The neuroprotective effects of GPR55 against hippocampal neuroinflammation and impaired adult neurogenesis in CSDS mice.

Depression is one of the most prevalent mental illnesses in the world today, and the onset of depression is usually accompanied by neuroinflammation and impaired adult neurogenesis. As a new potential member of the endocannabinoid (eCB) system, G protein coupled receptor 55 (GPR55) has been associated with mood regulation. However, the role of GPR55 in the pathophysiology of depression remains poorly understood. Thus, a 10-day chronic social defeat stress (CSDS) paradigm was utilized as an animal model of depression to explore the potential role of GPR55 in depression. After CSDS, the protein level of GPR55 decreased significantly, but the mRNA expression did not change significantly, highlighting that although the GPR55 protein was involved in the progression of the depression- and anxiety-like phenotypes, its mRNA was not. Additionally, depression- and anxiety-like behaviors were also accompanied by neuroinflammation and impaired adult neurogenesis in the hippocampus. Interestingly, O-1602, a GPR55 agonist, remarkably prevented the development of depression- and anxiety-like behaviors as well as hippocampal neuroinflammation and neurogenesis deficits induced by CSDS. However, after electroacupuncture (EA) alleviated depression- and anxiety-like behaviors induced by CSDS, treatment with a GPR55 antagonist (CID16020046) reversed this effect. Our research demonstrated that downregulation of GPR55 expression in the hippocampus might mediate CSDS-induced depression- and anxiety-like phenotypes, and activation and upregulation of GPR55, which might be correlated with its anti-inflammatory and subsequent neuroprotective effects, could be a potential treatment for depression.

Triggering receptor expressed on myeloid cells 2 (TREM2) dependent microglial activation promotes cisplatin-induced peripheral neuropathy in mice.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common adverse side effect of many antineoplastic agents. Patients treated with chemotherapy often report pain and paresthesias in a "glove-and-stocking" distribution. Diverse mechanisms contribute to the development and maintenance of CIPN. However, the role of spinal microglia in CIPN is not completely understood. In this study, cisplatin-treated mice displayed persistent mechanical allodynia, sensory deficits and decreased density of intraepidermal nerve fibers (IENFs). In the spinal cord, activation of microglia, but not astrocyte, was persistently observed until week five after the first cisplatin injection. Additionally, mRNA levels of inflammation related molecules including IL-1ß, IL-6, tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS) and CD16, were increased after cisplatin treatment. Intraperitoneal (i.p.) or intrathecal (i.t.) injection with minocycline both alleviated cisplatin-induced mechanical allodynia and sensory deficits, and prevented IENFs loss. Furthermore, cisplatin enhanced triggering receptor expressed on myeloid cells 2 (TREM2) /DNAX-activating protein of 12 kDa (DAP12) signaling in the spinal cord microglia. The blockage of TREM2 by i.t. injecting anti-TREM2 neutralizing antibody significantly attenuated cisplatin-induced mechanical allodynia, sensory deficits and IENFs loss. Meanwhile, anti-TREM2 neutralizing antibody prominently suppressed the spinal IL-6, TNF-α, iNOS and CD16 mRNA level, but it dramatically up-regulated the anti-inflammatory cytokines IL-4 and IL-10. The data demonstrated that cisplatin triggered persistent activation of spinal cord microglia through strengthening TREM2/DAP12 signaling, which further resulted in CIPN. Functional blockage of TREM2 or inhibition of microglia both benefited for cisplatin-induced peripheral neuropathy. Microglial TREM2/DAP12 may serve as a potential target for CIPN intervention.

Early single Aspirin-triggered Lipoxin blocked morphine anti-nociception tolerance through inhibiting NALP1 inflammasome: Involvement of PI3k/Akt signaling pathway.

