The Intriguing Effects of Substituents in the N-Phenethyl Moiety of Norhydromorphone: A Bifunctional Opioid from a Set of "Tail Wags Dog" Experiments.

(-)-N-Phenethyl analogs of optically pure N-norhydromorphone were synthesized and pharmacologically evaluated in several in vitro assays (opioid receptor binding, stimulation of [35S]GTPγS binding, forskolin-induced cAMP accumulation assay, and MOR-mediated ß-arrestin recruitment assays). "Body" and "tail" interactions with opioid receptors (a subset of Portoghese's message-address theory) were used for molecular modeling and simulations, where the "address" can be considered the "body" of the hydromorphone molecule and the "message" delivered by the substituent (tail) on the aromatic ring of the N-phenethyl moiety. One compound, N-p-chloro-phenethynorhydromorphone ((7aR,12bS)-3-(4-chlorophenethyl)-9-hydroxy-2,3,4,4a,5,6-hexahydro-1H-4,12-methanobenzofuro[3,2-e]isoquinolin-7(7aH)-one, 2i), was found to have nanomolar binding affinity at MOR and DOR. It was a potent partial agonist at MOR and a full potent agonist at DOR with a δ/µ potency ratio of 1.2 in the ([35S]GTPγS) assay. Bifunctional opioids that interact with MOR and DOR, the latter as agonists or antagonists, have been reported to have fewer side-effects than MOR agonists. The p-chlorophenethyl compound 2i was evaluated for its effect on respiration in both mice and squirrel monkeys. Compound 2i did not depress respiration (using normal air) in mice or squirrel monkeys. However, under conditions of hypercapnia (using air mixed with 5% CO2), respiration was depressed in squirrel monkeys.

Coadministration of Chemokine Receptor Antagonists with Morphine Potentiates Morphine's Analgesic Effect on Incisional Pain in Rats.

Crossdesensitization between opioid and chemokine receptors and involvement of chemokines in pain modulation are well established. We investigated if coadministration of chemokine receptor antagonists (CRAs) with morphine would enhance the analgesic potency of morphine on incisional pain in rats. Animals underwent incisional surgery on the left hind paw and pain responses were evaluated using von Frey filaments at various time points postsurgery between 15 and 360 minutes and daily between 24 and 72 hours. Dose-response curves for morphine, maraviroc (a CCR5 antagonist), and AMD3100 (a CXCR4 antagonist) alone were established. While morphine significantly reduced pain in a time- and dose-dependent manner, maraviroc and AMD3100 had no effect by themselves. Coadministration of either maraviroc or AMD3100 with morphine significantly increased morphine's analgesic effect on incisional pain, shifting the dose-response curve to the left 2.3- and 1.8-fold, respectively. Coadministration of both CRAs with morphine significantly shifted further the morphine dose-response curve to the left 3.3-fold. The effect of treatments on mRNA levels in the draining popliteal lymph node for a panel of chemokines and cytokines showed that message for many of these mediators was upregulated by the incision, and the combination of morphine with the CRAs markedly downregulated them. The data show that combining morphine with CRAs potentiates morphine's analgesic effect on incisional pain. Thus, the same analgesic effect of morphine alone can be achieved with lower doses of morphine when combined with CRAs. Using morphine in lower doses could reduce unwanted side effects and possibly block development of tolerance and dependence.

Potentiation of morphine antinociception and inhibition of diabetic neuropathic pain by the multi-chemokine receptor antagonist peptide RAP-103.

