A peripheral cannabinoid mechanism suppresses spinal fos protein expression and pain behavior in a rat model of inflammation

The present studies were conducted to test the hypothesis that systemically inactive doses of cannabinoids suppress inflammation-evoked neuronal activity in vivo via a peripheral mechanism. We examined peripheral cannabinoid modulation of spinal Fos protein expression, a marker of neuronal activity, in a rat model of inflammation. Rats received unilateral intraplantar injections of carrageenan (3%). In behavioral studies, carrageenan induced allodynia and mechanical hyperalgesia in response to stimulation with von Frey monofilaments. The cannabinoid agonist WIN55,212-2 (30 microg intraplantarly), administered concurrently with carrageenan, attenuated carrageenan-evoked allodynia and hyperalgesia relative to control conditions. In immunocytochemical studies, WIN55,212-2 suppressed the development of carrageenan-evoked Fos protein expression in the lumbar dorsal horn of the spinal cord relative to vehicle treatment. The same dose administered systemically or to the noninflamed contralateral paw failed to alter either carrageenan-evoked allodynia and hyperalgesia or carrageenan-evoked Fos protein expression, consistent with a peripheral site of action. The suppressive effects of WIN55,212-2 (30 microg intraplantarly) on carrageenan-evoked Fos protein expression and pain behavior were blocked by local administration of either the CB(2) antagonist SR144528 (30 microg intraplantarly) or the CB(1) antagonist SR141716A (100 microg intraplantarly). WIN55,212-3, the enantiomer of the active compound, also failed to suppress carrageenan-evoked Fos protein expression. These data provide direct evidence that a peripheral cannabinoid mechanism suppresses the development of inflammation-evoked neuronal activity at the level of the spinal dorsal horn and implicate a role for CB(2) and CB(1) in peripheral cannabinoid modulation of inflammatory nociception.     

Activation of cannabinoid CB2 receptors suppresses C-fiber responses and windup in spinal wide dynamic range neurons in the absence and presence of inflammation

Effects of the CB2-selective cannabinoid agonist AM1241 on activity evoked in spinal wide dynamic range (WDR) neurons by transcutaneous electrical stimulation were evaluated in urethane-anesthetized rats. Recordings were obtained in both the absence and the presence of carrageenan inflammation. AM1241, administered intravenously or locally in the paw, suppressed activity evoked by transcutaneous electrical stimulation during the development of inflammation. Decreases in WDR responses resulted from a suppression of C-fiber-mediated activity and windup. Abeta- and Adelta-fiber-mediated responses were not reliably altered. The AM1241-induced suppression of electrically evoked responses was blocked by the CB2 antagonist SR144528 but not by the CB1 antagonist SR141716A. AM1241 (33 microg/kg intraplantar [i.p.l.]), administered to the carrageenan-injected paw, suppressed activity evoked in WDR neurons relative to groups receiving vehicle in the same paw or AM1241 in the opposite (noninflamed) paw. The electrophysiological effects of AM1241 (330 microg/kg intravenous [i.v.]) were greater in rats receiving i.p.l. carrageenan compared with noninflamed rats receiving an i.p.l. injection of vehicle. AM1241 failed to alter the activity of purely nonnociceptive neurons recorded in the lumbar dorsal horn. Additionally, AM1241 (330 microg/kg i.v. and i.p.l.; 33 microg/kg i.p.l.) reduced the diameter of the carrageenan-injected paw. The AM1241-induced decrease in peripheral edema was blocked by the CB2 but not by the CB1 antagonist. These data demonstrate that activation of cannabinoid CB2 receptors is sufficient to suppress neuronal activity at central levels of processing in the spinal dorsal horn. Our findings are consistent with the ability of AM1241 to normalize nociceptive thresholds and produce antinociception in inflammatory pain states.     

Selective cannabinoid CB1 receptor activation inhibits spinal nociceptive transmission in vivo.