Clinical usage of opioids in pain relief is dampened by analgesic tolerance after chronic exposure, which is related to opioid-associated neuroinflammation. In the current study, which is based on a chronic morphine tolerance rat model and sustained morphine treatment on primary neuron culture, it was observed that Akt phosphorylation, cleaved-Caspase-1-dependent NALP1 inflammasome activation and IL-1ß maturation in spinal cord neurons were significantly enhanced by morphine. Moreover, treatment with LY294002, a specific inhibitor of PI3k/Akt signaling, significantly reduced Caspase-1 cleavage, NALP1 inflammasome activation and attenuated morphine tolerance. Tail-flick tests demonstrated that pharmacological inhibition on Caspase-1 activation or antagonizing IL-1ß dramatically blocked the development of morphine tolerance. The administration of an exogenous analogue of lipoxin, Aspirin-triggered Lipoxin (ATL), caused a decline in Caspase-1 cleavage, inflammasome activation and mature IL-1ß production and thus attenuated the development of morphine tolerance by inhibiting upstream Akt phosphorylation. Additionally, treatment with DAMGO, a selective µ-opioid receptor peptide, significantly induced Akt phosphorylation, Caspase-1 cleavage and anti-nociception tolerance, all of which were attenuated by ATL treatment. Taken together, the present study revealed the involvement of spinal NALP1 inflammasome activation in the development of morphine tolerance and the role of the µ-receptor/PI3k-Akt signaling/NALP1 inflammasome cascade in this process. By inhibiting this signaling cascade, ATL blocked the development of morphine tolerance.

Cannabinoid type 2 receptors play a crucial role in social defeat-induced depression.

BACKGROUND: The endocannabinoid system plays a crucial role in regulating mood, but the specific involvement of cannabinoid receptor type 2 (CB2R) in depression remains poorly understood. Similarly, the mechanisms by which electroacupuncture (EA) provides therapeutic benefits for depression are not clearly defined. This research aims to explore the function of CB2R in depression and examine if the therapeutic effects of EA are associated with the hippocampal CB2R system. METHODS: Mice experiencing social defeat stress (SDS) were used to model depression and anxiety behaviors. We quantified hippocampal CB2R and N-arachidonoylethanolamide (AEA) levels. The efficacy of a CB2R agonist, JWH133, in mitigating SDS-induced behaviors was evaluated. Additionally, EA's impact on CB2R and AEA was assessed, along with the influence of CB2R antagonist AM630 on EA's antidepressant effects. RESULTS: SDS led to depressive and anxiety-like behaviors, with corresponding decreases in hippocampal CB2R and AEA. Treatment with JWH133 ameliorated these behaviors. EA treatment resulted in increased CB2R and AEA levels, while AM630 blocked these antidepressant effects. LIMITATIONS: The study mainly focused on the SDS model, which may not entirely reflect other depression models. Besides, further investigation is needed to understand the precise mechanisms by which CB2R and AEA contribute to EA's effects. CONCLUSIONS: The study suggests hippocampal downregulation of CB2R and AEA contributes to depression. Upregulation of CB2R and AEA in response to EA suggests their involvement in EA's antidepressant effects. These findings provide insights into the role of the hippocampal CB2R system in depression and the potential mechanisms underlying EA's therapeutic effects.

Lipoxins and aspirin-triggered lipoxin alleviate bone cancer pain in association with suppressing expression of spinal proinflammatory cytokines.

BACKGROUND: The neuroinflammatory responses in the spinal cord following bone cancer development have been shown to play an important role in cancer-induced bone pain (CIBP). Lipoxins (LXs), endogenous lipoxygenase-derived eicosanoids, represent a unique class of lipid mediators that possess a wide spectrum of anti-inflammatory and pro-resolving actions. In this study, we investigated the effects of intrathecal injection with lipoxin and related analogues on CIBP in rats. METHODS: The CIBP model was induced by intra-tibia inoculation of Walker 256 mammary gland carcinoma cells. Mechanical thresholds were determined by measuring the paw withdrawal threshold to probing with a series of calibrated von Frey filaments. Lipoxins and analogues were administered by intrathecal (i.t.) or intravenous (i.v.) injection. The protein level of LXA4 receptor (ALX) was tested by western blot. The localization of lipoxin receptor in spinal cord was assessed by fluorescent immunohistochemistry. Real-time PCR was carried out for detecting the expression of pro-inflammatory cytokines. RESULTS: Our results demonstrated that: 1) i.t. injection with the same dose (0.3 nmol) of lipoxin A4 (LXA4), lipoxin B4 (LXB4) or aspirin-triggered-15-epi-lipoxin A4 (ATL) could alleviate the mechanical allodynia in CIBP on day 7 after surgery. ATL showed a longer effect than the others and the effect lasted for 6 hours. ATL administered through i.v. injection could also attenuate the allodynia in cancer rats. 2) The results from western blot indicate that there is no difference in the expression of ALX among the naive, sham or cancer groups. 3) Immunohistochemistry showed that the lipoxin receptor (ALX)-like immunoreactive substance was distributed in the spinal cord, mainly co-localized with astrocytes, rarely co-localized with neurons, and never co-localized with microglia. 4) Real-time PCR analysis revealed that, compared with vehicle, i.t. injection with ATL could significantly attenuate the expression of the mRNA of proinflammatory cytokines (IL-1ß and TNF-α) in the spinal cord in CIBP. CONCLUSIONS: Taken together, the results of our study suggest that LXs and analogues exert strong analgesic effects on CIBP. These analgesic effects in CIBP are associated with suppressing the expression of spinal proinflammatory cytokines.

