dynorphins and Disease-Models--Animal

Kappa-opioid receptor (KOR) antagonists are currently being considered for the treatment of a variety of neuropsychiatric conditions, including depressive, anxiety, and substance abuse disorders. A general ability to mitigate the effects of stress, which can trigger or exacerbate these conditions, may explain their putative efficacy across such a broad array of conditions. The discovery of their potentially therapeutic effects evolved from preclinical research designed to characterize the molecular mechanisms by which experience causes neuroadaptations in the nucleus accumbens (NAc), a key element of brain reward circuitry. This research established that exposure to drugs of abuse or stress increases the activity of the transcription factor CREB (cAMP response element binding protein) in the NAc, which leads to elevated expression of the opioid peptide dynorphin that in turn causes core signs of depressive- and anxiety-related disorders. Disruption of KORs-the endogenous receptors for dynorphin-produces antidepressant- and anxiolytic-like actions in screening procedures that identify standard drugs of these classes, and reduces stress effects in tests used to study addiction and stress-related disorders. Although interest in this target is high, prototypical KOR antagonists have extraordinarily persistent pharmacodynamic effects that complicate clinical trials. The development of shorter acting KOR antagonists together with more rapid designs for clinical trials may soon provide insight on whether these drugs are efficacious as would be predicted by preclinical work. If successful, KOR antagonists would represent a unique example in psychiatry where the therapeutic mechanism of a drug class is understood before it is shown to be efficacious in humans.

Topics: Animals; Anti-Anxiety Agents; Antidepressive Agents; Anxiety Disorders; Brain; CREB-Binding Protein; Disease Models, Animal; Dynorphins; Gene Expression Regulation; Humans; Narcotic Antagonists; Nucleus Accumbens; Receptors, Opioid, kappa; Reward; Stress, Psychological; Substance-Related Disorders; Translational Research, Biomedical

This review represents the focus of a symposium that was presented at the "Alcoholism and Stress: A Framework for Future Treatment Strategies" conference in Volterra, Italy on May 3-6, 2011 and organized/chaired by Dr. Brendan M. Walker. The primary goal of the symposium was to evaluate and disseminate contemporary findings regarding the emerging role of kappa-opioid receptors (KORs) and their endogenous ligands dynorphins (DYNs) in the regulation of escalated alcohol consumption, negative affect and cognitive dysfunction associated with alcohol dependence, as well as DYN/KOR mediation of the effects of chronic stress on alcohol reward and seeking behaviors. Dr. Glenn Valdez described a role for KORs in the anxiogenic effects of alcohol withdrawal. Dr. Jay McLaughlin focused on the role of KORs in repeated stress-induced potentiation of alcohol reward and increased alcohol consumption. Dr. Brendan Walker presented data characterizing the effects of KOR antagonism within the extended amygdala on withdrawal-induced escalation of alcohol self-administration in dependent animals. Dr. Georgy Bakalkin concluded with data indicative of altered DYNs and KORs in the prefrontal cortex of alcohol dependent humans that could underlie diminished cognitive performance. Collectively, the data presented within this symposium identified the multifaceted contribution of KORs to the characteristics of acute and chronic alcohol-induced behavioral dysregulation and provided a foundation for the development of pharmacotherapeutic strategies to treat certain aspects of alcohol use disorders.

Topics: Alcoholism; Amygdala; Animals; Disease Models, Animal; Dynorphins; Ethanol; Humans; Neurotransmitter Agents; Receptors, Opioid, kappa; Stress, Psychological; Substance Withdrawal Syndrome

Addictions to cocaine or heroin/prescription opioids [short-acting μ-opioid receptor (MOPr) agonists] involve relapsing cycles, with experimentation/escalating use, withdrawal/abstinence, and relapse/re-escalation. κ-Opioid receptors (KOPr; encoded by OPRK1), and their endogenous agonists, the dynorphins (encoded by PDYN), have counter-modulatory effects on reward caused by cocaine or MOPr agonist exposure, and exhibit plasticity in addictive-like states. KOPr/dynorphin activation is implicated in depression/anxiety, often comorbid with addictions. In this opinion article we propose that particular stages of the addiction cycle are differentially affected by KOPr/dynorphin systems. Vulnerability and resilience can be due to pre-existing (e.g., genetic) factors, or epigenetic modifications of the OPRK1 or PDYN genes during the addiction cycle. Pharmacotherapeutic approaches limiting changes in KOPr/dynorphin tone, especially with KOPr partial agonists, may hold potential for the treatment of specific drug addictions and psychiatric comorbidity.

Topics: Adaptation, Biological; Animals; Behavior, Addictive; Disease Models, Animal; Drug Discovery; Dynorphins; Enkephalins; Genetic Predisposition to Disease; Humans; Illicit Drugs; Narcotic Antagonists; Polymorphism, Genetic; Protein Precursors; Receptors, Opioid, kappa; Recurrence

Central nervous system (CNS) injury is a complex in which numerous neurochemicals and other vasoactive agents actively contribute towards the development of posttraumatic brain pathology and/or repair mechanisms. A focal trauma to the brain or spinal cord releases several endogenous neurodestructive agents within the CNS, resulting in adverse cellular reactions. Our laboratory is engaged in identifying these endogenous neurodestructive signals in the CNS following injury caused by trauma or hyperthermia. Our observations show that serotonin (5-HT), dynorphin A (Dyn A 1-17), nitric oxide synthase (NOS), and tumor necrosis factor-α (TNF-α) could be potential neurodestructive signals in the CNS injury. Thus, neutralization of these agents using monoclonal antibodies directed against 5-HT, NOS, Dyn A (1-17), and TNF-α in vivo will result in marked neuroprotection and enhance neurorepair after trauma. In addition, a suitable combination of monoclonal antibodies, for example, NOS and TNF-α, when applied 60-90 min after trauma, is capable to enhance neuroprotective ability and thwart cell and tissue injury after spinal cord insult. Taken together, our novel observations suggest a potential use of monoclonal antibodies as suitable therapeutic agents in CNS injuries to achieve neuroprotection and/or neurorepair.

