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The present study was performed to explore the role of galanin and galanin receptor 1 (GalR 1) in nociceptive modulation in the central nucleus of amygdala (CeA) in normal rats and rats with neuropathy, and the involvement of GalR 1 and PKC was also investigated. The hindpaw withdrawal latencies (HWLs) to thermal and mechanical stimulations were increased in a dose-dependent manner after intra-CeA injection of galanin in both normal rats and rats with neuropathy. The increased HWLs were significantly attenuated by intra-CeA injection of galanin receptor antagonist M40, indicating an involvement of galanin receptor in nociceptive modulation in CeA. Furthermore, intra-CeA administration of the GalR 1 agonist M 617 induced increases in HWLs in normal rats, suggesting that GalR 1 may be involved in galanin-induce antinociception in CeA. Additionally, intra-CeA injection of the PKC inhibitor inhibited galanin-induced antinociception, showing an involvement of PKC in galanin-induced antinociception in CeA of normal rats. Moreover, there was a significant increase in GalR1 content in CeA in rats with neuropathy than that in normal rats. These results illustrated that galanin induced antinociception in CeA in normal rats and rats with neuropathy, and there is an up-regulation of GalR1 expression in rats with neuropathy.

The expression of galanin and galanin receptors are up-regulated in the brains from patients with Alzheimer's disease (AD). However, the role of galanin in the progress of AD is still controversial. Here we demonstrated that galanin increased the protein expression of M1 muscarinic acetylcholine receptor (M1) in the primary cultured prefrontal cortical neurons by ELISA and Western Blot. Moreover, we showed that the mRNA expression of M1 was also up-regulated by galanin treatment. We further explored the mechanism of the galanin induced up-regulation of M1. We found that galanin activated the ERK signaling pathway in the primary cultured prefrontal cortical neurons. In addition, our results showed that the up-regulation of M1 mRNA was blocked by an ERK inhibitor, U0126. Taken together, our results demonstrated that the ERK signaling pathway mediated the galanin induced up-regulation of M1 in the primary cultured prefrontal cortical neurons, supporting the hypothesis that galanin plays a beneficial role in the development of AD.

The present study was performed to explore the role of galanin and galanin receptor 2 in nociceptive modulation in anterior cingulate cortex (ACC) of normal rats and rats with mononeuropathy. Intra-ACC injection of galanin induced significant increases in hindpaw withdrawal latencies (HWLs) to thermal and mechanical stimulations in both normal rats and rats with mononeuropathy, the increased HWLs were attenuated significantly by intra-ACC injection of galanin receptor 2 antagonist M871, indicating an involvement of galanin receptor 2 in nociceptive modulation in ACC. Interestingly, the galanin-induced HWL was significant higher in rats with mononeuropathy than that in normal rats tested by Randall Selitto test. Furthermore, both the galanin mRNA expression and galanin content increased significantly in ACC in rats with mononeuropathy than that in normal rats. Moreover, both the mRNA levels of galanin receptor 2 and the content of galanin receptor 2 in ACC increased significantly in rats with mononeuropathy than that in normal rats. These results found that galanin induced antinociception in ACC in both normal rats and rats with mononeuropathy. And there may be plastic changes in the expression of galanin and galanin receptor 2 in rats with mononeuropathy, as well as in the galanin-induced antinociception.

The present study explored the effect of 2,3,5,4′-tetrahydroxystilbene-2-O-β- d -glucoside (THSG) on the staurosporine (STS)-induced toxicity in cultured rat hippocampal neurons. The results showed that administration of 200 μM of THSG significantly protected against 0.3 μM of STS-induced apoptosis in cultured rat hippocampal neurons tested by methyl thiazolyl tetrazolium (MTT) and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assays. Furthermore, when the Akt signaling pathway was blocked by LY294002, an inhibitor of Phosphatidyl Inositol 3-kinase (PI3K), the protective effects of THSG against STS-induced neurotoxicity were abrogated. We further examined the involvement of PI3K/Akt signaling pathway in THSG protection against STS-induced cytotoxicity on cultured neurons and found that administration of THSG significantly inhibited the STS-induced decreases in the content of phosphorylated AKt (p-Akt). Moreover, we found that THSG rescued the down-regulation of B cell lymphoma/lewkmia-2 (Bcl 2 ) and pro-caspase-3 (pro-Csp3) caused by STS in the neurons. These results indicate that THSG protect the cultured rat hippocampal neurons against STS-induced cytotoxicity and the PI3K/Akt signaling and mitochondrial apoptotic pathways are involved in the THSG-induced protective effects.

Calcitonin gene-related peptide (CGRP) has been implicated in pain transmission and morphine tolerance. Although the release of CGRP is well observed in pain-related regions after chronic morphine treatment, a lack of evidence is obvious for the expression of CGRP-receptor subunits, calcitonin receptor-like receptor (CRLR) and receptor activity-modifying protein 1 (RAMP1) in the brain. In this study, we investigated in vitro the time-course of the expression of CRLR and RAMP1 following morphine administration, and compared their changes among striatal, cerebellar, cortical and the spinal cord neurons. The levels of CRLR showed no significant changes in striatal cultures during morphine treatment, while the levels of RAMP1 time-dependently increased. Decrease in the content of RAMP1 was detected in cerebellar cultures. Apart from these facts, no significant changes of CRLR or RAMP1 were detected in any other cultures. The changes in the level of CRLR on the cell membrane were examined. Significant increases of surface CRLR were detected in both striatal and cerebellar neurons after morphine treatment, while cortical neurons not. It is concluded that the expression of gross and surface CRLR and RAMP1 were differentially regulated in primary neuronal cultures following morphine treatments, indicating the involvement of CGRP receptor in morphine associated functions.