Involvement of spinal orexin A in the electroacupuncture analgesia in a rat model of post-laparotomy pain.

BACKGROUND: Orexin A (OXA, hypocretin/hcrt 1) is a newly discovered potential analgesic substance. However, whether OXA is involved in acupuncture analgesia remains unknown. The present study was designed to investigate the involvement of spinal OXA in electroacupuncture (EA) analgesia. METHODS: A modified rat model of post-laparotomy pain was adopted and evaluated. Von Frey filaments were used to measure mechanical allodynia of the hind paw and abdomen. EA at 2/15 Hz or 2/100 Hz was performed once on the bilateral ST36 and SP6 for 30 min perioperatively. SB-334867, a selective orexin 1 receptor (OX1R) antagonist with a higher affinity for OXA than OXB, was intrathecally injected to observe its effect on EA analgesia. RESULTS: OXA at 0.3 nmol and EA at 2/15 Hz produced respective analgesic effects on the model (P<0.05). Pre-surgical intrathecal administered of SB-334867 30 nmol antagonized OXA analgesia and attenuated the analgesic effect of EA (P<0.05). However, SB-334867 did not block fentanyl-induced analgesia (P>0.05). In addition, naloxone, a selective opioid receptor antagonist, failed to antagonize OXA-induced analgesia (P>0.05). CONCLUSIONS: The results of the present study indicate the involvement of OXA in EA analgesia via OX1R in an opioid-independent way.

Enhanced NMDA receptor NR1 phosphorylation and neuronal activity in the arcuate nucleus of hypothalamus following peripheral inflammation.

UNLABELLED: AbstractAim:To investigate the role of glutamate and N-methyl-D-aspartate (NMDA) receptors in central sensitization following peripheral inflammation in the arcuate nucleus (ARC) of the mediobasal hypothalamus. METHODS: Mediobasal hypothalamic slices were prepared from rats undergoing peripheral inflammation, which was induced by a unilateral injection of complete Freund's adjuvant (CFA) into hind paw. Neuronal activation levels in the ARC were monitored by recording extracellular unit discharges. The NMDA receptor NR1 subunit (NR1) was measured using Western blot analysis. RESULTS: Enhanced NR1 phosphorylation was observed in the ARC of CFA-inflamed rats. Compared with the control rats, the firing rate of spontaneous discharges in ARC neurons of inflamed rats was significantly higher, and it was significantly reduced both by an NMDA receptor antagonist (MK-801, 300 µmol/L) and by a non-NMDA receptor antagonist (CNQX, 30 µmol/L). Application of exogenous glutamate (200 µmol/L) or NMDA (25 µmol/L) resulted in increased neuronal discharges for ARC neurons, which was enhanced to a greater extent in inflamed rats than in control rats. CONCLUSION: Glutamate receptor activation in the hypothalamic ARC plays a crucial role in central sensitization associated with peripheral inflammation.

The antidepressant and anxiolytic effect of GPER on translocator protein (TSPO) via protein kinase a (PKA) signaling in menopausal female rats.

