Browsing by Subject "Morphine"

Now showing 1 - 9 of 9
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    AMPA Glutamate Receptor Trafficking in Models of Disease
    (2013-08) Miller, Eric
    The signaling between neurons in the brain underlies many crucial processes - from the beating of the heart to remembering that you have a meeting at noon. Indeed, it is proposed that changes in the way neurons communicate with each other form the basis of learning and memory. In this dissertation I will explore the ways in which neurological diseases can affect these integral neuronal functions and explain the changes on a cellular and molecular level. Deficits in AMPAR signaling are found in numerous diseases. I will explore the mechanisms of these deficits using in vitro models of three diseases: opioid-related cognitive deficits and addiction, Alzheimer's disease, and Parkinson's disease-related dementia. Background information is presented in the first chapter. In the second chapter the signaling pathways underlying morphine-induced synaptic deficits are delineated. I found that calcineurin is necessary for both functional and structural deficits in AMPAR signaling, while CaMKII is necessary for only the structural deficits. The role of tau in synaptic deficits cause by soluble Abeta; oligomers, which are found at elevated levels in Alzheimer's disease patients, are probed in the third chapter. I found that treatment with soluble Abeta; oligomers leads to phosphorylation- dependent mislocalization of tau to dendritic spines. Furthermore, treatment with soluble Abeta; oligomers leads to decreases in AMPAR signaling that require calcineurin activity and GluR1 residue S845, much like the mechanisms of AMPAR internalization in neurons treated with morphine. The fourth chapter unveils a novel role of tau and GSK3 in synaptic deficits found in neurons expressing A53T alpha-synuclein. In fact, I discovered that tau is involved in AMPAR signaling deficits found in neurons expressing A53T alpha- synuclein. Furthermore, both mislocalization of tau and synaptic deficits require phosphorylation of tau by GSK3. This dissertation shows that divergent pathways mediate structural and functional plasticity found in neurons exposed to morphine. Also, I show that deficits in AMPAR signaling in both Alzheimer's disease and Parkinson's disease involve tau mislocalization. These findings shed new light on the signaling pathways involved in AMPAR signaling deficits found in neurological diseases and provide new therapeutic targets for pharmacological interventions.
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    Chronic morphine treatment-modulated trafficking of AMPA receptors: a potential mechanism for drug addiction
    (2012-08) Kam, Yuet Fong
    Morphine is the benchmark analgesic for treating chronic pain. However, its clinical uses are hindered by its highly addictive nature, as chronic treatment with the drug will produce physical and psychological dependence upon the cessation of use. Drug craving is the main driving force for relapse after prolonged periods of abstinence, and represents an enormous challenge for the treatment of drug addiction. Since addiction is a long-term behavioral alteration, it is believed that addictive drugs produce reorganization of specific neural circuits and adjustment of synaptic strength. The underlying mechanisms of these neural adaptations may represent a promising target for prevention and/or treatment of addiction, but the detailed mechanisms of these processes remain unclear. Therefore, the main goals of this work are to delineate signaling pathways controlling morphine-induced neural adaptations and investigate their functional role in opiate addictive behaviors. Alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors are postsynaptic glutamate receptors, and are responsible for mediating most excitatory synaptic transmission under normal conditions. More importantly, the dynamic localization of AMPA receptors plays a critical role for modifying synaptic strength and synaptic morphology. Here, I hypothesized that regulation of AMPA receptor trafficking by morphine treatment underlies the drug-induced neural modulation implicated in the addiction process. Hence, the first part of this dissertation research examined whether and how chronic exposure to morphine modulates trafficking of surface GluR1 (a subunit of AMPA receptors) in primary hippocampal neurons. Using live-cell imaging techniques together with biochemical studies, I demonstrated that chronic exposure to morphine