Role of ERK in gender difference of fibromyalgia pain.

This study investigated the ERK pathway of the peripheral nervous system and discovered a gender-specific pattern of ERK activation in the dorsal root ganglion of an acid-induced chronic widespread muscular pain model. We employed a twice acid-induced chronic musculoskeletal pain model in rats to evaluate mechanical pain behavior in both male and female groups. We further conducted protein analysis of dissected dorsal root ganglions from both genders. Both male and female rats exhibited a similar pain behavior trend, with females demonstrating a lower pain threshold. Protein analysis of the dorsal root ganglion (DRG) showed a significant increase in phosphorylated ERK after the second acid injection in all groups. However, phosphorylation of ERK was observed in the dorsal root ganglion, with higher levels in the male ipsilateral group compared to the female group. Moreover, there was a no difference between the left and right sides in males, whereas the significant difference was observed in females. In conclusions, the administration of acid injections induced painful behavior in rats, and concurrent with this, a significant upregulation of pERK was observed in the dorsal root ganglia, with a greater magnitude of increase in males than females, and in the contralateral side compared to the ipsilateral side. Our findings shed light on the peripheral mechanisms underlying chronic pain disorders and offer potential avenues for therapeutic intervention.

Acetyltransferase Enok regulates transposon silencing and piRNA cluster transcription.

The piRNA pathway is a highly conserved mechanism to repress transposon activation in the germline in Drosophila and mammals. This pathway starts from transcribing piRNA clusters to generate long piRNA precursors. The majority of piRNA clusters lack conventional promoters, and utilize heterochromatin- and HP1D/Rhino-dependent noncanonical mechanisms for transcription. However, information regarding the transcriptional regulation of piRNA clusters is limited. Here, we report that the Drosophila acetyltransferase Enok, which can activate transcription by acetylating H3K23, is critical for piRNA production from 54% of piRNA clusters including 42AB, the major piRNA source. Surprisingly, we found that Enok not only promotes rhino expression by acetylating H3K23, but also directly enhances transcription of piRNA clusters by facilitating Rhino recruitment. Taken together, our study provides novel insights into the regulation of noncanonical transcription at piRNA clusters and transposon silencing.

Behavioral Survey of Effects of Pulsed Radiofrequency on Neuropathic and Nociceptive Pain in Rats: Treatment Profile and Device Implantation.

OBJECTIVES: Pulsed radiofrequency (PRF) stimulation is widely used for intractable pain; however, there is no consensus on treatment protocols and appropriate types of pain. We compared effectiveness of bipolar and unipolar PRF on neuropathic or inflammatory pains, and of targets at the dorsal root ganglion (DRG) and sciatic nerve (SN). We also examined efficacy of repetitive PRF stimulations. This preclinical study could serve as an extensive survey before human trials. MATERIALS: Spare nerve injury (SNI)-induced neuropathic pain and complete Freund's adjuvant (CFA) injection-induced inflammatory pain were used. Behavioral responses were measured using von Frey test, acetone test, and Hargreave's test at preinjury and postinjury time points. In both models, we evaluated results of DRG stimulation with unipolar PRF (45 V) versus bipolar PRF (5 V), stimulation at DRG vs. SN, and repetitive stimulations. RESULTS: Both unipolar and bipolar PRFs reduced SNI- or CFA-induced pain for a similar duration. In the SNI model, PRF-DRG had a stronger effect on tactile pain than PRF-SN but lower effect on cold allodynia, whereas in the CFA model PRF-DRG and PRF-SN showed similar effects. Repetitive PRF stimulation, by open technique or implantation method, produced analogous effect by each stimulus, and no evident analgesic tolerance or neurological deficit was shown. CONCLUSIONS: PRF temporarily attenuates neuropathic and inflammatory pain. Bipolar PRF generates significant analgesia with a much lower electrical power than unipolar PRF. Meanwhile, the minor variant effects between PRF-DRG and PRF-SN may indicate distinct mechanisms. The sustained-analgesia by repetitive treatments suggests implantation technique could be a promising choice.

Laser acupuncture-induced analgesic effect and molecular alterations in an incision pain model: a comparison with electroacupuncture-induced effects.