Postmenopausal depression is mainly caused by the deprivation of ovarian hormones during menopausal transition, it is of great importance to study on the treatment that could effectively relieve symptoms of menopausal depression with fewer side effects. Activation of G-protein-coupled estrogen receptor (GPER) has long been reported to facilitate neuronal plasticity and improve cognition in animals. Meanwhile, it could participate in regulation of intracellular signaling pathways through the characteristic of GPER, ameliorate intracellular mitochondrial function and oxidative stress. However, the impact of GPER on regulating estrogen deprived-depressant and anxious behaviors is still largely unknown. Here we used the ovariectomized female rats to imitate the condition of menopause. Owing to the lateral ventricle administration of G-1 which specifically react with GPER receptor intracerebrally, Ovariectomized (OVX) female rats showed depressive- or anxiety-like phenotypes with attenuated mitochondrial function. In addition, G-1 facilitated PKA activation, which further accelerated TSPO phosphorylation and alleviated menopausal depression- and anxiety-like behaviors. Moreover, PKA inhibitor PKI could partially antagonized the anti-anxiety and anti-depression effects of G-1. Taken together, we concluded that GPER activation might exhibit antidepressant and anxiolytic effect by elevating TSPO phosphorylation via protein kinase A signaling and rescuing the redox status in menopausal female rats.

[Electroacupuncture promotes regeneration and repair of myelin sheath of corpus callosum in demyelination mice].

OBJECTIVE: To explore the mechanism of electroacupuncture (EA) in accelerating the aggregation of microglia and promoting the remyelination at the location of demyelination. METHODS: C57BL/6 mice were randomly divided into 4 groups: normal, control, model (LPC) and LPC＋EA. The demyelination model was established by microinjection of Lysolecithin (LPC, 1 µL) into the left corpus callosum. EA (2 Hz/15 Hz, 2－4 mA) was applied to "Baihui"(GV20)and "Zhiyang"(GV9)for 30 min，once daily for 3 days, then, once every other day for 18 days. Immuno-fluorescence staining was used to observe the expression of myelin basic protein (MBP) and Axl tyrosine kinase receptor (Axl), Iba1 and numbers of Olig2-positive oligodendrocytes in the corpus callosum. Western blot was employed to detect the expression of MBP in the corpus callosum, and Oil Red O staining was used to observe changes of number of myelin pieces. RESULTS: Following modeling, the expression levels of MBP on day 5 and 10 after modeling were significantly decreased (P<0.05, P<0.01), Iba1 expression and Olig2-positive oligodendrocyte numbers on day 10 apparently increased (P<0.001, P<0.01). On day 21 after modeling, the levels of the above mentioned indexes returned to normal. After EA intervention, the levels of MBP expression on day 5 and 10, Axl, Iba1 protein expression and Olig2-positive oligodendrocyte numbers on day 5 were markedly increased (P<0.001，P<0.01，P<0.05), while Iba1 expression on day 10 was considerably decreased in comparison with the model group (P<0.01)．Oil Red O staining showed that on day 5 after modeling, the number of red lipid droplets were obviously increased in the corpus callosum tissue on the injection side, and apparently reduced in the EA group, suggesting a clearance of the accumulated myelin fragments by EA. CONCLUSION: EA intervention may reduce myelin debris and promote the aggregation of microglial cells and oligodendrocytes to the injured site, accelerate the myelin regeneration and up-regulate the expression of MBP and Axl of corpus callosum in demyelination mice.

miR-124/VAMP3 is a novel therapeutic target for mitigation of surgical trauma-induced microglial activation.

