The identity of cortical circuits mediating nociception and pain is largely unclear. The cingulate cortex is consistently activated during pain, but the functional specificity of cingulate divisions, the roles at distinct temporal phases of central plasticity and the underlying circuitry are unknown. Here we show in mice that the midcingulate division of the cingulate cortex (MCC) does not mediate acute pain sensation and pain affect, but gates sensory hypersensitivity by acting in a wide cortical and subcortical network. Within this complex network, we identified an afferent MCC–posterior insula pathway that can induce and maintain nociceptive hypersensitivity in the absence of conditioned peripheral noxious drive. This facilitation of nociception is brought about by recruitment of descending serotonergic facilitatory projections to the spinal cord. These results have implications for our understanding of neuronal mechanisms facilitating the transition from acute to long-lasting pain.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.


  1. 1.

    , & Structural and functional dichotomy of human midcingulate cortex. Eur. J. Neurosci. 18, 3134–3144 (2003).

  2. 2.

    Pain and emotion interactions in subregions of the cingulate gyrus. Nat. Rev. Neurosci. 6, 533–544 (2005).

  3. 3.

    et al. The integration of negative affect, pain and cognitive control in the cingulate cortex. Nat. Rev. Neurosci. 12, 154–167 (2011).

  4. 4.

    et al. An fMRI-based neurologic signature of physical pain. N. Engl. J. Med. 368, 1388–1397 (2013).

  5. 5.

    et al. Presynaptic and postsynaptic amplifications of neuropathic pain in the anterior cingulate cortex. J. Neurosci. 28, 7445–7453 (2008).

  6. 6.

    , & The role of the midcingulate cortex in monitoring others' decisions. Front. Neurosci. 7, 251 (2013).

  7. 7.

    & Cytoarchitecture of mouse and rat cingulate cortex with human homologies. Brain Struct. Funct. 219, 185–192 (2014).

  8. 8.

    , , & Contributions of the anterior cingulate cortex and amygdala to pain- and fear-conditioned place avoidance in rats. Pain 110, 343–353 (2004).

  9. 9.

    & Glutamatergic activation of anterior cingulate cortex produces an aversive teaching signal. Nat. Neurosci. 7, 398–403 (2004).

  10. 10.

    et al. Lesion of the rostral anterior cingulate cortex eliminates the aversiveness of spontaneous neuropathic pain following partial or complete axotomy. Pain 152, 1641–1648 (2011).

  11. 11.

    et al. Genetic elimination of behavioral sensitization in mice lacking calmodulin-stimulated adenylyl cyclases. Neuron 36, 713–726 (2002).

  12. 12.

    et al. Alleviating neuropathic pain hypersensitivity by inhibiting PKMζ in the anterior cingulate cortex. Science 330, 1400–1404 (2010).

  13. 13.

    et al. In vivo whole-cell patch-clamp recording of sensory synaptic responses of cingulate pyramidal neurons to noxious mechanical stimuli in adult mice. Mol. Pain 6, 62 (2010).

  14. 14.

    et al. Bidirectional modulation of hyperalgesia via the specific control of excitatory and inhibitory neuronal activity in the ACC. Mol. Brain 8, 81 (2015).

  15. 15.

    et al. Distinct roles of the anterior cingulate cortex in spinal and supraspinal bee venom-induced pain behaviors. Neuroscience 153, 268–278 (2008).

  16. 16.

    et al. Pain inhibition by optogenetic activation of specific anterior cingulate cortical neurons. PLoS One 10, e0117746 (2015).

  17. 17.

    & Pain imaging in health and disease—how far have we come? J. Clin. Invest. 120, 3788–3797 (2010).

  18. 18.

    et al. The primary somatosensory cortex and the insula contribute differently to the processing of transient and sustained nociceptive and non-nociceptive somatosensory inputs. Hum. Brain Mapp. 36, 4346–4360 (2015).

  19. 19.

    & Optogenetic investigation of neural circuits underlying brain disease in animal models. Nat. Rev. Neurosci. 13, 251–266 (2012).

  20. 20.

    et al. Presynaptically localized cyclic GMP-dependent protein kinase 1 is a key determinant of spinal synaptic potentiation and pain hypersensitivity. PLoS Biol. 10, e1001283 (2012).

  21. 21.

    , , & Peripheral mechanism of cutaneous nociception. in Wall and Melzack's Textbook of Pain (eds. McMahon, S.B. & Koltzenburg, M.) 3–34 (Elsevier Churchill Livingstone, 2006).

  22. 22.

    et al. Principles for applying optogenetic tools derived from direct comparative analysis of microbial opsins. Nat. Methods 9, 159–172 (2011).

  23. 23.

    , , , & Biophysical constraints of optogenetic inhibition at presynaptic terminals. Nat. Neurosci. 19, 554–556 (2016).

  24. 24.

    et al. Mechanical, thermal and formalin-induced nociception is differentially altered in 5-HT1A−/−, 5-HT1B−/−, 5-HT2A−/−, 5-HT3A−/− and 5-HTT−/− knock-out male mice. Pain 130, 235–248 (2007).