Cannabinoid1 (CB1) receptors are located at CNS sites, including the spinal cord, involved in somatosensory processing. Analgesia is one of the tetrad of behaviors associated with cannabinoid agonists. Here, effects of a potent cannabinoid CB1 receptor agonist arachidonyl-2-chloroethylamide (ACEA) on evoked responses of dorsal horn neurons in anesthetized rats were investigated. Extracellular recordings of convergent dorsal horn neurons were made in halothane anesthetized Sprague-Dawley rats (n = 16). Effects of spinal application of ACEA on electrically evoked responses of dorsal horn neurons were studied. Mean maximal effects of 0.5, 5, 50, and 500 ng/50 microl ACEA on the C-fiber-mediated postdischarge response were 79 +/- 6, 62 +/- 10, and 54 +/- 7% (P < 0.01), 45 +/- 6% (P < 0.01), of control, respectively. ACEA (500 ng/50 microl) also reduced the C-fiber-evoked nonpotentiated responses of neurons (59 +/- 9% of control, P < 0.05) and Adelta-fiber-evoked responses of neurons (68 +/- 10% of control, P < 0.01). Minor effects of ACEA on Abeta-fiber-evoked responses were observed. Spinal pre-administration of the selective CB1 receptor antagonist SR141716A (0.01 microg/50 microl) significantly reduced effects of ACEA (500 ng/50 microl) on postdischarge responses of dorsal horn neurons. This study demonstrates that spinal CB1 receptors modulate the transmission of C- and Adelta-fiber-evoked responses in anesthetized rats; this may reflect pre- and/or postsynaptic effects of cannabinoids on nociceptive transmission. CB1 receptors inhibit synaptic release of glutamate in rat dorsolateral striatum, a similar mechanism of action may underlie the effects of ACEA on noxious evoked responses of spinal neurons reported here.     

大麻素CB_1受体活化对神经病理性疼痛大鼠脊髓背角嘌呤能P2X_2受体表达的抑制作用

目的:观察脊髓背角大麻素CB_1受体(CB_1R)在坐骨神经缩窄性损伤(CCI)所致的神经病理性疼痛中的作用及其对嘌呤能P2X_2受体表达的调节。方法:7~8周龄SD大鼠分为4组:(1)sham组;(2)CCI组;(3)CP55940+CCI组;(4)AM251+CP55940+CCI组。分别于CCI术前1 d,术后1、3、5、7、10、14 d测定热缩足反射潜伏期(TWL);免疫印迹技术检测各组大鼠损伤侧L_4~L_6段脊髓背角P2X_2受体表达。结果:CCI术后大鼠出现热痛敏,TWL明显缩短;鞘内给予非选择性大麻素受体激动剂CP55940可明显延长CCI大鼠TWL(P0.05);预先鞘内注射CB_1R拮抗剂AM251(0.05 mg/kg)可显著降低CP55940的镇痛效果(P0.05)。免疫印迹实验结果显示:CCI大鼠脊髓背角P2X_2受体在术后7、14 d表达明显增加(P0.05);鞘内给予CP55940可显著降低P2X_2受体表达(P0.05),而预先给予AM251可降低CP55940抑制P2X_2受体表达的效应(P0.05)。结论:脊髓背角CB_1受体激活对CCI所致的神经病理性疼痛具有良好的镇痛作用,其镇痛效应可能与抑制CCI大鼠嘌呤能P2X_2受体表达有关。     

Beneficial effects of cannabinoids (CB) in a murine model of allergen-induced airway inflammation: role of CB1/CB2 receptors

The endocannabinoid system (ECS) consists of two cannabinoid (CB) receptors, namely CB₁ and CB₂ receptor, and their endogenous (endocannabinoids) and exogenous (cannabinoids, e.g. delta-9-tetrahydrocannabinol (THC)) ligands which bind to these receptors. Based on studies suggesting a role of THC and the ECS in inflammation, the objective of this study was to examine their involvement in type I hypersensitivity using a murine model of allergic airway inflammation. THC treatment of C57BL/6 wildtype mice dramatically reduced airway inflammation as determined by significantly reduced total cell counts in bronchoalveolar lavage (BAL). These effects were greatest when mice were treated during both, the sensitization and the challenge phase. Furthermore, systemic immune responses were significantly suppressed in mice which received THC during sensitization phase. To investigate a role of CB_1/2 receptors in this setting, we used pharmacological blockade of CB₁ and/or CB₂ receptors by the selective antagonists and moreover CB₁/CB₂ receptor double-knockout mice (CB₁^−/−/CB₂^−/−) and found neither significant changes in the cell patterns in BAL nor in immunoglobulin levels as compared to wildtype mice. Our results indicate that the activation of the ECS by applying the agonist THC is involved in the development of type I allergies. However, CB₁/CB₂ receptor-independent signalling seems likely in the observed results.     

