Social touch promotes interfemale communication via activation of parvocellular oxytocin neurons

Abstract

Oxytocin (OT) is a great facilitator of social life but, although its effects on socially relevant brain regions have been extensively studied, OT neuron activity during actual social interactions remains unexplored. Most OT neurons are magnocellular neurons, which simultaneously project to the pituitary and forebrain regions involved in social behaviors. In the present study, we show that a much smaller population of OT neurons, parvocellular neurons that do not project to the pituitary but synapse onto magnocellular neurons, is preferentially activated by somatosensory stimuli. This activation is transmitted to the larger population of magnocellular neurons, which consequently show coordinated increases in their activity during social interactions between virgin female rats. Selectively activating these parvocellular neurons promotes social motivation, whereas inhibiting them reduces social interactions. Thus, parvocellular OT neurons receive particular inputs to control social behavior by coordinating the responses of the much larger population of magnocellular OT neurons.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: In vivo recording of individual OT neurons in the PVN.
Fig. 2: Gentle non-nociceptive mechanical stimuli trigger OT neuron activation.
Fig. 3: Intra-PVN connectivity of parvOT and magnOT neurons.
Fig. 4: Magnocellular neurons and their release of OT into blood are controlled by parvOT neurons.
Fig. 5: ParvOT neurons receive more inputs than magnOT neurons.
Fig. 6: Modulation of parvocellular OT neurons alters social behavior.

Data and code availability

Python code (used for ex vivo calcium-imaging data analysis in Fig. 4a–d) and MATLAB code (used for in vivo fiber photometry data analysis in Fig. 4e–o and Extended Data Fig. 7a–n) can be found in Supplementary Software. All data that support the findings of the present study, as well as MATLAB codes for the analysis of extracellular recording data, are available from the corresponding author upon reasonable request.

References

  1. 1.

    Lee, H.-J., Macbeth, A. H., Pagani, J. & Scott Young, W. 3rd. Oxytocin: the great facilitator of life. Prog. Neurobiol. 88, 127–151 (2010).

    Google Scholar 

  2. 2.

    Jurek, B. & Neumann, I. D. The oxytocin receptor: from intracellular signaling to behavior. Physiol. Rev. 98, 1805–1908 (2018).

    CAS  PubMed  Google Scholar 

  3. 3.

    Walum, H. & Young, L. J. The neural mechanisms and circuitry of the pair bond. Nat. Rev. Neurosci. 19, 643–654 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Russell, J. A., Leng, G. & Douglas, A. J. The magnocellular oxytocin system, the fount of maternity: adaptations in pregnancy. Front. Neuroendocrinol. 24, 27–61 (2003).

    CAS  PubMed  Google Scholar 

  5. 5.

    Knobloch, H. S. et al. Evoked axonal oxytocin release in the central amygdala attenuates fear response. Neuron 73, 553–566 (2012).

    CAS  PubMed  Google Scholar 

  6. 6.

    Marlin, B. J. & Froemke, R. C. Oxytocin modulation of neural circuits for social behavior. Dev. Neurobiol. 77, 169–189 (2017).

    CAS  PubMed  Google Scholar 

  7. 7.

    Grinevich, V. & Stoop, R. Interplay between oxytocin and sensory systems in the orchestration of socio-emotional behaviors. Neuron 99, 887–904 (2018).

    CAS  PubMed  Google Scholar 

  8. 8.

    Dumais, K. M., Alonso, A. G., Immormino, M. A., Bredewold, R. & Veenema, A. H. Involvement of the oxytocin system in the bed nucleus of the stria terminalis in the sex-specific regulation of social recognition. Psychoneuroendocrinology 64, 79–88 (2016).

    CAS  PubMed  Google Scholar 

  9. 9.

    Dumais, K. M., Alonso, A. G., Bredewold, R. & Veenema, A. H. Role of the oxytocin system in amygdala subregions in the regulation of social interest in male and female rats. Neuroscience 330, 138–149 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Resendez, S. L. et al. Social stimuli induce activation of oxytocin neurons within the paraventricular nucleus of the hypothalamus to promote social behavior in male mice. J. Neurosci. 40, 2282–2295 (2020).

    CAS  PubMed  Google Scholar 

  11. 11.

    Bobrov, E., Wolfe, J., Rao, R. P. & Brecht, M. The representation of social facial touch in rat barrel cortex. Curr. Biol. 24, 109–115 (2014).

    CAS  PubMed  Google Scholar 

  12. 12.

    Chen, P. & Hong, W. Neural circuit mechanisms of social behavior. Neuron 98, 16–30 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Eliava, M. et al. A new population of parvocellular oxytocin neurons controlling magnocellular neuron activity and inflammatory pain processing. Neuron 89, 1291–1304 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Lima, S. Q., Hromádka, T., Znamenskiy, P. & Zador, A. M. PINP: a new method of tagging neuronal populations for identification during in vivo electrophysiological recording. PLoS ONE 4, e6099 (2009).

    PubMed  PubMed Central  Google Scholar 

  15. 15.

    Leng, T., Leng, G. & MacGregor, D. J. Spike patterning in oxytocin neurons: capturing physiological behaviour with Hodgkin–Huxley and integrate-and-fire models. PLoS ONE 12, e0180368 (2017).

    PubMed  PubMed Central  Google Scholar 

  16. 16.

    Netser, S., Haskal, S., Magalnik, H. & Wagner, S. A novel system for tracking social preference dynamics in mice reveals sex- and strain-specific characteristics. Mol. Autism 8, 53 (2017).