induced a significant loss of synaptic and extrasynaptic GluR1 by internalization. In mechanistic studies, I found that the GluR1 internalization was attributed to dephosphorylation of the receptor subunit at Ser845 following morphine treatment, but it did not result from altered neural network or NMDA receptor activation. Moreover, dephosphorylation of GluR1 at Ser845 was found to require morphine-evoked calcineurin activation. Therefore, calcineurin-dependent dephosphorylation of AMPA receptor and subsequent AMPA receptor internalization provides a novel mechanism for opioid-induced neural adaptations. The second part of this dissertation research attempted to link morphine's effects on GluR1 phosphorylation and endocytosis to addictive behavior, especially formation of memory for the environmental context of the drug experience, because recall of this memory by encountering the drug-paired cues triggers relapse to drug seeking. In this approach, a mutant mouse line was used, in which GluR1 at Ser845 was mutated to Ala (S845A) leading to an absence of morphine-induced GluR1 endocytosis. A behavioral test, conditioned place preference (CPP), was carried out to assess the ability of morphine to produce a positive association with environmental cues. I found that S845A mice were significantly slower to acquire morphine-induced CPP when compared to wild types (WT). This decreased sensitivity to morphine CPP in mutants was neither related to contextual memory deficits or abnormal locomotor activity, as there was no difference between WT and S845A mice in the contextual memory acquisition in the Morris water maze test or locomotion with or without morphine injection. To examine the persistence of morphine-associated contextual memory in the mutant mice, I also performed extinction tests on mice conditioned with 10 mg/kg morphine for four sessions, by which both WT and S845A mice exhibited similar CPP responses. Interestingly, a prolonged extinction was observed in S845A mutant mice, suggesting the S845A mutation either impaired the learning of the new conditioning or prolonged the retention of the old conditioning. Nevertheless, these results suggest that an alteration in GluR1 phosphorylation at Ser845 and subsequent receptor endocytosis/insertion are involved in acquisition and extinction of morphine CPP. Altogether, the present findings indicate that calcineurin-mediated GluR1-S845 dephosphorylation is required for morphine-induced internalization of GluR1-containing AMPA receptors, providing a molecular basis for the drug-induced neural modulation. This work also suggests that this regulation of GluR1 phosphorylation and trafficking by morphine is involved in the modulation of the drug-associated contextual memory, which reflects the involvement of AMPA receptor trafficking in the mechanisms underlying opiate-seeking behaviors.
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    Differential regulation of opioid receptors during inflammation
    (2009-07) Satterfield, Catherine Suzanne
    Properties of the opium poppy have been exploited for centuries for the alleviation of pain and to induce euphoria. Classically thought to produce its effects solely in the central nervous system, peripheral opioid analgesic systems are now widely accepted. The activation of these systems leads to a reduction in primary afferent fiber excitability leading to the inhibition of sensory transduction. Opioid receptors function is modulated by a variety of mechanisms. An example of this is enhanced peripheral opioid receptor function following inflammation. The present study examined peripheral opioid receptor regulation in early and late stages of CFA inflammation. Additionally, a new model of UVB of inflammation was characterized. Peripheral MOR receptors are differentially regulated in late and early CFA inflammation. Peripheral MOR is not responsible for attenuated responses of nociceptors to mechanical stimuli 18 hours after CFA inflammation. DAMGO reduced mechanical responsiveness of nociceptors at 72 hours after CFA inflammation in a concentration and antagonist reversible manner indicating that MOR efficacy is enhanced during later stages of CFA inflammation. UVB produced severe but localized inflammation that differed from inflammation produced by CFA. This inflammation sensitizes nociceptor units innervating irradiated skin and results in enhanced peripheral opioid receptor efficacy.
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    Effects of chronic morphine treatment on tumor angiogenesis and growth.