Activation of microglia and the subsequently elevated inflammatory cytokine release in the brain during surgery predispose individuals to cognitive dysfunction, also known as postoperative cognitive dysfunction (POCD). miR-124 is one of the most abundant microRNAs in the brain that regulates microglial function. Elucidating the role of miR-124 in microglial activation in the context of surgery may therefore promote understanding of as well as therapeutic development for post-surgical disorders involving microglial activation. The downstream targets of miR-124 were investigated using bioinformatic screening and dual-luciferase reporter assay validation, and vesicle-associated membrane protein 3 (VAMP3) was identified as a potential target. The kinetics of miR-124/VAMP3 expression was first examined in vitro in microglial cells (primary microglia and BV2 microglial cells) following lipopolysaccharide (LPS) stimulation. LPS induced a time-dependent decrease of miR-124 and upregulated the expression of VAMP3. Manipulating miR-124/VAMP3 expression by using miR-124 mimics or VAMP3-specific siRNA in LPS-stimulated BV2 microglial cells inhibited BV2 microglial activation-associated inflammatory cytokine release. To further examine the role of miR-124/VAMP3 in a surgical setting, we employed a rat surgical trauma model. Significant microglial activation and altered miR-124/VAMP3 expression were observed following surgical trauma. We also altered miR-124/VAMP3 expression in the rat surgical trauma model by administration of exogenous miR-124 and by using electroacupuncture, which is a clinically applicable treatment that modulates microglial function and minimizes postoperative disorders. We determined that electroacupuncture treatment specifically increases the expression of miR-124 in the hypothalamus and hippocampus. Increased miR-124 expression with a concomitant decrease in VAMP3 expression resulted in decreased inflammatory cytokine release related to microglial activation post-surgery. Our study indicates that miR-124/VAMP3 is involved in surgery-induced microglial activation and that targeting miR-124/VAMP3 could be a potential therapeutic strategy for postoperative disorders involving microglial activation.

Prevention and Treatment for Chemotherapy-Induced Peripheral Neuropathy: Therapies Based on CIPN Mechanisms.

BACKGROUND: Chemotherapy-induced peripheral neuropathy (CIPN) is a progressive, enduring, and often irreversible adverse effect of many antineoplastic agents, among which sensory abnormities are common and the most suffering issues. The pathogenesis of CIPN has not been completely understood, and strategies for CIPN prevention and treatment are still open problems for medicine. OBJECTIVES: The objective of this paper is to review the mechanism-based therapies against sensory abnormities in CIPN. METHODS: This is a literature review to describe the uncovered mechanisms underlying CIPN and to provide a summary of mechanism-based therapies for CIPN based on the evidence from both animal and clinical studies. RESULTS: An abundance of compounds has been developed to prevent or treat CIPN by blocking ion channels, targeting inflammatory cytokines and combating oxidative stress. Agents such as glutathione, mangafodipir and duloxetine are expected to be effective for CIPN intervention, while Ca/Mg infusion and venlafaxine, tricyclic antidepressants, and gabapentin display limited efficacy for preventing and alleviating CIPN. And the utilization of erythropoietin, menthol and amifostine needs to be cautious regarding to their side effects. CONCLUSIONS: Multiple drugs have been used and studied for decades, their effect against CIPN are still controversial according to different antineoplastic agents due to the diverse manifestations among different antineoplastic agents and complex drug-drug interactions. In addition, novel therapies or drugs that have proven to be effective in animals require further investigation, and it will take time to confirm their efficacy and safety.

The protective effects of Ghrelin/GHSR on hippocampal neurogenesis in CUMS mice.

Ghrelin is an orexigenic hormone that also plays an important role in mood disorders. Our previous studies demonstrated that ghrelin administration could protect against depression-like behaviors of chronic unpredictable mild stress (CUMS) in rodents. However, the mechanism related to the effect of ghrelin on CUMS mice has yet to be revealed. This article shows that ghrelin (5 nmol/kg/day for 2 weeks, i.p.) decreased depression-like behaviors induced by CUMS and increased hippocampal integrity (neurogenesis and spine density) measured via Ki67, 5-bromo-2-deoxyuridine (BrdU), doublecortin (DCX) labeling and Golgi-cox staining, which were decreased under CUMS. The behavioral phenotypes of Growth hormone secretagogue receptor (Ghsr)-null and wild type (WT) mice were evaluated under no stress condition and after CUMS exposure to determine the effect of Ghsr knockout on the behavioral phenotypes and stress susceptibility of mice. Ghsr-null mice exhibited depression-like behaviors under no stress condition. CUMS induced similar depression- and anxiety-like behavioral manifestations in both Ghsr-null and WT mice. A similar pattern of behavioral changes was observed after hippocampal GHSR knockdown. Additionally, both Ghsr knockout as well as CUMS exhibited deleterious effects on neurogenesis and spine density in the dentate gyrus (DG). Besides, CCK8 assay and 5-Ethynyl-2'-deoxyuridine (EdU) incorporation assay showed that ghrelin has a proliferative effect on primary cultured hippocampal neural stem cells (NSCs) and this proliferation was blocked by D-Lys3-GHRP-6 (DLS, the antagonist of GHSR, 100 µM) pretreatment. Ghrelin-induced proliferation is associated with the inhibition of G1 arrest, and this inhibition was blocked by LY294002 (specific inhibitor of PI3K, 20 µM). Furthermore, the in vivo data displayed that LY294002 (50 nmol, i.c.v.) can significantly block the antidepressant-like action of exogenous ghrelin treatment. All these results suggest that ghrelin/GHSR signaling maintains the integrity of hippocampus and has an inherent neuroprotective effect whether facing stress or not.

