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Mechanisms resulting in abdominal pain include altered neuro-immune interactions in the gastrointestinal tract, but the signaling processes that link immune activation with visceral hypersensitivity are unresolved. We hypothesized that enteric glia link the neural and immune systems of the gut and that communication between enteric glia and immune cells modulates the development of visceral hypersensitivity. To this end, we manipulated a major mechanism of glial intercellular communication that requires connexin-43 and assessed the effects on acute and chronic inflammation, visceral hypersensitivity, and immune responses. Deleting connexin-43 in glia protected against the development of visceral hypersensitivity following chronic colitis. Mechanistically, the protective effects of glial manipulation were mediated by disrupting the glial-mediated activation of macrophages through the macrophage colony-stimulating factor. Collectively, our data identified enteric glia as a critical link between gastrointestinal neural and immune systems that could be harnessed by therapies to ameliorate abdominal pain.
Learn More >The effective prevention of postoperative cognitive dysfunction (POCD) needs to be explored, and the effect of preoperative pain on POCD remains unclear. We established a chronic pain model induced by chronic constriction injury (CCI) and models of acute pain and anxiety without pain in mice that were subsequently subjected to partial hepatectomy surgery. Morris water maze (MWM) tests were performed to evaluate the learning and memory abilities of the mice. ELISA was used to measure IL-1β, IL-6, and TNF-α in serum, and HPLC-MS was used to detect neurotransmitters in the prefrontal cortices and hippocampi of the mice. The results indicated that chronic pain induced by CCI might have significantly impaired the learning and memory abilities of mice, while acute pain and anxiety without pain only affected the memory abilities of mice. Perioperative acute pain increased the level of IL-1β in serum, and CCI might have increased the level of IL-6. CCI and acute pain increased dopamine (DA) levels in the cortex, similar to anxiety. Like anxiety, CCI increased 5-hydroxytryptamine (5-HT) levels in the prefrontal cortex and hippocampus. Acute pain led to a decrease in the acetylcholine (ACH) level in the hippocampus. Our results suggest that acute pain and CCI-induced chronic pain might aggravate postoperative cognitive dysfunction via neurotransmitters and by changing the levels of inflammatory factors such as IL-1β and IL-6.
Learn More >Nalfurafine, a moderately selective kappa opioid receptor (KOR) agonist, is used in Japan for treatment of itch without causing dysphoria or psychotomimesis. Here we characterized pharmacology of compound 42B, a 3-dehydroxy analog of nalfurafine and compared with that of nalfurafine. Nalfurafine and 42B acted as full KOR agonists and partial opioid receptor (MOR) agonists, but 42B showed much lower potency for both receptors and lower KOR/MOR selectivity, different from previous reports. Molecular modeling revealed that water-mediated hydrogen-bond formation between 3-OH of nalfurafine and KOR accounted for its higher KOR potency than 42B. The higher potency of both at KOR over MOR may be due to hydrogen-bond formation between non-conserved Y7.35 of KOR and their carbonyl groups. Both showed modest G protein signaling biases. In mice, like nalfurafine, 42B produced antinociceptive and anti-scratch effects and did not cause conditioned place aversion (CPA) in the effective dose ranges. Unlike nalfurafine, 42B caused motor incoordination and hypolocomotion. As both agonists showed G protein biases, yet produced different effects on locomotor activity and motor incoordination, the findings and those in the literature suggest caution in correlating in vitro biochemical data with in vivo behavior effects. The factors contributing to the disconnect, including pharmacodynamic and pharmacokinetic issues, are discussed. In addition, our results suggest that among the KOR-induced adverse behaviors, CPA can be separated from motor incoordination and hypolocomotion.
Learn More >Chronic pain is a major comorbidity of chronic inflammatory diseases. Here, we report that the cytokine IL-1β, which is abundantly produced during multiple sclerosis (MS), arthritis (RA), and osteoarthritis (OA) both in humans and in animal models, drives pain associated with these diseases. We found that the type 1 IL-1 receptor (IL-1R1) is highly expressed in the mouse and human by a subpopulation of TRPV1+ dorsal root ganglion neurons specialized in detecting painful stimuli, termed nociceptors. Strikingly, deletion of the Il1r1 gene specifically in TRPV1+ nociceptors prevented the development of mechanical allodynia without affecting clinical signs and disease progression in mice with experimental autoimmune encephalomyelitis and K/BxN serum transfer-induced RA. Conditional restoration of IL-1R1 expression in nociceptors of IL-1R1-knockout mice induced pain behavior but did not affect joint damage in monosodium iodoacetate-induced OA. Collectively, these data reveal that neuronal IL-1R1 signaling mediates pain, uncovering the potential benefit of anti-IL-1 therapies for pain management in patients with chronic inflammatory diseases.