Accumulating evidence indicates that α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors (AMPARs) are involved in the relapse to abused drugs. However, the role of AMPARs containing the GluR2 subunit in opiate addiction is still unclear. GluR2-3Y, an interfering peptide, prevents the endocytosis of AMPARs containing the GluR2 subunit. In this study, we explored the effect of intravenous injection of GluR2-3Y on the acquisition, expression, and reinstatement of morphine-induced conditioned place preference (mCPP) in rats. We found that infusion of GluR2-3Y (1.5 nmol/g) one hour before morphine during the conditioning phase inhibited the acquisition of mCPP, while an identical injection one hour before the post-conditioning test had no influence on the expression of mCPP. Injection of GluR2-3Y (1.5 nmol/g) after mCPP extinction blocked the morphine-induced reinstatement of mCPP. Our results strongly support the hypothesis that inhibition of AMPAR endocytosis provides a new target for the treatment of opiate addiction.

The present study was performed to explore the antinociceptive effects of M617, a selective galanin receptor 1 agonist, in the central nucleus of amygdala (CeA) of rats. Intra-CeA injection of 0.1 nmol, 0.5 nmol and 1 nmol of M617 induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulations in rats. Furthermore, rats received intra-CeA administration of M617 and galanin. The HWL to noxious thermal and mechanical stimulations increased markedly, and there were no significant differences in HWLs of rats received intra-CeA administration of M617 and galanin. The results demonstrated that intra-CeA injection of M617 induced significant antinociceptive effects in CeA of rats, indicating that galanin receptor 1 may be involved in M617-induced antinociception in the CeA of rats.

Addiction is a pathological usurpation of the neural processes that normally serve reward-related learning and memory. Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is an important molecule involved in the mechanisms of learning and memory, suggesting its roles in drug addiction. In this study, we detected the changes of CaMKII protein levels in the nucleus accumbens (NAc), a key nucleus involved in drug-reward, during the reinstatement of morphine-seeking behavior with animal model of morphine self-administration in rats. Moreover, considering that the NAc is also involved in the natural reward-related learning and memory, we detected the changes of CaMKII protein levels in the NAc during the reinstatement of natural reward-seeking with animal model of saccharin self-administration as a control. We found that the level of αCaMKII phosphorylated on Thr286 increased in the NAc shell subregion but not the NAc core during the reinstatement of morphine-seeking, compared with that after extinction. However, during the reinstatement of saccharin-seeking, the protein level of αCaMKII phosphorylated on Thr286 did not change in the NAc shell. Surprisingly, both αCaMKII phosphorylated on Thr286 and βCaMKII phosphorylated on Thr287 decreased in the NAc core during the reinstatement of saccharin-seeking. These results suggest that increased phosphorylation of CaMKII (Thr286) in the NAc shell is involved in the relapse to opioids-seeking and the mechanisms underlying the reinstatement of morphine-seeking are different from those involved in the reinstatement of natural reward-seeking.

It has been demonstrated that galanin plays important roles in the modulation of nociceptive information in rats. The present study is performed to investigate the regulating role of galanin in nociception in the nucleus accumbens (NAc) of rats. Intra-NAc administration of galanin induces dose-dependent increases in the hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation in rats. Furthermore, the galanin-induced antinociceptive effects are blocked by following intra-NAc injection of the galanin receptor antagonist galantide. The results demonstrate that galanin induces antinociceptive effects in the NAc of rats, and galanin receptors are involved in the galanin-induced antinociception effects.

The Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) may be a core component in the common molecular pathways for drug addiction. Moreover, studies using animal models of drug addiction have demonstrated that changing CaMKII activity or expression influences animals' responses to the drugs of abuse. Here, we explored the roles of CaMKII in the nucleus accumbens (NAc) shell in the extinction and reinstatement of morphine-seeking behavior. Rats were trained to obtain intravenous morphine infusions through poking hole on a fixed-ratio one schedule. Selective CaMKII inhibitor myristoylated autocamtide-2-inhibitory peptide (myr-AIP) was injected into the NAc shell of rats after the acquisition of morphine self-administration (SA) or before the reinstatement test. The results demonstrated that injection of myr-AIP after acquisition of morphine SA did not influence morphine-seeking in the following extinction days and the number of days spent for reaching extinction criterion. However, pretreatment with myr-AIP before the reinstatement test blocked the reinstatement of morphine-seeking behavior induced by morphine-priming. Our results strongly indicate that CaMKII activity in the NAc shell is essential to the relapse to morphine-seeking.

It has been demonstrated that calcitonin gene-related peptide (CGRP) plays important roles in the modulation of nociception in the nucleus accumbens (NAc) of rats. The present study is performed to explore the possible involvement of opioid receptors in the CGRP-induced antinociception in the NAc of rats. Intra-NAc administration of CGRP induces significant increases in the hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation in rats. Interestingly, the CGRP-induced antinociceptive effects are inhibited by following intra-NAc injection of the opioid receptor antagonist naloxone, suggesting that the opioid receptors are involved in the CGRP-induced antinociception in the NAc of rats. Furthermore, the CGRP-induced antinociception is attenuated by intra-NAc injection of mu-opioid receptor (MOR) antagonist β-funaltrexamine (β-FNA) and kappa-opioid receptor (KOR) antagonist nor-binaltorphimine (nor-BNI), but not by delta-receptor (DOR) antagonist naltrindole. In the present study, we also demonstrated that there was no significant difference between the CGRP-induced antinociception and the morphine-induced antinociception in the NAc in rats. The results of the present study demonstrate that both mu- and kappa-opioid receptors are involved in the CGRP-induced antinociception in the NAc of rats.