Low-level laser acupuncture (LLLA) produces photobiomodulation through acupuncture point and is an alternative to low-level laser therapy. Although the analgesic effect of LLLA on chronic pain has been proven, its effect on acute postincisional pain has yet to be investigated. A plantar incision (PI) model was used to mimic human postsurgical pain. Male adult rats received GaAlAs laser irradiation at the right ST36 acupoint immediately after operation and on the following 4 days. Three laser treatment groups (two red laser groups with a 30- or 15-min treatment duration and one 30-min near-infrared laser group) were compared with sham LLLA and naive groups and an electroacupuncture (EA) group (separate study). Behavioral withdrawal thresholds of both hind paws were measured before and after incision. Expression of mitogen-activated protein kinases (p-ERK and p-p38), inducible nitric oxide synthase (iNOS), and tumor necrosis factor (TNF) in the spinal cord was analyzed. All three LLLA treatments attenuated post-PI tactile allodynia in the ipsilateral paw, but only the 30-min red laser treatment affected the contralateral paw and had similar efficacy to that of EA. All laser treatments barely reduced heat hyperalgesia in both hind paws. At 3 days after PI, the 30-min red laser group showed reversed increases of PI-induced p-ERK, p-p38, and iNOS but not TNF expression in the spinal cord. Repetitive LLLA treatments ameliorated PI-induced mechanical pain. The inhibition of multiple sensitization signals highlights the unique clinical role of LLLA. Thus, LLLA is an alternative to EA as an adjuvant for postoperative pain control.

Hair follicles modulate skin barrier function.

Our skin provides a protective barrier that shields us from our environment. Barrier function is typically associated with interfollicular epidermis; however, whether hair follicles influence this process remains unclear. Here, we utilize a potent genetic tool to probe barrier function by conditionally ablating a quintessential epidermal barrier gene, Abca12, which is mutated in the most severe skin barrier disease, harlequin ichthyosis. With this tool, we deduced 4 ways by which hair follicles modulate skin barrier function. First, the upper hair follicle (uHF) forms a functioning barrier. Second, barrier disruption in the uHF elicits non-cell autonomous responses in the epidermis. Third, deleting Abca12 in the uHF impairs desquamation and blocks sebum release. Finally, barrier perturbation causes uHF cells to move into the epidermis. Neutralizing Il17a, whose expression is enriched in the uHF, partially alleviated some disease phenotypes. Altogether, our findings implicate hair follicles as multi-faceted regulators of skin barrier function.

Hair follicles modulate skin barrier function.

Our skin provides a protective barrier that shields us from our environment. Barrier function is typically associated with the interfollicular epidermis; however, whether hair follicles influence this process remains unclear. Here, we utilize a potent genetic tool to probe barrier function by conditionally ablating a quintessential epidermal barrier gene, Abca12, which is mutated in the most severe skin barrier disease, harlequin ichthyosis. With this tool, we deduced 4 ways by which hair follicles modulate skin barrier function. First, the upper hair follicle (uHF) forms a functioning barrier. Second, barrier disruption in the uHF elicits non-cell-autonomous responses in the epidermis. Third, deleting Abca12 in the uHF impairs desquamation and blocks sebum release. Finally, barrier perturbation causes uHF cells to move into the epidermis. Neutralizing IL-17a, whose expression is enriched in the uHF, partially alleviated some disease phenotypes. Altogether, our findings implicate hair follicles as multi-faceted regulators of skin barrier function.

Distinct mechanisms for sebaceous gland self-renewal and regeneration provide durability in response to injury.

Sebaceous glands (SGs) release oils that protect our skin, but how these glands respond to injury has not been previously examined. Here, we report that SGs are largely self-renewed by dedicated stem cell pools during homeostasis. Using targeted single cell RNA-sequencing, we uncovered both direct and indirect paths by which these resident SG progenitors ordinarily differentiate into sebocytes, including transit through a PPARγ+Krt5+ transitional cell state. Upon skin injury, however, SG progenitors depart their niche, reepithelialize the wound, and are replaced by hair follicle-derived stem cells. Furthermore, following targeted genetic ablation of >99% of SGs from dorsal skin, these glands unexpectedly regenerate within weeks. This regenerative process is mediated by alternative stem cells originating from the hair follicle bulge, is dependent upon FGFR signaling, and can be accelerated by inducing hair growth. Altogether, our studies demonstrate that stem cell plasticity promotes SG durability following injury.

