Naloxone-induced Morphine Withdrawal Research Articles

NMDA Receptors in the Rat Paraventricular Thalamic Nucleus Reduce the Naloxone-induced Morphine Withdrawal.

NMDA receptors have a significant role in the development of opioid physical dependence. Evidence demonstrated that a drug of abuse enhances neuronal excitability in the Paraventricular Nucleus (PVT). The current research studied whether blocking NMDA receptors through the administration of MK801 in the PVT nucleus could affect the development of Morphine (Mor) dependence and hence the behavioral indices induced by morphine withdrawal in rats. Male Wistar rats weighing 250-300 g were used. For induction of drug dependence, we injected Mor subcutaneously (s.c.) (6, 16, 26, 36, 46, 56, and 66 mg/kg, 2 ml/kg) at an interval of 24 hours for 7 days. Animals were divided into two groups in which the NMDA receptor antagonist, MK801 (20 mM in 0.1 ml), or its vehicle were applied into the PVT nucleus for 7 days before each Mor administration. On day 8, after injection of naloxone (Nal, 2.5 mg/kg, i.p.), withdrawal behaviors were checked for 25 min. The current results demonstrated that the blockade of the NMDA receptor in the PVT nucleus significantly increased withdrawal behaviors provoked by the application of Nal in morphinedependent (Mor-d) rats. We concluded that the NMDA receptor in the PVT nucleus changes the development of Mor dependence.

The effect of acute topiramate administration on morphine withdrawal syndrome and brain-derived neurotrophic factor in central nervous system

ABSTRACT Objectives Nucleus accumbens plays an important role in opioid addiction. Topiramate, increases postsynaptic gamma-aminobutyric acid receptor activity and antagonizes glutamatergic activity. Brain-derived neurotrophic factor (BDNF), which plays a key role in synaptic plasticity, is produced from proBDNF. The aim of this study is to investigate the effects of 100 µM topiramate applied into the lateral ventricle or nucleus accumbens on naloxone-induced morphine withdrawal and the BDNF/proBDNF ratio in the frontal cortex. Methods In the study, 36 adult male Wistar rats weighing 250–350 g were used. Morphine dependence was created with morphine pellets following guide cannula implantations. Withdrawal findings were evaluated in naloxone-induced morphine withdrawal syndrome following topiramate administration, and locomotor activity measurements were performed simultaneously. The brains of sacrificed animals were removed for determination of BDNF/proBDNF ratio. Results Topiramate administered by either route significantly suppressed the number of jumps in morphine withdrawal. Topiramate applied into the nucleus accumbens significantly reduced stereotypical behavior in morphine withdrawal, but did not cause any changes in other locomotor activity behaviors. Topiramate applied into the lateral ventricle significantly decreased the BDNF/proBDNF ratio, whereas administered into the nucleus accumbens significantly increased this ratio. Conclusion The findings of this study indicate that topiramate administered into the lateral ventricle and nucleus accumbens reduces naloxone-induced morphine withdrawal symptoms, stereotypical locomotor activity, and changes the BDNF/proBDNF ratio.

Thymol Nanopolymer Synthesis and Its Effects on Morphine Withdrawal Syndrome in Comparison With Clonidine in Rats.

