Abstract
The spinal cord is the initial stage that integrates temperature information from peripheral inputs. Here we used molecular genetics and in vivo calcium imaging to investigate the coding of cutaneous temperature in the spinal cord in mice. We found that heating or cooling the skin evoked robust calcium responses in spinal neurons, and their activation threshold temperatures distributed smoothly over the entire range of stimulation temperatures. Once activated, heat-responding neurons encoded the absolute skin temperature without adaptation and received major inputs from transient receptor potential (TRP) channel V1 (TRPV1)-positive dorsal root ganglion (DRG) neurons. By contrast, cold-responding neurons rapidly adapted to ambient temperature and selectively encoded temperature changes. These neurons received TRP channel M8 (TRPM8)-positive DRG inputs as well as novel TRPV1+ DRG inputs that were selectively activated by intense cooling. Our results provide a comprehensive examination of the temperature representation in the spinal cord and reveal fundamental differences in the coding of heat and cold.
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References
Julius, D. TRP channels and pain. Annu. Rev. Cell Dev. Biol. 29, 355–384 (2013).
Vriens, J., Nilius, B. & Voets, T. Peripheral thermosensation in mammals. Nat. Rev. Neurosci. 15, 573–589 (2014).
Basbaum, A.I., Bautista, D.M., Scherrer, G. & Julius, D. Cellular and molecular mechanisms of pain. Cell 139, 267–284 (2009).
Palkar, R., Lippoldt, E.K. & McKemy, D.D. The molecular and cellular basis of thermosensation in mammals. Curr. Opin. Neurobiol. 34, 14–19 (2015).
Caterina, M.J. et al. Impaired nociception and pain sensation in mice lacking the capsaicin receptor. Science 288, 306–313 (2000).
Caterina, M.J. et al. The capsaicin receptor: a heat-activated ion channel in the pain pathway. Nature 389, 816–824 (1997).
McKemy, D.D., Neuhausser, W.M. & Julius, D. Identification of a cold receptor reveals a general role for TRP channels in thermosensation. Nature 416, 52–58 (2002).
Peier, A.M. et al. A TRP channel that senses cold stimuli and menthol. Cell 108, 705–715 (2002).
Bautista, D.M. et al. The menthol receptor TRPM8 is the principal detector of environmental cold. Nature 448, 204–208 (2007).
Dhaka, A. et al. TRPM8 is required for cold sensation in mice. Neuron 54, 371–378 (2007).
Colburn, R.W. et al. Attenuated cold sensitivity in TRPM8 null mice. Neuron 54, 379–386 (2007).
Dhaka, A., Viswanath, V. & Patapoutian, A. Trp ion channels and temperature sensation. Annu. Rev. Neurosci. 29, 135–161 (2006).
Story, G.M. et al. ANKTM1, a TRP-like channel expressed in nociceptive neurons, is activated by cold temperatures. Cell 112, 819–829 (2003).
Kobayashi, K. et al. Distinct expression of TRPM8, TRPA1, and TRPV1 mRNAs in rat primary afferent neurons with aδ/c-fibers and colocalization with trk receptors. J. Comp. Neurol. 493, 596–606 (2005).
Dhaka, A., Earley, T.J., Watson, J. & Patapoutian, A. Visualizing cold spots: TRPM8-expressing sensory neurons and their projections. J. Neurosci. 28, 566–575 (2008).
Takashima, Y. et al. Diversity in the neural circuitry of cold sensing revealed by genetic axonal labeling of transient receptor potential melastatin 8 neurons. J. Neurosci. 27, 14147–14157 (2007).
Todd, A.J. Neuronal circuitry for pain processing in the dorsal horn. Nat. Rev. Neurosci. 11, 823–836 (2010).
Ma, Q. Labeled lines meet and talk: population coding of somatic sensations. J. Clin. Invest. 120, 3773–3778 (2010).
Craig, A.D., Krout, K. & Andrew, D. Quantitative response characteristics of thermoreceptive and nociceptive lamina I spinothalamic neurons in the cat. J. Neurophysiol. 86, 1459–1480 (2001).
Burton, H. Responses of spinal cord neurons to systematic changes in hindlimb skin temperatures in cats and primates. J. Neurophysiol. 38, 1060–1079 (1975).
Bester, H., Chapman, V., Besson, J.M. & Bernard, J.F. Physiological properties of the lamina I spinoparabrachial neurons in the rat. J. Neurophysiol. 83, 2239–2259 (2000).
Andrew, D. & Craig, A.D. Spinothalamic lamina I neurones selectively responsive to cutaneous warming in cats. J. Physiol. (Lond.) 537, 489–495 (2001).
Stosiek, C., Garaschuk, O., Holthoff, K. & Konnerth, A. In vivo two-photon calcium imaging of neuronal networks. Proc. Natl. Acad. Sci. USA 100, 7319–7324 (2003).