Distinct mechanisms for sebaceous gland self-renewal and regeneration provide durability in response to injury.

Sebaceous glands (SGs) release oils that protect our skin, but how these glands respond to injury has not been previously examined. Here, we report that SGs are largely self-renewed by dedicated stem cell pools during homeostasis. Using targeted single-cell RNA sequencing, we uncovered both direct and indirect paths by which resident SG progenitors ordinarily differentiate into sebocytes, including transit through a Krt5+PPARγ+ transitional basal cell state. Upon skin injury, however, SG progenitors depart their niche, reepithelialize the wound, and are replaced by hair-follicle-derived stem cells. Furthermore, following targeted genetic ablation of >99% of SGs from dorsal skin, these glands unexpectedly regenerate within weeks. This regenerative process is mediated by alternative stem cells originating from the hair follicle bulge, is dependent upon FGFR2 signaling, and can be accelerated by inducing hair growth. Altogether, our studies demonstrate that stem cell plasticity promotes SG durability following injury.

Effects of far infrared rays irradiated from ceramic material (BIOCERAMIC) on psychological stress-conditioned elevated heart rate, blood pressure, and oxidative stress-suppressed cardiac contractility.

The present study examined the effects of BIOCERAMIC on psychological stress-conditioned elevated heart rate, blood pressure and oxidative stress-suppressed cardiac contractility using in vivo and in vitro animal models. We investigated the effects of BIOCERAMIC on the in vivo cardiovascular hemodynamic parameters of rats by monitoring their heart rates, systolic blood pressure, mean blood pressure and diastolic blood pressure. Thereafter, we assayed its effects on the heart rate in an isolated frog heart with and without adrenaline stimulation, and on cardiac contractility under oxidative stress. BIOCERAMIC caused significant decreases in heart rates and systolic and mean blood pressure in the stress-conditioned heart rate rat models (P < 0.05), as well as in the experimental models of an isolated frog heart with and without adrenaline stimulation (P < 0.05), and normalized cardiac contractility under oxidative stress (P < 0.05). BIOCERAMIC may, therefore, normalize the effects of psychological stress and oxidative stress conditions.

Basal cell carcinomas acquire secondary mutations to overcome dormancy and progress from microscopic to macroscopic disease.

Basal cell carcinomas (BCCs) frequently possess immense mutational burdens; however, the functional significance of most of these mutations remains unclear. Here, we report that loss of Ptch1, the most common mutation that activates upstream Hedgehog (Hh) signaling, initiates the formation of nascent BCC-like tumors that eventually enter into a dormant state. However, rare tumors that overcome dormancy acquire the ability to hyperactivate downstream Hh signaling through a variety of mechanisms, including amplification of Gli1/2 and upregulation of Mycn. Furthermore, we demonstrate that MYCN overexpression promotes the progression of tumors induced by loss of Ptch1. These findings suggest that canonical mutations that activate upstream Hh signaling are necessary, but not sufficient, for BCC to fully progress. Rather, tumors likely acquire secondary mutations that further hyperactivate downstream Hh signaling in order to escape dormancy and enter a trajectory of uncontrolled expansion.

Temporal changes in glutamate, glutamate transporters, basilar arteries wall thickness, and neuronal variability in an experimental rat model of subarachnoid hemorrhage.

BACKGROUND: Glutamate and glutamate transporters (GTs) (including glutamate/aspartate transporter, glutamate transporter-1, and excitatory amino acid carrier 1) have important roles in the pathogenesis of ischemic neurological injury. The changes in glutamate, GTs, and neuronal injury after subarachnoid hemorrhage (SAH) have not been widely investigated. In this study, we examined the changes in extracellular glutamate concentration, GTs, wall thickness of basilar arteries (BAs), and neuronal degeneration in experimental SAH rats. METHODS: An intrathecal microdialysis probe was inserted into male Sprague Dawley rats. SAH was induced using a double-hemorrhage model. To measure glutamate concentrations, extracellular dialysates were collected for 30 minutes before, and daily for 7 days after SAH. Changes in neurological scores, body weight, and BA wall thickness were measured. The neuron degeneration in the hippocampus and the changes of GTs in the cerebral cortex and hippocampus were measured. RESULTS: Glutamate concentrations were significantly higher in SAH rats from day (D)1 to D7 after SAH compared with the sham rats, especially at D1. A significant body weight reduction and neurological defects were observed at D3 after SAH. The walls of BAs in SAH rats were significantly thicker compared with those of sham rats; the maximum change was observed at D7. Hippocampal neuronal degeneration was observed after SAH and the highest severity was at D7. The expression of GTs was downregulated after SAH and persisted for 7 days. CONCLUSIONS: SAH induced in the double-hemorrhage rat model may produce an excessive and prolonged increase of extracellular glutamate concentrations and downregulation of GTs, which are accompanied by BA wall thickness, and hippocampal neuronal degeneration.