The drug delivery system is valuable in the treatment of the disease. A nanopolymer as a thymol and Thymbra spicata release system was synthesized and its effects on morphine withdrawal syndrome in comparison with clonidine in rats were studied. The nanopolymer was characterized by different methods, namely, IR, HNMR, CNMR, GPC, DLS, and AFM. Thymol in T. spicata extract was assessed. The loading and release rate of thymol and T. spicata extract on the nanopolymer were evaluated by HPLC. The median lethal dose (LD50) of the T. spicata extract, thymol, extract nanopolymer, and thymol nanopolymer was studied. The frequency of jumping, rearing, and teeth chattering in naloxone-induced morphine withdrawal syndrome was studied. Synthesized nanopolymer was desirable as a carrier for the drug. The loaded amount of extract and thymol on nanopolymer was estimated 55 ± 3.2% and 48 ± 2.6% and the drug released was 71 and 68%, respectively. LD50 of the T. spicata extract, thymol, extract nanopolymer, and thymol nanopolymer was 975, 580, 1,250, and 650 mg/kg, respectively. This study showed that thymol nanopolymer was more effective than clonidine to reduce the frequency of morphine withdrawal symptoms. Our results suggest that T. spicata extract, thymol, extract nanopolymer, and thymol nanopolymer are mighty in reducing the narcotic withdrawal signs. The mechanism of action and therapeutic potential is maybe similar to clonidine.

Linagliptin, a Selective Dipeptidyl Peptidase-4 Inhibitor, Reduces Physical and Behavioral Effects of Morphine Withdrawal.

(1) Background: Recent data indicate that receptors for GLP-1 peptide are involved in the activity of the mesolimbic system. Thus, the purpose of the present study was to examine the effect of the selective dipeptidyl peptidase-4 (DPP-4) inhibitor, linagliptin, on morphine dependence in mice. (2) Methods: Morphine dependence in mice was obtained by administration of increasing doses of morphine for eight consecutive days, twice a day. On the 9th day of the experiment, the naloxone-induced (2 mg/kg, ip) morphine withdrawal signs (jumping) were assessed. Moreover, behavioral effects of short-term (60 h after morphine discontinuation) and long-term (14 days after morphine discontinuation) morphine withdrawal were observed. In terms of behavioral effects, the depressive effect in the forced swim test and anxiety in the elevated plus maze test were investigated. Locomotor activity of mice was also studied. (3) Results: The administration of linagliptin (10 and 20 mg/kg, ip) for 8 consecutive days before morphine injections significantly diminished the number of naloxone-induced morphine withdrawal signs (jumping) in mice. In addition, the cessation of morphine administration induced depressive behavior in mice which were observed during short- and long-term morphine withdrawal. Linagliptin administered during morphine withdrawal significantly reduced the depressive behavior in studied mice. Furthermore, the short-term morphine withdrawal evoked anxiety which also was reduced by linagliptin in mice. (4) Conclusions: The present study reveals that GLP-1 receptors are involved in morphine dependence. What is more, linagliptin might be a valuable drug in attenuating the physical symptoms of morphine dependence. It might be also a useful drug in reducing emotional disturbances which may develop during the morphine withdrawal period.

The Effects of Hydroalcoholic Extract of Satureja khuzestanica on Naloxone-Induced Morphine Withdrawal Symptoms in Wistar Rats

Introduction: The inflammatory system, oxidative stress, and cholinergic pathways are some important factors in long-term opioid dependence withdrawal. The adverse effects of some adjunctive medications on withdrawal symptoms treatment limit their clinical efficiency and finding compounds with inhibitory effects on opioid dependence can be helpful. The antioxidant, antinociceptive, and anti-inflammatory properties of Satureja khuzestanica extract (SKE) have been noted. Additionally, it could reduce morphine analgesic tolerance. Given that no study has assessed the effect of SKE on morphine-induced withdrawal symptoms, we have tried to investigate it in this study. Methods: Male Wistar rats were used in this study. The rats were treated with morphine for 7 days. The control group received saline, and intervention groups received SKE intragastrically by gavage (100 mg/kg, 50 mg/kg, and 25 mg/kg) 15 minutes before morphine injections. Five hours after the last injection, naloxone was used and withdrawal symptoms were assessed for one hour. SPSS software version 16.0 was used for statistical analysis, and P≤0.05 was considered statistically significant. Results: SKE diminished weight loss and jumping (P≤0.001). It decreased grooming behaviors and cramps (P≤0.01). SKE was found to decrease morphine withdrawal symptoms and 100 mg/kg was the most effective dose. Additionally, different doses of SKE were able to eliminate ptosis, diarrhea, and teeth chattering in animals; however, different doses had different effects on withdrawal symptoms. Conclusion: Naloxone significantly increased the frequency of jumping, cramps, weight loss, grooming, and induced ptosis, diarrhea, and teeth chattering. Treatment with SKE can significantly reduce morphine withdrawal symptoms. This can be done through its ability to decrease inflammation.

