psychiatric inpatients. Chronobiol. Int. 18, 503–511 (2001).

  2. 2.

    Bachman, D. & Rabins, P. “Sundowning” and other temporally associated agitation states in dementia patients. Annu. Rev. Med. 57, 499–511 (2006).

  3. 3.

    Bronsard, G. & Bartolomei, F. Rhythms, rhythmicity and aggression. J. Physiol. Paris 107, 327–334 (2013).

  4. 4.

    Jagannath, A., Peirson, S. N. & Foster, R. G. Sleep and circadian rhythm disruption in neuropsychiatric illness. Curr. Opin. Neurobiol. 23, 888–894 (2013).

  5. 5.

    Tordjman, S. et al. Autism as a disorder of biological and behavioral rhythms: toward new therapeutic perspectives. Front Pediatr. 3, 1 (2015).

  6. 6.

    Miczek, K. A., Maxson, S. C., Fish, E. W. & Faccidomo, S. Aggressive behavioral phenotypes in mice. Behav. Brain Res. 125, 167–181 (2001).

  7. 7.

    Miczek, K. A. et al. Neurobiology of escalated aggression and violence. J. Neurosci. 27, 11803–11806 (2007).

  8. 8.

    Nelson, R. J. & Trainor, B. C. Neural mechanisms of aggression. Nat. Rev. Neurosci. 8, 536–546 (2007).

  9. 9.

    Sternson, S. M. Hypothalamic survival circuits: blueprints for purposive behaviors. Neuron 77, 810–824 (2013).

  10. 10.

    Yang, T. et al. Social control of hypothalamus-mediated male aggression. Neuron 95, 955–970.e4 (2017).

  11. 11.

    Reppert, S. M. & Weaver, D. R. Coordination of circadian timing in mammals. Nature 418, 935–941 (2002).

  12. 12.

    Lin, D. et al. Functional identification of an aggression locus in the mouse hypothalamus. Nature 470, 221–226 (2011).

  13. 13.

    Yang, C. F. et al. Sexually dimorphic neurons in the ventromedial hypothalamus govern mating in both sexes and aggression in males. Cell 153, 896–909 (2013).

  14. 14.

    Lee, H. et al. Scalable control of mounting and attack by Esr1+ neurons in the ventromedial hypothalamus. Nature 509, 627–632 (2014).

  15. 15.

    Falkner, A. L., Grosenick, L., Davidson, T. J., Deisseroth, K. & Lin, D. Hypothalamic control of male aggression-seeking behavior. Nat. Neurosci. 19, 596–604 (2016).

  16. 16.

    Welsh, D. K., Logothetis, D. E., Meister, M. & Reppert, S. M. Individual neurons dissociated from rat suprachiasmatic nucleus express independently phased circadian firing rhythms. Neuron 14, 697–706 (1995).

  17. 17.

    Jin, X. et al. A molecular mechanism regulating rhythmic output from the suprachiasmatic circadian clock. Cell 96, 57–68 (1999).

  18. 18.

    Gall, A. J., Todd, W. D. & Blumberg, M. S. Development of SCN connectivity and the circadian control of arousal: a diminishing role for humoral factors? PLoS One 7, e45338 (2012).

  19. 19.

    Saper, C. B. The central circadian timing system. Curr. Opin. Neurobiol. 23, 747–751 (2013).

  20. 20.

    Watts, A. G., Swanson, L. W. & Sanchez-Watts, G. Efferent projections of the suprachiasmatic nucleus: I. Studies using anterograde transport of Phaseolus vulgaris leucoagglutinin in the rat. J. Comp. Neurol. 258, 204–229 (1987).

  21. 21.

    Watts, A. G. & Swanson, L. W. Efferent projections of the suprachiasmatic nucleus: II. Studies using retrograde transport of fluorescent dyes and simultaneous peptide immunohistochemistry in the rat. J. Comp. Neurol. 258, 230–252 (1987).

  22. 22.