Recently the neuronal toxicity of intracellular amyloid beta (iAbeta) in Alzheimer's disease is attracting more and more attention. The present study explored the effects of curcumin on the iAbeta-induced toxicity in primary cultured rat prefrontal cortical neurons. The cell viability of primary cultured prefrontal cortical neurons decreased significantly after virus driven transfection of Abeta from 1 day to 7 days. Interestingly, administration of 1 microM, 10 microM or 20 microM of curcumin significantly inhibited the iAbeta-induced toxicity in primary cultured rat prefrontal cortical neurons tested by MTT and LDH release assays. We further studied the involvements of apoptotic or neuroprotective pathway proteins in curcumin protection against iAbeta-induced cytotoxicity in primary cultured rat prefrontal cortical neurons. The results demonstrated that the contents of activated caspase-3 increased significantly by iAbeta, while curcumin significantly inhibited the iAbeta-induced increases of activated caspase-3 tested by Western blot. And the contents of p-AKT decreased significantly after iAbeta treatment, while administration of curcumin significantly inhibited the iAbeta-induced decreases in the contents of p-AKT. The results suggest that curcumin may play a protective effect in primary cultured rat prefrontal cortical neurons against iAbeta-induced cytotoxicity, and both AKT and caspase-3 are involved in the curcumin-induced protective effects.

The present study explored the effect of curcumin against amyloid beta (Abeta)-induced toxicity on cultured rat primary prefrontal cortical neurons. The results showed that administration of 10 microM of curcumin induced significantly protection against 20 microM of Abeta(25-35)-induced toxicity on the cultured rat primary prefrontal cortical neurons tested by MTT and TUNEL assays. We further examined the involvements of the apoptotic or anti-apoptotic proteins in curcumin protection against Abeta(25-35)-induced cytotoxicity on cultured neurons and found that the content of apoptotic protein caspase-3 was increased and the content of anti-apoptotic factor Bcl2 was decreased significantly after Abeta(25-35) treatments, while administration of curcumin significantly inhibited the Abeta(25-35)-induced increases in the content of caspase-3 and inhibited the Abeta(25-35)-induced decreases in the content of Bcl2 tested by Western blot. The results suggest that curcumin protects cultured rat primary prefrontal cortical neurons against Abeta-induced cytotoxicity, and both Bcl2 and caspase-3 are involved in the curcumin-induced protective effects.

Opiates, including morphine, are widely used drugs for antinociception in clinics. Prolonged treatments of opioids induce both tolerance and dependence, which are the major side effects of opioid therapy. One of the mechanisms for the development of tolerance and dependence is implicated to be opioid-receptor trafficking. Here we review the current understandings of opioid-receptor phosphorylation, endocytosis and desensitization after repeated agonist treatments. Also, the role of G-protein coupled receptor kinases in opioid-receptor phosphorylation is discussed. How to associate these observations to physiological and behavioral changes in animal models and clinics is still under investigation.

Morphine has been used for pain treatment with a long history. Some data suggest that morphine is toxic to neurons and induces apoptosis, while other evidence shows that morphine could have beneficial effects against cell death. To determine how morphine affects pro-apoptotic protein Bax and molecular chaperone Hsp70, different concentrations of morphine were examined. Our results show that prolonged morphine administration for 5 days at 1microM concentration protects against serum deprivation induced cell death in rat primary neurons. Morphine treatment decreases Bax and Hsp70 levels in cultured rat primary neurons, suggesting morphine may have a protective role in staurosporine and serum deprivation induced cytotoxicity.

The central nervous system (CNS) is highly dependent on adequate supply of oxygen and is sensitive to hypoxia. It is known that hypoxia induces injuries on the brain tissue and the neuronal activity. Curcumin, a yellow pigment obtained from the rhizome of C. longa Linn., has been regarded as a multi-functional drug with antioxidative activity. In the present study, we first demonstrated a significant decrease in the content of beta-III tubulin protein in rat prefrontal cortex (PFC) tissues induced by repeated hypoxia, but not in rat cerebellum tissue. These suggest a relatively higher sensitivity and probably a higher vulnerability of rat PFC tissue to hypoxia in vivo. We reconfirmed the effect of hypoxia to primary cultured neurons from rat PFC and found a significant decrease in the contents of beta-III tubulin protein after chronic exposure to hypoxia. Moreover, we demonstrated that the hypoxia-induced decrease in beta-III tubulin protein content could be restored by curcumin, suggesting a potential protection of curcumin against hypoxia-induced decreases in beta-III tubulin content in rat PFC neurons.

We have previously shown that microinjection of galanin into the arcuate nucleus of hypothalamus (ARC) produced antinociceptive effects in rats (Sun et al., 2003a). In this study, the neural pathway of galanin from ARC to midbrain periaqueductal gray (PAG) in nociceptive modulation was investigated. The hindpaw withdrawal latencies (HWLs) with noxious thermal and mechanical stimulation were assessed by the hotplate and the Randall Selitto tests. Intra-ARC administration of 0.1, 0.5, or 1 nmol of galanin induced significant increases in HWLs of rats. The galanin-induced increases in HWLs were inhibited by injection of 10 μg of the opioid receptor antagonist naloxone or 1 nmol of the mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA) into PAG, suggesting that the antinociceptive effects induced by intra-ARC injection of galanin occur via the neural pathway from ARC to PAG. Furthermore, our results demonstrate that the galaninergic fibers directly innervated the beta-endorphinergic neurons in ARC by immunofluorescent methods. Taken together, our results suggest that galanin produces antinociceptive effects in the ARC of rats by activating the beta-endorphinergic pathway from ARC to PAG. © 2007 Wiley-Liss, Inc.

Acute morphine exposure induces antinociceptive activity, but the underlying mechanisms in the central nervous system are unclear. Using whole-cell patch clamp recordings, we explore the role of morphine in the modulation of excitatory synaptic transmission in lateral amygdala neurons of rats. The results demonstrate that perfusion of 10 μM of morphine to the lateral amygdala inhibits the discharge frequency significantly. We further find that there are no significant influences of morphine on the amplitude of spontaneous excitatory postsynaptic currents (sEPSCs). Interestingly, morphine shows no marked influence on the evoked excitatory postsynaptic currents (eEPSCs) in the lateral amygdala neurons. These results indicate that acute morphine treatment plays an important role in the modulation on the excitatory synaptic transmission in lateral amygdala neurons of rats.