PKC inhibitor reversed the suppressive effect of orexin-A on IPSCs of locus coeruleus neurons in naloxone-induced morphine withdrawal

The locus coeruleus (LC) as a target of addictive drugs receives a dense projection of orexinergic fibres from the lateral hypothalamus (LH) and is accordingly a candidate site for the expression of the somatic aspects of morphine withdrawal. Recently it has been shown that the inhibitory synaptic currents of LC neurons decrease partly through orexin type 1 receptors in the context of naloxone-induced morphine withdrawal; however, its cellular mechanism remains unclear. In this study, whole-cell patch clamp recordings of LC neurons in brainstem slices were used to investigate the impact of protein kinase C (PKC) on GABAergic inhibitory post-synaptic currents (IPSCs) in the context of naloxone-induced morphine withdrawal. Male Wistar rats (P14-P21) received morphine (20mg/kg, i.p.) daily for 7 consecutive days to induce morphine dependency. Our results showed that the application of PKC inhibitor (Go 6983; 1µM) alone did not decrease the probability of GABA release in the LC neurons of the morphine-treated rats in the presence of naloxone. Although, Go 6983 reversed the reduction of the amplitude of evoked IPSCs (eIPSCs) and spontaneous IPSCs (sIPSCs) frequency induced by orexin-A but did not change the sIPSCs amplitude. These results indicate that the suppressive effect of orexin-A on IPSCs is probably reversed by PKC inhibitor in the LC neurons of morphine-treated rats in the context of naloxone withdrawal.

Conditioned aversive memory associated with morphine withdrawal increases brain-derived neurotrophic factor in dentate gyrus and basolateral amygdala.

Morphine has been shown to increase the expression of brain-derived neurotrophic factor (BDNF) in the brain. However, little is known about the effect of conditioned naloxone-precipitated morphine withdrawal on BDNF and its precursor protein, proBDNF. We used the conditioned place aversion (CPA) paradigm to evaluate the role of corticotropin-releasing factor (CRF)/CRF1 receptor signaling on the BDNF expression and corticosterone plasma levels after CPA expression and extinction. Male mice were rendered dependent on morphine and injected acutely with naloxone before paired to confinement in a naloxone-associated compartment. The expression of BDNF and proBDNF in the dentate gyrus (DG) and basolateral amygdala (BLA) was measured in parallel with the corticosterone plasma levels with and without CRF1 receptor blockade. Mice subjected to conditioned naloxone-induced morphine withdrawal showed an increased expression of BDNF (in DG and BLA) in parallel with an enhancement of corticosterone plasma levels. These results demonstrated that BDNF expression together with the increased activity of hypothalamic-pituitary-adrenocortical (HPA) axis are critical to the acquisition of aversive memory. However, we have observed a decrease in corticosterone plasma levels and BDNF expression after CPA extinction reaffirming the importance of BDNF in the maintenance of aversive memory. In addition, the pre-treatment with the CRF1 receptor antagonist CP-154526 before naloxone conditioning session impaired morphine withdrawal-induced aversive memory acquisition, the increased corticosterone plasma levels, and the expression of BDNF observed after CPA expression in the DG and BLA. Altogether, present results are suggesting a clear connection between HPA axis and BDNF in the formation and extinction of aversive memory.

Morphine Dependence is Attenuated by Treatment of 3,4,5-Trimethoxy Cinnamic Acid in Mice and Rats.