    Lu, J. et al. Contrasting effects of ibotenate lesions of the paraventricular nucleus and subparaventricular zone on sleep-wake cycle and temperature regulation. J. Neurosci. 21, 4864–4874 (2001).

  23. 23.

    Vujovic, N., Gooley, J. J., Jhou, T. C. & Saper, C. B. Projections from the subparaventricular zone define four channels of output from the circadian timing system. J. Comp. Neurol. 523, 2714–2737 (2015).

  24. 24.

    Tong, Q., Ye, C. P., Jones, J. E., Elmquist, J. K. & Lowell, B. B. Synaptic release of GABA by AgRP neurons is required for normal regulation of energy balance. Nat. Neurosci. 11, 998–1000 (2008).

  25. 25.

    Kaur, S. et al. Glutamatergic signaling from the parabrachial nucleus plays a critical role in hypercapnic arousal. J. Neurosci. 33, 7627–7640 (2013).

  26. 26.

    Engeland, W. C. & Arnhold, M. M. Neural circuitry in the regulation of adrenal corticosterone rhythmicity. Endocrine 28, 325–332 (2005).

  27. 27.

    Vong, L. et al. Leptin action on GABAergic neurons prevents obesity and reduces inhibitory tone to POMC neurons. Neuron 71, 142–154 (2011).

  28. 28.

    Chou, T. C. et al. Critical role of dorsomedial hypothalamic nucleus in a wide range of behavioral circadian rhythms. J. Neurosci. 23, 10691–10702 (2003).

  29. 29.

    Krashes, M. J. et al. An excitatory paraventricular nucleus to AgRP neuron circuit that drives hunger. Nature 507, 238–242 (2014).

  30. 30.

    Anaclet, C. et al. The GABAergic parafacial zone is a medullary slow wave sleep-promoting center. Nat. Neurosci. 17, 1217–1224 (2014).

  31. 31.

    Fan, J. et al. Vasoactive intestinal polypeptide (VIP)-expressing neurons in the suprachiasmatic nucleus provide sparse GABAergic outputs to local neurons with circadian regulation occurring distal to the opening of postsynaptic GABAA ionotropic receptors. J. Neurosci. 35, 1905–1920 (2015).

  32. 32.

    Lynagh, T. & Lynch, J. W. An improved ivermectin-activated chloride channel receptor for inhibiting electrical activity in defined neuronal populations. J. Biol. Chem. 285, 14890–14897 (2010).

  33. 33.

    Berridge, K. C. Motivation concepts in behavioral neuroscience. Physiol. Behav. 81, 179–209 (2004).

  34. 34.

    Kennedy, A. et al. Internal states and behavioral decision-making: toward an integration of emotion and cognition. Cold Spring Harb. Symp. Quant. Biol. 79, 199–210 (2014).

  35. 35.

    LeDoux, J. Rethinking the emotional brain. Neuron 73, 653–676 (2012).

  36. 36.

    Silva, B. A. et al. Independent hypothalamic circuits for social and predator fear. Nat. Neurosci. 16, 1731–1733 (2013).

  37. 37.

    Kunwar, P.S. et al. Ventromedial hypothalamic neurons control a defensive emotion state. eLife 4, (2015).

  38. 38.

    Bilu, C. & Kronfeld-Schor, N. Effects of circadian phase and melatonin injection on anxiety-like behavior in nocturnal and diurnal rodents. Chronobiol. Int. 30, 828–836 (2013).

  39. 39.

    Albrecht, A. & Stork, O. Circadian rhythms in fear conditioning: an overview of behavioral, brain system, and molecular interactions. Neural Plast. 2017, 3750307 (2017).

  40. 40.

    Nakamura, W. et al. In vivo monitoring of circadian timing in freely moving mice. Curr. Biol. 18, 381–385 (2008).

  41. 41.