  25. 25.

    , , & Parallel processing of nociceptive A-delta inputs in SII and midcingulate cortex in humans. J. Neurosci. 28, 944–952 (2008).

  26. 26.

    , & Towards a theory of chronic pain. Prog. Neurobiol. 87, 81–97 (2009).

  27. 27.

    et al. Postsynaptic potentiation of corticospinal projecting neurons in the anterior cingulate cortex after nerve injury. Mol. Pain 10, 33 (2014).

  28. 28.

    , & Descending facilitatory modulation of a behavioral nociceptive response by stimulation in the adult rat anterior cingulate cortex. Eur. J. Pain 4, 83–96 (2000).

  29. 29.

    , & The affective component of pain in rodents: direct evidence for a contribution of the anterior cingulate cortex. Proc. Natl. Acad. Sci. USA 98, 8077–8082 (2001).

  30. 30.

    et al. Pavlovian fear memory induced by activation in the anterior cingulate cortex. Mol. Pain 1, 6 (2005).

  31. 31.

    et al. Empathy for pain involves the affective but not sensory components of pain. Science 303, 1157–1162 (2004).

  32. 32.

    & Structural plasticity and reorganisation in chronic pain. Nat. Rev. Neurosci. 18, 20–30 (2016).

  33. 33.

    et al. Human dorsal anterior cingulate cortex neurons mediate ongoing behavioural adaptation. Nature 488, 218–221 (2012).

  34. 34.

    et al. Role of Prelimbic GABAergic Circuits in Sensory and Emotional Aspects of Neuropathic Pain. Cell Rep. 12, 752–759 (2015).

  35. 35.

    et al. Activation of corticostriatal circuitry relieves chronic neuropathic pain. J. Neurosci. 35, 5247–5259 (2015).

  36. 36.

    How do you feel—now? The anterior insula and human awareness. Nat. Rev. Neurosci. 10, 59–70 (2009).

  37. 37.

    , , , & The dorsal posterior insula subserves a fundamental role in human pain. Nat. Neurosci. 18, 499–500 (2015).

  38. 38.

    , , , & Evidence against pain specificity in the dorsal posterior insula. F1000Res. 4, 362 (2015).

  39. 39.

    , , & Pain anticipation: an activation likelihood estimation meta-analysis of brain imaging studies. Hum. Brain Mapp. 36, 1648–1661 (2015).

  40. 40.

    & A role for midcingulate cortex in the interruptive effects of pain anticipation on attention. Clin. Neurophysiol. 119, 2370–2379 (2008).

  41. 41.

    et al. Anterior insula integrates information about salience into perceptual decisions about pain. J. Neurosci. 30, 16324–16331 (2010).

  42. 42.

    & Neuroimaging evidence of motor control and pain processing in the human midcingulate cortex. Cereb. Cortex 25, 1906–1919 (2015).

  43. 43.

    , & Parsing pain perception between nociceptive representation and magnitude estimation. J. Neurophysiol. 101, 875–887 (2009).

  44. 44.

    , , & Optogenetic activation of brainstem serotonergic neurons induces persistent pain sensitization. Mol. Pain 10, 70 (2014).

  45. 45.

    & A sore spot: central or peripheral generation of chronic neuropathic spontaneous pain? Pain 155, 1189–1191 (2014).

  46. 46.

    et al. Lentivirus-based genetic manipulations of cortical neurons and their optical and electrophysiological monitoring in vivo. Proc. Natl. Acad. Sci. USA 101, 18206–18211 (2004).

  47. 47.

    et al. Faithful expression of multiple proteins via 2A-peptide self-processing: a versatile and reliable method for manipulating brain circuits. J. Neurosci. 29, 8621–8629 (2009).

  48. 48.

    , & A simplified baculovirus-AAV expression vector system coupled with one-step affinity purification yields high-titer rAAV stocks from insect cells. Mol. Ther. 17, 1888–1896 (2009).

  49. 49.

    , , , & Development and optimization of adeno-associated virus vector transfer into the central nervous system. Methods Mol. Med. 76, 221–236 (2003).

  50. 50.

    & The Mouse Brain in Stereotaxic Coordinates (Academic, 2001).

  51. 51.

    , , & Afferents to anterior cingulate areas 24a and 24b and midcingulate areas 24a′ and 24b′ in the mouse. Brain Struct. Funct. 222, 1509–1532 (2017).

  52. 52.

    , & In vivo siRNA transfection and gene knockdown in spinal cord via rapid noninvasive lumbar intrathecal injections in mice. J. Vis. Exp. 85, 51229 (2014).

  53. 53.

    et al. The anterior cingulate cortex is a critical hub for pain-induced depression. Biol. Psychiatry 77, 236–245 (2015).

  54. 54.

    , , , & Brainstorm: a user-friendly application for MEG/EEG analysis. Comput. Intell. Neurosci. 2011, 879716 (2011).