The effect of 3, 4, 5-trimethoxy cinnamic acid (TMCA) against morphine-induced dependence in mice and rats was investigated. Mice were pretreated with TMCA and then morphine was injected intraperitoneally; whereas rats were treated with TMCA (i.p.) and infused with morphine into the lateral ventricle of brain. Naloxone-induced morphine withdrawal syndrome and conditioned place preference test were performed. Moreover, western blotting and immunohistochemistry were used to measure protein expressions. Number of naloxone-precipitated jumps and conditioned place preference score in mice were attenuated by TMCA. Likewise, TMCA attenuated morphine dependent behavioral patterns such as diarrhea, grooming, penis licking, rearing, teeth chattering, and vocalization in rats. Moreover, the expression levels of pNR1and pERK in the frontal cortex of mice and cultured cortical neurons were diminished by TMCA. In the striatum, pERK expression was attenuated despite unaltered expression of pNR1 and NR1. Interestingly, morphine-induced elevations of FosB/ΔFosB+ cells were suppressed by TMCA (50, 100mg/kg) in the nucleus accumbens sub-shell region of mice. In conclusion, TMCA could be considered as potential therapeutic agent against morphine-induced dependence.

Decrease of inhibitory synaptic currents of locus coeruleus neurons via orexin type 1 receptors in the context of naloxone-induced morphine withdrawal.

Acute opioid withdrawal syndrome is a series of neurological symptoms caused by the abrupt cessation of the chronic administration of opioids such as morphine. The locus coeruleus (LC) in the brain stem receives a dense projection of orexinergic fibers from the hypothalamus and is a candidate site for the expression of the somatic aspects of morphine withdrawal. Previous studies have shown that orexin-A contributes to the behavioral symptoms of naloxone-induced morphine withdrawal, partly by reducing the activity of GABAergic neurons, suggesting that orexin-A may negatively modulate fast GABAergic neurotransmission during morphine withdrawal. We used whole-cell patch-clamp recordings of LC neurons in brainstem slices to investigate the effect of orexin-A on bicuculline-sensitive GABAergic inhibitory postsynaptic currents (IPSCs) during naloxone-induced morphine withdrawal. Male Wistar rats (P14-P21) were given morphine (20mg/kg, i.p.) daily for seven consecutive days to create dependency on the drug. The application of naloxone (1µM) to brain slices of morphine-treated rats reduced the amplitude of evoked IPSCs (eIPSCs) as well as spontaneous IPSCs (sIPSCs) frequency but did not change sIPSCs amplitude. Orexin-A (100nM) significantly enhanced the suppressive effect of naloxone on eIPSCs amplitude and sIPSCs frequency but had no effect on the presence of the orexin type 1 receptor (OX1R) antagonist, SB-334867. Orexin-A alone had no significant effect on eIPSCs and sIPSCs in the absence of naloxone. In summary, our results show that orexin-A, via OX1R, potentiates the suppressive effect of naloxone on GABAergic IPSCs of LC neurons in morphine-treated rats. We conclude that orexins may have a critical role in regulating GABAergic neurotransmission to LC neurons during naloxone-induced morphine withdrawal.

The involvement of CRF1 receptor within the basolateral amygdala and dentate gyrus in the naloxone-induced conditioned place aversion in morphine-dependent mice