Compartment-specific localization of cannabinoid 1 (CB1) and mu-opioid receptors in rat nucleus accumbens

Interactions between cannabinoid and opioid systems have been implicated in reward and drug seeking behaviors involving neuronal circuitry in the nucleus accumbens (Acb) shell and core. To determine the relevant sites, we examined the electron microscopic localization of cannabinoid type-1 (CB1) receptors and mu-opioid receptors in each Acb compartment in rat brain. CB1 receptor immunogold labeling was seen on the plasma membrane and within the cytoplasm of neuronal and glial profiles throughout the Acb. These neuronal profiles included somata and dendrites as well as axon terminals, many of which formed excitatory-type, asymmetric synapses with notable perforations that are often associated with synaptic plasticity. The number of CB1-labeled terminals within the neuropil of the Acb shell was significantly greater than in the core. Mu-opioid receptors were also detected in axonal and dendritic profiles. These dendrites were most prevalent in the Acb shell, where mu-receptors also were located in 21% of the dendritic profiles and 3% of the axon terminals containing CB1 receptors. More of the CB1-labeled terminals contacted dendrites expressing mu-opioid receptors in the shell (19%) compared with the core (13%). Conversely, of the synaptic mu-labeled terminals, 20% in the shell and 10% in the core contacted dendrites containing CB1 receptors. These findings provide ultrastructural evidence that cannabinoid-opioid interactions are mediated by activation of CB1 and mu-opioid receptors within the same or synaptically linked neurons in the Acb shell and core. They also suggest a particularly important role for presynaptic CB1 receptors in the reward circuit of the Acb shell.     

Functional expression of cell surface cannabinoid CB(1) receptors on presynaptic inhibitory terminals in cultured rat hippocampal neurons

At present, little is known about the mechanisms by which cannabinoids exert their effects on the central nervous system. In this study, fluorescence imaging and electrophysiological techniques were used to investigate the functional relationship between cell surface cannabinoid type 1 (CB(1)) receptors and GABAergic synaptic transmission in cultured hippocampal neurons. CB(1) receptors were labelled on living neurons using a polyclonal antibody directed against the N-terminal 77 amino acid residues of the rat cloned CB(1) receptor. Highly punctate CB(1) receptor labelling was observed on fine axons and at axonal growth cones, with little somatic labelling. The majority of these sites were associated with synaptic terminals, identified either with immunohistochemical markers or by using the styryl dye FM1-43 to label synaptic vesicles that had undergone active turnover. Dual labelling of neurons for CB(1) receptors with either the inhibitory neurotransmitter GABA or its synthesising enzyme glutamate decarboxylase, demonstrated a strong correspondence. The immunocytochemical data was supported by functional studies using whole-cell patch-clamp recordings of miniature inhibitory postsynaptic currents (mIPSCs). The cannabinoid agonist WIN55,212-2 (100nM) markedly inhibited (by 77+/-6.3%) the frequency of pharmacologically-isolated GABAergic mIPSCs. The effects of WIN55,212-2 were blocked in the presence of the selective CB(1) receptor antagonist SR141716A (100nM).In conclusion, the present data show that cell surface CB(1) receptors are expressed at presynaptic GABAergic terminals, where their activation inhibits GABA release. Their presence on growth cones could indicate a role in the targeting of inhibitory connections during development.     

Electroacupuncture suppresses spinal expression of neurokinin-1 receptors induced by persistent inflammation in rats

It has been demonstrated that electroacupuncture (EA) significantly suppresses behavioral hyperalgesia in a rat model of persistent inflammatory pain and that neurokinin-1 (NK-1)/substance P (SP) receptors play important roles in nociception and hyperalgesia at the spinal cord level. The present study investigated spinal NK-1 receptor involvement in EA-produced suppression of hyperalgesia in a rat model of persistent inflammatory pain. The results showed that hind paw inflammation induced a significant increase of NK-1 receptor expression in the spinal dorsal horn and that this effect was significantly suppressed by EA. This suggests that EA-induced suppression of hyperalgesia is involved, at least partly, in the suppression of the spinal NK-1 receptors induced by sustained peripheral nociceptive input.     