Topics: Animals; Antibodies, Monoclonal; Antigen-Antibody Reactions; Brain Injuries; Disease Models, Animal; Dynorphins; History, 18th Century; History, 19th Century; History, 20th Century; Humans; Nerve Regeneration; Neuroprotective Agents; Nitric Oxide Synthase Type I; Serotonin; Spinal Cord Injuries; Tumor Necrosis Factor-alpha; Wound Healing

Stress is most often associated with aversive states. It rapidly induces the release of hormones and neuropeptides including dynorphin, which activates kappa opioid receptors (KORs) in the central and peripheral nervous systems. In animal models, many aversive effects of stress are mimicked or exacerbated by stimulation of KORs in limbic brain regions. Although KOR signaling during acute stress may increase physical ability (by producing analgesia) and motivation to escape a threat (by producing aversion), prolonged KOR signaling in response to chronic or uncontrollable stress can lead to persistent expression of behavioral signs that are characteristic of human depressive disorders (i.e., "prodepressive-like" signs). Accumulating evidence suggests that KORs contribute to the progressive amplification (sensitization) of stress-induced behaviors that occurs with repeated exposure to stress. Many of the aversive effects of stress are blocked by KOR antagonists, suggesting that these agents may have potential as therapeutics for stress-related conditions such as depression and anxiety disorders. This review summarizes current data on how KOR systems contribute to the acute (rapid), delayed, and cumulative molecular and behavioral effects of stress. We focus on behavioral paradigms that provide insight on interactions between stress and KOR function within each of these temporal categories. Using a simplified model, we consider the time course and mechanism of KOR-mediated effects in stress and suggest future directions that may be useful in determining whether KOR antagonists exert their therapeutic effects by preventing the development of stress-induced behaviors, the expression of stress-induced behaviors, or both.

Topics: Animals; Brain; Chronic Disease; Comorbidity; Depressive Disorder; Disease Models, Animal; Dynorphins; Fear; Humans; Narcotic Antagonists; Receptors, Opioid, kappa; Stress, Psychological

Since the first description of their opioid properties three decades ago, dynorphins have increasingly been thought to play a regulatory role in numerous functional pathways of the brain. Dynorphins are members of the opioid peptide family and preferentially bind to kappa opioid receptors. In line with their localization in the hippocampus, amygdala, hypothalamus, striatum and spinal cord, their functions are related to learning and memory, emotional control, stress response and pain. Pathophysiological mechanisms that may involve dynorphins/kappa opioid receptors include epilepsy, addiction, depression and schizophrenia. Most of these functions were proposed in the 1980s and 1990s following histochemical, pharmacological and electrophysiological experiments using kappa receptor-specific or general opioid receptor agonists and antagonists in animal models. However, at that time, we had little information on the functional relevance of endogenous dynorphins. This was mainly due to the complexity of the opioid system. Besides actions of peptides from all three classical opioid precursors (proenkephalin, prodynorphin, proopiomelanocortin) on the three classical opioid receptors (delta, mu and kappa), dynorphins were also shown to exert non-opioid effects mainly through direct effects on NMDA receptors. Moreover, discrepancies between the distribution of opioid receptor binding sites and dynorphin immunoreactivity contributed to the difficulties in interpretation. In recent years, the generation of prodynorphin- and opioid receptor-deficient mice has provided the tools to investigate open questions on network effects of endogenous dynorphins. This article examines the physiological, pathophysiological and pharmacological implications of dynorphins in the light of new insights in part obtained from genetically modified animals.

Topics: Animals; Brain Diseases; Disease Models, Animal; Dynorphins; History, 20th Century; Humans; Mental Disorders; Mice; Substance-Related Disorders

Epilepsy is a significant health problem. Despite the widespread use of both classic and newer pharmacological agents that target ion channels, amino acid transmission or receptors, there are numerous examples of mono- or polytherapy being ineffective. Seizures that are secondary to CNS infections are among the most refractory medically, and thus insult-specific agents are desirable. Recently, the study of the neuropharmacological actions of dynorphin in CNS viral injury has yielded new insights into epileptogenesis and epilepsy treatment. The opioid neuropeptide dynorphin modulates neuronal excitability in vitro in hippocampal slices and potentiates endogenous anti-ictal (i.e. protective) processes in animal models and humans. This work has renewed interest in the role of dysregulation of dynorphin in the pathogenesis of refractory seizures, including encephalitic seizures. The important role of dynorphin in epilepsy is also supported by new models of symptomatic epilepsies based on viral-induced seizures.

Topics: Animals; Central Nervous System Diseases; Disease Models, Animal; Dynorphins; Epilepsy; Hippocampus; Humans; Rats; Seizures

Alzheimer's disease (AD) occurs against a background of cognitive and neurobiological aging. Animal models of normal aging may be used to study the neurobiological structures that are most involved in AD pathology, i.e. hippocampal/cortical systems. For example, spatial learning is dependent upon the integrity of the hippocampus, a structure that is much affected in humans with AD. Spatial learning tasks, such as the Morris water maze, have been used to screen aged rats for cognitive status prior to neurobiological assessment of hippocampal circuitry. Manifestations of the aging process, which are often minimal or entirely obscured in studies comparing young and aged brains, become apparent when the cognitive status of aged animals is taken into account. For example, studies examining the septohippocampal cholinergic system in behaviorally-characterized rodents have shown that there is a decline in many markers for these cholinergic neurons that coincides with severity of spatial learning impairment. Another advantage of cognitive assessment in animal models used to study aging is that it may help to distinguish between those neurobiological changes that are functionally detrimental and those that may represent compensatory adaptations to maintain cognitive function. Age-related changes in two neurobiological measures in the hippocampus are discussed in this report. Alterations in the opioid peptide dynorphin (increased peptide content and prodynorphin mRNA) in hippocampus may contribute to impairment in that the greatest changes occur in those aged rats with severe spatial learning deficits.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aging; Animals; Cellular Senescence; Cerebral Cortex; Cholinergic Fibers; Cognition; Disease Models, Animal; Dynorphins; Hippocampus; Long-Term Potentiation; Nerve Net; Neural Pathways; Orientation; Receptors, Kainic Acid; Reference Values; Septum Pellucidum

While there is high comorbidity of stress-related disorders and alcohol use disorder, few effective treatments are available and elucidating underlying neurobiological mechanisms has been hampered by a general lack of reliable animal models. Here, we use a novel mouse model demonstrating robust and reproducible stress-enhanced alcohol drinking to examine the role of dynorphin/kappa opioid receptor (DYN/KOR) activity within the extended amygdala in mediating this stress-alcohol interaction.. Mice received repeated weekly cycles of chronic intermittent ethanol exposure alternating with weekly drinking sessions ± forced swim stress exposure. Pdyn messenger RNA expression was measured in the central amygdala (CeA), and DYN-expressing CeA neurons were then targeted for chemogenetic inhibition. Finally, a KOR antagonist was microinjected into the CeA or bed nucleus of the stria terminalis to examine the role of KOR signaling in promoting stress-enhanced drinking.. Stress (forced swim stress) selectively increased alcohol drinking in mice with a history of chronic intermittent ethanol exposure, and this was accompanied by elevated Pdyn messenger RNA levels in the CeA. Targeted chemogenetic silencing of DYN-expressing CeA neurons blocked stress-enhanced drinking, and KOR antagonism in the CeA or bed nucleus of the stria terminalis significantly reduced stress-induced elevated alcohol consumption without altering moderate intake in control mice.. Using a novel and robust model of stress-enhanced alcohol drinking, a significant role for DYN/KOR activity within extended amygdala circuitry in mediating this effect was demonstrated, thereby providing further evidence that the DYN/KOR system may be a valuable target in the development of more effective treatments for individuals presenting with comorbidity of stress-related disorders and alcohol use disorder.