    PubMed  PubMed Central  Google Scholar 

  17. 17.

    Maícas-Royo, J., Leng, G. & MacGregor, D. J. A predictive, quantitative model of spiking activity and stimulus-secretion coupling in oxytocin neurons. Endocrinology 159, 1433–1452 (2018).

    PubMed  PubMed Central  Google Scholar 

  18. 18.

    Portfors, C. V. Types and functions of ultrasonic vocalizations in laboratory rats and mice. J. Am. Assoc. Lab. Anim. Sci. 46, 28–34 (2007).

    CAS  PubMed  Google Scholar 

  19. 19.

    Lenschow, C. et al. Sexually monomorphic maps and dimorphic responses in rat genital cortex. Curr. Biol. 26, 106–113 (2016).

    CAS  PubMed  Google Scholar 

  20. 20.

    Althammer, F. & Grinevich, V. Diversity of oxytocin neurones: beyond magno- and parvocellular cell types? J. Neuroendocrinol. 30, e12549 (2018).

    Google Scholar 

  21. 21.

    Johnson, Z. V. et al. Central oxytocin receptors mediate mating-induced partner preferences and enhance correlated activation across forebrain nuclei in male prairie voles. Horm. Behav. 79, 8–17 (2016).

    CAS  PubMed  Google Scholar 

  22. 22.

    Hung, L. W. et al. Gating of social reward by oxytocin in the ventral tegmental area. Science 357, 1406–1411 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. 23.

    Okabe, S., Yoshida, M., Takayanagi, Y. & Onaka, T. Activation of hypothalamic oxytocin neurons following tactile stimuli in rats. Neurosci. Lett. 600, 22–27 (2015).

    CAS  PubMed  Google Scholar 

  24. 24.

    Bru, T., Salinas, S. & Kremer, E. J. An update on canine adenovirus type 2 and its vectors. Viruses 2, 2134–2153 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Gunaydin, L. A. et al. Natural neural projection dynamics underlying social behavior. Cell 157, 1535–1551 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. 26.

    VanRyzin, J. W. et al. Microglial phagocytosis of newborn cells is induced by endocannabinoids and sculpts sex differences in juvenile rat social play. Neuron 102, 435–449.e6 (2019).

    CAS  PubMed  Google Scholar 

  27. 27.

    Erny, D. et al. Host microbiota constantly control maturation and function of microglia in the CNS. Nat. Neurosci. 18, 965–977 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Wickersham, I. R. et al. Monosynaptic restriction of transsynaptic tracing from single, genetically targeted neurons. Neuron 53, 639–647 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Manning, M., Stoev, S., Cheng, L. L., Ching Wo, N. & Chan, W. Y. Design of oxytocin antagonists, which are more selective than atosiban. J. Pept. Sci. 7, 449–465 (2001).

    CAS  PubMed  Google Scholar 

  30. 30.

    Grund, T. et al. Neuropeptide S activates paraventricular oxytocin neurons to induce anxiolysis. J. Neurosci. 37, 12214–12225 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Rhodes, C. H., Morriell, J. I. & Pfaff, D. W. Immunohistochemical analysis of magnocellular elements in rat hypothalamus: Distribution and numbers of cells containing neurophysin, oxytocin, and vasopressin. J. Comp. Neurol. 198, 45–64 (1981).

    CAS  PubMed  Google Scholar 

  32. 32.

    Uvnäs-Moberg, K., Handlin, L. & Petersson, M. Self-soothing behaviors with particular reference to oxytocin release induced by non-noxious sensory stimulation. Front. Psychol. 5, 1529 (2015).

    PubMed  PubMed Central  Google Scholar 

  33. 33.

    Walker, S. C., Trotter, P. D., Swaney, W. T., Marshall, A. & Mcglone, F. P. C-tactile afferents: Cutaneous mediators of oxytocin release during affiliative tactile interactions? Neuropeptides 64, 27–38 (2017).

    CAS  PubMed  Google Scholar 

  34. 34.

    Brown, C. H., Bains, J. S., Ludwig, M. & Stern, J. E. Physiological regulation of magnocellular neurosecretory cell activity: integration of intrinsic, local and afferent mechanisms. J. Neuroendocrinol. 25, 678–710 (2013).

    CAS  PubMed  Google Scholar 

  35. 35.

    Hoffman, G. E. & Lyo, D. Anatomical markers of activity in neuroendocrine systems: are we all ‘Fos-ed out’? J. Neuroendocrinol. 14, 259–268 (2002).

    CAS  PubMed  Google Scholar 

  36. 36.

    Hoffman, G. E., Smith, M. S. & Verbalis, J. G. c-Fos and related immediate early gene products as markers of activity in neuroendocrine systems. Front. Neuroendocrinol. 14, 173–213 (1993).

    CAS  PubMed  Google Scholar 

  37. 37.

    Hasan, M. T. et al. A fear memory engram and its plasticity in the hypothalamic oxytocin system. Neuron 103, 133–146.e8 (2019).

    CAS  PubMed  Google Scholar 

  38. 38.

    Stern, J. E. Electrophysiological and morphological properties of pre-autonomic neurones in the rat hypothalamic paraventricular nucleus. J. Physiol. 537, 161–177 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Bosch, O. J. Brain oxytocin correlates with maternal aggression: link to anxiety. J. Neurosci. 25, 6807–6815 (2005).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Fenelon, V. S., Poulain, D. A. & Theodosis, D. T. Fos synthesis and neuronal activation: analysis of Fos immunoreactivity in identified magnocellular neurons during lactation. Ann. N.Y. Acad. Sci. 689, 508–511 (1993).