Aryl-hydrocarbon receptor-dependent alteration of FAK/RhoA in the inhibition of HUVEC motility by 3-methylcholanthrene.

We previously demonstrated the antiproliferative and antiangiogenic effects of 3-methylcholanthrene (3MC), an aryl-hydrocarbon receptor (AhR) agonist, in human umbilical vascular endothelial cells (HUVECs). Herein, we unraveled its molecular mechanisms in inhibiting HUVEC motility. 3MC down-regulated FAK, but up-regulated RhoA, which was rescued by AhR knockdown. It led us to identify novel AhR binding sites in the FAK/RhoA promoters. Additionally, 3MC increased RhoA activity via suppression of a negative feedback pathway of FAK/p190RhoGAP. With an increase in membrane-bound RhoA, subsequent stress fiber and focal adhesion complex formation was observed in 3MC-treated cells, and this was reversed by a RhoA inhibitor and AhR antagonists. Notably, these compounds significantly reversed 3MC-mediated anti-migration in a transwell assay. The in vitro findings were further confirmed using an animal model of Matrigel formation in Balb/c mice. Collectively, AhR's genomic regulation of FAK/RhoA, together with RhoA activation, is ascribable to the anti-migration effect of 3MC in HUVECs.

Ovarian Hormone-dependent and Spinal ERK Activation-regulated Nociceptive Hypersensitivity in Female Rats with Acid Injection-induced Chronic Widespread Muscle Pain.

Symptoms of chronic widespread muscle pain (CWP) meet most of the diagnostic criteria for fibromyalgia syndrome, which is prevalent in females. We used an acid injection-induced muscle pain (AIMP) model to mimic CWP. After female rats received an ovariectomy (OVX), acid saline solution was injected into the left gastrocnemius muscle. Time courses of changes in pain behaviours and p-ERK in the spinal cord were compared between groups. Intrathecal injections of oestradiol (E2) to the OVX group before two acid injections and E2 or progesterone (P4) injections in male rats were compared to evaluate hormone effects. We found that repeated acid injections produced mechanical hypersensitivity and enhanced p-ERK expression in the spinal dorsal horn. OVX rats exhibited significantly less tactile allodynia than did the rats in the other groups. The ERK inhibitor U0126 alleviated mechanical allodynia with lower p-ERK expression in the sham females but did not affect the OVX rats. Intrathecal E2 reversed the attenuated mechanical hypersensitivity in the OVX group, and E2 or P4 induced transient hyperalgesia in male rats. Accordingly, our results suggested that ovarian hormones contribute to AIMP through a spinal p-ERK-mediated pathway. These findings may partially explain the higher prevalence of fibromyalgia in females than males.

Methylprednisolone inhibits the expression of glial fibrillary acidic protein and chondroitin sulfate proteoglycans in reactivated astrocytes.

Reactive gliosis caused by post-traumatic injury often results in marked expression of chondroitin sulfate proteoglycan (CSPG), which inhibits neurite outgrowth and regeneration. Methylprednisolone (MP), a synthetic glucocorticoid, has been shown to have neuroprotective and anti-inflammatory effects for the treatment of acute spinal cord injury (SCI). However, the effect of MP on CSPG expression in reactive glial cells remains unclear. In our study, we induced astrocyte reactivation using alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and cyclothiazide to mimic the excitotoxic stimuli of SCI. The expression of glial fibrillary acidic protein (GFAP), a marker of astrocyte reactivation, and CSPG neurocan and phosphacan were significantly elevated by AMPA treatment. The conditioned media from AMPA-treated astrocytes strongly inhibited neurite outgrowth of rat dorsal root ganglion neurons, and this effect was reversed by pretreatment with MP. Furthermore, MP downregulated GFAP and CSPG expression in adult rats with SCI. Additionally, both the glucocorticoid receptor (GR) antagonist RU486 and GR siRNA reversed the inhibitory effects of MP on GFAP and neurocan expression. Taken together, these results suggest that MP may improve neuronal repair and promote neurite outgrowth after excitotoxic insult via GR-mediated downregulation of astrocyte reactivation and inhibition of CSPG expression.