Ghrelin exhibited antidepressant and anxiolytic effect via the p38-MAPK signaling pathway in hippocampus.

Ghrelin, a peptide derived from stomach, is an endogenous ligand for growth hormone secretagogue receptor (GHSR). So far, the exact role of ghrelin in depression and anxiety is still being debated. The p38 mitogen-activated protein kinase (p38-MAPK) is known to be activated in response to various stress stimuli. Thus, we hypothesize that ghrelin has an antidepressant effect, to which the p38-MAPK signaling pathway significantly contributes. To test this hypothesis, chronic social defeat stress (CSDS) was used as a model of depression. We employed the adeno-associated virus-mediated siRNA approach to down-regulate GHSR expression in the hippocampus of mice in vivo. Both ghrelin and the p38 inhibitor, SB203580, were administered to identify the effect of ghrelin on depressive-like behavior of stressed mice and to better assess the role of the p38-MAPK signaling pathway in this process. We found that CSDS activated the endogenous ghrelin-GHSR in hippocampal neurons, which possibly resulted in opposing the formation of depression- and anxiety-like behaviors in mice. Furthermore, the p38-MAPK signaling pathway had an important role in the antidepressant effect of ghrelin. Therefore, we conclude that ghrelin may reduce CSDS-induced depression- and anxiety-like behaviors via inhibiting the p38-MAPK signaling pathway in hippocampal neurons of mice.

Involvement of serotonin 2A receptors in the analgesic effect of tramadol in mono-arthritic rats.

The analgesic effects of tramadol are considered to be mediated by both the opioid system and the serotonergic system. This study investigated the involvement of a subtype of serotonin receptors, 5-hydroxytryptamine (5-HT)2A receptor, in the analgesic effect of tramadol. The intraperitoneal (i.p.) injection of tramadol reduced the paw withdrawal latency (PWL) to radiant heat testing in mono-arthritic rats. The antagonistic effect of i.p. ketanserin (a 5-HT2A receptor antagonist) on tramadol analgesia was observed. The expression of the 5-HT2A receptor mRNA in the nucleus of raphe magnus (NRM), ventrolateral periaqueductal gray (vlPAG) and spinal dorsal horn of mono-arthritic rats after a ten-day treatment with tramadol was measured with in situ hybridization. Either single injections or 10 days of tramadol treatment dose-dependently elevated PWL of arthritic rats while ketanserin could partially antagonize the tramadol analgesic effect. Expression of the 5-HT2A receptor mRNA in NRM, ipsilateral vlPAG, and the ipsilateral spinal dorsal horn of arthritic rats was significantly increased after tramadol treatment. These results suggest that 5-HT2A receptors are involved in the analgesic effect of tramadol. This study provides evidence for involvement of 5-HT2A receptors in the tramadol analgesia of inflammatory pain. The increase in this receptor mRNA in the chronic study may contribute to the sustaining effect of tramadol long-term treatments in clinical practice.

[Effects of intracerebroventricular injection of delta-opioid receptor agonist TAN-67 or antagonist naltrindole on acute cerebral ischemia in rat].

This work was performed to determine the role of delta-opioid receptor (DOR) in protection against acute ischemia/reperfusion injury. Transient (1 h) focal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO). DOR agonist TAN-67 (30 nmol, 60 nmol, 200 nmol), DOR antagonist naltrindole (20 nmol, 50 nmol, 100 nmol) or artificial cerebral spinal fluid (aCSF) was injected respectively into the lateral cerebroventricle of the rat 30 min before the induction of brain ischemia. Neurological deficits were assessed by the five-grade system (Longa's methods). The brain infarct was measured by cresyl violet (CV) staining and infarct volume was analyzed by an image processing and analysis system. The expression of DOR was detected by Western blot. The results showed that 60 nmol TAN-67 significantly reduced the infarct volume (P<0.05), attenuated neurological deficits (P<0.05) and tended to increase the expression of about 60 kDa DOR protein (P>0.05), while 100 nmol naltrindole aggravated ischemic damage and decreased about 60 kDa DOR protein expression (P<0.05). These results suggest that DOR activation protects the brain against acute ischemia/reperfusion injury in rat.