Glial expression of cannabinoid CB(2) receptors and fatty acid amide hydrolase are beta amyloid-linked events in Down's syndrome

Recent data suggest that the endocannabinoid system (ECS) may be involved in the glial response in different types of brain injury. Both acute and chronic insults seem to trigger a shift in the pattern of expression of some elements of this system from neuronal to glial. Specifically, data obtained in human brain tissue sections from Alzheimer's disease patients showed that the expression of cannabinoid receptors of the CB(2) type is induced in activated microglial cells while fatty acid amide hydrolase (FAAH) expression is increased in reactive astrocytes. The present study was designed to determine the time-course of the shift from neuronal to glial induction in the expression of these proteins in Down's syndrome, sometimes referred to as a human model of Alzheimer-like beta-amyloid (Abeta) deposition. Here we present immunohistochemical evidence that both CB(2) receptors and FAAH enzyme are induced in Abeta plaque-associated microglia and astroglia, respectively, in Down's syndrome. These results suggest that the induction of these elements of the ECS contributes to, or is a result of, amyloid deposition and subsequent plaque formation. In addition, they confirm a striking differential pattern of distribution of FAAH and CB(2) receptors.     

Modulation of the balance between cannabinoid CB(1) and CB(2) receptor activation during cerebral ischemic/reperfusion injury

Cannabinoid receptor activation has been shown to modulate both neurotransmission (CB(1)) and neuroinflammatory (CB(2)) responses. There are conflicting reports in the literature describing the influence of cannabinoid receptor activation on ischemic/reperfusion injury. The goal of this study was to evaluate how changing the balance between CB(1) and CB(2) activation following cerebral ischemia influences outcome. CB(1) and CB(2) expression were tested at different times after transient middle cerebral artery occlusion (MCAO) in mice by real-time RT-PCR. Animals subjected to 1 h MCAO were randomly assigned to receive different treatments: a CB(1) antagonist, a CB(2) antagonist, a CB(2) agonist, a CB(1) antagonist plus CB(2) agonist, a CB(2) antagonist plus CB(2) agonist or an equal volume of vehicle as control. Cerebral blood flow was continuously monitored during ischemia; cerebral infarction and neurological deficit were tested 24 h after MCAO. Cerebral CB(1) and CB(2) mRNA expression undertook dynamic changes during cerebral ischemia. The selective CB(1) antagonist significantly decreased cerebral infarction by 47%; the selective CB(2) antagonist increased infarction by 26% after 1 h MCAO followed by 23 h reperfusion in mice. The most striking changes were obtained by combining a CB(1) antagonist with a CB(2) agonist. This combination elevated the cerebral blood flow during ischemia and reduced infarction by 75%. In conclusion, during cerebral ischemia/reperfusion injury, inhibition of CB(1) receptor activation is protective while inhibition of CB(2) receptor activation is detrimental. The greatest degree of neuroprotection was obtained by combining an inhibitor of CB(1) activation with an exogenous CB(2) agonist.     

Stimulation of cannabinoid receptor 2 (CB2) suppresses microglial activation

Background  

Activated microglial cells have been implicated in a number of neurodegenerative disorders, including Alzheimer's disease (AD), multiple sclerosis (MS), and HIV dementia. It is well known that inflammatory mediators such as nitric oxide (NO), cytokines, and chemokines play an important role in microglial cell-associated neuron cell damage. Our previous studies have shown that CD40 signaling is involved in pathological activation of microglial cells. Many data reveal that cannabinoids mediate suppression of inflammation in vitro and in vivo through stimulation of cannabinoid receptor 2 (CB₂).     