Rapid and Delayed Effects of Pulsed Radiofrequency on Neuropathic Pain: Electrophysiological, Molecular, and Behavioral Evidence Supporting Long-Term Depression.

BACKGROUND: Pulsed radiofrequency (PRF) has been widely employed for ameliorating clinical neuropathic pain. How PRF alters electrophysiological transmission and modulates biomolecular functions in neural tissues has yet to be clarified. We previously demonstrated that an early application of low-voltage bipolar PRF adjacent to the dorsal root ganglion (DRG) reduced acute neuropathic pain in animals. By contrast, the present study investigated how PRF alters postsynaptic sensitization to produce early and delayed effects on neuropathic pain. OBJECTIVES: Our objective was to test the hypothesis that a 5-minute session of PRF could rapidly produce selective long-term depression (LTD) on C-fiber-mediated spinal sensitization and sustain the effect through the long-lasting inhibition of injury-induced ERK-MAPK activation. This may explain the prolonged analgesic effect of PRF on chronic neuropathic pain. STUDY DESIGN: Experiments were conducted on both normal rats and neuropathic pain rats that received spinal nerve ligation (SNL) 8 days prior. SETTING: An animal laboratory in a medical center of a university in Taiwan. METHODS: We first compared changes in field potentials in the L5 superficial spinal dorsal horn (SDH) that were evoked by conditioning electrical stimuli in the sciatic nerve in male adult rats before (as the baseline) and after PRF stimulation for at least 2 hours. Bipolar PRF was applied adjacent to the L5 DRG at an intensity of 5 V for 5 minutes, whereas the control rats were treated with sham applications. The electrophysiological findings were tested for any correlation with induction of spinal phospho-ERK (p-ERK) in normal and neuropathic pain rats. We then investigated the delayed effect of PRF on SNL-maintained pain behaviors for 2 weeks as well as p-ERK in SDH among the control, SNL, and PRF groups. Finally, potential injury in the DRGs after PRF stimulation was evaluated through behavioral observations and ATF-3, a neuronal stress marker. RESULTS: In the evoked field-potential study, the recordings mediated through A- and C-afferent fibers were identified as A-component and C-component, respectively. PRF significantly reduced the C-components over 2 hours in both the normal and SNL rats, but it did not affect the A-components. In the SNL rats, the C-component was significantly depressed in the PRF group compared with the sham group. PRF also inhibited acute p-ERK induced by mechanical nociception in both the control and SNL rats. For a longer period, PRF ameliorated SNL-maintained mechanical allodynia for 10 days and thermal analgesia for 14 days, and it significantly reduced late ERK activation within spinal neurons and astrocytes 14 days afterward. Moreover, PRF in the normal rats did not alter basal withdrawal thresholds or increase the expression and distribution of ATF-3 in the DRGs. LIMITATIONS: Several issues should be considered before translating the animal results to clinical applications. CONCLUSIONS: Low-voltage bipolar PRF produces LTD through selective suppression on the C-component, but not on the A-component. It also inhibits ERK activation within neurons and astrocytes in SDHs. The findings suggest that PRF alleviates long-lasting neuropathic pain by selectively and persistently modulating C-fiber-mediated spinal nociceptive hypersensitivity.Key words: Pulsed radiofrequency (PRF), dorsal root ganglion (DRG), neuropathic pain, ERK activation, evoked field potential, ATF-3, long-term depression (LTD), spinal nerve ligation (SNL).

GAGA factor, a positive regulator of global gene expression, modulates transcriptional pausing and organization of upstream nucleosomes.