AIMS: We determined the ability of the multi-chemokine receptor (CCR2/CCR5/CCR8) antagonist RAP-103 to modulate pain behaviors in an acute model of surgical pain, with and without an added opioid (morphine), and by itself in a chronic model of Streptozotocin (STZ)-induced diabetic peripheral neuropathy (DPN). MATERIALS AND METHODS: Pain behaviors were assessed by mechanical and thermal tests in rats. Cytokine and chemokine biomarkers in sciatic nerve and spinal cord were assessed by in situ qPCR. KEY FINDINGS: In the incisional pain assay, RAP-103 (0.01-1 mg/kg, i.p.) alone had no antiallodynic effect post-surgery. RAP-103 (0.5 mg/kg) when co-administered with morphine (0.5-5 mg/kg), reduced the ED50 of morphine from 3.19 mg/kg to 1.42 mg/kg. In a DPN model, rats exhibited persistent mechanical and cold allodynia. Oral administration of RAP-103 (0.5-0.02 mg/kg/day) resulted in a complete reversal of established hypersensitivity in DPN rats (P < .001), which gradually returned to pain hypersensitivity after the cessation of the treatment. The mRNA expression of cytokines, IL-1ß, TNFα; chemokines CCL2, CCL3; and chemokine receptors CCR2 and CCR5 in DPN rat sciatic nerve, but not spinal cord, were significantly increased. RAP-103 resulted in significant reductions in sciatic nerve expression of IL-1ß, TNFα and CCL3 in STZ-induced diabetic rats with trends toward lower levels for CCL2 and CCR5, while CCR2 was unchanged. SIGNIFICANCE: In acute pain, co-administration of RAP-103 with morphine provided the same antinociceptive effect with a reduced dose of morphine, reducing opioid side-effects and risks. RAP-103 by itself is an effective non-opioid antinociceptive treatment for diabetic neuropathic pain.

The effect of gp120 on morphine's antinociceptive and neurophysiological actions.

Recently, we have shown that morphine's analgesic activity can be attenuated by chemokines, specifically CCL5 and CXCL12. Because the HIV-1 coat protein, glycoprotein 120 (gp120), binds to the same receptors as do CCL5 and CXCL12, experiments were designed to investigate the effect of gp120 in the brain on antinociception induced by morphine in the cold-water (-3°C) tail-flick (CWT) and hot-plate (+54°C) tests. In addition, mu-opioid-receptor-mediated effects in brain periaqueductal grey (PAG) slices were examined with whole-cell patch-clamp recordings. The results showed that (1) pretreatment with gp120 itself (10, 25, 50, 100 or 133 ng, PAG) had no nociceptive effect in the CWT; (2) pretreatment with gp120 (25 or 100 ng) dose-dependently reduced antinociception induced by subcutaneous (sc) injection of morphine (3 or 6 mg/kg) or PAG injection of morphine (100 ng) in the CWT; (3) a PAG injection of gp120 (133 ng), given 30 min before sc injection of morphine (6 mg/kg), similarly reduced morphine antinociception in the hot-plate test; (4) the inhibitory effect of gp120 on morphine-induced antinociception in the CWT was reversed by AMD3100, an antagonist of CXCR4; (5) pretreatment of slices with gp120 (200 pM) prevented morphine (10 µM)-induced hyperpolarization and reduction of input resistance in PAG neurons. Electrophysiology studies paralleled gp120-induced desensitization of a mu-opioid-receptor-mediated response in PAG neurons at the single-cell level. These studies are the first to demonstrate that the analgesic activity of morphine can be reduced by the presence of gp120 in the PAG and that pretreatment with AMD3100 is able to restore the analgesic effects of morphine.

A Cannabinoid 2-Selective Agonist Inhibits Allogeneic Skin Graft Rejection In Vivo.

Previous work from our laboratory showed that a CB2 selective agonist, O-1966, blocked the proliferative response of C57BL/6 mouse spleen cells exposed to spleen cells of C3HeB/FeJ mice in vitro in the mixed lymphocyte reaction (MLR). The MLR is widely accepted as an in vitro correlate of in vivo grant rejection. Mechanisms of the immunosuppression induced by the cannabinoid were explored, and it was shown that O-1966 in this in vitro assay induced CD25+Foxp3+ Treg cells and IL-10, as well as down-regulated mRNA for CD40 and the nuclear form of the transcription factors NF-κB and NFAT in T-cells. The current studies tested the efficacy of O-1966 in prolonging skin grafts in vivo. Full thickness flank skin patches (1-cm2) from C3HeB/FeJ mice were grafted by suturing onto the back of C57BL/6 mice. O-1966 or vehicle was injected intraperitoneally into treated or control groups of animals beginning 1 h pre-op, and then every other day until 14 days post-op. Graft survival was scored based on necrosis and rejection. Treatment with 5 mg/kg of O-1966 prolonged mean graft survival time from 9 to 11 days. Spleens harvested from O-1966 treated mice were significantly smaller than those of vehicle control animals based on weight. Flow cytometry analysis of CD4+ spleen cells showed that O-1966 treated animals had almost a 3-fold increase in CD25+Foxp3+ Treg cells compared to controls. When dissociated spleen cells were placed in culture ex vivo and stimulated with C3HeB/FeJ cells in an MLR, the cells from the O-1966 treated mice were significantly suppressed in their proliferative response to the allogeneic cells. These results support CB2 selective agonists as a new class of compounds to prolong graft survival in transplant patients.