    CAS  PubMed  Google Scholar 

  41. 41.

    Neumann, I., Douglas, A. J., Pittman, Q. J., Russell, J. A. & Landgraf, R. Oxytocin released within the supraoptic nucleus of the rat brain by positive feedback action is involved in parturition‐related events. J. Neuroendocrinol. 8, 227–233 (1996).

    CAS  PubMed  Google Scholar 

  42. 42.

    Augustine, R. A. et al. Prolactin regulation of oxytocin neurone activity in pregnancy and lactation. J. Physiol. 595, 3591–3605 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Kennett, J. E. & McKee, D. T. Oxytocin: an emerging regulator of prolactin secretion in the female rat. J. Neuroendocrinol. 24, 403–412 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. 44.

    Dölen, G., Darvishzadeh, A., Huang, K. W. & Malenka, R. C. Social reward requires coordinated activity of nucleus accumbens oxytocin and serotonin. Nature 501, 179–184 (2013).

    PubMed  PubMed Central  Google Scholar 

  45. 45.

    McGlone, F., Wessberg, J. & Olausson, H. Discriminative and affective touch: sensing and feeling. Neuron 82, 737–755 (2014).

    CAS  PubMed  Google Scholar 

  46. 46.

    Leng, G. & Ludwig, M. Reply to: Improving research standards to restore trust in intranasal oxytocin. Biol. Psychiatry 79, e55–e56 (2016).

    PubMed  Google Scholar 

  47. 47.

    Meyer-Lindenberg, A., Domes, G., Kirsch, P. & Heinrichs, M. Oxytocin and vasopressin in the human brain: social neuropeptides for translational medicine. Nat. Rev. Neurosci. 12, 524–538 (2011).

    CAS  PubMed  Google Scholar 

  48. 48.

    Grinevich, V. & Neumann, I. D. How puzzle stones from animal studies translate into psychiatry. Mol. Psychiatry https://doi.org/10.1038/s41380-020-0802-9 (2020).

  49. 49.

    Menon, R. et al. Oxytocin signaling in the lateral septum prevents social fear during lactation. Curr. Biol. 28, 1066–1078.e6 (2018).

    CAS  PubMed  Google Scholar 

  50. 50.

    Grinevich, V. et al. Somatic transgenesis. Viral Vectors 3, 243–274 (2016).

  51. 51.

    Paxinos, G. & Watson, C. The Rat Brain in Stereotaxic Coordinates, 7th edn (Elsevier Acad. Press, 2014).

  52. 52.

    Tasker, J. G. & Dudek, F. E. Electrophysiological properties of neurones in the region of the paraventricular nucleus in slices of rat hypothalamus. J. Physiol. 434, 271–293 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  53. 53.

    Chu, C.-P. et al. Effects of stresscopin on rat hypothalamic paraventricular nucleus neurons in vitro. PLoS ONE 8, e53863 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  54. 54.

    Luther, J. A. & Tasker, J. G. Voltage-gated currents distinguish parvocellular from magnocellular neurones in the rat hypothalamic paraventricular nucleus. J. Physiol. 523, 193–209 (2000).

    CAS  PubMed  PubMed Central  Google Scholar 

  55. 55.

    Luther, J. A. et al. Neurosecretory and non-neurosecretory parvocellular neurones of the hypothalamic paraventricular nucleus express distinct electrophysiological properties. J. Neuroendocrinol. 14, 929–932 (2002).

    CAS  PubMed  Google Scholar 

  56. 56.

    Yuill, E. A., Hoyda, T. D., Ferri, C. C., Zhou, Q.-Y. & Ferguson, A. V. Prokineticin 2 depolarizes paraventricular nucleus magnocellular and parvocellular neurons. Eur. J. Neurosci. 25, 425–434 (2007).

    PubMed  PubMed Central  Google Scholar 

  57. 57.

    Tang, Y., Benusiglio, D., Grinevich, V. & Lin, L. Distinct types of feeding related neurons in mouse hypothalamus. Front. Behav. Neurosci. 10, 91 (2016).

    PubMed  PubMed Central  Google Scholar 

  58. 58.

    Maícas Royo, J., Brown, C. H., Leng, G. & MacGregor, D. J. Oxytocin neurones: intrinsic mechanisms governing the regularity of spiking activity. J. Neuroendocrinol. 28, 28 (2016).

  59. 59.

    Grund, T. et al. Chemogenetic activation of oxytocin neurons: temporal dynamics, hormonal release, and behavioral consequences. Psychoneuroendocrinology 106, 77–84 (2019).

    CAS  PubMed  Google Scholar 

  60. 60.

    de Jong, T. R. et al. Salivary oxytocin concentrations in response to running, sexual self-stimulation, breastfeeding and the TSST: the Regensburg Oxytocin Challenge (ROC) study. Psychoneuroendocrinology 62, 381–388 (2015).

    CAS  PubMed  Google Scholar 

  61. 61.

    Landgraf, R., Neumann, I., Holsboer, F. & Pittman, Q. J. Interleukin-1β stimulates both central and peripheral release of vasopressin and oxytocin in the rat. Eur. J. Neurosci. 7, 592–598 (1995).

    CAS  PubMed  Google Scholar 

  62. 62.