BACKGROUND: Genome-wide studies in higher eukaryotes have revealed the presence of paused RNA polymerase II (RNA-Pol) at about 30-50 bp downstream of the transcription start site of genes involved in developmental control, cell proliferation and intercellular signaling. Promoter-proximal pausing is believed to represent a critical step in transcriptional regulation. GAGA sequence motifs have frequently been found in the upstream region of paused genes in Drosophila, implicating a prevalent binding factor, GAF, in transcriptional pausing. RESULTS: Using newly isolated mutants that retain only ~3 % normal GAF level, we analyzed its impacts on transcriptional regulation in whole animals. We first examined the abundance of three major isoforms of RNA-Pol on Hsp70 during heat shock. By cytogenetic analyses on polytene chromosomes and chromatin immunoprecipitation (ChIP), we show that paused RNA-Pol of Hsp70 is substantially reduced in mutants. Conversely, a global increase in paused RNA-Pol is observed when GAF is over-expressed. Coupled analyses of transcriptome and GAF genomic distribution show that 269 genes enriched for upstream GAF binding are down-regulated in mutants. Interestingly, ~15 % of them encode transcriptional factors, which might control ~2000 additional genes down-regulated in mutants. Further examination of RNA-Pol distribution in GAF targets reveals that a positive correlation exists between promoter-proximal RNA-Pol density and GAF occupancy in WT, but not in mutants. Comparison of nucleosome profiles indicates that nucleosome occupancy is preferentially attenuated by GAF in the upstream region that strongly favors nucleosome assembly. Using a dominant eye phenotype caused by GAF over-expression, we detect significant genetic interactions between GAF and the nucleosome remodeler NURF, the pausing factor NELF, and BAB1 whose binding sites are enriched specifically in genes displaying GAF-dependent pausing. CONCLUSION: Our results provide direct evidence to support a critical role of GAF in global gene expression, transcriptional pausing and upstream nucleosome organization of a group of genes. By cooperating with factors acting at different levels, GAF orchestrates a series of events from local nucleosome displacement to paused transcription. The use of whole animals containing broad tissue types attests the physiological relevance of this regulatory network.

Bcl-2 gene family expression in the brain of rat offspring after gestational and lactational dioxin exposure.

Recent epidemiological studies have shown that dioxin, a persistent organic pollutant, is related to cognitive and behavioral abnormalities in the offspring of exposed cohort. In order to investigate the possible impact of dioxin in survival gene expression during brain development, we established an animal model of gestational and lactational dioxin-exposed rat offspring. The expressions of dioxin-responsive gene cytochrome P450 1A1 (CYP1A1), apoptotic gene Bax, and anti-apoptotic genes Bcl-2 and Bcl-xL were examined in rat liver and brains using Western blot analysis and RT-PCR. The results showed that treatment of pregnant rats with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (2 microg/kg body weight through oral delivery) at gestation day 15 resulted in an increase of Bcl-xL in offspring male liver and cerebral cortex, but a decrease in female offspring. In contrast, the expression of Bcl-xL in the cerebellum was decreased in male, but increased in female. Bcl-2, another anti-apoptotic gene, was also downregulated in P0 female liver, cerebral cortex, but was not observed in male. In the 4-month-old offspring, however, the Bcl-2 protein levels in the liver and cerebellum of both male and female pups were higher in the TCDD group as compared with the control group. However, the Bcl-2 level in the cerebral cortex of TCDD-treated groups was higher than the control group only in female but not male offspring at 4 months old. The expression of Bax showed no significant changes upon TCDD exposure at P0 stage, but was significantly reduced in the 4-month-old male cortex. These results indicate that early exposure of dioxin could affect the development of certain brain regions with gender difference, in terms of its differential effect on expressions of Bcl-xL, Bcl-2, and Bax.

Spinal p38 activity and analgesic effect after low- and high-intensity electroacupuncture stimulation in a plantar incision rat model.