    (2009-06) Koodie, Lisa
    Morphine is one of the most effective analgesics commonly prescribed for the treatment of severe to moderate cancer pain. To date very little is known regarding the effect of long-term morphine treatment on tumor angiogenesis. At this time, the effect of morphine on tumor growth is contradictory and still inconclusive. As solid tumors grow, the formation of a blood supply or angiogenesis is essential. In previous studies, morphine inhibited vascular endothelial growth factor (VEGF) secretion from mice cardiomyocytes and human umbilical vein endothelial cells. VEGF is a highly potent pro-angiogeneic molecule and we therefore hypothesized morphine would also inhibit angiogenesis associated with tumor growth. In the first part of these studies we show that morphine inhibited the hypoxia-induced tumor cell expression of VEGF to significantly reduce tumor cell angiogenesis, and suppress tumor growth in vivo. Additional investigations supported the view that the effect of morphine was not due to a direct effect on tumor cell apoptosis, but instead indirectly through angiogenesis. Tumor, stromal and inflammatory cells within the tumor microenvironment all contribute to a large pool of chemoattractants that increase the recruitment of myeloid cells from peripheral blood circulation into the tumor tissues. These cells mature and differentiate into neutrophils, and macrophages that eventually result in a pro-inflammatory-like environment to support and maintain tumor growth. Considering that morphine is highly immuno-suppressive, we also hypothesized that morphine will inhibit immune cell recruitment and thus angiogenesis. In an in vivo model of cell migration and recruitment we found that morphine inhibited not only CD11b+ progenitors of inflammatory cells but also the recruitment of Tie2+/CD14+ endothelial cell precursors known to actively participate in vessel formation to tumor sites. These studies have allowed us to further understand the effects of a potent analgesic such as morphine in cancer growth. Our data support the use of morphine for pain associated with cancer. Our results support the view that morphine may not cause any further detriment in the cancer patients' quality of life but further suppress angiogenesis associated with tumor growth and progression.
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    The Effects of Morphine Tolerance on the PI3K/AKT Intracellular Signaling Pathway During Acute and Chronic Pain in the Central and Peripheral Nervous System of Mice
    (2019-09) Okerman, Travis
    The management of chronic pain with opioids can cause opioid-induced analgesic tolerance and hyperalgesia, complicating clinical pain-management treatments. Mu opioid receptors (MOPs) are inhibitory G-protein coupled receptors. MOPs have been studied for years; however, the intracellular signaling pathways triggered by their activation are not well known. Research presented here sought to determine if opioid induced tolerance is linked to a decreased activity in the PI3K/AKT intracellular signaling pathway. To assess gene expression within this pathway and cGMP nucleotide levels, C57Bl/6 wild type male mice were divided into saline , morphine tolerant (MT), and morphine tolerant with spinal nerve ligation (MT+SNL) groups. Injections were given subcutaneously twice a day for a total of five days. MT mice without SNL developed opioid induced tolerance by day 3 and opioid induced hyperalgesia by day 5. MT+SNL mice had lower TPWL responses for the ipsi (injured) side compared to their contra (uninjured) side. Brainstem, spinal cord, dorsal root ganglia, and sciatic nerves were harvested from mice on day 6 of the behavior testing and were used for qPCR gene expression analysis. Genes chosen for qPCR analysis were Akt1, Akt2, Akt3, Pik3cg (splice variants v1-v3), Pten, Abcc8, Abcc9, Kcnj11 (splice variants v1 and v2), Kcnj8, Oprm1, Jnk3, and nNos1. There were few significant gene expression changes of the PI3K/AKT intracellular signaling pathway for MT mice compared to saline mice in both the central and peripheral nervous system. The addition of the MT+SNL model saw larger gene expression increases or decreases in almost all genes, with no clear trend for gene expression changes both between and within the central and peripheral nervous system tissues. Additionally, there was decreased gene expression in all genes except Pten in the MT+SNL spinal cord. The daily administration of PI3K/AKT pathway inhibitors, thalidomide, SP600125, and quercetin to MT+SNL mice attenuated the development of morphine tolerance, suggesting PI3K/AKT pathway activity is positively correlated with morphine tolerance. Thus, our initial hypothesis that stated PI3K/AKT pathway downregulation may contribute to morphine tolerance was not supported by our data collected from this study. Overall, the PI3K/AKT intracellular signaling pathway is a potential target for reducing the development of morphine tolerance. Continued research into this pathway, including further protein analysis and studies utilizing knockout mice, will advance understanding of morphine tolerance and potentially contribute to the development of new analgesic drug therapies.