    Neumann, I. D., Maloumby, R., Beiderbeck, D. I., Lukas, M. & Landgraf, R. Increased brain and plasma oxytocin after nasal and peripheral administration in rats and mice. Psychoneuroendocrinology 38, 1985–1993 (2013).

    CAS  PubMed  Google Scholar 

  63. 63.

    Ishiyama, S. & Brecht, M. Neural correlates of ticklishness in the rat somatosensory cortex. Science 354, 757–760 (2016).

    CAS  PubMed  Google Scholar 

  64. 64.

    Althammer, F., Ferreira-Neto, H. C., Rubaharan, M., Roy, K. R. & Stern, J. E. Three-dimensional morphometric analysis reveals time-dependent structural changes in microglia and astrocytes in the central amygdala and hypothalamic paraventricular nucleus of heart failure rats. Res. Sq. https://doi.org/10.21203/rs.3.rs-22630/v1 (2020).

  65. 65.

    Kim, E. J., Jacobs, M. W., Ito-Cole, T. & Callaway, E. M. Improved monosynaptic neural circuit tracing using engineered rabies virus glycoproteins. Cell Rep. 15, 692–699 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  66. 66.

    Zhang, B. et al. Reconstruction of the hypothalamo-neurohypophysial system and functional dissection of magnocellular oxytocin neurons in the brain. Preprint at bioRxiv https://doi.org/10.1101/2020.03.26.007070 (2020).

Download references

Acknowledgements

We thank T. Grund and X. Liu for initial contribution to this study, R. Stoop for valuable comments on the manuscript, J. Müller for packaging viral vectors, E. Kremer for the canine virus, J. Maicos-Roya for contributing to the modeling of OT release, S. Netser for his comments on the manuscript, C. Pitzer and the Interdisciplinary Neurobehavioral Core Facility of Heidelberg University for some of the behavioral experiments performed there, and T. Splettstoesser (www.scistyle.com) for composing Extended Data Fig. 10. The work was supported by Chinese Scholarship Council No. 201406140043 (to Y.T.), the German Research Foundation (DFG) within the Collaborative Research Center (SFB) 1158 seed grant for young researchers and Fyssen Foundation (to A.L.), DFG postdoctoral fellowship AL 2466/1-1 (to F.A.), Alexander von Humboldt research fellowship (to D.H.), Human Frontier Science Program RGP0019/2015 (to V.G. and S.W.), Israel Science Foundation (grant nos. 1350/12 and 1361/17), Milgrom Foundation and the Ministry of Science, Technology and Space of Israel (grant no. 3-12068, to S.W.), NIH grant (no. R01NS094640, to J.E.S.), BBSRC grant (no. BB/S000224/1, to G.L.), DFG grant (nos. NE 465/27, NE 465/31 and NE 465/34, to I.D.N.), ANR-DFG grant and PICS grant (no. GR 3619/701 and no. GR 07882, to A.C. and V.G.), NARSAD Young Investigator grant (no. 24821) and ANR JCJC grant (no. GR 19-CE16-0011-01, to A.C.), DFG grant (no. GR 3619/4-1), SFB 1158, SNSF-DFG grant (no. GR 3619/8-1) and Fritz Thyssen Foundation grant (no. 10.16.2.018 MN) (all to V.G.).

Author information

Affiliations

Authors

Contributions

Y.T., D.B., A.L., A.C. and V.G. designed and conceived the project. L.H. and P.D. performed the ex vivo electrophysiology. L.H. and A. Baudon did the ex vivo calcium imaging. Y.T., D.B. and S.W. performed the in vivo electrophysiology. Y.T. and A.L. did the fiber photometry. Y.T., D.B. and S.W. performed the behavioral experiments and analyses. D.B., M.E., D.H. and F.A. did the immunohistochemistry and confocal microscopy. A.L. and J.S. performed the trans-synaptic labeling of OT neuron inputs. M.S., M.O. and K.K.C. assisted with virus design for trans-synaptic labeling. F.A., M.K.K., R.K.R. and J.E.S. did the 3D reconstruction and analysis. A. Bludau and I.D.N. calculated the plasma OT dosages. G.L. did the modeling. Y.T., D.B., A.L., L.H., F.A., I.D.N., A.C. and V.G. prepared the manuscript. I.D.N., A.C. and V.G. supervised and administered the project, and acquired the funding.

Corresponding authors

Correspondence to Alexandre Charlet or Valery Grinevich.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Peer review information Nature Neuroscience thanks Dayu Lin, Jeffrey Tasker and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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

Extended data

Extended Data Fig. 1 Recording of optogenetically identified oxytocin neurons, local field potential, phase locking, synchronization and spike distribution.