Chemokine receptor antagonists enhance morphine's antinociceptive effect but not respiratory depression.

AIMS: We have shown that chemokines injected into the periaqueductal gray region of the brain blocks opioid-induced analgesia in the rat cold-water tail flick test (CWTF). The present experiments tested whether chemokine receptor antagonists (CRAs), in combination with sub-analgesic doses of morphine, would provide maximal analgesia in the CWTF test and the mouse formalin pain assay. The effect of CRAs on respiratory depression was also evaluated. MAIN METHODS: One, two or four CRAs (AMD3100/CXCR4, maraviroc/CCR5, RS504393/CCR2 orAZD8797/CX3CR1) were used in combination with sub-analgesic doses of morphine, all given systemically. Pain was assessed using the rat CWTF test or formalin injection into the paw of mice scored by licking. Respiration and oxygen saturation were measured in rats using a MouseOX® Plus - pulse oximeter. KEY FINDINGS: In the CWTF test, a sub-maximal dose of morphine in combination with maraviroc alone, maraviroc plus AMD3100, or with the four chemokine receptor antagonists, produced synergistic increases in antinociception. In the formalin test, the combination of four CRAs plus a sub-maximal dose of morphine resulted in increased antinociception in both male and female mice. AMD3100 had an additive effect with morphine in both sexes. Coadministration of CRAs with morphine did not potentiate the opioid respiratory depressive effect. SIGNIFICANCE: These results support the conclusion that combinations of CRAs can increase the potency of sub-analgesic doses of morphine analgesia without increasing respiratory depression. The results support an "opioid sparing" strategy for alleviation of pain using reduced doses of opioids in combination with CRAs to achieve maximal analgesia.

Targeting of the orphan receptor GPR35 by pamoic acid: a potent activator of extracellular signal-regulated kinase and ß-arrestin2 with antinociceptive activity.

Known agonists of the orphan receptor GPR35 are kynurenic acid, zaprinast, 5-nitro-2-(3-phenylproplyamino) benzoic acid, and lysophosphatidic acids. Their relatively low affinities for GPR35 and prominent off-target effects at other pathways, however, diminish their utility for understanding GPR35 signaling and for identifying potential therapeutic uses of GPR35. In a screen of the Prestwick Library of drugs and drug-like compounds, we have found that pamoic acid is a potent GPR35 agonist. Pamoic acid is considered by the Food and Drug Administration as an inactive compound that enables long-acting formulations of numerous drugs, such as the antihelminthics oxantel pamoate and pyrantel pamoate; the psychoactive compounds hydroxyzine pamoate (Vistaril) and imipramine pamoate (Tofranil-PM); and the peptide hormones triptorelin pamoate (Trelstar) and octreotide pamoate (OncoLar). We have found that pamoic acid induces a G(i/o)-linked, GPR35-mediated increase in the phosphorylation of extracellular signal-regulated kinase 1/2, recruitment of ß-arrestin2 to GPR35, and internalization of GPR35. In mice, it attenuates visceral pain perception, indicating an antinociceptive effect, possibly through GPR35 receptors. We have also identified in collaboration with the Sanford-Burnham Institute Molecular Libraries Probe Production Center new classes of GPR35 antagonist compounds, including the nanomolar potency antagonist methyl-5-[(tert-butylcarbamothioylhydrazinylidene)methyl]-1-(2,4-difluorophenyl)pyrazole-4-carboxylate (CID2745687). Pamoic acid and potent antagonists such as CID2745687 present novel opportunities for expanding the chemical space of GPR35, elucidating GPR35 pharmacology, and stimulating GPR35-associated drug development. Our results indicate that the unexpected biological functions of pamoic acid may yield potential new uses for a common drug constituent.