The present study was performed to explore the effect of the galanin receptor 2 (GalR2) antagonist M871 on the galanin-induced antinociception in periaqueductal grey (PAG), and an involvement of Ca(2+)/calmodulin-dependent kinase II (CaMKII) in the galanin-induced antinociception. Intra-PAG injection of galanin induced marked increases in HWLs to noxious thermal and mechanical stimulation. The increased HWLs to thermal and mechanical stimulation decreased significantly after intra-PAG administration of the GalR2 antagonist M871, indicating an involvement of GalR2 in the galanin-induced antinociception in PAG of rats. Furthermore, rats received intra-PAG injection of galanin, followed 5min later by intra-PAG administration of the CaMKII inhibitor MAP. The galanin-induced increases in HWLs to thermal and mechanical stimulation decreased significantly after intra-PAG administration of MAP, indicating that there is an involvement of CaMKII in the galanin-induced antinociception in PAG, blockade the activity of CaMKII by MAP inhibits the galanin-induced antinociception in PAG of rats. Our results strongly indicate that the galanin-induced antinociception is mediated by GalR2 in the PAG, and CaMKII may be involved in the galanin-induced antinociception in PAG of rats.

Calcium/calmodulin-dependent protein kinase II (CaMKII) is a serine/threonine- dependent protein kinase, which has been implicated in pain modulation at different levels of the central nervous system. The present study was performed in rats with mononeuropathy induced by left common sciatic nerve ligation. Unilateral sciatic nerve loose ligation produced decreases in the hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation. Intra-nucleus accumbens (NAc) injection of 3 μg, 6 μg and 12 μg of myristoylated autocamtide-2-inhibitory peptide (AIP), the CaMKII inhibitor, dose-dependently increased the HWL to noxious thermal and mechanical stimulation in rats with mononeuropathy. Furthermore, intra-NAc administration of morphine, the HWL to noxious thermal and mechanical stimulation increased markedly, and there were no significant differences between morphine group and AIP group. Taken together, the results showed that intra-NAc injection of AIP induced significant antinociceptive effects in rats with mononeuropathy, indicating that CaMKII may play an important role in the transmission and/or modulation of nociceptive information in the NAc in rats with mononeuropathy.

Cholecystokinin (CCK) is an endogenous anti-opioid peptide in the central nervous system. The present study investigated the effects of endogenous CCK on tolerance to morphine antinociception in the nucleus accumbens (NAc) of rats. Chronic administration of morphine to NAc induced marked tolerance to antinociception. Intra-NAc administration of the CCK2 receptor antagonist LY225910 inhibited not only the development but also the expression of chronic morphine-induced antinociceptive tolerance. However, intra-NAc injection of LY225910 did not influence the antinociception induced by intra-NAc administration of morphine in the intact rats. The results indicate that endogenous CCK plays an important role in morphine-induced antinociceptive tolerance in the NAc of rats.

The present study investigated the alteration in intracerebroventricular administration of substance P-induced anti-nociception after morphine tolerance in rats. The nociceptive response latencies to noxious thermal and mechanical stimulation increased significantly after intracerebroventricular administration of SP in normal rats. The SP-induced anti-nociception decreased significantly in morphine-tolerant rats than that in opioid naive rats. The results demonstrated that SP induced significant anti-nociceptive effects in the brain in normal rats, and the SP-induced anti-nociception was attenuated significantly in morphine-tolerant rats than that in opioid naive rats, indicating an alteration in SP-induced anti-nociception in the brain after chronic morphine treatments.

The fact that galanin, beta-endorphin and their receptors are present in the arcuate nucleus of hypothalamus (ARC), coupled with our previous observation that both beta-endorphin and galanin play antinociceptive roles in pain modulation in the ARC, made it of interest to study their interactions. The hindpaw withdrawal latency (HWL) in response to noxious thermal and mechanical stimulation was assessed by the hot-plate test and the Randall Selitto Test. We showed that the antinociceptive effect induced by intra-ARC injection of galanin was dose-dependently attenuated by the following intra-ARC injection of naloxone. Furthermore, intra-ARC administration of the selective mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA) attenuated the increased HWL induced by intra-ARC injection of galanin in a dose-dependent manner, while the delta-opioid receptor antagonist naltrindole or the kappa-opioid receptor antagonist nor-binaltorphimine (nor-BNI) did not. Moreover, intra-ARC injection of a galanin receptor antagonist galantide attenuated intraperitoneal morphine-induced increases in HWLs. These results demonstrate that the antinociceptive effect of galanin was related to the opioid system, especially mu-opioid receptor was involved in, and that systemic morphine induced antinociception involves galanin in the ARC.

Neuropeptide nociceptin/orphanin FQ is the endogenous ligand for the opioid-receptor-like receptor 1 (ORL1), mediating essential functions in the central and peripheral nervous systems. The present study was performed to investigate the role of nociceptin and ORL1 receptor in nociception and morphine-induced antinociception in the arcuate nucleus of hypothalamus in rats. Hindpaw withdrawal latencies (HWL) were measured by hot-plate and Randall Selitto tests. The HWL to both thermal and mechanical stimulation decreased significantly after intra-arcuate nucleus injection of nociceptin in a dose-dependent manner. The effect of nociceptin was blocked significantly by subsequent intra-arcuate nucleus administration of [Nphe1]nociceptin(1–13)-NH2, an ORL1 receptor antagonist. Furthermore, an intra-arcuate nucleus injection of nociceptin dramatically attenuated the antinociceptive effect induced by morphine either injected in the same site or applied intraperitoneally. These results suggest that nociceptin in the arcuate nucleus induces a hyperalgesic effect by acting on ORL1 receptors. The present study also demonstrates an interaction between nociceptin and opioids in the arcuate nucleus of the hypothalamus.