Drug withdrawal-associated aversive memories trigger relapse to drug-seeking behavior. Corticotrophin-releasing factor (CRF) is an important mediator of the reinforcing properties of drugs of abuse. However, the involvement of CRF1 receptor (CRF1R) in aversive memory induced by opiate withdrawal has yet to be elucidated. We used the conditioned-place aversion (CPA) paradigm to evaluate the role of CRF1R on opiate withdrawal memory acquisition, along with plasticity-related processes that occur after CPA within the basolateral amygdala (BLA) and dentate gyrus (DG). Male mice were rendered dependent on morphine and injected acutely with naloxone before paired to confinement in a naloxone-associated compartment. The CPA scores as well as the number of TH-positive neurons (in the NTS-A2 noradrenergic cell group), and the expression of the transcription factors Arc and pCREB (in the BLA and DG) were measured with and without CRF1R blockade.Mice subjected to conditioned naloxone-induced morphine withdrawal robustly expressed CPA. Pre-treatment with the selective CRF1R antagonist CP-154,526 before naloxone conditioning session impaired morphine withdrawal-induced aversive memory acquisition. CP-154,526 also antagonized the enhanced number of TH-positive neurons in the NTS-A2 that was seen after CPA. Increased Arc expression and Arc-pCREB co-localization were seen in the BLA after CPA, which was not modified by CP-154,526. In the DG, CPA was accompanied by a decrease of Arc expression and no changes in Arc-pCREB co-localization, whereas pre-treatment with CP-154,526 induced an increase in both parameters. These results indicate that CRF-CRF1R pathway could be a critical factor governing opiate withdrawal memory storage and retrieval and might suggest a role for TH-NA pathway in the effects of withdrawal on memory. Our results might indicate that the blockade of CRF1R could represent a promising pharmacological treatment strategy approach for the attenuation of the relapse to drug-seeking/taking behavior triggered by opiate withdrawal-associated aversive memories.

Effects of Injection of Gamma-Aminobutyric Acid Agonists into the Nucleus Accumbens on Naloxone-Induced Morphine Withdrawal

Aims: This study was to investigate the effects of local administration of gamma-aminobutyric acid (GABA) agonists into the nucleus accumbens (NAc) on naloxone-induced morphine withdrawal symptoms. Methods: Bilateral guide cannulas were stereotaxically implanted in the shell or core regions of the NAc of Sprague-Dawley rats. After a recovery period, 3 morphine pellets, each consisting of 75 mg morphine base, were placed subcutaneously on the first and third days of the study with the rats under mild ether anaesthesia. The GABA agonists, baclofen hydrochloride or muscimol hydrobromide, were injected into the NAc, and morphine withdrawal was induced by naloxone on the fifth day. Results: Administration of baclofen to the shell or core regions of the NAc of Sprague-Dawley rats led to statistically significant decreases in both behavioural and locomotor activity parameters during the morphine withdrawal period, compared to the control group. However, there were no statistically significant changes in locomotor activity or withdrawal behavioural parameters, with the exception of wet dog shakes, between control and muscimol-treated groups. Conclusion: These findings show that GABAergic conduction in the NAc is effective on the morphine withdrawal symptoms, and that both the shell and core regions of the NAc are associated with this effect.

Morphine dependence is attenuated by red ginseng extract and ginsenosides Rh2, Rg3, and compound K

BackgroundRed ginseng and ginsenosides have shown plethoric effects against various ailments. However, little is known regarding the effect of red ginseng on morphine-induced dependence and tolerance. We therefore investigated the effect of red ginseng extract (RGE) and biotransformed ginsenosides Rh2, Rg3, and compound K on morphine-induced dependence in mice and rats. MethodsWhile mice were pretreated with RGE and then morphine was injected intraperitoneally, rats were infused with ginsenosides and morphine intracranially for 7 days. Naloxone-induced morphine withdrawal syndrome was estimated and conditioned place preference test was performed for physical and psychological dependence, respectively. Western blotting was used to measure protein expressions. ResultsWhereas RGE inhibited the number of naloxone-precipitated jumps and reduced conditioned place preference score, it restored the level of glutathione in mice. Likewise, ginsenosides Rh2, Rg3, and compound K attenuated morphine-dependent behavioral patterns such as teeth chattering, grooming, wet-dog shake, and escape behavior in rats. Moreover, activated N-methyl-D-aspartate acid receptor subunit 1 and extracellular signal-regulated kinase in the frontal cortex of rats, and cultured cortical neurons from mice were downregulated by ginsenosides Rh2, Rg3, and compound K despite their differential effects. ConclusionRGE and biotransformed ginsenosides could be considered as potential therapeutic agents against morphine-induced dependence.