a, The cell type specificity of rAAV-OTp-ChR2-mCherry expression in OT cells. OT-immunosignal (green) colocalizes with ChR2-mCherry (red) signal; the overlay appears in yellow. Scale bar = 100 µm. Bottom, right: quantification of the colocalization mCherry/ChR2 (red bar, 90.8 ± 3.9 %) and mCherry/OT immnoreactive cells (green bar, 82.6 ± 7.7 %). Bar plots show mean ± SEM. b, Post-hoc verification of implanted optoprobe location in the PVN in a representative animal (one of five rats). Scale bar = 100 µm. c, Sorted extracellular spike waveforms (n = 175 action potentials) of a representative single unit optically-identified as OT neurons. d, Silicon probe (NeuroNexus) with 32-channel single shank were used in acute (anesthetized) single units recording. Sorted units and their location in the channels map were visualized with Phy-GUI (klusta, Python). Spike sorting was manually done in Plexon Offline Sorter 4.0 (Plexon, Inc.). e, PSTHs illustrating two optogenetically-identified oxytocin neurons by their response to blue light pulses (1 Hz, 5 Hz and 10 Hz laser stimulation, 10 ms, λ=473 nm, 10 mW/mm2). In both neurons, low frequency stimulation evoked spikes with a relatively constant short latency of 2-10 ms. f, Power spectrogram of the local field potential (LFP) in the PVN recorded before (open field, OF), during and after (OF) a free social interaction (FSI) session. g, Average theta (5-10 Hz) power recorded during and FSI session is significantly higher than before FSI session (p = 0.03, n = 5 rats, paired two-sided t test). All data represented as mean + SEM. h, P value distribution of phase-locking between theta (5-10 Hz) oscillations and OT cells spikes during exploratory (OF) or social (FSI) behavior (** p = 0.0089, n = 15 cells from 5 rats, paired two-sided t test; box plot shows median 10th, 25th, 75th, and 90th percentiles; min/max: OF, 0.07/0.99; SI, 0.002/0.08). i, j, Example traces (black) of LFP in the PVN and band-pass (5-10 Hz) filtered theta oscillations (red) during exploratory (OF) or social (FSI) behavior. Examples of distribution of four OT neurons firing in relation to LFP theta oscillations; OT neurons spikes are phase-locked with theta oscillations (** P = 0.0014, n = 15 cells) during social interaction (FSI, j), but not during exploratory behavior (i). Circular representation of OT neurons firing in relation to theta oscillation phase shows phase locking during social behavior (l) exclusively. No significant difference of spike-phase coupling between social behavior subtypes (P = 0.28). Significance of phase locking are determined by Rayleigh test for circular uniformity. k, l, Cross-correlation of pairs of oxytocin neurons recorded simultaneously. During open field (k) test there is no detectable correlation between oxytocin neurons spiking activity, but during social interaction (l) there is a significant increase (P = 0.0038, n = 12 cell pairs) of temporally correlated spikes within a time window of τ = 1.2 ± 0.5 s (mean correlation half-time). m, n, Examples of interspike interval (ISI, time bins = 10 ms) histograms of four OT neurons recorded during exploratory behavior (OF, m) and during social interaction (FSI, n). During FSI there is a prominent increase of spikes with short intervals due to increased spike clustering.

Extended Data Fig. 2 Analysis of social interactions, the firing rate of oxytocin and non-oxytocin PVN neurons, and activity-dependent model of oxytocin secretion.

a-c, Examples of manually classified social interaction behaviors: ano-genital sniffing, chasing, head-to-head events. PSTHs of a single unit identified as oxytocin neurons aligned to the onset of the specific behavior. Averaged (10 s time bin) PSTHs responses before, during, and after each behavior (0 to +10 s vs. basal P = 0.12, P = 0.45, P = 0.88, respectively, n = 6 cells per behavior, two-sided Wilcoxon test). d, Example firing rate of four PVN OT neurons recorded during exploratory (OF) behavior. e, Linear regression of PVN OT and non-OT neuronal firing rate in relation to rat’s distance in the area during FSI. OT neuronal activity shows a moderate negative correlation with distance (r = -0.3, p = 0.0092), while non-OT neurons do not show any significant correlation (r = −0.04, P = 0.11). f, Example firing rate of four PVN non-OT neurons recorded during social interaction. g, Average firing rate of 21 non-OT neurons from five rats (OF baseline 1.2 ± 0.2 Hz, not socially interacting (not SI) firing rate 1.2 ± 0.2 Hz, and social interacting (SI) firing rate 1.4 ± 0.2 Hz; P = 0.83, P = 0.23, P = 0.34, one-way ANOVA). Average index of dispersion on 1-s time bins of 21 non-OT cells (OF 1.1 ± 0.2, not SI 1.1 ± 0.2, SI 1.4 ± 0.3, P = 0.78, P = 0.07, P = 0.11, one-way ANOVA). Average pairwise Pearson correlation of spiking activity (1-s time bins) of 21 non-OT neurons’ pairs recorded in OF and FSI (P = 0.98, paired two-sided t test). All data show average + SEM. h, Schematic illustration of the theoretical model used to estimate the amount of secreted oxytocin from axonal terminals given the measured neuronal spike pattern. i, Average firing rate of OT cells from rats that underwent open field (OF), CSI, and FSI tests. j, Predicted OT secretion rate for a representative OT cell in each condition (left). Average OT predicted secretion rate (right) in each condition (OF-CSI, P = 0.11, OF-FSI, ** P = 0.005, OF-CSI, ** P = 0.007, spike pattern data used for prediction are from n = 15 cells from 5 rats, one-way ANOVA followed by Tukey’s post hoc test). Data represented as mean + SEM.

Extended Data Fig. 3 Ultra Sound Vocalizations (USVs) and OT neuronal activity.

a, Histograms of USVs peak frequency distribution during CSI (top) and FSI (bottom). b, Examples of sound spectrograms showing USVs events classified as whistles, calls, or complex vocalizations. c, PSTHs of oxytocin neurons spiking activity aligned to USVs onset (red dashed line) show no significant time-locked correlation between them. d, Firing rate of oxytocin neurons 0-5 s before USV events versus 0-5 s after USV events showing no significant correlation (P = 0.24, n = 53 vocalisations, two-sided Wilcoxon test). e, Total number of USVs registered in 5 pairs (experimental and stimulus animal) of rats during OF, CSI, and FSI tests divided - according to their frequency and duration – in non-social (< 25 kHz), trills (< 10 ms), calls (> 10 ms, not modulated), and complex vocalizations (> 10 ms, frequency modulated or mixed). All data represented as mean + SEM.