WIN 55212-2 impairs contextual fear conditioning through the activation of CB1 cannabinoid receptors

The memory deficits induced by cannabinoid agonists have been found in several behavioral paradigms. Nevertheless, there is evidence that not all types of memory are impaired after cannabinoid administration. The aim of this study was to investigate whether the cannabinoid agonist WIN 55212-2 (WIN) is able to influence the acquisition of fear conditioning using tone and contextual versions. For tone-fear conditioning, male Wistar rats were placed in the conditioning chamber and after 3 min, a sound (CS) was presented for 10s that terminated with a 1-s electric footshock (1.5 mA). For contextual-fear conditioning, a similar procedure was used but no sound was presented. Twenty-four hours after, the animals were re-exposed to the respective CS (tone or conditioning chamber) and the freezing behavior was registered. A subsequent experiment investigated a possible state-dependent effect of WIN by administering WIN or control solution 30 min before conditioning and before testing. WIN (2.5 and 5.0 mg/kg) administered i.p. 30 min before impaired contextual fear conditioning but did not modify the freezing behavior elicited by tone presentation. These animals did not show any state-dependent effects of WIN. Further, the impaired contextual conditioning was prevented by preadministration of SR141716A (1.0 mg/kg, i.p.) or SR147778 (1.0 mg/kg, i.p.), selective cannabinoid CB1 receptor antagonists. The present findings highlight that cannabinoid agonists effects are selective for the hippocampus-dependent aversive memories in rats. This effect appears to be related to the activation of CB1 cannabinoid receptors and confirms that cannabinoids might provide a novel approach for the treatment of unpleasant memories.     

Effects of activation of group III metabotropic glutamate receptors on spinal synaptic transmission in a rat model of neuropathic pain

Chronic neuropathic pain remains an unmet clinical problem because it is often resistant to conventional analgesics. Metabotropic glutamate receptors (mGluRs) are involved in nociceptive processing at the spinal level, but their functions in neuropathic pain are not fully known. In this study, we investigated the role of group III mGluRs in the control of spinal excitatory and inhibitory synaptic transmission in a rat model of neuropathic pain induced by L5/L6 spinal nerve ligation. Whole-cell recording of lamina II neurons was performed in spinal cord slices from control and nerve-ligated rats. The baseline amplitude of glutamatergic EPSCs evoked from primary afferents was significantly larger in nerve-injured rats than in control rats. However, the baseline frequency of GABAergic and glycinergic inhibitory postsynaptic currents (IPSCs) was much lower in nerve-injured rats than in control rats. The group III mGluR agonist l(+)-2-amino-4-phosphonbutyric acid (l-AP4) produced a greater inhibition of the amplitude of monosynaptic and polysynaptic evoked EPSCs in nerve-injured rats than in control rats. l-AP4 inhibited the frequency of miniature EPSCs in 66.7% of neurons in control rats but its inhibitory effect was observed in all neurons tested in nerve-injured rats. Furthermore, l-AP4 similarly inhibited the frequency of GABAergic and glycinergic IPSCs in control and nerve-injured rats. Our study suggests that spinal nerve injury augments glutamatergic input from primary afferents but decreases GABAergic and glycinergic input to spinal dorsal horn neurons. Activation of group III mGluRs attenuates glutamatergic input from primary afferents in nerve-injured rats, which could explain the antinociceptive effect of group III mGluR agonists on neuropathic pain.     

CB1 and CB2 cannabinoid receptor expression during development and in epileptogenic developmental pathologies

Recent data support the involvement of the endocannabinoid signaling in early brain development, as well as a key role of cannabinoid receptors (CBR) in pathological conditions associated with unbalanced neuronal excitability and inflammation. Using immunocytochemistry, we explored the expression and cellular pattern of CBR 1 and 2 (CB1 and CB2) during prenatal human cortical development, as well as in focal malformations of cortical development associated with intractable epilepsy (focal cortical dysplasia; cortical tubers in patients with the tuberous sclerosis complex and glioneuronal tumors). Strong CB1 immunoreactivity was detected in the cortical plate in developing human brain from the earliest stages tested (gestational week 9) and it persisted throughout prenatal development. Both cannabinoid receptors were not detected in neural progenitor cells located in the ventricular zone. Only CB1 was expressed in the subventricular zone and in Cajal–Retzius cells in the molecular zone of the developing neocortex. CB2 was detected in cells of the microglia/macrophage lineage during development. In malformations of cortical development, prominent CB1 expression was demonstrated in dysplastic neurons. Both CBR were detected in balloon/giant cells, but CB2 appeared to be more frequently expressed than CB1 in these cell types. Reactive astrocytes were mainly stained with CB1, whereas cells of the microglia/macrophage lineage were stained with CB2. These findings confirm the early expression pattern of cannabinoid receptors in the developing human brain, suggesting a function for CB1 in the early stages of corticogenesis. The expression patterns in malformations of cortical development highlight the role of cannabinoid receptors as mediators of the endocannabinoid signaling and as potential pharmacological targets to modulate neuronal and glial cell function in epileptogenic developmental pathologies.     