Download references


We thank R. LeFaucheur for secretarial help, as well as N. Gehrig, V. Buchert, L. Brenner, H.-J. Wrede, D. Baumgartl-Ahlert and K. Meyer for technical assistance. We are grateful to the Interdisciplinary Neurobehavioral Core Facility in Heidelberg for support with behavioral experiments. We gratefully acknowledge funding in form of SFB1158 grants from the Deutsche Forschungsgemeinschaft (DFG) to R.K. (project B01), T.K. (project B08), R.S. (project A05) and H.F. (project B07), European Research Council (ERC) Advanced Investigator grants to R.K. (Pain Plasticity 294293) and H.F. (Phantommind 230249) and DFG funding via the Excellence Cluster CellNetworks (Ectop funding to R.K. and H.F.). We acknowledge support from the European Molecular Biology Organization (EMBO) to L.L.T. in the form of an EMBO long-term postdoctoral fellowship.

Author information

Author notes

    • Patric Pelzer
    •  & Wannan Tang

    Present addresses: Letten Centre and GliaLab, Department of Physiology, Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway (W.T.) and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany (P.P.).


  1. Institute of Pharmacology, Heidelberg University, Heidelberg, Germany.

    • Linette Liqi Tan
    • , Céline Heinl
    • , Vijayan Gangadharan
    •  & Rohini Kuner
  2. Department of Functional Neuroanatomy, Institute for Anatomy and Cell Biology, Heidelberg University, Heidelberg, Germany.

    • Patric Pelzer
    •  & Thomas Kuner
  3. Max Planck Institute for Medical Research, Department of Molecular Neurobiology, Heidelberg, Germany.

    • Wannan Tang
    •  & Rolf Sprengel
  4. Department of Cognitive and Clinical Neuroscience, Central Institute of Mental Health, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany.

    • Herta Flor
  5. CellNetworks Cluster of Excellence, Heidelberg University, Heidelberg, Germany.

    • Herta Flor
    • , Thomas Kuner
    •  & Rohini Kuner
  6. Max Planck Research Group at the Institute for Anatomy and Cell Biology, Heidelberg University, Heidelberg, Germany.

    • Rolf Sprengel


  1. Search for Linette Liqi Tan in:

  2. Search for Patric Pelzer in:

  3. Search for Céline Heinl in:

  4. Search for Wannan Tang in:

  5. Search for Vijayan Gangadharan in:

  6. Search for Herta Flor in:

  7. Search for Rolf Sprengel in:

  8. Search for Thomas Kuner in:

  9. Search for Rohini Kuner in:


L.L.T., R.S., H.F., T.K. and R.K. were involved in manuscript preparation. L.L.T. conducted the experiments and analyzed data. R.K. designed the study and wrote the manuscript. W.T. generated the viruses; V.G. helped with behavioral experiments; C.H. and P.P. performed electrophysiology experiments.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Rohini Kuner.

Integrated supplementary information

Supplementary figures

  1. 1.

    Atlas representations of the midcingulate (MCC) region targeted in this study compared against the anterior cingulate (ACC) region commonly reported.

  2. 2.

    Cellular properties of opsin- and control GFP-expressing neurons obtained from patch clamp whole-cell recordings performed in slices from the MCC.

  3. 3.

    Lack of phototoxicity in morphological analyses on opsin-expressing cortical sections.

  4. 4.

    Fos expression in the MCC after capsaicin injection in the hindpaw.

  5. 5.

    Optogenetic modulation of activity in the MCC and/or the hind limb region of the primary somatosensory cortex (S1HL) on capsaicin-evoked nocifensive behaviors and basal mechanical sensitivities of the hindpaws.

  6. 6.

    Effects of cortical illumination on paw mechanical sensitivity of control animals.

  7. 7.

    Effects of silencing the MCC or hind limb area of the primary somatosensory (S1HL) activity on capsaicin-evoked secondary mechanical hypersensitivity in the paw.

  8. 8.

    Effects of optogenetic inhibition and activation in the MCC on paw mechanical withdrawal responses.

  9. 9.

    Functional characterization of rAVV-CaMKII-ChR2 expression in the cortex.

  10. 10.

    Effects of MCC stimulation on mechanical withdrawal responses in hindpaws.

  11. 11.

    Quantification of Fos-positive cells in various cortical regions.

  12. 12.

    Example of Fos downregulation in the PI upon inhibition of the MCC in the capsaicin model.

  13. 13.

    Quantification and examples of Fos expression in the thalamus upon manipulation of activity in the MCC.

  14. 14.

    Mechanical behavioral responses of hindpaws upon silencing activity of the MCC–PI pathway or MCC–NAc pathway, or in the PI directly.

  15. 15.

    Viral tracing examples showing projections of excitatory neurons from the PI within the raphe nucleus (RMg) regions.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–15

  2. 2.

    Life Sciences Reporting Summary

About this article

Publication history






Further reading