Extended Data Fig. 4 Oxytocin neurons response to airpuffs and to socially-related olfactory stimuli.

a, Illustration of single-unit recordings of oxytocin neurons during airpuffs applied to 1. anterior, 2. central, 3. posterior (n = 23 cells from 8 rats) part of the dorsal body region. Average increase response of oxytocin neurons compared to baseline for different stimulations regions (*P = 0.017, *P = 0.025, *P = 0.021 respectively, n = 23 cells from 8 rats, one-way ANOVA followed by Bonferroni post hoc comparison). PSTHs showing single oxytocin neurons (left) and averaged (right) and response to repeated airpuffs in all stimulation sites. b, Combined PSTHs of 1., 2., 3., showing different response latency of oxytocin neuron to airpuff stimulations on the dorsal body area. c, Single-unit recordings of oxytocin neurons during airpuff stimulations on: whiskers (n = 10 cells), abdomen (n = 14 cells), and anogenital area (n = 12 cells). Average increase response of oxytocin neurons compared to baseline for different stimulation regions. PSTHs showing single oxytocin neurons (left) and averaged (right) and response to repetitive airpuff stimulations in all stimulation sites. All data represented as mean + SEM. d, Illustration of the experimental setup for recording oxytocin neurons activity (opto-electrodes) during presentation of olfactory stimuli. e, Average firing rate and index of dispersion of six oxytocin neurons 10 s before presentation of the olfactory stimuli vs 10 s after; no significant changes are detected (P = 0.34, P = 0.48, n = 6 cells from 3 rats, paired two-sided t test). Data represented as average + SEM. f, PSTHs of 4 (out of 6 recorded) oxytocin neurons spiking activity aligned to onset of olfactory cues (red dashed line) - urinated bedding smell; no significant changes in firing rate are detected.

Extended Data Fig. 5 Ex vivo effects of excitatory hM3D(Gq) and inhibitory hM4D(Gi) DREADD in parvOT neurons.

a, Schema of viral vectors injected for ex vivo recording of parvOT neurons with concomitant DREADD-Gq activation. The picture shows hM3D(Gq) (purple) and OT (blue) immunoreactivities in the PVN of one out five rats. Scale bar = 100 µm and (inset) 10 µm. b-c, Spontaneous response from parvOT neurons expressing hM3D(Gq) in before and after the CNO bath application; b example traces, c quantification (baseline 0.85 ± 0.39 Hz vs CNO 1.31 ± 0.51 Hz, n = 9 cells from 5 rats; P = 0.0039). d-f, Evoked responses from parvOT neurons expressing hM3D(Gq) to current injections before and after the CNO bath application; (d) example traces, (e) quantification per step (before CNO current step 80 pA 1.727 ± 0.428 nAP vs after CNO current step 80 pA 3.182 ± 0.772 nAP, n = 11; **P < 0.01); (f) quantification of the average response (16.18 ± 3.89 AP vs CNO 22.55 ± 5.66 AP, n = 11 cells from 5 rats; P = 0.0314). g, Schema of viral vectors injected for ex-vivo recording of parvOT neurons with concomitant DREADD-Gi excitation. The picture shows hM4D(Gi) (purple) and OT (blue) immunoreactivities in the PVN of one out seven rats. Scale bar = 100 µm and (inset) 10 µm. h-i, Spontaneous response from parvOT neurons expressing hM4D(Gi) in before and after the CNO bath application; b example traces, c quantification (baseline 1.38 ± 0.38 Hz vs CNO 0.36 ± 0.18 Hz, n = 7 cells from 7 rats; P = 0.0469). j-l, Evoked responses from parvOT neurons expressing hM4D(Gi) to current injections before and after the CNO bath application; (j) example traces, (k) quantification per step (before CNO current step 20 pA 1.625 ± 0.263 nAP vs after CNO current step 20 pA 0.5 ± 0.189 nAP, n = 8; ***P < 0.001); l, quantification of the average response (baseline 13 ± 2.02 AP vs CNO 7.75 ± 2.03 AP, n = 8 cells from 7 rats; p = 0.0007). All results are expressed as average ± SEM. The statistical significances: * P < 0.05; ** P < 0.01; *** P < 0.001 (two-sided Wilcoxon test: c and i; Two-way ANOVA followed by a Holm-Sidak post hoc test: e and k; Paired two-sided t test: f and l). Open circles indicate individual cells.