    Todd, W. D., Gall, A. J., Weiner, J. A. & Blumberg, M. S. Distinct retinohypothalamic innervation patterns predict the developmental emergence of species-typical circadian phase preference in nocturnal Norway rats and diurnal Nile grass rats. J. Comp. Neurol. 520, 3277–3292 (2012).

  42. 42.

    Hermes, M. L., Kolaj, M., Doroshenko, P., Coderre, E. & Renaud, L. P. Effects of VPAC2 receptor activation on membrane excitability and GABAergic transmission in subparaventricular zone neurons targeted by suprachiasmatic nucleus. J. Neurophysiol. 102, 1834–1842 (2009).

  43. 43.

    Khachiyants, N., Trinkle, D., Son, S. J. & Kim, K. Y. Sundown syndrome in persons with dementia: an update. Psychiatry Investig. 8, 275–287 (2011).

  44. 44.

    Bedrosian, T. A. & Nelson, R. J. Sundowning syndrome in aging and dementia: research in mouse models. Exp. Neurol. 243, 67–73 (2013).

  45. 45.

    Canevelli, M. et al. Sundowning in dementia: clinical relevance, pathophysiological determinants, and therapeutic approaches. Front. Med. (Lausanne) 3, 73 (2016).

  46. 46.

    Hope, T., Keene, J., Gedling, K., Fairburn, C. G. & Jacoby, R. Predictors of institutionalization for people with dementia living at home with a carer. Int. J. Geriatr. Psychiatry 13, 682–690 (1998).

  47. 47.

    Bedrosian, T. A. et al. Nocturnal light exposure impairs affective responses in a wavelength-dependent manner. J. Neurosci. 33, 13081–13087 (2013).

  48. 48.

    Oishi, Y. et al. Role of the medial prefrontal cortex in cataplexy. J. Neurosci. 33, 9743–9751 (2013).

  49. 49.

    Hattori, T. et al. Self-exposure to the male pheromone ESP1 enhances male aggressiveness in mice. Curr. Biol. 26, 1229–1234 (2016).

  50. 50.

    Padilla, S. L. et al. Agouti-related peptide neural circuits mediate adaptive behaviors in the starved state. Nat. Neurosci. 19, 734–741 (2016).

  51. 51.

    Hashikawa, K. et al. Esr1+ cells in the ventromedial hypothalamus control female aggression. Nat. Neurosci. 20, 1580–1590 (2017).

  52. 52.

    Burgos-Artizzu, X.P., Dollar, P., Lin, D., Anderson, D.J. & Perona, P. in IEEE Conference on Computer Vision and Pattern Recognition, Providence, Rhode Island, 1322–1329 (2012).

  53. 53.

    Zhang, R. et al. Loss of hypothalamic corticotropin-releasing hormone markedly reduces anxiety behaviors in mice. Mol. Psychiatry 22, 733–744 (2017).

  54. 54.

    Pei, H., Sutton, A. K., Burnett, K. H., Fuller, P. M. & Olson, D. P. AVP neurons in the paraventricular nucleus of the hypothalamus regulate feeding. Mol. Metab. 3, 209–215 (2014).

  55. 55.

    Cheong, R. Y., Czieselsky, K., Porteous, R. & Herbison, A. E. Expression of ESR1 in glutamatergic and GABAergic neurons is essential for normal puberty onset, estrogen feedback, and fertility in female mice. J. Neurosci. 35, 14533–14543 (2015).

  56. 56.

    Guillery, R. W. On counting and counting errors. J. Comp. Neurol. 447, 1–7 (2002).

  57. 57.

    Venner, A., Anaclet, C., Broadhurst, R. Y., Saper, C. B. & Fuller, P. M. A novel population of wake-promoting GABAergic neurons in the ventral lateral hypothalamus. Curr. Biol. 26, 2137–2143 (2016).

  58. 58.