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    Morphine inhibition of macrophage phagocytosis and bactericidal functions.
    (2011-01) Ninkovic, Jana
    For centuries, opioids have been implicated in increasing susceptibility to infection, reducing bacterial clearance, and increasing bacterial dissemination. Macrophages as key cells of innate immunity play an essential role in pathogen clearance and antigen presentation. Macrophage phagocytosis is a key mechanism responsible for host defense against bacterial pathogens. Although it is known that opioid addicts are prone to both bacterial and viral infections, the molecular and cellular mechanisms underlying these processes remain to be elucidated. Therefore the goal of this research was to investigate mechanisms of decreased bacterial clearance as a contributing factor in the increased susceptibility to infection in opiate drug abusers. To this end, first set of studies examined the role of morphine on inhibition of key mechanisms involved in Fc-gamma receptor mediated phagocytosis. It was demonstrated that morphine inhibits phagocytosis by inhibiting actin polymerization through a cAMP, PKA and MAPK dependant pathways. By superactivation of adenylyl cyclase morphine increases intracellular cAMP leading to inhibition of actin polymerization. Furthermore, morphine by inhibiting p38 MAPK and ERK 1/2 MAPK causes inhibition of actin polymerization and phagocytosis. By modulating TLR4 receptor function morphine was also able to increase macrophage phagocytosis, indicating that morphine might have a differential effect on internalization of Gram-positive, versus Gram-negative pathogens. These effects were mediated through a MyD88 and p38 MAPK dependant pathways leading to changes in actin polymerization and phagocytosis. In addition to macrophage's ability to internalize pathogens, elimination of internalized pathogen is essential for effective bacterial clearance. We therefore set out to investigate morphine's modulation of macrophage bactericidal mechanisms. We note that morphine inhibits bacterial killing by inhibiting essential mechanisms involved in this process such as formation of reactive oxygen intermediates, reactive nitrogen intermediates, as well as phago-lysosomal fusion. Morphine by inhibiting these essential mechanisms impedes eradication of bacterial infections and leads to detrimental consequences for the host. These series of studies have extended our knowledge in an underrepresented yet clinically significant field of study, however many questions still remain to be addressed and it is crucial to investigate the answers given the prevalence of morphine use today.
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    Neuroligin-3 Modulates Opioid-Evoked Changes in Behavior and Brain Function
    (2022-05) Brandner, Dieter
    Chronic opioid exposure causes structural and functional changes in brain circuits, which may contribute to opioid use disorders. Synaptic cell-adhesion molecules are prime candidates for mediating this opioid-evoked plasticity. Neuroligin-3 is a postsynaptic adhesion protein that shapes synaptic function at multiple sites in the mesolimbic dopamine system. We therefore studied how genetic knockout of neuroligin-3 alters responses to chronic morphine in male mice. Constitutive neuroligin-3 knockout caused a persistent reduction in psychomotor sensitization after chronic morphine exposure, as well as a change in the topography of locomotor stimulation produced by morphine. This latter change was recapitulated by conditional genetic deletion of neuroligin-3 from cells expressing the Drd1 dopamine receptor, whereas the reduction in psychomotor sensitization was recapitulated by conditional genetic deletion from dopamine neurons. In the absence of neuroligin-3 expression, dopamine neurons within the ventral tegmental area showed diminished activation following chronic morphine exposure, as measured by in vivo calcium imaging with fiber photometry. This altered pattern of dopamine neuron activity may be driven by aberrant forms of opioid-evoked synaptic plasticity in the absence of neuroligin-3: dopamine neurons lacking neuroligin-3 showed weaker synaptic inhibition at baseline, which was subsequently strengthened after chronic morphine. In total, our study highlights neurobiological adaptations in dopamine neurons of the ventral tegmental area that correspond with increased behavioral sensitivity to opioids, and further suggests that neuroligin-3 expression by dopamine neurons provides a molecular substrate for opioid-evoked adaptations in brain function and behavior.