Intrahypothalamic injection of the HIV-1 envelope glycoprotein induces fever via interaction with the chemokine system.

Wasting syndrome is a common complication of HIV infection and is marked by progressive weight loss and weakness, often associated with fever. The mechanisms involved in the pathogenesis of these syndromes are not well defined, and neither are the brain areas involved. The present study tests a new hypothesis: that the preoptic anterior hypothalamus (POAH), the main brain area for thermoregulation and fever, has a role in the pathogenesis of fever induced by glycoprotein 120 (gp120), the surface envelope protein used by the HIV to gain access into immune cells, and that the CXC chemokine receptors (CXCR4) that serve as a coreceptor for HIV entry mediate the effect. A sterilized stainless steel C313G cannula guide was implanted into the POAH, and a biotelemetry system was used to monitor the body temperature (Tb) changes. The administration of gp120 into the POAH induced fever in a dose-dependent manner. To demonstrate possible links between the gp120 and CXCR4 in generating the fever, we pretreated the rats with 1,1'-[1,4-phenylenebis(methylene)]bis[1,4,8,11-tetraazacyclotetradecane] octohydrobromide dihydrate (AMD 3100), an antagonist of stromal cell-derived growth factor (SDF)-1alpha/CXCL12, acting at its receptor, CXCR4, 30 min before administration of gp120. AMD 3100 significantly reduced the gp120-induced fever. The present studies show that the presence of HIV-1 envelope glycoprotein gp120 in the POAH provokes fever via interaction CXCR4 pathway.

Potentiating effect of morphine on oral Salmonella enterica serovar Typhimurium infection is µ-opioid receptor-dependent.

Previous studies from our laboratory demonstrated that mice treated with morphine pellets are sensitized to Salmonella enterica, serovar Typhimurium infection. However, the opioid receptor antagonist, naltrexone, only partially blocked the effect of morphine, raising the possibility that the opioid might have some of its effects through a nonopioid receptor. To further clarify whether sensitization to infection is an opioid receptor-dependent phenomenon, µ-opioid receptor knockout (MORKO) mice were used in the present study. Wild-type (WT) and MORKO mice were treated with morphine and their sensitivity to oral Salmonella infection was assessed by mortality, bacterial burdens in gut associated lymphoid tissue and in blood and peritoneal fluid, and by levels of pro-inflammatory cytokines in plasma. MORKO animals treated with morphine were refractory to a sublethal dose of Salmonella, while similar treatment of WT animals resulted in 100% mortality. WT animals treated with morphine had high bacterial loads in all organs tested, while morphine-treated MORKO animals had no culturable Salmonella in any organs. Pro-inflammatory cytokine levels were elevated in morphine-treated WT but not MORKO mice infected with Salmonella. These results provide definitive evidence that the morphine-mediated enhancement of oral Salmonella infection is dependent on the µ-opioid receptor.

Chemokine Receptor Antagonists in Combination with Morphine as a Novel Strategy for Opioid Dose Reduction in Pain Management.

INTRODUCTION: Although opioids are widely prescribed for pain, in many circumstances, they have only modest efficacy. Preclinical studies have shown that chemokines, immune mediators released during tissue injury and inflammation, can desensitize opioid receptors and block opioid analgesia by a process termed "heterologous desensitization." The present studies tested the hypothesis that in evoked pain, certain chemokine receptor antagonists (CRAs), given with a submaximal dose of morphine, would result in enhanced morphine potency. METHODS: Three rodent pain assays were used: incisional pain in rats, the cold-water tail flick test in rats, and the formalin test in mice. The FDA-approved, commercially available CRAs, maraviroc and AMD3100, were used. They block the chemokine receptors and ligands, CCR5/CCL5 (RANTES) and CXCR4/CXCL4 (SDF-1α), respectively. RESULTS: In the incisional pain assay, it was found that the combination of a single CRA, or of both CRAs, with morphine significantly shifted the morphine dose-response curve to the left, as much as 3.3-fold. In the cold-water tail flick and formalin tests, significant increases of the antinociceptive effects of morphine were also observed when combined with CRAs. CONCLUSIONS: These results support the potential of a new "opioid-sparing" approach for pain treatment, which combines CRAs with reduced doses of morphine.