Electro-Acupuncture Alleviates Chronic Unpredictable Stress-Induced Depressive- and Anxiety-Like Behavior and Hippocampal Neuroinflammation in Rat Model of Depression.

Depression is the second leading cause of disability worldwide. The effects of clinical depression may be mediated by neuroinflammation such as activation of microglia and high levels of proinflammatory cytokines in certain brain areas. Traditional Chinese medicine techniques such as electro-acupuncture (EA) are used extensively in Asia to treat mental health disorders. However, EA has not been rigorously studied in treatment of depression. This study was designed to assess the effectiveness of EA on depressive-like behavior and explore the role of hippocampal neuroinflammation in the potential antidepressant effect of EA. In this study, we used six chronic unpredictable stressors daily in a random sequence for 10 weeks. EA were performed on "Bai-Hui" (Du-20) (+) and "Yang-Ling-Quan" (GB-34, the right side; -) acupoints by an EA apparatus (HANS Electronic Apparatus, LH202H, 2/100 Hz, 0.3 mA) for 30 min once every other day for last 4 weeks. The behavior tests including open field test and forced swimming test, which are widely used to assess depressive and anxiety-like behavior were performed on the Monday and Tuesday of the eleventh week. The results showed that 10 week of chronic unpredictable stress (CUS) caused behavioral deficits in rats and neuroinflammation in hippocampus, such as increased expression of NLRP3 inflammasome components, upregulated mRNA level of IL-1ß and the protein level of IL-1ß mature form (p17) and activation of microglia. Moreover, 4 weeks of EA treatment significantly attenuated behavioral deficits caused by CUS. EA's antidepressant effect was accompanied by markedly decreased expression of certain NLRP3 inflammasome components and matured IL-1ß. Meanwhile, EA treatment can significantly reverse CUS-induced increases in P2X7 receptor, Iba-1, IL-18, TNFα and IL-6 expression and decreases in GFAP expression. In conclusion, EA exhibited the antidepressant effect and alleviated the hippocampal neuroinflammation. These findings may provide insight into the role of hippocampal neuroinflammation in the antidepressant effect of EA.

Changes in expression of nociceptin/orphanin FQ and its receptor in spinal dorsal horn during electroacupuncture treatment for peripheral inflammatory pain in rats.

The neuropeptide nociceptin/orphanin FQ (N/OFQ), the endogenous agonist of the N/OFQ peptide receptor (NOP receptor), has been demonstrated to be involved in many physiological and pathological functions including pain modulation. It was reported that electroacupuncture (EA) had a potent analgesic effect on inflammatory pain by activating various endogenous transmitters such as the opioid peptides. In the present study, we investigated the effect of EA on peripheral inflammatory pain and the expression of N/OFQ and the NOP receptor in the spinal dorsal horn of rats, using a behavioral test, RT-PCR, immunohistochemistry and Western blot analysis techniques. The results showed: (1) EA had an accumulative analgesic effect on chronic inflammatory pain; (2) in the superficial layers of the spinal dorsal horn, the level of mRNA of the precursor protein for N/OFQ (preproN/OFQ, ppN/OFQ) was increased and the N/OFQ immunoreactivity was decreased after peripheral inflammation, and could be significantly increased by EA treatment; (3) both mRNA and protein levels of the NOP receptor in the spinal dorsal horn were significantly increased after chronic inflammatory pain and could be further enhanced by EA treatment. The present data demonstrated that EA could activate the endogenous N/OFQ-NOP receptor system, and this might underlie the effectiveness of EA in the treatment of inflammatory pain.

Lipopolysaccharide-induced protein kinase D activation mediated by interleukin-1beta and protein kinase C.