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    Repeated morphine exposure activates synaptogenesis and other neuroplasticity-related gene networks in the dorsomedial prefrontal cortex of male and female rats
    (2023-04) Liu, Shirelle
    Opioid abuse is a chronic disorder likely involving stable neuroplastic modifications. While a number of molecules contributing to these changes have been identified, the broader spectrum of genes and gene networks that are affected by repeated opioid administration remain understudied.In this study, Next-Generation RNA-sequencing (RNA-seq) was employed followed by quantitative chromatin immunoprecipitation to investigate changes in gene expression and their regulation in adult male and female rats’ dorsomedial prefrontal cortex (dmPFC) after a regimen of daily injection of morphine (5.0 mg/kg; 10 days). Ingenuity Pathway Analysis (IPA) was used to analyze affected molecular pathways, gene networks, and associated regulatory factors. A complementary behavioral study evaluated the effects of the same morphine injection regimen on locomotor activity, pain sensitivity, and somatic withdrawal signs. Behaviorally, repeated morphine injection induced locomotor hyperactivity and hyperalgesia in both sexes. 90% of differentially expressed genes (DEGs) in morphine-treated rats were upregulated in both males and females, with a 35% overlap between sexes. A substantial number of DEGs play roles in synaptic signaling and neuroplasticity. Chromatin immunoprecipitation revealed enrichment of H3 acetylation, a transcriptionally activating chromatin mark. Although broadly similar, some differences were revealed in the gene ontology networks enriched in females and males. The results cohere with findings from previous studies based on a priori gene selection. This study also reveals novel genes and molecular pathways that are upregulated by repeated morphine exposure, with some common to males and females and others that are sex-specific.
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    Temporal Modulation of Gut Microbiome and Metabolome by Morphine
    (2015-12) Wang, Fuyuan
    Opioids such as morphine have many beneficial properties as analgesics, however, opioids may induce multiple adverse gastrointestinal symptoms. It has been recently demonstrated that morphine treatment results in significant disruption in gut barrier function leading to increased translocation of gut commensal bacteria. However, it is unclear how opioids modulate gut homeostasis. A mouse model was used to investigate the effects of morphine treatment on gut microbiome and metabolome. When phylogenetic profiles of gut microbes were characterized, the results revealed a significant shift in the colonic microbiome following morphine treatment when compared to placebo. At the species level, Enterococcus faecalis was associated with morphine-modulated gut microbiome alteration. Morphine treatment also resulted in dramatic changes in the fecal metabolomic profile. Through LC-MS based metabolomics profiling analysis, fatty acids and bile acids metabolism and in particular, deoxycholic acid (DCA) and phosphatidylethanolamines (PEs) was identified to be greatly affected by morphine treatment, implicating that changes in the microbiome community has functional consequences. In a longitudinal study, naltrexone, an opioid receptor antagonist, reversed the effect of morphine on bile acid metabolism, indicating morphine induced changes are opioid receptor dependent. Cross-correlation between gut microbiome and metabolome indicated association between bacterial communities and functional metabolites. Furthermore, morphine induced dysbiosis disrupts morphine metabolism and its enterohepatic recirculation. This study shed light on the effects of morphine on the microbiome-metabolome-host axis, and its role in gut homeostasis. In a mouse model of Citrobacter rodentium infection, morphine treatment resulted in 1) the promotion of C. rodentium systemic dissemination, 2) increase in virulence factors expression with C. rodentium colonization in intestinal contents, 3) altered gut microbiome, 4) damaged integrity of gut epithelial barrier function, 5) inhibition of C. rodentium-induced increase of goblet cells, and 6) dysregulated IL-17A immune response. This is the first study to demonstrate that morphine promotes pathogen dissemination in the context of intestinal C. rodentium infection, indicating morphine modulates virulence factor-mediated adhesion of pathogenic bacteria and induces disruption of mucosal host defense during C. rodentium intestinal infection in mice.