Accepted

Ghrelin has been shown to alleviate neuropathic pain by inhibiting the release of proinflammatory cytokines. The purpose of this study was to investigate the role of GSK-3β/β-catenin signaling in mediating the effect of ghrelin on neuropathic pain and to understand the associated mechanisms. Chronic constriction injury (CCI) of the sciatic nerve was used to establish a rat model of neuropathic pain. Hyperalgesia and allodynia were evaluated by observing the mechanical withdrawal threshold and the thermal withdrawal latency. Wnt3a and β-catenin protein expression and GSK-3β phosphorylation were detected by western blotting analysis. The levels of tumor necrosis factor-α and IL-1β were determined using an enzyme-linked immunosorbent assay. In addition, we used immunohistochemical analysis to determine the levels of GSK-3β phosphorylation in the dorsal horn of the spinal cord. Intrathecal delivery of ghrelin effectively ameliorated CCI-induced mechanical allodynia and thermal hyperalgesia at 7 and 14 days and reduced the levels of tumor necrosis factor-α. Ghrelin inhibited CCI-induced GSK-3β activation and β-catenin overexpression in the spinal dorsal horn. Moreover, intrathecal injection of ghrelin suppressed the activation of GSK-3β in the spinal dorsal horn of CCI rats, as assessed by immunohistochemical analysis. Our data indicated that ghrelin could markedly alleviate neuropathic pain by inhibiting the expression of β-catenin, via the suppression of GSK-3β activation, in the spinal cord of CCI rats.

The cornea is extensively innervated by trigeminal ganglion cold thermoreceptor neurons expressing TRPM8. These neurons respond to cooling, hyperosmolarity and wetness of the corneal surface. Surgical injury of corneal nerve fibers alters tear production and often causes dry eye sensation. The contribution of TRPM8-expressing corneal cold-sensitive neurons (CCSNs) to these symptoms is unclear. Using extracellular recording of CCSNs nerve terminals combined with confocal tracking of re-innervation, Ca imaging and patch-clamp recordings of fluorescent retrogradely labeled corneal neurons in culture, we analyzed the functional modifications of CCSNs induced by peripheral axonal damage in male mice. After injury, the percentage of CCSNs, the cold- and menthol-evoked intracellular [Ca] rises and TRPM8-current density in CCSNs were larger than in sham animals, with no differences in the brake K current I Active and passive membrane properties of CCSNs from both groups were alike, and corresponded mainly to those of canonical low- and high-threshold cold thermoreceptor neurons. Ongoing firing activity and menthol-sensitivity were higher in CCSN terminals of injured mice, an observation accounted by mathematical modeling. These functional changes developed in parallel with a partial re-innervation of the cornea by TRPM8(+) fibers and with an increase in basal tearing in injured animals compared to sham mice. Our results unveil key TRPM8-dependent functional changes in CCSNs in response to injury, suggesting that increased tearing rate and ocular dryness sensation derived from deep surgical ablation of corneal nerves are due to enhanced functional expression of TRPM8 channels in these injured trigeminal primary sensory neurons.This paper unveils a key role of TRPM8 in the sensory and autonomic disturbances associated with surgical damage of eye surface nerves. We studied the damage-induced functional alterations of corneal cold-sensitive neurons using confocal tracking of re-innervation, extracellular corneal nerve terminal recordings, tearing measurements Ca imaging and patch-clamp recordings of cultured corneal neurons, and mathematical modeling. Corneal nerve ablation upregulates TRPM8 mainly in canonical cold thermoreceptors, enhancing their cold- and menthol-sensitivity, inducing a rise in the ongoing firing activity of TRPM8(+) nerve endings and an increase in basal tearing. Our results suggest that unpleasant dryness sensations together with augmented tearing rate following corneal nerve injury are largely due to upregulation of TRPM8 in cold thermoreceptor neurons.

Pain perception is essential for survival and can be amplified or suppressed by expectations, experiences, and context. The neural mechanisms underlying bidirectional modulation of pain remain largely unknown. Here, we demonstrate that the central nucleus of the amygdala (CeA) functions as a pain rheostat, decreasing or increasing pain-related behaviors in mice. This dual and opposing function of the CeA is encoded by opposing changes in the excitability of two distinct subpopulations of GABAergic neurons that receive excitatory inputs from the parabrachial nucleus (PB). Thus, cells expressing protein kinase C-delta (CeA-PKCδ) are sensitized by nerve injury and increase pain-related responses. In contrast, cells expressing somatostatin (CeA-Som) are inhibited by nerve injury and their activity drives antinociception. Together, these results demonstrate that the CeA can amplify or suppress pain in a cell-type-specific manner, uncovering a previously unknown mechanism underlying bidirectional control of pain in the brain.