Topics: Alcohol Drinking; Alcoholism; Animals; Central Amygdaloid Nucleus; Disease Models, Animal; Dynorphins; Ethanol; Mice; Receptors, Opioid, kappa; RNA, Messenger

COVID-19 is a respiratory infection caused by the SARS-CoV-2 virus that can rapidly escalate to life-threatening pneumonia and acute respiratory distress syndrome (ARDS). Recently, extracellular high mobility group box 1 (HMGB1) has been identified as an essential component of cytokine storms that occur with COVID-19; HMGB1 levels correlate significantly with disease severity. Thus, the modulation of HMGB1 release may be vital for treating COVID-19. HMGB1 is a ubiquitous nuclear DNA-binding protein whose biological function depends on posttranslational modifications, its redox state, and its cellular localization. The acetylation of HMGB1 is a prerequisite for its translocation from the nucleus to the cytoplasm and then to the extracellular milieu. When released, HMGB1 acts as a proinflammatory cytokine that binds primarily to toll-like receptor 4 (TLR4) and RAGE, thereby stimulating immune cells, endothelial cells, and airway epithelial cells to produce cytokines, chemokines, and other inflammatory mediators. In this study, we demonstrate that inhaled [D-Ala

Topics: Acetylation; Acute Lung Injury; Animals; COVID-19; COVID-19 Drug Treatment; Disease Models, Animal; Dynorphins; HMGB1 Protein; Mice; Mice, Inbred C57BL; Receptors, Opioid; Sirtuin 1

Opiates produce a strong rewarding effect, but abstinence from opiate use emerges with severe negative emotions. Depression is one of the most frequent emotion disorders associated with opiate abstinence, which is thought to be a main cause for relapse. However, neurobiological bases of such an aversive emotion processing are poorly understood. Here, we find that morphine abstinence activates κ-opioid receptors (KORs) by increasing endogenous KOR ligand dynorphin expression in the amygdala, which in turn facilitates glutamate transporter 1 (GLT1) expression by activation of p38 mitogen-activated protein kinase (MAPK). Upregulation of GLT1 expression contributes to opiate-abstinence-elicited depressive-like behaviors through modulating amygdalar glutamatergic inputs to the nucleus accumbens (NAc). Intra-amygdala injection of GLT1 inhibitor DHK or knockdown of GLT1 expression in the amygdala significantly suppresses morphine-abstinence-induced depressive-like behaviors. Pharmacological and pharmacogenetic activation of amygdala-NAc projections prevents morphine-abstinence-induced behaviors. Overall, our study provides key molecular and circuit insights into the mechanisms of depression associated with opiate abstinence.

Topics: Amygdala; Animals; Behavior, Animal; Depression; Disease Models, Animal; Dynorphins; Excitatory Postsynaptic Potentials; Glucose Transporter Type 1; Glutamic Acid; Male; Mice, Inbred C57BL; Mice, Knockout; Morphine; Neural Pathways; Nucleus Accumbens; p38 Mitogen-Activated Protein Kinases; Receptors, Opioid, kappa; Signal Transduction; Substance Withdrawal Syndrome

Polycystic ovarian syndrome (PCOS), the most common endocrinopathy affecting women worldwide, is characterized by elevated luteinizing hormone (LH) pulse frequency due to the impaired suppression of gonadotrophin-releasing hormone (GnRH) release by steroid hormone negative feedback. Although neurons that co-express kisspeptin, neurokinin B, and dynorphin (KNDy cells) were recently defined as the GnRH/LH pulse generator, little is understood about their role in the pathogenesis of PCOS. We used a prenatal androgen-treated (PNA) mouse model of PCOS to determine whether changes in KNDy neurons or their afferent network underlie altered negative feedback. First, we identified elevated androgen receptor gene expression in KNDy cells of PNA mice, whereas progesterone receptor and dynorphin gene expression was significantly reduced, suggesting elevated androgens in PCOS disrupt progesterone negative feedback via direct actions upon KNDy cells. Second, we discovered GABAergic and glutamatergic synaptic input to KNDy neurons was reduced in PNA mice. Retrograde monosynaptic tract-tracing revealed a dramatic reduction in input originates from sexually dimorphic afferents in the preoptic area, anteroventral periventricular nucleus, anterior hypothalamic area and lateral hypothalamus. These results reveal 2 sites of neuronal alterations potentially responsible for defects in negative feedback in PCOS: changes in gene expression within KNDy neurons, and changes in synaptic inputs from steroid hormone-responsive hypothalamic regions. How each of these changes contribute to the neuroendocrine phenotype seen in in PCOS, and the role of specific sets of upstream KNDy afferents in the process, remains to be determined.

Topics: Afferent Pathways; Androgens; Animals; Disease Models, Animal; Dynorphins; Female; Kisspeptins; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neurokinin B; Neurons; Neurons, Afferent; Neurosecretory Systems; Polycystic Ovary Syndrome; Pregnancy; Prenatal Exposure Delayed Effects

When Zika virus emerged as a public health emergency there were no drugs or vaccines approved for its prevention or treatment. We used a high-throughput screen for Zika virus protease inhibitors to identify several inhibitors of Zika virus infection. We expressed the NS2B-NS3 Zika virus protease and conducted a biochemical screen for small-molecule inhibitors. A quantitative structure-activity relationship model was employed to virtually screen ∼138,000 compounds, which increased the identification of active compounds, while decreasing screening time and resources. Candidate inhibitors were validated in several viral infection assays. Small molecules with favorable clinical profiles, especially the five-lipoxygenase-activating protein inhibitor, MK-591, inhibited the Zika virus protease and infection in neural stem cells. Members of the tetracycline family of antibiotics were more potent inhibitors of Zika virus infection than the protease, suggesting they may have multiple mechanisms of action. The most potent tetracycline, methacycline, reduced the amount of Zika virus present in the brain and the severity of Zika virus-induced motor deficits in an immunocompetent mouse model. As Food and Drug Administration-approved drugs, the tetracyclines could be quickly translated to the clinic. The compounds identified through our screening paradigm have the potential to be used as prophylactics for patients traveling to endemic regions or for the treatment of the neurological complications of Zika virus infection.