Overexpression of cannabinoid receptors CB1 and CB2 correlates with improved prognosis of patients with hepatocellular carcinoma

CB1 and CB2 are multifunctional cannabinoid-specific receptors considered to be involved in inhibition of tumor development. To elucidate their roles in hepatocarcinogenesis, we analyzed the expression of these receptors in tumor and matched nontumorous tissues of human hepatocellular carcinoma (HCC) samples. In situ hybridization analysis showed overexpression of CB1 mRNAs in 8 of 13 (62%) HCC samples, and of CB2 mRNAs in 7 of 13 (54%). Immunohistochemical analysis of 64 HCC samples showed the expression of CB1 and CB2 receptors to increase from normal liver to chronic hepatitis to cirrhosis. Marked expression of CB1 and CB2 receptors was noted in the majority of cirrhotic liver samples (86 and 78%, respectively). In HCC, high expression of CB1 and CB2 receptors was observed in 29 (45%) and 33 (52%) cases, respectively. Clinicopathological evaluation indicated a significant correlation between CB1 and CB2 expression and two clinicopathological parameters such as the histopathological differentiation (P = 0.021 and 0.001, respectively), portal vein invasion (P = 0.015 and 0.004, respectively). Univariate analysis indicated that disease-free survival was significantly better in HCC patients with high versus those with low CB1 and CB2 expression levels (P = 0.010 and 0.037, respectively). Our results indicate that CB1 and CB2 have potential as prognostic indicators and suggest possible beneficial effects of cannabinoids on prognosis of patients with HCC.     

Evoked pain behavior and spinal glia activation is dependent on tumor necrosis factor receptor 1 and 2 in a mouse model of bone cancer pain

Bone–cancer-related pain is one of the most disabling factors in patients suffering from primary bone cancer or bone metastases. Recent studies point toward an important role of proinflammatory cytokines, example tumor necrosis factor-α (TNF), for tumor growth and bone–cancer-associated pain. Mechanisms by which TNF, through its receptor subtypes, TNF receptor 1 (TNFR1) and −2 (TNFR2), elicits altered sensation and pain behavior, are still incompletely understood. To look for a potential role of TNF in bone cancer pain, cancer-related pain was analyzed in fibrosarcoma-bearing C57Bl/6J wild type mice after systemic antagonism of TNF. To further clarify the role of TNF receptor (TNFR) in bone-cancer pain, naive and fibrosarcoma-bearing C57Bl/ 6J wild type and transgenic mice with a deficiency of TNFR1 (TNFR1ko), TNFR2 (TNFR2ko), and TNFR1+2 (TNFR1+2ko) were compared regarding cancer-related pain and hyperalgesia, tumor growth, osteoclast activation, and spinal astrogliosis. Systemic antagonism of TNF significantly alleviated tactile hypersensitivity and spontaneous bone–cancer-related pain behavior. Most interestingly, combined deletion of the TNFR1 and TNFR2, but not of either gene alone, almost completely inhibited the development of tactile hypersensitivity, whereas spontaneous pain behavior was transiently increased. Accordingly, spinal astrogliosis was markedly reduced, whereas tumor growth was significantly increased in TNFR1+2ko mice. In contrast, deletion of the TNFR1 or TNFR2 gene alone did not change tumor growth or spinal astrogliosis. Our findings suggest that the combined absence of TNFR1 and TNFR2 is necessary for the attenuation of cancer-related tactile hypersensitivity and concomitant spinal astrogliosis, whereas tumor growth seems to be inhibited by combined TNFR activation. These findings support the hypothesis of cytokine-dependent pain development in cancer pain. Differential targeting of TNFR activation could be an interesting strategy in bone–cancer-related pain conditions.     