Evaluation of 3, 4, 5-Trimethoxycinnamic Acid Derivatives as Potential Antinarcotic and Anxiolytic Agents

To determine the biological activity of natural products for antinarcotics and anxiolytics, Polygala tenuifolia was selected as an interesting natural resources. 3, 4, 5-Trimethoxycinnamic acid (TMCA) was an ingredient from Polygala tenuifolia and a series of TMCA derivatives was prepared and evaluated for antinarcotic effects on morphine dependence in mice and binding affinities on serotonergic receptors. The key synthetic strategies involve generation of ketones, esters through condensation reaction, and amides via coupling reaction using HOBt/EDCI system in high yield. We found that the naloxone-induced morphine withdrawal syndrome was significantly suppressed by new synthetic TMCA derivatives (20 mg/kg/day). Most of TMCA derivatives were found to have high affinity to 5-HT1A receptor. Interestingly, the TMCA derivatives showed inhibitory action to the anxiety which was induced by restraint stress in mice. These results suggested that new synthetic TMCA derivatives have a potential antinarcotic and anxiolytic effect in mice.

The blockade of GABAA receptors attenuates the inhibitory effect of orexin type 1 receptors antagonist on morphine withdrawal syndrome in rats

The aim of present study was to investigate the involvement of orexin-A neuropeptide in naloxone-induced morphine withdrawal syndrome via modulating neurons bearing GABAA receptors. The locus coeruleus (LC) is a sensitive site for expression of the somatic aspects of morphine withdrawal. Intra-LC microinjection of GABAA receptor agonist attenuates morphine withdrawal signs in rats. Here we studied the influence of LC orexin type 1 receptors blockade by SB-334867 in presence of bicuculline, a GABAA receptor antagonist, on naloxone-induced morphine withdrawal syndrome.Adult male Wistar rats, weighing 250–300g, were rendered dependent on morphine by subcutaneous (s.c.) injection of increasing morphine doses (6, 16, 26, 36, 46, 56 and 66mg/kg, 2ml/kg) at set intervals of 24h for 7 days. On 8th day, naloxone (3mg/kg, s.c.) was injected and the somatic signs of morphine withdrawal were evaluated. Intra-LC microinjections (0.2μl) of either bicuculline (15μM) or SB-334867 (3mM) or a combination of both chemicals were done immediately before naloxone injection. Intra-LC microinjection of bicuculline (15μM) had no significant effect on morphine withdrawal signs, whereas intra-LC microinjection of SB-334867 considerably attenuated morphine withdrawal signs. However, the effect of SB-334867 in attenuating naloxone-induced morphine withdrawal signs was blocked in presence of bicuculline. This finding, for the first time, indicated that orexin-A may participate in expression of naloxone-induced morphine withdrawal syndrome partly through decreasing the activity of neurons bearing GABAA receptors.

Expression of heat shock protein 27 and troponin T and troponin I after naloxone-precipitated morphine withdrawal

Heat shock protein (Hsp27) renders cardioprotection from stress situations but little is known about its role in myofilaments. In this study we have evaluated the relationship between Hsp27 and troponin response after naloxone-induced morphine withdrawal. Rats were treated with two morphine (75mg) pellets during six days. Precipitated withdrawal was induced by naloxone on day seven. Hsp27 expression, Hsp27 phosphorylated at serine 82 (Ser82), cardiac troponin T (cTnT), cardiac troponin I (cTnI) and µ-calpain were evaluated by immunoblotting in left ventricle. Hsp, cTnT and cTnI was also evaluated by immunofluorescence procedure. Our results show that enhancement in Hsp27 expression and phosphorylation induced by naloxone-precipitated morphine withdrawal occurs with concomitant increases of cTnT and µ-calpain expression, whereas cTnI was decreased. We also observed co-localization of Hsp27 with cTnT in cardiac tissues. These findings provide new information into the possible role of Hsp27 in the protection of cTnT degradation by µ-calpain (a protease mediating proteolysis of cTnT and cTnI) after morphine withdrawal.