Topics: Animals; Antiviral Agents; Artificial Intelligence; Chlorocebus aethiops; Disease Models, Animal; Drug Evaluation, Preclinical; High-Throughput Screening Assays; Immunocompetence; Inhibitory Concentration 50; Methacycline; Mice, Inbred C57BL; Protease Inhibitors; Quantitative Structure-Activity Relationship; Small Molecule Libraries; Vero Cells; Zika Virus; Zika Virus Infection

Polycystic ovary syndrome (PCOS), a common reproductive disorder in women, is characterized by hyperandrogenemia, chronic anovulation, cystic ovarian follicles, and luteinizing hormone (LH) hyper-pulsatility, but the pathophysiology isn't completely understood. We recently reported a novel mouse model of PCOS using chronic letrozole (LET; aromatase inhibitor). Letrozole-treated females demonstrate multiple PCOS-like phenotypes, including polycystic ovaries, anovulation, and elevated circulating testosterone and LH, assayed in "one-off" measures. However, due to technical limitations, in vivo LH pulsatile secretion, which is elevated in PCOS women, was not previously studied, nor were the possible changes in reproductive neurons. Here, we used recent technical advances to examine in vivo LH pulse dynamics of freely moving LET female mice versus control and ovariectomized (OVX) mice. We also determined whether neural gene expression of important reproductive regulators such as kisspeptin, neurokinin B (NKB), and dynorphin, is altered in LET females. Compared to controls, LET females exhibited very rapid, elevated in vivo LH pulsatility, with increased pulse frequency, amplitude, and basal levels, similar to PCOS women. Letrozole-treated mice also had markedly elevated Kiss1, Tac2, and Pdyn expression and increased Kiss1 neuronal activation in the hypothalamic arcuate nucleus. Notably, the hyperactive LH pulses and increased kisspeptin neuron measures of LET mice were not as elevated as OVX females. Our findings indicate that LET mice, like PCOS women, have markedly elevated LH pulsatility, which likely drives increased androgen secretion. Increased hypothalamic kisspeptin and NKB levels may be fundamental contributors to the hyperactive LH pulse secretion in the LET PCOS-like condition and, perhaps, in PCOS women.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Aromatase Inhibitors; Disease Models, Animal; Dynorphins; Female; Gene Expression; Kisspeptins; Letrozole; Luteinizing Hormone; Mice; Neurokinin B; Neurons; Polycystic Ovary Syndrome

Prenatal testosterone (T)-treated female sheep display reproductive deficits similar to women with polycystic ovarian syndrome (PCOS), including an increase in LH pulse frequency due to actions of the central GnRH pulse generator. In this study, we used multiple-label immunocytochemistry to investigate the possibility of changes in the γ-aminobutyric acid (GABA) neurotransmitter system at two key components of the GnRH pulse generator in prenatal T-treated sheep: kisspeptin/neurokinin B/dynorphin (KNDy) neurons of the arcuate nucleus, and GnRH neurons in the preoptic area (POA) and mediobasal hypothalamus (MBH). We observed a significant decrease and increase, respectively, in the number of GABAergic synapses onto POA and MBH GnRH neurons in prenatal T-treated ewes; additionally, there was a significant increase in the number of GABAergic inputs onto KNDy neurons. To determine the actions of GABA on GnRH and KNDy neurons, we examined colocalization with the chloride transporters NKCC1 and KCC2, which indicate stimulatory or inhibitory activation of neurons by GABA, respectively. Most GnRH neurons in both POA and MBH colocalized NKCC1 cotransporter whereas none contained the KCC2 cotransporter. Most KNDy neurons colocalized either NKCC1 or KCC2, and 28% of the KNDy population contained NKCC1 alone. Therefore, we suggest that, as in the mouse, GABA in the sheep is stimulatory to GnRH neurons, as well as to a subset of KNDy neurons. Increased numbers of stimulatory GABAergic inputs to both MBH GnRH and KNDy neurons in prenatal T-treated animals may contribute to alterations in steroid feedback control and increased GnRH/LH pulse frequency seen in this animal model of PCOS.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Disease Models, Animal; Dynorphins; Female; GABAergic Neurons; Gonadotropin-Releasing Hormone; K Cl- Cotransporters; Kisspeptins; Neurokinin B; Polycystic Ovary Syndrome; Pregnancy; Prenatal Exposure Delayed Effects; Preoptic Area; Sheep; Solute Carrier Family 12, Member 2; Symporters; Testosterone

Focal epilepsy represents one of the most common chronic CNS diseases. The high incidence of drug resistance, devastating comorbidities, and insufficient responsiveness to surgery pose unmet medical challenges. In the quest of novel, disease-modifying treatment strategies of neuropeptides represent promising candidates. Here, we provide the "proof of concept" that gene therapy by adeno-associated virus (AAV) vector transduction of preprodynorphin into the epileptogenic focus of well-accepted mouse and rat models for temporal lobe epilepsy leads to suppression of seizures over months. The debilitating long-term decline of spatial learning and memory is prevented. In human hippocampal slices obtained from epilepsy surgery, dynorphins suppressed seizure-like activity, suggestive of a high potential for clinical translation. AAV-delivered preprodynorphin expression is focally and neuronally restricted and release is dependent on high-frequency stimulation, as it occurs at the onset of seizures. The novel format of "release on demand" dynorphin delivery is viewed as a key to prevent habituation and to minimize the risk of adverse effects, leading to long-term suppression of seizures and of their devastating sequel.

Topics: Animals; Dependovirus; Disease Models, Animal; Dynorphins; Epilepsy, Temporal Lobe; Gene Expression; Gene Expression Regulation; Genetic Therapy; Genetic Vectors; Humans; Mice; Models, Theoretical; Neurotransmitter Agents; Organ Culture Techniques; Rats; Transduction, Genetic; Treatment Outcome