Learn More >Although opioids have been used as treatment of neuropathic pain, opioids have abuse potential in humans. Since soluble epoxide hydrolase (sEH) in the metabolism of polyunsaturated fatty acids plays a key role in the pain, sEH inhibitors would be promising new therapeutic drugs for neuropathic pain. In this study, we examined the effect of the sEH inhibitor TPPU on rewarding effects in mice using the conditioned place preference (CPP) paradigm.
Learn More >Innocuous mechanical stimuli, such as rubbing or stroking the skin, relieve itch through the activation of low-threshold mechanoreceptors (LTMRs). However, the mechanisms behind this inhibition remain unknown. We presently investigated whether stroking the skin reduces the responses of superficial dorsal horn neurons to pruritogens in male C57BL/6J mice. Single-unit recordings revealed that neuronal responses to chloroquine were enhanced during skin stroking, and this was followed by suppression of firing below baseline levels after the termination of stroking. Most of these neurons additionally responded to capsaicin. Stroking did not suppress neuronal responses to capsaicin, indicating state-dependent inhibition. Vesicular glutamate transporter 3 (VGLUT3)-lineage sensory nerves compose a subset of LTMRs. Stroking-related inhibition of neuronal responses to chloroquine was diminished by optogenetic inhibition of VGLUT3-lineage sensory nerves in male and female / mice. Conversely, in male and female mice, optogenetic stimulation of VGLUT3-lineage sensory nerves inhibited firing responses of spinal neurons to pruritogens after the termination of stimulation. This inhibition was nearly abolished by spinal delivery of the κ-opioid receptor antagonist nor-binaltorphimine dihydrochloride, but not the neuropeptide Y receptor Y1 antagonist BMS193885. Optogenetic stimulation of VGLUT3-lineage sensory nerves inhibited pruritogen-evoked scratching without affecting mechanical and thermal pain behaviors. Therefore, VGLUT3-lineage sensory nerves appear to mediate inhibition of itch by tactile stimuli.Rubbing or stroking the skin is known to relieve itch. We investigated the mechanisms behind touch-evoked inhibition of itch in mice. Stroking the skin reduced the activity of itch-responsive spinal neurons. Optogenetic inhibition of VGLUT3-lineage sensory nerves diminished stroking-evoked inhibition, and optogenetic stimulation of VGLUT3-lineage nerves inhibited pruritogen-evoked firing. Together, our results provide a mechanistic understanding of touch-evoked inhibition of itch.
Learn More >Microglia can modulate spinal nociceptive transmission. Yet, their role in spinal cord stimulation (SCS)-induced pain inhibition is unclear. Here, we examined how SCS affects microglial activation in the lumbar cord of rats with chronic constriction injury (CCI) of the sciatic nerve. Male rats received conventional SCS (50 Hz, 80% motor threshold, 180 min, 2 sessions/day) or sham stimulation on days 18-20 post-CCI. SCS transiently attenuated the mechanical hypersensitivity in the ipsilateral hind paw and increased OX-42 immunoreactivity in the bilateral dorsal horns. SCS also upregulated the mRNAs of M1-like markers, but not M2-like markers. Inducible NOS protein expression was increased, but brain-derived neurotrophic factor was decreased after SCS. Intrathecal minocycline (1 μg-100 μg), which inhibits microglial activation, dose-dependently attenuated the mechanical hypersensitivity. Pretreatment with low-dose minocycline (1 μg, 30 min) prolonged the SCS-induced pain inhibition. These findings suggest that conventional SCS may paradoxically increase spinal M1-like microglial activity and thereby compromise its own ability to inhibit pain.
Learn More >Chronic low back pain (LBP) has high prevalence in the adult population which is associated with enormous disability. Hence, our aim was to further characterise our LBP rat model by using immunohistological and immunohistochemical methods at an advanced stage (day 49) of the model. Male Sprague-Dawley rats were anaesthetised and their lumbar L4/L5 and L5/L6 intervertebral discs (IVDs) were punctured (0.5 mm outer diameter, 2 mm-deep) 10 times per disc. Sham-rats underwent similar surgery, but no discs were punctured. For LBP- but not sham-rats, noxious pressure hyperalgesia was fully developed in the lumbar axial deep tissues on day 21 post-surgery, which was maintained until at least day 49. In the lumbar (L4-L6) dorsal root ganglia (DRGs), somatostatin (SRIF) and the somatostatin receptor type 4 (SST receptor) were co-localised with substance P and IB4, markers of small diameter unmyelinated peptidergic and non-peptidergic C-fibres respectively as well as with NF200, a marker of medium to large diameter neurons. On day 49, there was increased expression of SRIF but not the somatostatin receptor type 4 (SST receptor) in the lumbar DRGs and the spinal dorsal horns. There was increased DRG expression levels of the putative pro-nociceptive mediators: phosphorylated p38 (pp38) mitogen-activated protein kinase (MAPK) and phosphorylated p44/p42 MAPK (pp44/pp42 MAPK) as well as pp38 MAPK expression levels in the lumbar spinal cord. Taken together, the increased expression of SRIF in the lumbar DRGs and spinal cord and its co-localization with nociceptive fibres in DRG sections suggest a potential role of SRIF in modulating chronic mechanical LBP.