CB2 receptor activation attenuates microcirculatory dysfunction during cerebral ischemic/reperfusion injury.

Previous studies from our laboratory indicated that selective cannabinoid CB(2) agonists were able to attenuate cerebral ischemia/reperfusion (I/R) injury. The goal of current study is to further test whether this attenuation involves cerebral microcirculatory function during I/R injury. Middle cerebral artery occlusion with reperfusion (MCAO/R) was performed in male mice. A selective CB(2) agonist was administered at different dosages and different times. Cerebral infarction volume, neurological function and cerebral microcirculatory function (leukocyte/endothelial interactions, cell adhesion molecule expression and blood-brain barrier disruption) were examined in vivo and in vitro. CB(2) knockout mice were subjected to MCAO/R following same procedures. Administration of the CB(2) agonist at middle dosage exerted optimal effects in reducing cerebral infarction and improving neurological function compared with other dosage groups and control group. Treatment with the CB(2) agonist at the optimal dose was still effective when given 3 h after MCAO. Transient ischemia significantly increased leukocyte/endothelial interactions, adhesion molecules expression and blood-brain barrier disruption which were all attenuated by pre-treatment with a CB(2) agonist. CB(2) knockout mice showed larger cerebral infarction and worse neurological function compared to wide type. In conclusion, CB(2) activation contributed to protecting the brain through the attenuation of cerebral microcirculatory dysfunction during cerebral I/R injury.

Opioid-sparing effects of cannabinoids on morphine analgesia: participation of CB1 and CB2 receptors.

BACKGROUND AND PURPOSE: Much of the opioid epidemic arose from abuse of prescription opioid drugs. This study sought to determine if the combination of a cannabinoid with an opioid could produce additive or synergistic effects on pain, allowing reduction in the opioid dose needed for maximal analgesia. EXPERIMENTAL APPROACH: Pain was assayed using the formalin test in mice and the carrageenan assay in rats. Morphine and two synthetic cannabinoids were tested: WIN55,212-2 (WIN), which binds to both CB1 and CB2 receptors, and possibly TRPV1 channels; and GP1a, which has activity at CB2 receptors and is reported to inhibit fatty acid amide hydrolase, thus raising levels of endogenous cannabinoids. KEY RESULTS: Morphine in combination with WIN in the formalin test gave synergistic analgesia. Studies with selective antagonists showed that WIN was acting through CB1 receptors. Morphine in combination with GP1a in the formalin test was sub-additive. In the carrageenan test, WIN had no added effect when combined with morphine, but GP1a with morphine showed enhanced analgesia. Both WIN and Gp1a used alone had analgesic activity in the formalin pain test, but not in the carrageenan pain test. CONCLUSIONS AND IMPLICATIONS: The ability of a cannabinoid to produce an additive or synergistic effect on analgesia when combined with morphine varies with the pain assay and may be mediated by CB1 or CB2 receptors. These results hold the promise of using cannabinoids to reduce the dose of opioids for analgesia in certain pain conditions.

First in vivo evidence for a functional interaction between chemokine and cannabinoid systems in the brain.