Although previous research has demonstrated a role for kappa opioid receptor-mediated signaling in escalated alcohol consumption associated with dependence and stress exposure, involvement of the dynorphin/kappa opioid receptor (DYN/KOR) system in binge-like drinking has not been fully explored. Here we used pharmacological and chemogenetic approaches to examine the influence of DYN/KOR signaling on alcohol consumption in the drinking-in-the-dark (DID) model of binge-like drinking. Systemic administration of the KOR agonist U50,488 increased binge-like drinking (Experiment 1) while, conversely, systemic administration of the KOR antagonist nor-BNI reduced drinking in the DID model (Experiment 2). These effects of systemic KOR manipulation were selective for alcohol as neither drug influenced consumption of sucrose in the DID paradigm (Experiment 3). In Experiment 4, administration of the long-acting KOR antagonist nor-BNI into the central nucleus of the amygdala (CeA) decreased alcohol intake. Next, targeted "silencing" of DYN+ neurons in the CeA was accomplished using a chemogenetic strategy. Cre-dependent viral expression in DYN+ neurons was confirmed in CeA of Pdyn-IRES-Cre mice and functionality of an inhibitory (hM4Di) DREADD was validated (Experiment 5). Activating the inhibitory DREADD by CNO injection reduced binge-like alcohol drinking, but CNO injection did not alter alcohol intake in mice that were treated with control virus (Experiment 6). Collectively, these results demonstrate that DYN/KOR signaling in the CeA contributes to excessive alcohol consumption in a binge-drinking model.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Behavior, Animal; Binge Drinking; Central Amygdaloid Nucleus; Disease Models, Animal; Dynorphins; Genetic Techniques; Male; Mice; Mice, Inbred C57BL; Naltrexone; Receptors, Opioid, kappa

Kisspeptin-neurokinin B (NKB)-dynorphin neurons are critical regulators of the hypothalamic-pituitary-gonadal axis. NKB and dynorphin are hypothesized to influence the frequency of GnRH pulses, whereas kisspeptin is hypothesized to be a generator of the GnRH pulse. How these neuropeptides interact remains unclear.. To probe the role of NKB in GnRH pulse generation and to determine the interactions between NKB, kisspeptin, and dynorphin in humans and mice with a complete absence of NKB.. Case/control.. Academic medical center.. Members of a consanguineous family bearing biallelic loss-of-function mutations in the gene encoding NKB and NKB-deficient mice.. Frequent blood sampling to characterize neuroendocrine profile and administration of kisspeptin, GnRH, and naloxone, a nonspecific opioid receptor antagonist used to block dynorphin.. LH pulse characteristics.. Humans lacking NKB demonstrate slow LH pulse frequency, which can be increased by opioid antagonism. Mice lacking NKB also demonstrate impaired LH secretion, which can be augmented with an identical pharmacologic manipulation. Both mice and humans with NKB deficiency respond to exogenous kisspeptin.. The preservation of LH pulses in the absence of NKB and dynorphin signaling suggests that both peptides are dispensable for GnRH pulse generation and kisspeptin responsiveness. However, NKB and dynorphin appear to have opposing roles in the modulation of GnRH pulse frequency.

Topics: Academic Medical Centers; Adolescent; Adult; Animals; Case-Control Studies; Child; Disease Models, Animal; Dynorphins; Female; Gonadotropin-Releasing Hormone; Humans; Hypogonadism; Kisspeptins; Luteinizing Hormone; Mice; Mice, Knockout; Narcotic Antagonists; Neurokinin B; Neurons; Signal Transduction; Substance P; Treatment Outcome; Young Adult

The serotonergic system is a well-established modulator of l-dopa-induced dyskinesia. To date, targeting serotonin transporters or serotonin receptor subtype 1A (5-HT. The goal of the present study was to characterize Vilazodone's effects on l-dopa-induced behaviors, neurochemistry and gene expression in unilateral 6-hydroxydopamine-lesioned hemi-parkinsonian rats.. In experiments 1 and 2, l-dopa-naïve and l-dopa-primed animals were coadministered Vilazodone and l-dopa daily for 3 weeks to model subchronic use, and behavioral, neurochemical, and messenger RNA (mRNA) expression changes were measured. In experiment 3, dyskinetic behavior was assessed following 5-HT. Vilazodone significantly suppressed developing and established l-dopa-induced dyskinesia without compromising the promotor effects of l-dopa therapy. In the dopamine-depleted striatum, Vilazodone-l-dopa cotreatment increased dopamine content, suggesting a normalization of dopamine kinetics in dyskinetic brain, and reduced l-dopa-induced c-Fos and preprodynorphin mRNA overexpression, indicative of attenuated dopamine D. Our findings show Vilazodone has a serotonin-dependent effect on rodent l-dopa-induced dyskinesia and implicate the potential for repositioning Vilazodone against l-dopa-induced dyskinesia development and expression in Parkinson's disease patients. © 2018 International Parkinson and Movement Disorder Society.

Topics: Animals; Antiparkinson Agents; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Gene Expression Regulation; Levodopa; Male; Oxidopamine; Parkinsonian Disorders; Piperazines; Protein Precursors; Proto-Oncogene Proteins c-fos; Pyridines; Rats; Rats, Sprague-Dawley; Receptors, Serotonin; RNA, Messenger; Selective Serotonin Reuptake Inhibitors; Serotonin; Serotonin Antagonists; Time Factors; Vilazodone Hydrochloride

Hyperalgesia that develops following nerve ligation corresponds temporally and in magnitude with the number of thalamic mast cells located contralateral to the ligature. We tested the possibility that mast cells modulate nociception centrally, similar to their role in the periphery.. We examined the central effect of two hyperalgesic compounds that induce mast cell degranulation and of stabilized mast cells using cromolyn.. Thermal hyperalgesia (tail flick) induced by nerve growth factor (NGF, a neurotrophic compound) and mechanical hyperalgesia (von Frey) induced by dynorphin A (1-17) (opioid compound) each correlated with the per cent of thalamic mast cells that were degranulated. Degranulation of these mast cells by the central injection of compound 48/80, devoid of neurotrophic or opioid activity, was sufficient to recapitulate thermal hyperalgesia. Stabilization of mast cells by central injections of cromolyn produced no analgesic effect on baseline tail flick or von Frey fibre sensitivity, but inhibited thermal hyperalgesia produced by compound 48/80 and tactile hyperalgesia induced by dynorphin and by Freund's complete adjuvant. Finally, chemical nociception produced by the direct activation of nociceptors by formalin (phase I) was not inhibited by centrally injected cromolyn whereas chemical nociception dependent on central sensitization (formalin-phase II and acetic acid-induced abdominal stretches) was.. These convergent lines of evidence suggest that degranulation of centrally located mast cells sensitizes central nociceptive pathways leading to hyperalgesia and tonic chemical sensitivity.. Hyperalgesia induced by spinal nerve ligation corresponds temporally and in magnitude with degranulation of thalamic mast cells. Here, we provide evidence that hyperalgesia induced by NGF, formalin and dynorphin also may depend on mast cell degranulation in the CNS whereas cromolyn, a mast cell stabilizer, blocks these effects in mice.