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

Download references

Acknowledgements

We thank Q. Ha, M. Thompson, S. Bandaru, C. Friedman, R. Thomas, and M. Ha for excellent technical assistance and C. Dulac for helpful comments during the early stages of this project. We thank J. Lynch (University of Queensland) for the hGlyR construct; and D. Anderson, X. Burgos-Artizzu, and P. Dollár (California Institute of Technology) for the Behavior Annotator MatLab script and code. This work was supported by the G. Harold and Leila Y. Mathers Foundation and the US National Institutes of Health (NIH) grants NS072337, NS085477, AG09975, and HL095491 to C.B.S.; NS073613, NS092652, and NS103161 to P.M.F.; and DK111401, DK075632, DK096010, DK089044, DK046200, and DK057521 to B.B.L. W.D.T. was supported by Alzheimer’s Association grant AARF-16-443613 and NIH grants NS084582-01A1 and HL00701-15. N.L.M. was supported by CNPq (National Health Council for Scientific and Technological Development/Brazil, grant 200881/2014-0), CAPES (Coordination for the Improvement of Higher Education Personnel).

Author information

Author notes

    • Henning Fenselau

    Present address: Center for Endocrinology, Diabetes and Preventive Medicine, University Hospital Cologne, Cologne, Germany

    • Rong Zhang

    Present address: Brain Research Center, College of Life Science, NorthWest University, Xi’an, China

  1. These authors contributed equally: William D. Todd and Henning Fenselau.

Affiliations

  1. Department of Neurology, Program in Neuroscience, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, USA

    • William D. Todd
    • , Joshua L. Wang
    • , Natalia L. Machado
    • , Anne Venner
    • , Rebecca Y. Broadhurst
    • , Satvinder Kaur
    • , Patrick M. Fuller
    •  & Clifford B. Saper
  2. Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, Program in Neuroscience, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA

    • Henning Fenselau
    •  & Bradford B. Lowell
  3. Division of Endocrinology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA

    • Rong Zhang
  4. Department of Physiology and Biophysics, Federal University of Minas Gerais, Belo Horizonte, Brazil

    • Natalia L. Machado
  5. Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark

    • Timothy Lynagh
  6. Department of Pediatrics, University of Michigan, Ann Arbor, MI, USA

    • David P. Olson
  7. Department of Synaptic Transmission in Energy Homeostasis, Max Planck Institute for Metabolism Research, Cologne, Germany

    • Henning Fenselau

Authors

  1. Search for William D. Todd in:

  2. Search for Henning Fenselau in:

  3. Search for Joshua L. Wang in:

  4. Search for Rong Zhang in:

  5. Search for Natalia L. Machado in:

  6. Search for Anne Venner in:

  7. Search for Rebecca Y. Broadhurst in:

  8. Search for Satvinder Kaur in:

  9. Search for Timothy Lynagh in:

  10. Search for David P. Olson in:

  11. Search for Bradford B. Lowell in:

  12. Search for Patrick M. Fuller in:

  13. Search for Clifford B. Saper in:

Contributions

W.D.T., H.F., and C.B.S. designed the experiments. W.D.T., H.F., J.L.W., R.Z., and N.L.M. carried out these experiments. J.L.W., A.V., S.K., T.L., D.P.O., B.B.L., and P.M.F. provided analytic tools and reagents. W.D.T., H.F., J.L.W., R.Z., A.V., and R.Y.B. analyzed the data. W.D.T., H.F., P.M.F., and C.B.S. wrote the paper.

Competing interests

The authors declare no competing interests.

Corresponding author

Correspondence to Clifford B. Saper.

Integrated supplementary information

  1. Supplementary Figure 1 Representative hypothalamic sections depicting Cre expression following AAV-iCre-2A-Venus in the SPZ.

    Sections were immunohistochemically double-labeled for Cre (black), to reveal cells transfected by the AAV, and VIP (brown) to reveal fibers from the SCN that delineate the SPZ. ot, optic tract. f, fornix. Representative of 16 mice.