The aims of the present study were to investigate, in rats, the behavioral effects of substance P (SP) microinjected into the ventrolateral periaqueductal gray (PAG) and the effects of the neurokinin 1 (NK-1) receptor antagonist [ d -Arg1, d -Trp7, 9, Leu11]-substance P (Spantide). The effect of morphine administration on the release of SP in the ventrolateral PAG was also investigated using microdialysis in awake rats. SP microinjected into the ventrolateral part of the PAG induced significant increases in the hindpaw withdrawal latencies (HWLs) to thermal and mechanical stimulation as an antinociceptive response. The NK-1 receptor antagonist blocked these effects but exhibited no antinociceptive effect alone. Subcutaneous administration of morphine increased basal SP-like immunoreactivity (SP-LI) release in the microdialysate obtained from the ventrolateral PAG of freely moving rats. Our results demonstrate that SP injected into the ventrolateral PAG induces an antinociceptive effect via activation of NK-1 receptors. Morphine administered systemically induces the release of SP in the ventrolateral PAG. We suggest that an increased release of SP in the PAG may contribute to opioid antinociception.

The present study investigated the effect of diazepam binding inhibitor (DBI) on nociception in the central nervous system of rats. There were dose-dependent increases in hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation after intrathecal injection of 1, 5 or 10 microg of DBI in rats, indicating a DBI-induced anti-nociceptive effect at the spinal levels of rats. Furthermore, it was found that there were no significant influences of intrathecal co-administration of gamma-aminobutyric acid (GABA) on the intrathecal DBI-induced increases in HWLs of rats. Intracerebroventricular administration of 1, 10 or 20 microg of DBI also induced dose-dependent increases in HWL to thermal and mechanical stimulation in rats, suggesting an anti-nociceptive effect of DBI in the brain. Moreover, there were no significant influences of intracerebroventricular co-administration of 2 microg of GABA on the intracerebroventricular DBI-induced increases in HWL of rats. The results of the present study demonstrated that DBI played anti-nociceptive effects in the central nervous system of rats.

The present study investigates the involvement of opioid receptors in the antinociceptive effects of nociceptin in the spinal cord of the rat. Intrathecal administrations of 5 and 10 nmol of nociceptin significantly increase the withdraw response latencies to noxious thermal and mechanical stimulations. This nociceptin-induced antinociceptive effect is significantly attenuated by intrathecal injection of (Nphe(1))nociceptin(1-13)-NH(2), a selective antagonist of the nociceptin receptor (opioid receptor-like receptor ORL1), indicating an ORL1 receptor-mediated mechanism. This antinociceptive effect is also significantly attenuated by intrathecal injections of naloxone (a nonselective opioid receptor antagonist), naltrindole (a selective delta-opioid receptor antagonist), and beta-funaltrexamine (a selective mu-opioid receptor antagonist) in a dose-dependent manner, but not by the selective kappa-opioid receptor antagonist norbinaltorphimine. Since it is unlikely that nociceptin acts by direct binding to opioid receptors, these results suggest a possible interaction between the nociceptin/ORL1 and opioid systems in the dorsal horn of the rat spinal cord.

The present study investigated the effect of neuropeptide Y on nociception in the nucleus accumbens of rats. Intra-nucleus accumbens administration of neuropeptide Y induced dose-dependent increases in the hindpaw withdrawal latency (HWL) to thermal and mechanical stimulation in rats. There were no significant changes in the HWL to both stimulation during 60 min after the administration of NPY to outside of the nucleus accumbens. The anti-nociceptive effect of NPY was blocked by subsequent intra-nucleus accumbens injection of the Y1 receptor antagonist neuropeptide Y 28-36, indicating that Y1 receptor is involved in the neuropeptide Y-induced anti-nociception in the nucleus accumbens. Furthermore, the anti-nociceptive effect of neuropeptide Y was attenuated by intra-nucleus accumbens administration of the opioid antagonist naloxone, suggesting an involvement of the endogenous opioid system in the neuropeptide Y-induced anti-nociception in the nucleus accumbens of rats. Moreover, the neuropeptide Y-induced anti-nociception was attenuated by following intra-nucleus accumbens injection of the selective opioid antagonists nor-binaltorphimine and beta-funaltrexamine, but not by naltrindole, illustrating that mu- and kappa-opioid receptors, not the delta-opioid receptor, were involved in the neuropeptide Y-induced anti-nociception in the nucleus accumbens of rats.

Intra-periaqueductal grey (PAG) injection of 1 nmol of oxytocin induced significant increases in hindpaw withdrawal latency (HWL) to thermal and mechanical stimulation in rats. The anti-nociceptive effect of oxytocin was attenuated significantly by subsequent intra-PAG injection of the mu opioid antagonist beta-funaltrexamine (beta-FNA) and the kappa opioid antagonist nor-binaltorphimine (nor-BNI), but not by the delta antagonist naltrindole. The results demonstrated that mu and kappa opioid receptors, not delta receptors, were involved in the oxytocin-induced anti-nociception in PAG of rats.

Previous studies in our laboratory demonstrated that galanin and its receptors play important roles in nociceptive modulation in the central nervous system. The present study was performed to explore the antinociceptive effects of the galanin receptor 1 agonist M 617 in the central nervous system of rats. Intracerebroventricular injection of 0.1 nmol, 0.5 nmol, 1 nmol or 2 nmol of M 617 induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulations in rats. Furthermore, both intracerebroventricular injection of M 617 and galanin induced significant increases in HWLs in rats. Interestingly, there were no significant differences between the antinociceptive effects induced by M 617 and galanin, indicating that galanin receptor 1 plays main roles in galanin-induced antinociceptive effects in the brain of rats.