Topics: Animals; Cell Count; Cromolyn Sodium; Disease Models, Animal; Dynorphins; Hyperalgesia; Male; Mast Cells; Mice; Nerve Growth Factor; Neurotransmitter Agents; Nociception; Nociceptors

Dynorphin 1-17 is an endogenous peptide that is released at sites of inflammation by leukocytes, binding preferentially to κ-opioid receptors (KOP) to mediate nociception. We have previously shown that dynorphin 1-17 is rapidly biotransformed to smaller peptide fragments in inflamed tissue homogenate. This study aimed to determine the efficacy and potency of selected dynorphin fragments produced in an inflamed environment at the KOP, μ and δ-opioid receptors (MOP and DOP respectively) and in a model of inflammatory pain. Functional activity of Dynorphin 1-17 and fragments (1-6, 1-7 and 1-9) were screened over a range of concentrations against forskolin stimulated human embryonic kidney 293 (HEK) cells stably transfected with one of KOP, MOP or DOP. The analgesic activity of dynorphin 1-7 in a unilateral model of inflammatory pain was subsequently tested. Rats received unilateral intraplantar injections of Freund's Complete Adjuvant to induce inflammation. After six days rats received either dynorphin 1-7, 1-17 or the selective KOP agonist U50488H and mechanical allodynia determined. Dynorphin 1-7 and 1-9 displayed the greatest activity across all receptor subtypes, while dynorphin 1-7, 1-9 and 1-17 displaying a potent activation of both KOP and DOP evidenced by cAMP inihibition. Administration of dynorphin 1-7 and U50488H, but not dynorphin 1-17 resulted in a significant increase in paw pressure threshold at an equimolar dose suggesting the small peptide dynorphin 1-7 mediates analgesia. These results show that dynorphin fragments produced in an inflamed tissue homogenate have changed activity at the opioid receptors and that dynorphin 1-7 mediates analgesia.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analgesia; Animals; Disease Models, Animal; Dynorphins; HEK293 Cells; Humans; Inflammation; Pain; Rats; Receptors, Opioid, delta; Receptors, Opioid, kappa; Receptors, Opioid, mu; Transfection

Long-term levodopa (l-dopa) treatment is associated with the development of l-dopa-induced dyskinesias in the majority of patients with Parkinson disease (PD). The etiopathogonesis and mechanisms underlying l-dopa-induced dyskinesias are not well understood.. We used striatal optogenetic stimulation to induce dyskinesias in a hemiparkinsonian model of PD in rats. Striatal dopamine depletion was induced unilaterally by 6-hydroxydopamine injection into the medial forebrain bundle. For the optogenetic manipulation, we injected adeno-associated virus particles expressing channelrhodopsin to stimulate striatal medium spiny neurons with a laser source.. Simultaneous optical activation of medium spiny neurons of the direct and indirect striatal pathways in the 6-hydroxydopamine lesion but l-dopa naïve rats induced involuntary movements similar to l-dopa-induced dyskinesias, labeled here as optodyskinesias. Noticeably, optodyskinesias were facilitated by l-dopa in animals that did not respond initially to the laser stimulation. In general, optodyskinesias lasted while the laser stimulus was applied, but in some instances remained ongoing for a few seconds after the laser was off. Postmortem tissue analysis revealed increased FosB expression, a molecular marker of l-dopa-induced dyskinesias, primarily in medium spiny neurons of the direct pathway in the dopamine-depleted hemisphere.. Selective optogenetic activation of the dorsolateral striatum elicits dyskinesias in the 6-hydroxydopamine rat model of PD. This effect was associated with a preferential activation of the direct striato-nigral pathway. These results potentially open new avenues in the understanding of mechanisms involved in l-dopa-induced dyskinesias. © 2017 International Parkinson and Movement Disorder Society.

Topics: Adrenergic Agents; Animals; Antiparkinson Agents; Brain; Channelrhodopsins; Corpus Striatum; Disease Models, Animal; Dynorphins; Dyskinesias; Functional Laterality; Levodopa; Male; Optogenetics; Oxidopamine; Parkinson Disease; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Transduction, Genetic; Tyrosine 3-Monooxygenase

Topics: Animals; Autism Spectrum Disorder; Behavior, Animal; Disease Models, Animal; Dynorphins; Glutamic Acid; Mice, Inbred C57BL; Mice, Inbred Strains; Social Behavior

Aconitine and its structurally-related diterpenoid alkaloids have been shown to interact differentially with neuronal voltage-dependent sodium channels, which was suggested to be responsible for their analgesia and toxicity. Bulleyaconitine A (BAA) is an aconitine analogue and has been prescribed for the management of pain. The present study aimed to evaluate the inhibitory effects of BAA on pain hypersensitivity and morphine antinociceptive tolerance, and explore whether the expression of dynorphin A in spinal microglia was responsible for its actions. Single intrathecal or subcutaneous (but not intraventricular or local) injection of BAA blocked spinal nerve ligation-induced painful neuropathy, bone cancer-induced pain, and formalin-induced tonic pain by 60 to 100% with the median effective dose values of 94 to 126 ng per rat (intrathecal) and 42 to 59 μg/kg (subcutaneous), respectively. After chronic treatment, BAA did not induce either self-tolerance to antinociception or cross-tolerance to morphine antinociception, and completely inhibited morphine tolerance. The microglial inhibitor minocycline entirely blocked spinal BAA (but not exogenous dynorphin A) antinociception, but failed to attenuate spinal BAA neurotoxicity. In a minocycline-sensitive and lidocaine- or ropivacaine-insensitive manner, BAA stimulated the expression of dynorphin A in the spinal cord, and primary cultures of microglia but not of neurons or astrocytes. The blockade effects of BAA on nociception and morphine tolerance were totally eliminated by the specific dynorphin A antiserum and/or κ-opioid receptor antagonist. Our results suggest that BAA eliminates pain hypersensitivity and morphine tolerance through directly stimulating dynorphin A expression in spinal microglia, which is not dependent on the interactions with sodium channels.. The newly illustrated mechanisms underlying BAA antinociception help us to better understand and develop novel dynorphin A expression-based painkillers to treat chronic pain.