Learn More >Chronic pain often predicts the onset of psychological distress. Symptoms including anxiety and depression after pain chronification reportedly are caused by brain remodeling/recruitment of the limbic and reward/aversion circuitries. Pain is the primary precipitating factor that has caused opioid overprescribing and continued overuse of opioids leading to the current opioid epidemic. Yet experimental pain therapies often fail in clinical trials. Better understanding of underlying pathologies contributing to pain chronification is needed to address these chronic pain related issues. In the present study, a chronic neuropathic pain model persisting 10 weeks was studied. The model develops both anxiety- and pain-related behavioral measures to mimic clinical pain. The manganese-enhanced magnetic resonance imaging (MEMRI) utilized improved MRI signal contrast in brain regions with higher neuronal activity in the rodent chronic constriction trigeminal nerve injury (CCI-ION) model. T1-weighted MEMRI signal intensity was increased compared to controls in supraspinal regions of the anxiety and aversion circuitry, including anterior cingulate gyrus (ACC), amygdala, habenula, caudate, and dorsomedial periaqueductal gray (PAG). Despite continuing mechanical hypersensitivity, MEMRI T1 signal intensity as the neuronal activity measure, was not significantly different in thalamus and decreased in somatosensory cortex (S1BF) of CCI-ION rats compared to naïve controls. This is consistent with decreased fMRI BOLD signal intensity in thalamus and cortex of patients with longstanding trigeminal neuropathic pain reportedly associated with grey matter volume decrease in these regions. Significant increase in MEMRI T2 signal intensity in thalamus of CCI-ION animals was indication of tissue water content, cell dysfunction and/or reactive astrogliosis. Decreased T2 signal intensity in S1BF cortex of rats with CCI-ION was similar to findings of reduced T2 signals in clinical patients with chronic orofacial pain indicating prolonged astrocyte activation. These findings support use of MEMRI and chronic rodent models for preclinical studies and therapeutic trials to reveal brain sites activated only after neuropathic pain has persisted in timeframes relevant to clinical pain and to observe treatment effects not possible in short-term models which do not have evidence of anxiety-like behaviors. Potential improvement is predicted in the success rate of preclinical drug trials in future studies with this model.
Learn More >Anticonvulsant drugs are used to treat a wide range of non-epileptic conditions, including chronic, neuropathic pain. We obtained a phenoxyalkylaminoalkanol derivative, KM-416 which had previously demonstrated a significant anticonvulsant activity and had also been shown to bind to 5-HT, α-receptors and SERT and not to exhibit mutagenic properties. As KM-416 is a promising compound in our search for drug candidates, in the present study we further assessed its pharmacological profile (analgesic, local anesthetic, and antidepressant-like activities) accompanied with patch-clamp studies. Considering the importance of drug safety, its influence on the cardiovascular system was also evaluated. Moreover, KM-416 was subjected to forced degradation and pharmacokinetic studies to examine its stability and pharmacokinetic parameters. KM-416 revealed a significant antinociceptive activity in the tonic – the formalin test, neurogenic – the capsaicin test, and neuropathic pain model – streptozotocin-induced peripheral neuropathy. Moreover, it exerted a local anesthetic effect. In addition, KM-416 exhibited anti-depressant like activity. The results from the patch-clamp studies indicated that KM-416 can inhibit currents elicited by activation of NMDA receptors, while it also exhibited a voltage-dependent inhibition of Na currents. KM-416 did not influence ventricular depolarization and repolarization. Following oral administration, pharmacokinetics of KM-416 was characterized by a rapid absorption in the rat. The brain-to-plasma AUC ratio was 6.7, indicating that KM-416 was well distributed to brain. The forced degradation studies showed that KM-416 was very stable under stress conditions. All these features made KM-416 a promising drug candidate for further development against neuropathic pain and epilepsy.
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