炎性痛致大鼠脊髓后角ProBDNF及其受体的表达变化

目的:探讨完全弗氏佐剂(complete Freund’s Adjuvant,CFA)致炎性疼痛后大鼠脊髓后角内ProBDNF及受体P75NTR和Sortilin的表达变化及意义。方法:大鼠随机分为正常组和实验组,实验组大鼠左侧足底皮下注射CFA和生理盐水混合溶剂100μl,建立炎性疼痛模型,实验组包括注射CFA后6 h、1、3、7和14 d组。采用Von Frey纤维测定不同时间点大鼠机械缩足阈值(pawwithdrawal threshold,PWT)的变化;采用免疫组织化学方法检测ProBDNF及P75NTR、Sortilin在脊髓后角的表达变化。结果:足底注射CFA后1 h PWT即下降,并在6 h后达到最低值,3 d后逐渐上调,至14 d仍低于基础值。ProBDNF在正常脊髓后角可见阳性表达;注射CFA后1 d注射侧脊髓后角较对侧其ProBDNF的表达明显上调,上调持续到7 d左右,至14 d逐渐恢复。正常大鼠脊髓后角P75NTR有较弱表达,主要集中在后角I、II层纤维;注射CFA后6 h开始,注射侧脊髓后角内P75NTR的表达明显上调,III-V层的阳性产物也逐渐增加,这种上调持续到7 d左右达高峰,至14 d逐渐下降。Sortilin在正常脊髓后角浅层仅有较弱阳性产物表达,注射CFA后不同时间点脊髓后角Sortilin的表达无明显差异。结论:CFA能诱导大鼠产生为期2周以上的炎性痛病程;脊髓后角ProBDNF和P75NTR的表达上调可能与炎性疼痛中外周痛觉信号的传导和中枢敏化有关。     

Effects of a cannabinoid agonist on spinal nociceptive neurons in a rodent model of neuropathic pain

The effects of the synthetic cannabinoid WIN 55,212-2 on heat-evoked firing of spinal wide dynamic range (WDR) neurons were examined in a rodent model of neuropathic pain. Fifty-eight WDR neurons (1 cell/animal) were recorded from the ipsilateral spinal dorsal horns of rats with chronic constriction injury (CCI) and sham-operated controls. Relative to sham-operated controls, neurons recorded in CCI rats showed elevations in spontaneous firing, noxious heat-evoked responses, and afterdischarge firing as well as increases in receptive field size. WIN 55,212-2 (0.0625, 0.125, and 0.25 mg/kg, intravenous) dose-dependently suppressed heat-evoked activity and decreased the receptive field areas of dorsal horn WDR neurons in both nerve injured and control rats with a greater inhibition in CCI rats. At the dose of 0.125 mg/kg iv, WIN 55,212-2 reversed the hyperalgesia produced by nerve injury. The effect of intravenous administration of WIN 55,212-2 appears to be centrally mediated because administration of the drug directly to the ligated nerve did not suppress the heat-evoked neuronal activity in CCI rats. Pretreatment with the cannabinoid CB(1) receptor antagonists SR141716A or AM251, but not the CB(2) antagonist SR144528, blocked the effects. These results provide a neural basis for reports of potent suppression by cannabinoids of the abnormal sensory responses that result from nerve injury.     

Unraveling the complexities of cannabinoid receptor 2 (CB2) immune regulation in health and disease

It has become clear that the endocannabinoid system is a potent regulator of immune responses, with the cannabinoid receptor 2 (CB2) as the key component due to its high expression by all immune subtypes. CB2 has been shown to regulate immunity by a number of mechanisms including development, migration, proliferation, and effector functions. In addition, CB2 has been shown to modulate the function of all immune cell types examined to date. CB2 is a G_i-protein-coupled receptor and thus exhibits a complex pharmacology allowing both stimulatory and inhibitory signaling that depends on receptor expression levels, ligand concentration, and cell lineage specificities. Here, we discuss both in vitro and in vivo experimental evidence that CB2 is a potent regulator of immune responses making it a prime target for the treatment of inflammatory diseases.