Spironolactone decreases the somatic signs of opiate withdrawal by blocking the mineralocorticoid receptors (MR)

Pharmacological evidence has accumulated showing that glucocorticoids and glucocorticoid receptor (GR) facilitate several responses to different drugs of abuse. Recent findings have attributed a prominent role to the mineralocorticoid receptor (MR) in modulating behavior during the addictive process. The purpose of this study was to investigate the effects of MR blockade on: brain stress system responses to naloxone-induced morphine withdrawal, the somatic signs of abstinence; the effects of morphine withdrawal on noradrenaline (NA) turnover in the paraventricular nucleus (PVN), c-Fos expression and tyrosine hydroxylase (TH) phosphorylated at Ser31 levels in the nucleus tractus solitarius noradrenergic cell group (NTS-A2); and finally, hypothalamus-pituitary-adrenocortical (HPA) axis activity. The role of MR signaling was assessed with i.p. pretreatment with the MR antagonist, spironolactone. Rats were implanted with two morphine (or placebo) pellets. Six days later rats were pretreated with spironolactone or vehicle 30min before naloxone. The physical signs of abstinence, NA turnover, TH activation, c-Fos expression and the HPA axis activity were measured using HPLC, immunoblotting and RIA. Spironolactone attenuated the somatic signs of withdrawal that were seen after naloxone administration to chronic morphine treated animals. On the other hand, pretreatment with spironolactone resulted in no significant modification of the increased NA turnover, TH activation, c-Fos expression or HPA axis activity that occurred during morphine withdrawal. These results suggest that somatic signs of opiate withdrawal are modulated by MR signaling. However, blockade of MR did not significantly alter the brain stress system response to morphine withdrawal.

Blockade of orexin type-1 receptors in locus coeruleus nucleus attenuates the development of morphine dependency in rats

The aim of this study was to evaluate the effects of orexin type-1 receptor (OX1R) antagonism in locus coeruleus (LC) nucleus on the development of morphine physical dependence in rats. Animals were rendered dependent on morphine by subcutaneous (s.c.) administration of morphine sulfate (6, 16, 26, 36, 46, 56 and 66mg/kg, 2ml/kg) at set intervals of 24h for 7 days. Immediately before each morphine administration, the animals received intra-LC administration of SB-334867 (3mM, 0.2μl), a selective orexin type-1 receptor antagonist. On day 8, naloxone (3mg/kg, i.p.) was injected and physical dependence was evaluated for 30min. Our results showed that administration of OX1R antagonist before each morphine injection significantly decreased somatic signs of naloxone-induced morphine withdrawal syndrome, including defecation, wet-dog shake, diarrhea, jumping, scratching, and teeth chattering. These results suggest that the activation of OX1R in LC nucleus might be involved in the development of morphine dependency.

Role of low-intensity laser therapy on naloxone-precipitated morphine withdrawal signs in mice: is nitric oxide a possible candidate mediator?

In the present study, the potential involvement of nitric oxide (NO) system in attenuating effects of low-intensity laser therapy (LILT) on naloxone-induced morphine withdrawal signs was evaluated. A hundred mice were rendered morphine-dependent using three escalating doses of morphine sulfate during three consecutive days. After the last dose on day 4, animals were given naloxone HCl (2 mg/kg s.c) to induce withdrawal signs. The effects of LILT (12.5 J/cm(2)) and L-NG-nitroarginine methyl ester (L-NAME) (10, 20, 50, and 100 mg/kg) and their coadministration on escape jump count and stool weight as typical withdrawal signs were assessed. LILT and L-NAME (20, 50, and 100 mg/kg) per se significantly decreased escape jump count and stool weight in morphine-dependent naloxone-treated mice (p < 0.01). Coadministration of LILT and L-NAME (20, 50, and 100 mg/kg) also reduced escape jump and stool weight (p < 0.05) but with no synergetic or additive response. Here, LILT at this fluence may show its maximal effects on NO and therefore no noticeable effects appeared during coadministration use. Moreover, LILT and L-NAME follow the same track of changes in escape jump and stool weight. Conceivably, it seems that LILT acts partly via NO system, but the exact path is still obscure and rather intricate. The precise mechanisms need to be clarified.