The present study investigated the effect of intraperiaqueductal grey injection of nociceptin/orphanin FQ (N/OFQ) and an antagonist (Nphe(1))nociceptin(1-13)NH(2) on the hindpaw withdrawal response to thermal and mechanical stimulation in rats. N/OFQ (5 nmol) significantly decreased the nociceptive thresholds in both tests and 1, 5 and 10 nmol of (Nphe(1))nociceptin(1-13)NH(2) significantly reversed this effect in a dose dependent way. Our results demonstrate, that N/OFQ has a nociceptive action, possibly through inhibition of PAG neurons. This effect is blocked by the antagonist (Nphe(1))nociceptin(1-13)NH(2) probably via ORL1 receptors in the periaqueductal grey.

Experimental inflammation was induced by subcutaneous injection of carrageenan into the left hindpaw of rats. Intra-periaqueductal grey (PAG) injection of 0.02 or 0.1 nmol of neuropeptide Y (NPY), but not 0.004 nmol, induced significant increases in hindpaw withdrawal latency (HWL) to thermal and mechanical stimulation in rats with inflammation. Furthermore, the anti-nociceptive effect of NPY was blocked partly by following intra-PAG injection of the Y1 receptor antagonist NPY28-36. The results demonstrated that NPY plays an anti-nociceptive role in PAG in rats with inflammation, in which Y1 receptor is involved.

The present study was performed to investigate the antinociceptive effects of M617, a selective galanin receptor 1 agonist, and M1145, a selective galanin receptor 2 agonist, in the periaqueductal grey (PAG) in rats with morphine tolerance. Intra-PAG injection of 0.1 nmol, 0.5 nmol and 1 nmol of M617 induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulations in rats with morphine tolerance. Nevertheless, intra-PAG injection of 5 nmol of the selective galanin receptor 2 agonist M1145 showed no significant influences on HWLs to noxious thermal and mechanical stimulations in rats with morphine tolerance. The results demonstrated that it is the selective galanin receptor 1 agonist M617, not the selective galanin receptor 2 agonist M1145, induced significant antinociceptive effects in morphine-tolerant rats, indicating that galanin receptor 1 is involved in nociceptive modulation in the PAG of morphine-tolerant rats.

The Mu opioid receptor (MOR) has been shown to participate in the analgesic effect of the calcitonin gene-related peptide (CGRP) in the nucleus accumbens (NAc) of adult rats. However, it is not clear whether and how CGRP regulates the MOR at the molecular levels. In the present study, it is found that the level of MORs on the cell membrane of NAc neurons was increased twice more than the control level following CGRP treatment (1μM, 30min), which is a phenomenon that was blocked by the peptidergic antagonist CGRP8-37. No direct physical interaction was observed between MORs and CGRP receptors, and neither brefeldin A nor dynosore preincubation affected such effects of CGRP. However, addition of 20μM monensin 1h before CGRP treatment significantly blocked the action of CGRP on surface MORs. In living animals, microinjection of CGRP (1nmol in 1μl) into the NAc partially restored morphine antinociception in morphine-tolerant rats, and the effect of CGRP on surface MORs extended beyond normal NAc neurons to chronic morphine-treated NAc neurons. To conclude, these results demonstrate that CGRP can act on MORs and increase the number of surface MORs in NAc neurons, partially explaining the involvement of opioid receptors in CGRP-induced antinociception in the rat NAc.

It is well known that hippocampus plays important roles in learning and memory. Both calcitonin gene-related peptide (CGRP) and CGRP receptors are found in hippocampus. In the present study we explored the influence of CGRP on long term potentiation (LTP) in hippocampal Schaffer collateral-CA1. Our results demonstrated that CGRP inhibited the LTP induced by high frequency stimulation (HFS) in hippocampal CA1 neurons in rat brain slices. The inhibitory effect was blocked by CGRP receptor-1 antagonist CGRP 8-37. The results indicate that both CGRP and CGRP receptor 1 are involved in the modulation process of LTP in hippocampus of rats.

The neuropeptide galanin and its receptors are found to be upregulated in brain associated with Alzheimer's disease (AD), while the role of galanin in AD is still unclear. The present study was performed to explore the neuroprotective role of galanin both in vitro and in vivo. Our results demonstrated that galanin inhibited the neurotoxicity induced by amyloid-beta(25-35) (Abeta(25-35)) or Abeta(1-42) in primary cultured hippocampal neurons of rats. Moreover, Gal(2-11) (an agonist of GalR(2/3)) also inhibited the neurotoxicity induced by Abeta(25-35) in the cultured neurons. We further found that galanin inhibited the activation of p53, Bax, and caspase-3 induced by Abeta(25-35) in the cultured hippocampal neurons. Moreover, galanin reversed the down regulation of Bcl-2 induced by Abeta(25-35) in the cultured neurons. Interestingly, in the Morris water maze task we found that intra-CA1 injection of Abeta(25-35)-induced spatial learning deficits in rats were blocked by galanin. In addition, galanin inhibited the Abeta(25-35)-induced dysregulation of p53, Bax, and MAP2 in rat hippocampus. Our results strongly demonstrate that galanin plays neuroprotective roles in nerve cells and in AD-induced learning and memory deficits.

The central nucleus of amygdala (CeA) is a very important brain structure involved in multiple physiological functions, especially in pain modulation. There are high densities of galanin and galanin receptors found in the CeA. The present study was performed to explore the antinociceptive effects of galanin in the CeA of rats, and possible involvements of opioid receptors in the galanin-induced antinociception. Intra-CeA injection of galanin induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulations in rats. Interestingly, the amtinociceptive effect induced by intra-CeA injection of galanin was blocked by intra-CeA injection of naloxone, a common opioid receptor antagonist, indicating an involvement of opioid receptors in the galanin-induced antinociception in the CeA of rats. Moreover, intra-CeA injection of either selective mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA) or delta-opioid receptor antagonist naltrindole, but not kappa-opioid receptor antagonist nor-binaltorphimine (nor-BNI), significantly attenuated the galanin-induced increases in HWLs in the CeA of rats. Taken together, the results demonstrate that galanin induces antinociceptive effects in the CeA of rats, and both mu- and delta-opioid receptors are involved in the galanin-induced antinociception.