Topics: Aconitine; Aconitum; Analgesics; Analysis of Variance; Animals; Animals, Newborn; Bone Neoplasms; CD11b Antigen; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Routes; Dynorphins; Female; Functional Laterality; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Hyperalgesia; Male; Microglia; Microscopy, Confocal; Morphine; Neurons; Pain; Pain Measurement; Phosphopyruvate Hydratase; Rats; Receptors, Opioid, kappa; RNA, Messenger; Spinal Cord; Time Factors

The opioidergic neuropeptides dynorphin (DYN) and enkephalin (ENK) and the D1 and D2 dopaminergic receptors (D1R, D2R) are involved in the striatal control of motor and behavioral function. In Parkinson's disease, motor disturbances such as "on-off" motor fluctuations and involuntary movements (dyskinesia) are severe complications that often arise after chronic l-dihydroxyphenylalanine (l-DOPA) treatment. Changes in the striatal expression of preproENK (PPENK), proDYN (PDYN), D1R, and D2R mRNA have been observed in parkinsonian animals treated with l-DOPA. Enhanced opioidergic transmission has been found in association with l-DOPA-induced dyskinesia, but the connection of PPENK, PDYN, D1R, and D2R mRNA expression with locomotor activity remains unclear. In this study, we measured PPENK, PDYN, D1R and D2R mRNA levels by in situ hybridization in the striatum of 6-OHDA hemi-parkinsonian rats treated with l-DOPA (PD+l-DOPA group), along with two control groups (PD+saline and naive+l-DOPA). We found different levels of expression of PPENK, PDYN, D1R and D2R mRNA across the experimental groups and correlated the changes in mRNA expression with dyskinesia and locomotor variables assessed by open field test during several phases of l-DOPA treatment. Both PDYN and PPENK mRNA levels were correlated with the severity of dyskinesia, while PPENK mRNA levels were also correlated with the frequency of contralateral rotational movements and with locomotor variables. Moreover, a strong correlation was found between D1R mRNA expression and D2R mRNA expression in the PD+l-DOPA group. These findings suggest that, in parkinsonian animals treated with l-DOPA, high levels of PPENK are a prerequisite for a locomotor sensitization to l-DOPA treatment, while PDYN overexpression is responsible only for the development of dyskinesia.

Topics: Analysis of Variance; Animals; Antiparkinson Agents; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Female; Gene Expression Regulation; Levodopa; Oxidopamine; Parkinson Disease; Protein Precursors; Rats; Rats, Sprague-Dawley; Receptors, Dopamine; RNA, Messenger; Statistics as Topic; Sympatholytics; Tyrosine 3-Monooxygenase

Aconiti brachypodi Radix (Xue-shang-yi-zhi-hao) has been prescribed to manage chronic pain, arthritis, and traumatic injuries. Bullatine A, a C20-diterpenoid alkaloid, is one of its principle effective compounds. This study aimed to investigate the anti-hypersensitivity of bullatine A in a variety of rat pain models and explore its mechanisms of action.. Rat neuropathic pain, inflammatory pain, diabetic neuropathic pain, and bone cancer pain models were used. Dynorphin A and pro-inflammatory cytokines were measured in the spinal cord and cultured primary microglia. Double immunofluorescence staining of dynorphin A and glial and neuronal cellular markers was also measured in the spinal cord.. Subcutaneous and intrathecal injection of bullatine A dose-dependently attenuated spinal nerve ligation-, complete Freud's adjuvant-, diabetes-, and bone cancer-induced mechanical allodynia and thermal hyperalgesia, with the efficacies of 45-70 % inhibition, and half-effective doses of 0.9-1.9 mg/kg for subcutaneous injection. However, bullatine A was not effective in blocking acute nociceptive response in the normal condition. Bullatine A specifically stimulated dynorphin A expression in microglia in the spinal cord in vivo and cultured primary microglia in vitro; the stimulatory effects were completely inhibited by the microglial inhibitor minocycline. In contrast, bullatine A did not have an inhibitory effect on peripheral nerve injury- or lipopolysaccharide-induced pro-inflammatory cytokine expression. The spinal anti-allodynic effects of bullatine A were entirely blocked by intrathecal injection of minocycline, the specific dynorphin A antiserum, and the selective k-opioid receptor antagonist.. We, for the first time, demonstrate that bullatine A specifically attenuates pain hypersensitivity, regardless of the pain models employed. The results also suggest that stimulation of spinal microglial dynorphin A expression mediates bullatine A anti-nociception in pain hypersensitivity conditions.

Topics: Alkaloids; Analgesics; Animals; Animals, Newborn; Cells, Cultured; Disease Models, Animal; Diterpenes; Dose-Response Relationship, Drug; Dynorphins; Female; Gene Expression; Hyperalgesia; Injections, Subcutaneous; Male; Neuralgia; Rats; Rats, Sprague-Dawley; Rats, Wistar; Spinal Cord

During the inflammation which occurs following nerve damage, macrophages are recruited to the site of injury. Phenotypic diversity is a hallmark of the macrophage lineage and includes pro-inflammatory M1 and anti-inflammatory M2 populations. Our aim in this study was to investigate the ability of polarized M0, M1, and M2 macrophages to secrete opioid peptides and to examine their relative contribution to the modulation of neuropathic pain.. Mouse bone marrow-derived cells were cultured as unstimulated M0 macrophages or were stimulated into an M1 phenotype using lipopolysaccharide and interferon-γ or into an M2 phenotype using interleukin-4. The macrophage phenotypes were verified using flow cytometry for surface marker analysis and cytokine bead array for cytokine profile assessment. Opioid peptide levels were measured by radioimmunoassay and enzyme immunoassay. As a model of neuropathic pain, a chronic constriction injury (CCI) of the sciatic nerve was employed. Polarized M0, M1, and M2 macrophages (5 × 10. Compared to M0 and M1 cells, M2 macrophages contained and released higher amounts of opioid peptides, including Met-enkephalin, dynorphin A (1-17), and β-endorphin. M2 cells transferred perineurally at the nerve injury site reduced mechanical, but not heat hypersensitivity following the second injection. The analgesic effect was reversed by the perineurally applied opioid receptor antagonist naloxone methiodide. M2 cells did not affect sensitivity following sham surgery. Neither M0 nor M1 cells altered mechanical and heat sensitivity in CCI or sham-operated animals. Tracing the fluorescently labeled M0, M1, and M2 cells ex vivo showed that they remained in the nerve and preserved their phenotype.. Perineural transplantation of M2 macrophages resulted in opioid-mediated amelioration of neuropathy-induced mechanical hypersensitivity, while M1 macrophages did not exacerbate pain. Therefore, rather than focusing on macrophage-induced pain generation, promoting opioid-mediated M2 actions may be more relevant for pain control.