Crosstalk between G protein-coupled receptors (GPCRs) and tyrosine kinase receptor (TXR) in the heart after morphine withdrawal

G protein-coupled receptors (GPCRs) comprise a large family of membrane receptors involved in signal transduction. These receptors are linked to a variety of physiological and biological processes such as regulation of neurotransmission, growth, and cell differentiation among others. Some of the effects of GPCRs are known to be mediated by the activation of mitogen-activated extracellular kinase (MAPK) pathways. Cross-talk among various signal pathways plays an important role in activation of intracellular and intranuclear signal transduction cascades. Naloxone-induced morphine withdrawal leads to an up-regulation of adenyl cyclase-mediated signaling, resulting in high expression of protein kinase (PK) A. In addition, there is also an increased expression of extracellular signal regulated kinase (ERK), one member of MAPK. For this reason, the crosstalk between these GPCRs and receptors with tyrosine kinase activity (TKR) can be considered a possible mechanism for adaptive changes that occurs after morphine withdrawal. Morphine withdrawal activates ERK1/2 and phosphorylated tyrosine hydroxylase (TH) at Ser31 in the right and left ventricle. When N-(2-guanidinoethyl)-5-isoquinolinesulfonamide (HA-1004), a PKA inhibitor was infused, the ability of morphine withdrawal to activate ERK, which phosphorylates TH at Ser31, was reduced. The present finding demonstrated that the enhancement of ERK1/2 expression and the phosphorylation state of TH at Ser31 during morphine withdrawal are dependent on PKA and suggest cross-talk between PKA and ERK1/2 transduction pathway mediating morphine withdrawal-induced activation of TH. Increasing understanding of the mechanisms that interconnect the two pathway regulated by GPCRs and TKRs may facilitate the design of new therapeutic strategies.

Pharmacological modulation of geranylgeranyltransferase and farnesyltransferase attenuates opioid withdrawal in vivo and in vitro

Geranylgeranyltransferase and farnesyltransferase I, are noted to mediate a number of signal transduction cascades which are known to be involved in the causation of opioid withdrawal syndrome. GGTI-2133 and FTI-276 are selective modulators of geranylgeranyltransferase and farnesyltransferase subtype 1 respectively. Therefore, the present study investigated the effect of GGTI-2133 and FTI-276 on propagation of morphine dependence and resultant withdrawal signs in vivo, in sub-chronic morphine mouse model, and in vitro, in isolated rat ileum. Morphine was administered twice daily for 5 days following which a single day 6 injection of naloxone (8 mg/kg, i.p.) precipitated opioid withdrawal syndrome in mice. Withdrawal syndrome was quantitatively assessed in terms of withdrawal severity score and the frequency of jumping, rearing, fore paw licking & circling. Naloxone induced contraction in morphine withdrawn isolated rat ileum was employed as an in vitro model of opioid withdrawal syndrome. An isobolographic study design was employed to assess a potential synergistic activity between GGTI-2133 and FTI-276. GGTI-2133 and FTI-276 dose dependently attenuated naloxone induced morphine withdrawal syndrome both in vivo and in vitro. GGTI-2133 was also observed to exert a synergistic interaction with FTI-276. It is concluded that GGTI-2133 and FTI-276 attenuate the propagation of morphine dependence and reduce withdrawal signs possibly by a geranylgeranyl transferase; farnesyltransferase activation pathway linked mechanisms potentially in an interdependent manner.