Growing evidence supports the idea that in addition to their well established role in the immune system, chemokines might play a role in both normal and pathological brain function, and the chemokine network could interact with other neuromodulators. The chemokine stromal cell-derived growth factor (SDF)-1alpha/CXCL12, a member of the CXC chemokine family, was tested for its possible effect on the analgesic responses of the cannabinoid receptor agonist aminoalkylindole 4,5-dihydro-2-methyl-4-(4-morpholinylmethyl)-1-(1-naphthalenyl-carbonyl)-6H-pyrrolo-[3,2,1ij]quinolin-6-one [(+)-WIN 55,212-2, hereafter WIN 55,212-2] at the level of the periaqueductal gray (PAG), a brain region critical to the processing of pain signals, and a primary site of action of many analgesic compounds. The administration of WIN 55,212-2 (0.1-0.4 microg/microl) into the PAG resulted in antinociception in a dose-dependent manner. The selective cannabinoid (CB)1 antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride (SR 141716A; 1-10 microg) given into the PAG blocked the WIN 55,212-2-induced antinociception. In contrast, the selective CB2 antagonist N-[(1S)-endo-1,3,3-trimethyl bicyclo heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528; 10 microg) did not alter the WIN 55,212-2-induced antinociception. Pretreatment with SDF-1alpha/CXCL12 (100 ng) caused a reduction in antinociceptive responses of WIN 55,212-2. The inhibitory effect of SDF-1alpha/CXCL12 on WIN 55,212-2-induced antinociception was reversed by octahydrochloride [corrected] hydrate (AMD 3100) (10-50 ng), an antagonist of the SDF-1alpha/CXCL12, acting at its receptor, CXCR4. This study reports the first in vivo evidence of a functional interaction between chemokine and cannabinoid systems in the brain, showing that the activation of SDF-1alpha/CXCL12 receptors (CXCR4) in the PAG interferes with the analgesic effects of WIN 55212-2.

Bi-directional heterologous desensitization between the major HIV-1 co-receptor CXCR4 and the kappa-opioid receptor.

We previously characterized multiple interactions between chemokine and opioid G protein-coupled receptors (GPCR), and we found both mu and delta-opioid receptors cross-desensitize CCR1, CCR2, CCR5, but not CXCR4. Here we report that the kappa-opioid receptor (KOR) is able to cross-desensitize CXCR4, and this phenomenon is bi-directional. Chemotactic responses by KOR activation are diminished with prior activation of CXCR4. Additionally, calcium mobilization assays show these cross-desensitization processes occur within seconds of receptor activation, and target receptor internalization is not responsible for desensitization between these receptors. These results have implications for several essential processes including neuronal and lymphocyte development, inflammatory responses, and pain/sensitivity.

Endomorphin 1 and endomorphin 2 suppress in vitro antibody formation at ultra-low concentrations: anti-peptide antibodies but not opioid antagonists block the activity.

Endomorphin 1 (EM-1) and endomorphin 2 (EM-2) were tested for their capacity to alter immune function. Addition of either of these peptides to murine spleen cells in vitro inhibited antibody formation to sheep red blood cells in a bi-phasic dose dependent manner. Maximal inhibition was achieved at doses in the range of 10(-13) to 10(-15)M. Neither naloxone (general opioid receptor antagonist) nor CTAP (selective mu opioid receptor antagonist) blocked the immunosuppressive effect. To show that there was specificity to the immunosuppressive activity of the peptides, affinity-purified rabbit antibodies were raised against each of the synthetic EM peptides haptenized to KLH and tested for capacity to inhibit immunosuppression. Antibody responses were monitored by a standard solid phase antibody capture ELISA, and antibodies were purified by immunochromatography using the synthetic peptides coupled to a Sepharose 6B resin. Verification of the specificity of affinity-purified antisera was performed by immunodot-blot and solid-phase RIA assays. The antisera specific for both EM-1 and EM-2 neutralized the immunosuppressive effects of their respective peptides in a dose-related manner. Control normal rabbit IgG had no blocking activity on either EM-1 or EM-2. These studies show that the endomorphins are immunomodulatory at ultra-low concentrations, but the data do not support a mechanism involving the mu-opioid receptor.

A new brain area affected by 3,4-methylenedioxymethamphetamine: A microdialysis-biotelemetry study.