Extended Data Fig. 6 Three-dimensional reconstruction and assessment of parvOT-magnOT connectivity.

a, Confocal images show specific labeling of parvOT neurons using CAV2-Cre in combination with OTp-DIO-GFP. Top panel shows a representative PVN (one out of three independent experiments, n = 3 rats) image with 2 labeled parvOT neurons. Bottom panel shows an image with higher magnification and overlay with synaptophysin (SYN). OT = purple, GFP = green, SYN = red. Scale bars = 200 µm (top panel) and 50 µm (bottom panel). b, Images show raw fluorescent confocal z-stacks and surface reconstruction of individual fluorescent channels (one out of three independent experiments, n = 3 rats). The top panel shows fluorescent signals of OT, GFP and merge (+SYN). The bottom panel shows the same channels with applied surface reconstruction. Bottom images have been vertically flipped to enhance visualization of the reconstructed channels. Scale bars = 75 µm. c, Images show the IMARIS three-dimensional surface reconstruction of OT, GFP and SYN. Boxes with dashed lines and asterisks indicate the overlap of GFP and SYN. In the right panel the overlap between OT (purple) and GFP (green) has been manually removed to visualize the GFP/SYN (green/red) overlap (n = 169 cells from 3 rats). Scale bar = 10 µm. d, Confocal images show synaptic contact of parvOT neurons with magnOT somata and dendrites. Top panel shows axo-somatic contact. Bottom panel shows axo-dendritic contact using a high magnification confocal z-stack. Asterisks indicate synaptic contact. Scale bars = 10 µm. e, Quantification of SON OT neuron chance to receive innervation by parvOT neurons in respect to their rostro-caudal location (n = 169 cells from 3 rats). All data are presented as mean ± SEM.

Extended Data Fig. 7 Fiber photometry recording of PVN parvOT neurons and SON magnOT neurons with chemogenetic manipulation of parvOT neurons.

a-b, Schema of viral vectors injected (CAV2-Cre in the SON and OTp-DIO-GCaMP6s in the PVN) and implanted optic fiber for fiber photometry recording of PVN parvOT neurons (a). No Ca2+ transient nor changes in Ca2+ signal upon ‘airpuff’ stimulation were detected when recording parvOT neurons exclusively (b, solid line: average, shaded area: SEM, n = 3 rats). c-d, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording of SON magnOT neurons (c). Examples of fiber photometry-based Ca2+ signals of SON magnOT population during airpuff stimulation (d); the graphic is an average of 33 airpuff responses (11 airpuffs per animal, n = 3 rats). e, Relative change in the area under the curve (AUC) 0-30 s after airpuffs with respect to 30 s before the stimuli, of SON magnOT vs. parvOT neurons (solid line: average, shaded area: SEM, n = 33 airpuffs from 3 rats, ** P = 0.008, unpaired two-sided t test). Data show mean + SEM. f-g-h, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording (f) of SON OT neurons with concomitant DREADD-Gq activation of parvOT neurons. Normalized area under the curve (AUC) of GCaMP6s signal (g, solid line: average, shaded area: SEM, 1 min bin size) of SON OT neurons showing increase of cells activities after parvOT activation mediated by CNO i.p. injection (indicated by arrow). 30-min averaged AUC (h) showing a gradual increase in cellular activity and lasting at least 120 min (*P = 0.0406, *P = 0.0274, *P = 0.0107, *P = 0.0301, n = 3 rats, two-way ANOVA Tukey’s corrected post-hoc comparison). Data show mean + SEM. i-j-k, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording (i) of SON OT neurons with concomitant DREADD-Gi inhibition of parvOT neurons. Normalized area under the curve (AUC) of GCaMP6s signal (j, solid line: average, shaded area: SEM, 1 min bin size) of PVN OT neurons showing a decrease of cellular activity after parvOT inhibition mediated by i.p. CNO injection (indicated by arrow). 30-min averaged AUC (k) shows a sharp decrease in cellular activity that lasts at least 60 min and then gradually recovers (***P = 0.0008, **P = 0.0051, *P = 0.0140, *P = 0.0168, n = 3 rats, two-way ANOVA Tukey’s corrected post-hoc comparison). Data show mean + SEM. l-m-n, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording (l) of SON OT neurons in control animals (DREADD-free) expressing GFP in parvOT neurons. Normalized area under the curve (AUC) of GCaMP6s signal (m, solid line: average, shaded area: SEM, 1 min bin size) of PVN OT neurons showing no significant changes in Ca2+ signal upon CNO injection. No significant changes are detected in 30-min averaged AUC (n) up to 120 min (P = 0.5715, P = 0.2429, P = 0.4673, P = 0.2848, n = 2 rats, two-way ANOVA Tukey’s corrected post-hoc comparison). Data show mean values. o, Panels of an immunostained section of the SON (one of out of eight independent experiments) showing post-hoc verification of implanted optic fiber above the SON and co-localization of immunoreactive, GCaMP6s (green), oxytocin (blue), and merged channels. Scale bar 100 μm and 10 μm (inset).

Extended Data Fig. 8 Inputs to parvOT and magnOT neurons.