Studies have demonstrated that oxytocin plays important roles in pain modulation in the central nervous system. Oxytocin-ergic neurons are found in paraventricular nucleus and supraoptic nucleus of the hypothalamus. The oxytocin-ergic neurons send fibers from hypothalamus to amygdala and high density of oxytocin receptors are found in the central nucleus of amygdala (CeA). The present study was performed to investigate the influences of oxytocin and its receptors on nociceptive responses in the CeA of rats. Intra-CeA injection of 0.1, 0.5 or 1nmol of oxytocin induced dose-dependent increases in the handpaw withdrawal latency induced by noxious thermal and mechanical stimulation in rats. The oxytocin-induced anti-nociception could be blocked by the selective oxytocin antagonist 1-deamino-2-d-Tyr-(Oet)-4-Thr-8-Orn-oxytocin. The present study demonstrated that oxytocin and its receptors are involved in nociceptive modulation in the CeA of rats.

This study explored the possibility of a relay at habenula for the descending neural pathway of antinociception. The latency of the escape response elicited by radiant heat on the snout of the rabbit was taken as index of nociception. (1) Microinjection of 20 micrograms of morphine into nucleus accumbens resulted in a one-fold increase in nociceptive threshold 20-40 min after the injection. This effect of morphine was markedly attenuated by naloxone or met-enkephalin antiserum administered to the nucleus habenula, suggesting that the release of met-enkephalin in habenula is essential for the antinociception induced by morphine injected into nucleus accumbens. (2) Injection of 10 micrograms of morphine into habenula produced a significant increase in escape response latency 20-40 min after the injection. This antinociceptive effect of morphine was attenuated by naloxone or muscimol, and enhanced by bicuculline methochloride administered to periaqueductal grey, suggesting that morphine may act on habenula to activate a descending neural pathway extending to periaqueductal grey to induce an antinociceptive effect, which seems to utilize endogenous opioid peptides and gamma-aminobutyric acid as its mediators. Taking together, the results suggest that habenula is an important relay in the descending neural pathway from nucleus accumbens to periaqueductal grey subserving antinociception.

The present study was performed to investigate the effects of valproic acid sodium (VPA), a widely utilized antiepileptic drug, on the establishment of chronic morphine-induced conditioned place preference (CPP). The rat model of morphine-induced CPP was conditioned with alternating intracerebroventricular (i.c.v.) injections of morphine (60 microg/6 microl) and saline for 5 days. To investigate the influence of VPA on morphine-induced CPP, rats received chronic pretreatment of i.c.v. VPA (500 microg/rat) 10 min previous to the daily morphine injection. The results demonstrated that the morphine-induced CPP was significantly attenuated by VPA pretreatment, while the VPA itself could not induce any CPP or conditioned place aversion (CPA) effects. The results of the present study not only confirmed the reliability of establishing morphine-induced CPP model by i.c.v. injection, but also suggest that the antiepileptic drug VPA may be utilized as potential therapeutic medications for drug abuse in the future.

The hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulation increased significantly after intra-hypothalamic arcuate nucleus (ARC) injection of galanin in mononeuropathic rats, while intra-ARC injection of the putative antagonist of galanin receptors markedly reduced the HWLs. The number of galaninergic neurons in the ARC increased in rats with mononeuropathy than that in normal rats. The results demonstrated that both endogenous and exogenous galanin were involved in the regulation of nociception in the ARC of rats with peripheral nerve injury.

Studies demonstrated that intrathecal 1,2,3,4-tetrahydro-6-nitro-2, 3-dioxo[f]quinoxaline-7-sulfonamide disodium (NBQX), an antagonist of AMPA/kainate receptors, induced antinociception in the spinal cord of rats. The present study demonstrated that the NBQX-induced increases in hindpaw withdrawal latencies (HWLs) were dose-dependently attenuated by intrathecal pretreatment of the AMPA receptor desensitization inhibitor, diazoxide. The effect was unrelated to the opening of K+ channels by diazoxide. On the other hand, intrathecal pretreatment of concanavalin A, which selectively inhibits the desensitization of kainate receptor, produced no significant influence on the NBQX-induced antinociception. The results suggest that the NBQX-induced antinociception was mediated by AMPA receptors, not by kainate receptors, in the spinal cord of rats.

The present study was performed to explore the involvement of opioid receptors in the antinociception induced by a-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor antagonist in rats. The hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation was assessed by hot plate test and the Randall Selitto Test. Intrathecal injection of 20 nmol of 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo[f]quinoxaline-7-sulfonamide (NBQX) disodium, a competitive AMPA receptor antagonist, increased significantly the HWLs to both thermal and mechanical stimulation in rats. The increased HWLs induced by NBQX were dose-dependently attenuated by the opioid receptor antagonist naloxone, while naloxone itself had no marked influences on the HWL of rats. Furthermore, the increased HWLs induced by NBQX were inhibited by the mu-opioid antagonist beta-funaltrexamine (β-FNA) or the delta-opioid antagonist naltrindole, but not by the kappa-opioid antagonist nor-binaltorphimine (nor-BNI). The results suggest that mu- and delta-opioid receptors, not kappa-opioid receptor, are involved in the antinociception induced by AMPA antagonist in the spinal cord of rats.

It is well known that there are three types of opioid receptors, mu- (MOR), delta- (DOR), and kappa-opioid receptor (KOR) in the central nervous system. The present study investigated the involvement of opioid receptors in morphine-induced antinociception in the nucleus accumbens (NAc) of rats. The hindpaw withdrawal latencies to thermal and mechanical stimulation increased markedly after intra-NAc administration of morphine. The antinociceptive effects induced by morphine were dose-dependently inhibited by intra-NAc administration of the non-selective opioid receptor antagonist naloxone. Furthermore, the morphine-induced antinociception was significantly attenuated by subsequent intra-NAc injection of the MOR antagonist beta-funaltrexamine or the KOR antagonist nor-binaltorphimine, but not the DOR antagonist naltrindole. The results indicate that MOR and KOR, but not DOR are involved in the morphine-induced antinociception in the NAc of rats.