AIMS: Postoperative pain is a major problem. Electroacupuncture (EA) has been accepted as a useful and low-risk complementary therapy for post-operative pain. Animal studies indicate that surgical incision activates p38 MAPK in the spinal microglia, which critically contributes to post-incisional nociceptive development. How EA affects incision-induced p38 activation is important but yet to be fully elucidated. METHODS: Male adult rats received plantar incision (PI) at the right hind paw followed by 30-min EA of 4-Hz, one of two intensities (3 and 10mA), and at right ST36 (Zusanli) acupoint immediately after PI and for 3 successive days. EA analgesia was evaluated by von Frey fibers and Hargreaves' tests. Spinal p38 activation was examined by immunostaining. In separate groups, SB203580, a p38 inhibitor, was intrathecally injected alone or with EA to test the combining effect on nociception and spinal phospho-p38. KEY FINDINGS: EA of 10-mA significantly ameliorated mechanical allodynia, but 3-mA did not. None of them altered thermal hyperalgesia. Repeated EA could not inhibit phospho-p38 in the PI rats, contrarily, EA per se significantly induced phospho-p38 in the normal rats. Intrathecal SB203580 injection dose-dependently prevented PI-induced allodynia. Combination of low-dose SB203580 and 3-mA EA, which were ineffective individually, profoundly reduce post-PI allodynia. SIGNIFICANCE: We demonstrated that 10-mA EA exerts a significant inhibition against post-PI mechanical hypersensitivity via a p38-independent pathway. Importantly, co-treatment with low-dose p38 inhibitor and 3-mA EA can counteract spinal phospho-p38 to exert strong analgesic effect. Our finding suggests a novel strategy to improve EA analgesic quality.

Coadministration of glycogen-synthase kinase 3 inhibitor with morphine attenuates chronic morphine-induced analgesic tolerance and withdrawal syndrome.

BACKGROUND: Glycogen-synthase kinase 3 (GSK3) is involved in many signaling pathways and is associated with a host of high-profile pathophysiological states. However, its role in morphine tolerance, especially naloxone-precipitated withdrawal syndrome, has not been well investigated. The present study was undertaken to study the role of GSK3 in chronic morphine exposure. METHODS: Adult male Sprague-Dawley rats were subjected to intraperitoneal (i.p.) injections of morphine (10 mg/kg) twice daily for 6 consecutive days, and tail-flick tests were conducted to evaluate changes of morphine-induced antinociception. GSK3 inhibitor, SB216763 or SB415286, was i.p. injected prior to morphine to investigate the influences on morphine tolerance. There were four groups receiving morphine plus vehicle (2% dimethyl sulfoxide), morphine plus SB216763 (0.6 mg/kg) or SB415286 (1.0 mg/kg), GSK3 inhibitor alone, or dimethyl sulfoxide: as the control group. On Day 7, naloxone (i.p., 1 mg/kg) was administered and naloxone-precipitated withdrawal behaviors were individually compared between groups. RESULTS: Repeated morphine exposure in this study led to progressive shortening of tail-flick latencies and produced six of nine observed naloxone-precipitated withdrawal behaviors. Coadministration with SB216763 or SB415286 significantly prevented antinociceptive tolerance and alleviated parts of withdrawal syndrome. Both inhibitors could similarly reverse withdrawal behaviors including grooming, chewing, and ptosis, but did not affect withdrawal behaviors of penis licking and defecation. CONCLUSION: The results demonstrate the importance of GSK3 in reducing chronic morphine-induced tolerance and withdrawal syndrome. Although GSK3 is involved in diverse physiological functions, aiming at GSK3-related pathway could still be a potential tool to improve therapeutic quality in clinical morphine treatment.

Comparison of electroacupuncture and morphine-mediated analgesic patterns in a plantar incision-induced pain model.

Electroacupuncture (EA) is a complementary therapy to improve morphine analgesia for postoperative pain, but underlying mechanism is not well-known. Herein, we investigated EA-induced analgesic effect in a plantar incision (PI) model in male Sprague-Dawley rats. PI was performed at the left hind paw. EA of 4 Hz and high intensity or sham needling was conducted at right ST36 prior to PI and repeated for another 2 days. Behavioral responses to mechanical and thermal stimuli, spinal phospho-ERK, and Fos expression were all analyzed. In additional groups, naloxone and morphine were administered to elucidate involvement of opioid receptors and for comparison with EA. EA pretreatment significantly reduced post-PI tactile allodynia for over 1 day; repeated treatments maintained analgesic effect. Intraperitoneal naloxone could reverse EA analgesia. Low-dose subcutaneous morphine (1 mg/kg) had stronger inhibitory effect on PI-induced allodynia than EA for 1 h. However, analgesic tolerance appeared after repeated morphine injections. Both EA and morphine could equally inhibit PI-induced p-ERK and Fos inductions. We conclude that though EA and morphine attenuate postincision pain through opioid receptor activations, daily EA treatments result in analgesic accumulation whereas daily morphine injections develop analgesic tolerance. Discrepant pathways and mechanisms underlying two analgesic means may account for the results.