  2. Supplementary Figure 2 Injection of AAV-iCre-2A-Venus into the SPZ successfully deleted Vgat.

    Representative section from a Vgat in situ hybridization demonstrating unilateral SPZ Vgat-deletion compared to an undeleted SPZ. White ovals depict the SPZ. Representative of 2 mice.

  3. Supplementary Figure 3 Rhythms in aggression propensity are not due to the direct effects of light.

    In constant darkness (free-running conditions), intact mice (C57Bl6/J, n=12) show differences in aggression propensity (total time attacking) between early subjective day [circadian time (CT)1] and early subjective night (CT13). Planned comparisons, paired t test between CT1 and CT13, two-tailed, t(11)=2.482, P=0.0305. Means ± s.e.m.

  4. Supplementary Figure 4 SPZ Vgat-deletions do not disrupt entrained or free-running rhythms of Tb.

    (a) Mean (± s.e.m.) Tb per hour in SPZ Vgat-deleted mice (red, n=8) and intact GFP-injected littermate controls (blue, n=8) (two-way repeated measures ANOVA). (b) Double plotted actogram for Tb in LD (12:12 light-dark cycle) and constant darkness (DD). Arrow denotes the beginning of the DD period (from the end of the normal dark period).

  5. Supplementary Figure 5 SPZ subregions have different projection patterns in mice.

    Darkfield photomicrographs depicting hrGFP-labeled neurons (orange/brown) in the SPZ and fibers (yellow/gold) in the DMH and VMH. (a, b) One Vgat-IRES-Cre mouse with an AAV-FLEX-hrGFP injection site that transfected neurons in both the ventral SPZ (a) and dorsal SPZ (b). Images are from the same mouse, but from adjacent sections, as depicted in Fig. 2d-e. Representative of 2 mice. (c, d) Two additional Vgat-IRES-Cre mice with different AAV-FLEX-hrGFP injection sites transfecting all but the ventral SPZ (c), representative of 2 mice, or dorsal SPZ (d), representative of 4 mice, [see white ovals, compared to (a) and (b)]. (e-f) A lack of transfected cells in the ventral SPZ results in less hrGFP-labeled fibers and terminals in the DMH (e, same mouse as c), representative of 2 mice, while a lack of transfected cells in the dorsolateral SPZ results in less hrGFP labeling in the VMH (f, same mouse as d), representative of 4 mice. Using subtraction methods we can deduce that, in mice, the ventral SPZGABA population projects more heavily to the DMH while the dorsal SPZGABA population projects more heavily to the VMH (arrows).

  6. Supplementary Figure 6 The dorsal SPZ shows high numbers of c-Fos+ neurons at ZT1 in intact, uninjected mice.

    C57BL6/J mice (n=6) were maintained under the same lighting and housing conditions as Vgat-IRES-Cre mice prior to perfusion (Fig. 4f-g), except they did not undergo prior surgery or IP injections of IVM/VEH. When perfused 90 minutes after ZT1, these mice showed intense Fos immunolabeling (black) within the dorsal SPZ (blue ovals). This area densely projects to the VMH (Fig. 3 and Fig. S3) and closely mirrors the anatomical region of SPZGABA neurons that we identified as critical for mediating changes in aggressive behavior (Fig. 1g and 3b).

  7. Supplementary Figure 7 IVM does not increase aggression at ZT1 in control mice.

    There were no significant differences between IVM and VEH in total time attacking [left; paired t tests, two-tailed: t(7)=0.44, nsP=0.67), number of attack bouts [center; paired t tests, two-tailed: t(7)=0.22, nsP=0.83), or attack latency [right; paired t tests, two-tailed: t(7)=0.22, nsP=0.83] at ZT1 in control Vgat-IRES-Cre mice (n=8) injected with ChR2 into the SPZ, which does not respond to IVM. Means ± s.e.m.

Supplementary information

  1. Supplementary Text and Figures

    Supplementary Figures 1–7

  2. Reporting Summary

  3. Supplementary Software

    Custom script for weighting heat map by aggression score