Johannssen, H.C. & Helmchen, F. In vivo Ca2+ imaging of dorsal horn neuronal populations in mouse spinal cord. J. Physiol. (Lond.) 588, 3397–3402 (2010).
Chen, X., Gabitto, M., Peng, Y., Ryba, N.J. & Zuker, C.S. A gustotopic map of taste qualities in the mammalian brain. Science 333, 1262–1266 (2011).
Chen, T.W. et al. Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 499, 295–300 (2013).
Nakai, J., Ohkura, M. & Imoto, K. A high signal-to-noise Ca(2+) probe composed of a single green fluorescent protein. Nat. Biotechnol. 19, 137–141 (2001).
Harrison, J.L. & Davis, K.D. Cold-evoked pain varies with skin type and cooling rate: a psychophysical study in humans. Pain 83, 123–135 (1999).
Kenshalo, D.R., Holmes, C.E. & Wood, P.B. Warm and cool thresholds as a function of rate of stimulus temperature change. Percept. Psychophys. 3, 81–84 (1968).
Hensel, H. Temperaturempfindung und intracutane Warmebewegung. Pflugers Arch. Gesamte Physiol. Menschen Tiere 252, 165–215 (1950).
Pogorzala, L.A., Mishra, S.K. & Hoon, M.A. The cellular code for mammalian thermosensation. J. Neurosci. 33, 5533–5541 (2013).
Knowlton, W.M. et al. A sensory-labeled line for cold: TRPM8-expressing sensory neurons define the cellular basis for cold, cold pain, and cooling-mediated analgesia. J. Neurosci. 33, 2837–2848 (2013).
Cavanaugh, D.J. et al. Distinct subsets of unmyelinated primary sensory fibers mediate behavioral responses to noxious thermal and mechanical stimuli. Proc. Natl. Acad. Sci. USA 106, 9075–9080 (2009).
Zhang, J., Cavanaugh, D.J., Nemenov, M.I. & Basbaum, A.I. The modality-specific contribution of peptidergic and non-peptidergic nociceptors is manifest at the level of dorsal horn nociresponsive neurons. J. Physiol. (Lond.) 591, 1097–1110 (2013).
Kandel, E.R., Schwartz, J.H. & Jessell, T.M. Principles of Neural Science 4th edn. (McGraw-Hill, 2000).
Hensel, H. & Iggo, A. Analysis of cutaneous warm and cold fibres in primates. Pflugers Arch. 329, 1–8 (1971).
Zeilhofer, H.U., Wildner, H. & Yévenes, G.E. Fast synaptic inhibition in spinal sensory processing and pain control. Physiol. Rev. 92, 193–235 (2012).
Urch, C.E. & Dickenson, A.H. In vivo single unit extracellular recordings from spinal cord neurones of rats. Brain Res. Brain Res. Protoc. 12, 26–34 (2003).
Rieke, F. & Rudd, M.E. The challenges natural images pose for visual adaptation. Neuron 64, 605–616 (2009).
Sarria, I. & Gu, J. Menthol response and adaptation in nociceptive-like and nonnociceptive-like neurons: role of protein kinases. Mol. Pain 6, 47 (2010).
Rohács, T., Lopes, C.M., Michailidis, I. & Logothetis, D.E. PI(4,5)P2 regulates the activation and desensitization of TRPM8 channels through the TRP domain. Nat. Neurosci. 8, 626–634 (2005).
Liu, B., Zhang, C. & Qin, F. Functional recovery from desensitization of vanilloid receptor TRPV1 requires resynthesis of phosphatidylinositol 4,5-bisphosphate. J. Neurosci. 25, 4835–4843 (2005).
Yao, J. & Qin, F. Interaction with phosphoinositides confers adaptation onto the TRPV1 pain receptor. PLoS Biol. 7, e46 (2009).
Tominaga, M. et al. The cloned capsaicin receptor integrates multiple pain-producing stimuli. Neuron 21, 531–543 (1998).
Tominaga, M. & Tominaga, T. Structure and function of TRPV1. Pflugers Arch. 451, 143–150 (2005).
Duclaux, R. & Kenshalo, D.R. Sr. Response characteristics of cutaneous warm receptors in the monkey. J. Neurophysiol. 43, 1–15 (1980).
Kenshalo, D.R. & Duclaux, R. Response characteristics of cutaneous cold receptors in the monkey. J. Neurophysiol. 40, 319–332 (1977).
Karashima, Y. et al. TRPA1 acts as a cold sensor in vitro and in vivo. Proc. Natl. Acad. Sci. USA 106, 1273–1278 (2009).
Bandell, M. et al. Noxious cold ion channel TRPA1 is activated by pungent compounds and bradykinin. Neuron 41, 849–857 (2004).
Braz, J., Solorzano, C., Wang, X. & Basbaum, A.I. Transmitting pain and itch messages: a contemporary view of the spinal cord circuits that generate gate control. Neuron 82, 522–536 (2014).