a, c, Virus injection strategy to retrotrace inputs from parvOT and magnOT neurons, respectively and to control injection sites of Cre (n = 5 rats). b, d, Injection site of CAV2-Cre in SON and of AAVretro-Cre in PP were confirmed by staining for Cre or mCherry in SON (n = 5 rats). e, Immunofluorescence for mCherry (red, fused to TVA), OT (blue) and overlay of the PVN from a rat injected with rAAV-OTp-TCB (n = 2 rats). f, Immunofluorescence for OT (blue), GFP (Green) and overlay of the PVN from a rat injected with rAAV-OTp-TCB, and Rb-GFP two weeks later (n = 2 rats). g, Injection site of rabies in parvOT (Top) and magnOT (bottom) groups (n = 5 rats). h, Epifluorescence microscope images showing neurons monosynaptically retrolabelled by Rabies-GFP in various brain areas projecting to both parvOT and magnOT neurons. Top line: neurons projecting to parvOT neurons; Bottom line: neurons projecting to magnOT neurons (n = 5 rats). i, j, Epifluorescence microscope images showing neurons monosynaptically retrolabelled by Rabies-GFP in brain areas projecting specifically to parvOT (i) or magnOT neurons (j) (n = 5 rats). All scale bars = 100 μm. All scale bars in insert = 10 μm. Brain areas legend: Amygdala: AMY, Arcuate hypothalamic nucleus: Arc, Bed nucleus of stria terminalis: NST, Cingulate cortex: CgC, Claustrum: Cl, Dorsal raphe nucleus: DRN, Dorsal tenia tecta: DTT, Dorsomedial hypothalamic area: DMH, Habenular nucleus: Hb, Horizontal limb of diagonal band nucleus: HDB, Infralimbic cortex: ILC, Insular cortex: Ins, Lateral hypothalamic area: LH, Lateral lemniscus nucleus: LMN, Lateral septal nucleus: SEP, Mammillary body: MMB, Medial preoptic area: MPO, Median raphe nucleus: MRN, Nucleus accumbens: NAC, Orbital cortex: OC, Parabrachial nucleus: PBN, Paraventricular thalamus: PVT, Pedunculopontine tegmental nucleus: PPT, Periaqueductal gray area: PAG, Posterior hypothalamic nucleus: PH, Posterior intralaminar thalamus: PIL, Prelimbic cortex: PLC, Raphe magnus nucleus: RMg, Subfornical organ:SFO, Substancia nigra: SN, Vascular organ of lamina terminalis: OVLT, Ventral Subiculum: VS, Zona incerta: ZI.

Extended Data Fig. 9 Effects of DREADD activation of parvocellular oxytocin neurons on social behaviors.

a, Parvocellular oxytocin-GFP control group: time spent by rats injected with saline or CNO socially interacting with conspecific stimulus in CSI (P = 0.34) and FSI session (P = 0.29, n = 6 rats, paired two-sided t test). b, CNO does not affect locomotor activity: average distance run by experimental rats (n = 6 rats GFP group, n = 15 rats Parvo-Gi group, n = 9 rats Parvo-Gq group) injected with saline or CNO during an open field (OF) test. No significant changes were detected, one-way ANOVA Tukey’s corrected post-hoc comparison. c, Quantification of ‘active social avoidance’ behavior of experimental rat (actively escaping from stimulus rat, which is trying to interact) expressing inhibitory DREADD hM4D(Gi) in parvOT neurons and injected either with saline (control) or CNO. The total time of free social interaction session is 5 minutes. ** P = 0.0096, n = 15 rats, paired two-sided t test). d-e, OT receptor antagonist (OTR-a), CNO, or both do not affect locomotor activity: average distance run by experimental rats (d: P = 0.79, n = 6 rats saline/CNO/saline, P = 0.92 n = 6 rats saline/CNO/OTR-a; e: P = 0.73, n = 5 rats saline/CNO/saline, p = 0.64, n = 5 rats saline/CNO/OTR-a) injected with saline/CNO/OTR-a during an open field (OF) test. One-way ANOVA Tukey’s corrected post-hoc comparison. f-g, Quantification of ‘active social avoidance’ behavior of experimental rat after administration of saline/CNO/OTR-a. Infusion of OTR-a induced an increase of avoidance behavior (Parvo-Gq: **P = 0.0027, **P = 0.0018, n = 6 rats per group, Parvo-GFP: **P = 0.0042, **P = 0.0013, n = 5 rats per group, one-way ANOVA Tukey’s corrected post-hoc comparison). h-i, Time spent in different subtype of social behavior of experimental rat expressing excitatory DREADD hM3D(Gq) (h) or GFP (i) in parvOT neurons after administration of saline/CNO/OTR-a. Crawling on top behavior was the subtype most affected by parvOT neurons chemogenetic activation and by infusion of OTR-a (Parvo-Gq: ** P = 0.01, **P = 0.004, *P = 0.023, n = 6 rats per group, Parvo-GFP: **P = 0.006, *P = 0.035, *P = 0.047, one-way ANOVA Tukey’s corrected post-hoc comparison). All data represented as mean + SEM.

Extended Data Fig. 10 Working hypothesis.

Non-nociceptive signals (‘social touch’) arising from stimulation of dorsal body parts of interacting virgin female rats converge onto hypothalamic parvocellular oxytocin neurons via ascending pathways. As a consequence, the somatosensory-driven activation of parvocellular oxytocin neurons is transmitted to magnocellular oxytocin neurons inducing central oxytocin release in the social-relevant forebrain areas (as schematically depicted by circles representing nine amino acids in the oxytocin molecule), to support motivated social communication between female conspecifics.

Supplementary information

Supplementary Information

Supplementary Videos 1 and 2 description, and Supplementary Tables 1–3.

Reporting summary

Supplementary Video 1

Simultaneous recording of four OT neurons during social interaction of female rats.

Supplementary Video 2

Silencing of parvOT neurons attenuates social interactions of female rats.

Supplementary Software 1

Python code used for ex vivo calcium-imaging data analysis in Fig. 4a–d. MATLAB code used for in vivo fiber photometry data analysis in Fig. 4e–o and Extended Data Fig. 7a–n.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Tang, Y., Benusiglio, D., Lefevre, A. et al. Social touch promotes interfemale communication via activation of parvocellular oxytocin neurons. Nat Neurosci (2020). https://doi.org/10.1038/s41593-020-0674-y

Download citation