Protein kinase D (PKD), a newly described serine/threonine kinase, has been implicated in many signal transduction pathways. The present study was designed to determine whether and how PKD is activated in inflammation. The results demonstrated that lipopolysaccharide (LPS, 30 microg/ml) stimulated PKD and protein kinase C (PKC) phosphorylation in spinal neurons within 0.5 h, and the activation reached a maximum at 3 or 8 h and declined at 12 h. The phosphorylation could be inhibited by the selective inhibitors for PKC (100 nM), mainly for PKCalpha and PKCbeta, suggesting the involvement of the PKC pathway. Particularly, PKCalpha might be critical for LPS-induced PKD activation since the PKCbeta inhibitor (100 nM) observed no effect on the phosphorylation of PKD. Furthermore, the expression of interleukin-1beta (IL-1beta) was significantly induced by LPS within 0.5 h, and reached a maximum at 8 h. IL-1 receptor antagonist inhibited PKD and PKCs activation induced by LPS at a concentration of 50 nM and achieved maximum at 1000 nM. These results demonstrated for the first time that PKD could be activated by LPS in spinal neurons, might via the IL-1beta/PKCalpha pathway. Additionally, immunostaining showed an increase in number of phosphorylated PKD-immunoreactive cells of adult spinal dorsal horn induced by intraplantar injected carrageenan (2 microg/100 microl), and antisense oligodeoxynucleotide to IL-1 receptor type I (50 microg/10 microl, intrathecal injected) inhibited the PKD activation, suggesting an involvement of IL-1beta/PKD pathway in inflammation in adult spinal cord.

Sympathetic nervous system mediates surgical trauma stress-induced splenocyte apoptosis in rats.

Surgical trauma stress has been reported to induce immunosuppression. The mechanisms involved are still unclear. The present study was designed to assess the role of the sympathetic nervous system in regulating splenocyte apoptosis induced by surgical trauma stress. Our results showed that the rats that underwent surgical trauma stress exhibited a significant reduction in splenic cellularity, the loss of splenocytes was likely mediated by apoptosis, for a substantial increase in apoptosis was observed by using DNA gel electrophoresis and TUNEL assay. At the same time, an increase in Fas(CD95/Apo-1) protein expression in splenocytes was also observed. These effects were significantly abolished by either chemical sympathectomy or beta-adrenergic receptor antagonist propranolol. The data clearly revealed that the sympathetic nervous system especially beta-adrenergic receptors was involved in surgical trauma-induced immune alterations via a mechanism of apoptotic cell death.

Electroacupuncture combined with clomipramine enhances antidepressant effect in rodents.

The present study was designed to evaluate the antidepressant effect of electroacupuncture (EA) and the potential additive or synergistic effects of EA and clomipramine (CLO, a tricyclic antidepressant) in the mouse forced swimming test (FST) and chronic mild stress (CMS) induced depression-model rats. The FST is an antidepressant screening procedure performed initially to observe the immediate effects of EA and/or CLO on the immobility time. CLO (2.5, 5, 10, 20 and 60mg/kg intraperitoneally) were administered at 23, 6 and 1h respectively prior to each test. EA was given at the 'Bai-Hui' (Du 20) and unilateral 'An-Mian' (EX 17) acupoints 1h before each test. Immobility time was significantly reduced by EA and CLO at 2.5, 5, 10, 20 or 60mg/kg, respectively. EA combined with 2.5mg/kg CLO exhibited additive effects on the immobility time. In addition, rats were exposed chronically (1st-11th week) to a variety of mild unpredictable stressors. Depressed mood and anhedonia were recognized as a decrease in sucrose intake in the CMS rats. CLO at 2.5, 5mg/kg and EA at the same acupoints and parameters were administrated on the CMS rats once every other day for 6 weeks (5th-11th week). The intake of 1% sucrose solution was reduced by CMS, which was restored to normal level after 6 weeks treatment with 5mg/kg CLO or EA combined with 2.5mg/kg CLO. However, neither the sucrose intake nor the sucrose preference in the depressive rats was significantly changed by the treatment with EA or 2.5mg/kg CLO alone. These results demonstrated that EA combined with CLO at low doses has an additive or synergistic antidepressant action, and this combination may provide an effective strategy for depression management.