Nociceptors, or pain‐sensitive receptors, are unique among sensory receptors in that their sensitivity is increased by noxious stimulation. This process, called sensitization or hyperalgesia, is mediated by a variety of proinflammatory factors, including bradykinin, ATP and NGF, which cause sensitization to noxious heat stimuli by enhancing the membrane current carried by the heat‐ and capsaicin‐gated ion channel, TRPV1. Several different mechanisms for sensitization of TRPV1 have been proposed. Here we show that NGF, acting on the TrkA receptor, activates a signalling pathway in which PI3 kinase plays a crucial early role, with Src kinase as the downstream element which binds to and phosphorylates TRPV1. Phosphorylation of TRPV1 at a single tyrosine residue, Y200, followed by insertion of TRPV1 channels into the surface membrane, explains most of the rapid sensitizing actions of NGF.

1 The effects of activation of protein kinase C (PKC) on membrane currents gated by capsaicin, protons, heat and anandamide were investigated in primary sensory neurones from neonatal rat dorsal root ganglia (DRG) and in HEK293 cells (human embryonic kidney cell line) transiently or stably expressing the human vanilloid receptor hVR1. 2 Maximal activation of PKC by a brief application of phorbol 12-myristate 13-acetate (PMA) increased the mean membrane current activated by a low concentration of capsaicin by 1.65-fold in DRG neurones and 2.18-fold in stably transfected HEK293 cells. Bradykinin, which activates PKC, also enhanced the response to capsaicin in DRG neurones. The specific PKC inhibitor RO31-8220 prevented the enhancement caused by PMA. 3 Activation of PKC did not enhance the membrane current at high concentrations of capsaicin, showing that PKC activation increases the probability of channel opening rather than unmasking channels. 4 Application of PMA alone activated an inward current in HEK293 cells transiently transfected with VR1. The current was suppressed by the VR1 antagonist capsazepine. PMA did not, however, activate a current in the large majority of DRG neurones nor in HEK293 cells stably transfected with VR1. 5 Removing external Ca²⁺ enhanced the response to a low concentration of capsaicin 2.40-fold in DRG neurones and 3.42-fold in HEK293 cells. Activation of PKC in zero Ca²⁺ produced no further enhancement of the response to capsaicin in either DRG neurones or HEK293 cells stably transfected with VR1. 6 The effects of PKC activation on the membrane current gated by heat, anandamide and low pH were qualitatively similar to those on the capsaicin-gated current. 7 The absence of a current activated by PMA in most DRG neurones or in stably transfected HEK293 cells suggests that activation of PKC does not directly open VR1 channels, but instead increases the probability that they will be activated by capsaicin, heat, low pH or anandamide. Removal of calcium also potentiates activation, and PKC activation then has no further effect. The results are consistent with a model in which phosphorylation of VR1 by PKC increases the probability of channel gating by agonists, and in which dephosphorylation occurs by a calcium-dependent process.

Pain differs from other sensations in many respects. Primary pain-sensitive neurons respond to a wide variety of noxious stimuli, in contrast to the relatively specific responses characteristic of other sensory systems, and the response is often observed to sensitize on repeated presentation of a painful stimulus, while adaptation is typically observed in other sensory systems. In most cases the cellular mechanisms of transduction and sensitization in response to painful stimuli are not understood. We report here that application of pulses of noxious heat to a subpopulation of isolated primary sensory neurons rapidly activates an inward current. The ion channel activated by heat discriminates poorly among alkali cations. Calcium ions both carry current and partially suppress the current carried by other ions. The current is markedly increased by bradykinin, a potent algogenic nonapeptide that is known to be released in vivo by tissue damage. Phosphatase inhibitors prolong the sensitization caused by bradykinin, and a similar sensitization is caused by activators of protein kinase C. We conclude that bradykinin sensitizes the response to heat by activating protein kinase C.

The rate of action potential firing in nociceptors is a major determinant of the intensity of pain. Possible modulators of action potential firing include the HCN ion channels, which generate an inward current, I_h, after hyperpolarization of the membrane. We found that genetic deletion of HCN2 removed the cyclic adenosine monophosphate (cAMP)–sensitive component of I_h and abolished action potential firing caused by an elevation of cAMP in nociceptors. Mice in which HCN2 was specifically deleted in nociceptors expressing Na_V1.8 had normal pain thresholds, but inflammation did not cause hyperalgesia to heat stimuli. After a nerve lesion, these mice showed no neuropathic pain in response to thermal or mechanical stimuli. Neuropathic pain is therefore initiated by HCN2-driven action potential firing in Na_V1.8-expressing nociceptors.

Thermally activated ion channels are known to detect the entire thermal range from extreme heat (TRPV2), painful heat (TRPV1, TRPM3 and ANO1), non-painful warmth (TRPV3 and TRPV4) and non-painful coolness (TRPM8) through to painful cold (TRPA1)^{1,2,3,4,5,6,7}. Genetic deletion of each of these ion channels, however, has only modest effects on thermal behaviour in mice^{6,7,8,9,10,11,12}, with the exception of TRPM8, the deletion of which has marked effects on the perception of moderate coolness in the range 10–25 °C¹³. The molecular mechanism responsible for detecting non-painful warmth, in particular, is unresolved. Here we used calcium imaging to identify a population of thermally sensitive somatosensory neurons which do not express any of the known thermally activated TRP channels. We then used a combination of calcium imaging, electrophysiology and RNA sequencing to show that the ion channel generating heat sensitivity in these neurons is TRPM2. Autonomic neurons, usually thought of as exclusively motor, also express TRPM2 and respond directly to heat. Mice in which TRPM2 had been genetically deleted showed a striking deficit in their sensation of non-noxious warm temperatures, consistent with the idea that TRPM2 initiates a ‘warm’ signal which drives cool-seeking behaviour.