The widespread abuse of 3,4-methylenedioxymethamphetamine (MDMA) has intensified the need to learn more about this drug (e.g. its effects, its mechanism of action, brain areas affected). MDMA-induced hyperthermia is a severe physiological event not only because it can produce severe adverse consequences in human as well as experimental animals, but also because it plays a major role in determining the severity of the long-term MDMA-induced neurotoxicity that occurs. However, the effects of MDMA on the preoptic anterior hypothalamus, the main brain area responsible for control of body temperature, are still unknown. In vivo microdialysis-biotelemetry and pharmacological testing were used to determine whether the preoptic anterior hypothalamus is among the brain areas affected by MDMA by investigating the role of the dopamine neurotransmitter system. We examined the effect of a hyperthermic dose of MDMA on the extracellular level of dopamine in the preoptic anterior hypothalamus, and whether this effect is related to the acute hyperthermic response. The administration of a hyperthermic dose of MDMA (20 mg/kg) is accompanied by an increase in the extracellular level of dopamine in the preoptic anterior hypothalamus. Both the hyperthermia and augmented level of dopamine in the preoptic anterior hypothalamus after intraperitoneal injection of MDMA were significantly reduced by the pretreatment with D(1)-selective dopamine receptor antagonist, R-(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzapine (SCH 23390). These data provide the first in vivo evidence that the effects of MDMA extend to preoptic anterior hypothalamus.

Elevated level of the proinflammatory chemokine, RANTES/CCL5, in the periaqueductal grey causes hyperalgesia in rats.

The present data provide the first in vivo evidence that the proinflammatory chemokine, Regulated on Activation Normal T cell Expressed and Secreted (RANTES/CCL5) microinjected directly into the periaqueductal grey in rats, a brain region critical to the processing of pain signals, and a primary site of action of many analgesic compounds, induced hyperalgesia. Pretreatment with antibodies against RANTES/CCL5 prevented the hyperalgesic response, indicating that RANTES/CCL5 is able to interfere with the control of hyperalgesia at the level of the periaqueductal grey and suggesting that chemokine blockers could have analgesic properties.

Cannabinoid CB(2) receptor activation decreases cerebral infarction in a mouse focal ischemia/reperfusion model.

Cannabinoid CB(2) Receptor (CB(2)) activation has been shown to have immunomodulatory properties without psychotropic effects. The hypothesis of this study is that selective CB(2) agonist treatment can attenuate cerebral ischemia/reperfusion injury. Selective CB(2) agonists (O-3853, O-1966) were administered intravenously 1 h before transient middle cerebral artery occlusion (MCAO) or 10 mins after reperfusion in male mice. Leukocyte/endothelial interactions were evaluated before MCAO, 1 h after MCAO, and 24 h after MCAO via a closed cranial window. Cerebral infarct volume and motor function were determined 24 h after MCAO. Administration of the selective CB(2) agonists significantly decreased cerebral infarction (30%) and improved motor function (P<0.05) after 1 h MCAO followed by 23 h reperfusion in mice. Transient ischemia in untreated animals was associated with a significant increase in leukocyte rolling and adhesion on both venules and arterioles (P<0.05), whereas the enhanced rolling and adhesion were attenuated by both selective CB(2) agonists administered either at 1 h before or after MCAO (P<0.05). CB(2) activation is associated with a reduction in white blood cell rolling and adhesion along cerebral vascular endothelial cells, a reduction in infarct size, and improved motor function after transient focal ischemia.

Deletion of mu-opioid receptor in mice alters the development of acute neuroinflammation.

The realization that the mu-opioid system plays a key role in the control of the process of neuroinflammation is a new concept that may lead to identification of novel therapies for this extremely widespread and intractable syndrome. Fever is the hallmark among the defense mechanisms evoked by the entry into the body of pathogens to initiate the innate immune responses. In an attempt to determine the possible involvement of mu-opioid receptors in the control of brain inflammation, we examined the effect of their deletion on the fever induced by i.c.v. injection of lipopolysaccharide (LPS). The first series of experiments examined the thermal consequence of the absence of mu-opioid receptors on circadian body temperature rhythm and basal body temperature. Mu-opioid receptor knockout mice (MOP-KO) showed a normal circadian body temperature rhythm and basal body temperature compared with the wild type (WT). The second series of experiments investigated i.c.v. administration of LPS on body temperature in WT and MOP-KO. In the WT, i.c.v. injection of 100 ng of LPS induced fever, but there was no increase in body temperature in the MOP-KO mice. Saline, given i.c.v., did not alter the body temperature, either in WT or MOP-KO. These results show that the mu-opioid system participates in the control of acute neuroinflammation, further reinforcing our earlier finding that the opioid system is involved in the pathogenesis of fever induced by bacterial LPS, and that mu-opioid receptors are the target for morphine-induced hyperthermia.