Acknowledgements
We thank L. Luo for his generous support during the entire project, Z. Shen for initial experiments and G. Kamalani for assistance; and B. Barres (Stanford University) and D. Julius (University of California, San Francisco) for Mgfap-cre and Trpa1 knockout mice. We are grateful to X. Gao, C. Guenthner, B. Weissbourd and members of the Chen laboratory for comments on the manuscript. This work was supported by grants from the intramural research program of NIDCR (M.A.H.), and the Whitehall Foundation, Terman Fellowship and start-up funding from Stanford University (X.C.).
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C.R. and X.C. designed the study. C.R. conducted imaging experiments. C.R. and X.C. analyzed data. M.A.H. provided TRPM8- and TRPV1-DTR mice, and performed in situ hybridization experiments. C.R. and X.C. wrote the paper with help from M.A.H. X.C. supervised the research.
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Integrated supplementary information
Supplementary Figure 1 Temperatures recorded on the surface of the skin and under the skin are similar.
Stimulation temperature recorded on the surface of the skin (black) and under the skin (red). The data are averaged from 4 measurements and are presented as mean (dark color) ± s.e.m. (light color).
Supplementary Figure 2 Properties of spinal responses to cooling.
(a) The percentage of total neurons at different depths below the surface that are activated by cooling to 29 °C (yellow) and 5 °C (blue) (25 μm, n = 2 mice; 35 μm, n = 5 mice; 45 μm, n = 5 mice; 55 μm, n = 5 mice; 65 μm, n = 4 mice; 75 μm, n = 3 mice; 85 μm, n = 2 mice.) (b) Heat maps (middle) and example traces (bottom) of neuronal responses to cooling (top) in the spinal cord before (left) and after (right) application of NBQX (an AMPA and kainite receptors antagonist, 60 μM) and APV (an NMDA receptor antagonist, 150 μM). Scale bars, 10 s and 10% ΔF/F. (c) Top: An example FOV illustrating OGB (green) labeling in the spinal cord of a Mgfap-cre; Ai14 (cre-dependent tdTomato, red) mouse. Scale bar, 100 μm. Bottom: example calcium traces of cold response from astrocytes (red) and neurons (green). Scale bars, 10 s and 10% ΔF/F.
Supplementary Figure 3 Cooling-response curves at different ATs.
(a) Neuronal responses to cooling stimuli with a series of ΔTs (3 °C, 6 °C, 10 °C and 16 °C) at two ATs (Top traces and heat maps: AT = 32 °C. Bottom traces and heat maps: AT = 27 °C). (b) Quantification of a (n = 5 mice). Scale bars, 10 s.
Supplementary Figure 4 Dorsoventral distribution and absolute temperature-coding property of heat-responding neurons.
(a) The percentage of total neurons that are activated by heating to 37 °C (orange) and 45 °C (red) at different depths below the surface (n = 2-5 mice). (25 μm, n = 2 mice; 35 μm, n = 5 mice; 45 μm, n = 5 mice; 55 μm, n = 5 mice; 65 μm, n = 4 mice; 75 μm, n = 3 mice; 85 μm, n = 2 mice.) (b) Quantification of the number of heat-responding neurons (squares, left Y axis) and the averaged peak ΔF/F (circles, right Y axis) in response to heating at different rates (n = 5 mice).
Supplementary Figure 5 Diphtheria toxin (DT) treatments of the TRPM8-DTR-GFP and TRPV1-DTR-GFP mice.
(a) The expression of TRPM8 (in situ hybridization) and TRPV1 (immunofluorescence) in the DRGs of wild type, TRPM8- and TRPV1-DTR mice after diphtheria toxin treatment. Similar observations were made in all mice (number specified below). Scale bar, 100 μm. (b) Quantification of a (Left: n = 2 mice for WT, n = 5 mice for TRPM8-DTR and n = 4 mice for TRPV1-DTR, right: n = 4 mice per genotype). (c) The expression of GFP (immunofluorescence) in the DRGs of saline- or DT-treated TRPM8- and TRPV1-DTR-GFP mice. Similar observations were made in all mice (number specified below). Scale bar, 100 μm. (d) Quantification of c (TRPM8-DTR-GFP: n = 2 mice per treatment group, TRPV1-DTR-GFP: n = 4 mice per treatment group).
Supplementary Figure 6 TRPA1 channel does not mediate spinal responses to cold.
(a) Heat maps of the activities of all cold-responding neurons in representative FOVs in WT (138 neurons) and Trpa1 knockout (146 neurons) mice. (b) The temperature−response relationship in WT (n = 12 mice, same data as in Fig. 2c) and Trpa1 knockout (n = 4 mice) mice. (c) The distribution of activation thresholds of cold-responding neurons in WT (same data as in Fig. 5e) and TRPA1 knockout mice. Scale bar, 10 s.
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Ran, C., Hoon, M. & Chen, X. The coding of cutaneous temperature in the spinal cord. Nat Neurosci 19, 1201–1209 (2016). https://doi.org/10.1038/nn.4350
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DOI: https://doi.org/10.1038/nn.4350
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