The present study was performed to investigate the alternation of galanin in nociceptive modulation and galanin-like immunoreactivity in the central nervous system of rats after morphine tolerance. The hindpaw withdrawal latencies to both thermal and mechanical stimulation increased significantly after intracerebroventricular injection of 3 nmol of galanin in opioid-naive rats. The antinociceptive effect induced by galanin was attenuated remarkably at the same dose in morphine-tolerant rats. Furthermore, an up-regulation of galanin-like immunoreactivity in the arcuate nucleus of hypothalamus of morphine-tolerant rat was observed by immunohistochemical methods, whereas no significant changes were detected in periaqueductal gray. The present study demonstrated that there are alternations in both galanin-induced antinociception and galanin-like immunoreactivity in the brain of rat after morphine tolerance. The results suggest an involvement of galanin in the central nervous system in morphine tolerance.

Neuropeptide Y (NPY) plays an important role in pain modulation at different levels in the central nervous system. In the brain, NPY and NPY receptors distribute abundantly in the arcuate nucleus of hypothalamus (ARC), a structure involved in pain processing. The present study was undertaken to investigate the role of NPY in nociceptive modulation in the ARC of intact rats and rats with carrageenan-induced inflammation. Intra-ARC administration of NPY induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to thermal and mechanical stimulation in intact rats, which was attenuated by the Y1 receptor antagonist NPY28-36. Intra-ARC administration of NPY also induced dose-dependent increases in HWLs to noxious stimulation in rats with inflammation. Furthermore, intra-ARC injection of either the antiserum against NPY or NPY28-36 induced decreases in HWLs in rats with inflammation, while both of them produced no effects in intact ones. Additionally, there were marked increases of Y1 receptor in the bilateral ARC of rats with inflammation tested by immunohistochemistry, while no significant changes of NPY were observed, implicating that the increased Y1 receptor has an important effect in the NPY-induced antinociception. We also found that intra-ARC injection of Y2 receptor agonist NPY3-36 produced no significant antinociception in either intact rats or rats with inflammation. Together, we demonstrate that NPY exerts an antinociceptive effect in the ARC of intact rats and rats with inflammation. Both Y1 receptor and endogenous released NPY in the ARC are involved in the nociceptive modulation during inflammation.

The present study was performed to explore the involvement of opioid receptors in the calcitonin gene-related peptide 8-37 (CGRP8-37, an antagonist of CGRP receptor)-induced inhibition of the activity of wide-dynamic-range (WDR) neurons in the spinal dorsal horn of rats. Extracellular recording was performed with a multibarrelled glass micropipette, and the chemicals were delivered by micro-iontophoresis. The discharge frequency of WDR neurons was evoked by subcutaneous electrical stimulation applied to the ipsilateral hindpaw. Iontophoretic application of CGRP8-37 by an ejection current of 160 nA induced significant inhibition of the discharge frequency of WDR neurons. The inhibitory effect of CGRP8-37 on the activity of WDR neurons was attenuated by later iontophoretic application of the opioid antagonist naloxone. Furthermore, the effect of CGRP8-37 was attenuated by either iontophoretic application of the kappa-receptor antagonist nor-binaltorphimine (nor-BNI) or the mu-receptor antagonist β-funaltrexamine (β-FNA) but not by the delta-receptor antagonist naltrindole. The results indicate that kappa- and mu-opioid receptors on the membrane of WDR neurons are involved in the modulation of CGRP8-37-induced antinociception in dorsal horn of the spinal cord in rats. © 2004 Wiley-Liss, Inc.

Corticotropin-releasing factor (CRF) plays important physiological functions in the central nervous system. The present study was performed to investigate the role of CRF and CRF receptor in nociceptive modulation in the central nucleus of amygdala (CeA) of rats. The hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation increased significantly after intra-CeA administration of 0.1 and 0.01 nmol of CRF, but not 0.001 nmol, indicating that CRF induces antinociceptive effects in the CeA of rats. The antinociceptive effect of CRF was attenuated by intra-CeA administration of 0.1 nmol a-hCRF9–41, a selective CRF receptor antagonist, suggesting that the CRF-induced antinociception is mediated by the CRF receptors in the CeA. Furthermore, the HWL to both thermal and mechanical stimulation decreased significantly after intra-CeA administration of a-hCRF9–41alone, suggesting an involvement of endogenous CRF in the CeA in nociceptive modulation. The present study demonstrated that both exogenous and endogenous CRF plays an antinociceptive effect in the CeA, the effect is mediated by CRF receptor. D 2003 Elsevier B.V. All rights reserved. Theme: Sensory system Topic: Pain modulation: pharmacology

In the arcuate nucleus of hypothalamus (ARC), galaninergic fibers form synaptic contacts with proopiomelanocortin neurons, which are involved in pain modulation. The present study assessed the role of exogenous and endogenous galanin in the modulation of nociception in the ARC of rats. The hindpaw withdrawal latency (HWL) to thermal and mechanical stimulation was assessed by the hot-plate test and the Randall Selitto Test. Intra-ARC injection of galanin dose-dependently increased the HWLs in intact rats, indicating an antinociceptive role of exogenous galanin in the ARC. The antinociceptive effect of galanin was blocked by following intra-ARC injection of galantide, a putative galanin receptor antagonist, suggesting that the antinociceptive effect of galanin is mediated by galanin receptors. Moreover, intra-ARC injection of galanin increased the HWL in rats with inflammation. Intra-ARC administration of galantide alone reduced the HWLs in rats with inflammation, while there were no influences of galantide on the HWL in intact rats. Taken together, the results show that galanin has an antinociceptive role in the ARC of intact rats and rats with inflammation.