Topics: Acyltransferases; Adoptive Transfer; Animals; beta-Endorphin; Cell Polarity; Cytokines; Disease Models, Animal; Dynorphins; Flow Cytometry; Histocompatibility Antigens Class II; Hyperalgesia; Lipopolysaccharides; Macrophages; Male; Mice; Mice, Inbred C57BL; Neuralgia; Opioid Peptides; Pain Threshold; Physical Stimulation

Infraorbital nerve constriction (CION) causes hypersensitivity to facial mechanical, heat and cold stimulation in rats and mice and is a reliable model to study trigeminal neuropathic pain. In this model there is evidence that mechanisms operated by kinin B1 and B2 receptors contribute to heat hyperalgesia in both rats and mice. Herein we further explored this issue and assessed the role of kinin receptors in mechanical hyperalgesia after CION. Swiss and C57Bl/6 mice that underwent CION or sham surgery or dynorphin A (1-17) administration were repeatedly submitted to application of either heat stimuli to the snout or mechanical stimuli to the forehead. Treatment of the animals on the fifth day after CION surgery with DALBK (B1 receptor antagonist) or HOE-140 (B2 receptor antagonist), both at 0.01-1μmol/kg (i.p.), effectively reduced CION-induced mechanical hyperalgesia. Knockout mice for kinin B1, B2 or B1/B2 receptors did not develop heat or mechanical hyperalgesia in response to CION. Subarachnoid dynorphin A (1-17) delivery (15nmol/5μL) also resulted in orofacial heat hyperalgesia, which was attenuated by post-treatment with DALBK (1 and 3μmol/kg, i.p.), but was not affected by HOE-140. Additionally, treatment with an anti-dynorphin A antiserum (200μg/5μL, s.a.) reduced CION-induced heat hyperalgesia for up to 2h. These results suggest that both kinin B1 and B2 receptors are relevant in orofacial sensory nociceptive changes induced by CION. Furthermore, they also indicate that dynorphin A could stimulate kinin receptors and this effect seems to contribute to the maintenance of trigeminal neuropathic pain.

Topics: Animals; Bradykinin; Bradykinin B1 Receptor Antagonists; Bradykinin B2 Receptor Antagonists; Disease Models, Animal; Dynorphins; Facial Pain; Hot Temperature; Hyperalgesia; Male; Mice, Inbred C57BL; Mice, Knockout; Neuralgia; Neurotransmitter Agents; Pain Measurement; Receptors, Bradykinin; Touch

Prenatal testosterone (T)-treated ewes display a constellation of reproductive defects that closely mirror those seen in PCOS women, including altered hormonal feedback control of GnRH. Kisspeptin/neurokinin B/dynorphin (KNDy) neurons of the arcuate nucleus (ARC) play a key role in steroid feedback control of GnRH secretion, and prenatal T treatment in sheep causes an imbalance of KNDy peptide expression within the ARC. In the present study, we tested the hypothesis that prenatal T exposure, in addition to altering KNDy peptides, leads to changes in the morphology and synaptic inputs of this population, kisspeptin cells of the preoptic area (POA), and GnRH cells. Prenatal T treatment significantly increased the size of KNDy cell somas, whereas POA kisspeptin, GnRH, agouti-related peptide, and proopiomelanocortin neurons were each unchanged in size. Prenatal T treatment also significantly reduced the total number of synaptic inputs onto KNDy neurons and POA kisspeptin neurons; for KNDy neurons, the decrease was partly due to a decrease in KNDy-KNDy synapses, whereas KNDy inputs to POA kisspeptin cells were unaltered. Finally, prenatal T reduced the total number of inputs to GnRH cells in both the POA and medial basal hypothalamus, and this change was in part due to a decreased number of inputs from KNDy neurons. The hypertrophy of KNDy cells in prenatal T sheep resembles that seen in ARC kisspeptin cells of postmenopausal women, and together with changes in their synaptic inputs and projections to GnRH neurons, may contribute to defects in steroidal control of GnRH observed in this animal model.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Disease Models, Animal; Dynorphins; Female; Gonadotropin-Releasing Hormone; Kisspeptins; Neurokinin B; Neurons; Polycystic Ovary Syndrome; Pregnancy; Prenatal Exposure Delayed Effects; Preoptic Area; Sheep; Testosterone

Spinocerebellar ataxia type 23 is caused by mutations in PDYN, which encodes the opioid neuropeptide precursor protein, prodynorphin. Prodynorphin is processed into the opioid peptides, α-neoendorphin, and dynorphins A and B, that normally exhibit opioid-receptor mediated actions in pain signalling and addiction. Dynorphin A is likely a mutational hotspot for spinocerebellar ataxia type 23 mutations, and in vitro data suggested that dynorphin A mutations lead to persistently elevated mutant peptide levels that are cytotoxic and may thus play a crucial role in the pathogenesis of spinocerebellar ataxia type 23. To further test this and study spinocerebellar ataxia type 23 in more detail, we generated a mouse carrying the spinocerebellar ataxia type 23 mutation R212W in PDYN. Analysis of peptide levels using a radioimmunoassay shows that these PDYN(R212W) mice display markedly elevated levels of mutant dynorphin A, which are associated with climber fibre retraction and Purkinje cell loss, visualized with immunohistochemical stainings. The PDYN(R212W) mice reproduced many of the clinical features of spinocerebellar ataxia type 23, with gait deficits starting at 3 months of age revealed by footprint pattern analysis, and progressive loss of motor coordination and balance at the age of 12 months demonstrated by declining performances on the accelerating Rotarod. The pathologically elevated mutant dynorphin A levels in the cerebellum coincided with transcriptionally dysregulated ionotropic and metabotropic glutamate receptors and glutamate transporters, and altered neuronal excitability. In conclusion, the PDYN(R212W) mouse is the first animal model of spinocerebellar ataxia type 23 and our work indicates that the elevated mutant dynorphin A peptide levels are likely responsible for the initiation and progression of the disease, affecting glutamatergic signalling, neuronal excitability, and motor performance. Our novel mouse model defines a critical role for opioid neuropeptides in spinocerebellar ataxia, and suggests that restoring the elevated mutant neuropeptide levels can be explored as a therapeutic intervention.

Topics: Action Potentials; Age Factors; Animals; Animals, Newborn; Cell Count; Cells, Cultured; Cerebellum; Disease Models, Animal; Dynorphins; Gene Expression Regulation; Mice; Mice, Inbred C57BL; Mice, Transgenic; Movement Disorders; Mutation; Patch-Clamp Techniques; Purkinje Cells; Signal Transduction; Spinocerebellar Degenerations; Synapses

Expression of dynorphin, an endogenous opioid peptide, increases with age and has been associated with cognitive deficits in rodents. Elevated dynorphin levels have been reported in postmortem samples from Alzheimer's disease (AD) patients, and prodynorphin (PDYN) gene polymorphisms might be linked to cognitive function in the elderly. Activation of κ-opioid receptors by dynorphins has been associated with stress-related memory impairments. Interestingly, these peptides can also modulate glutamate neurotransmission and may affect synaptic plasticity underlying memory formation. N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxy-5-methyl-4-isoxazol-propionate (AMPA) ionotropic glutamate receptor levels generally decrease with aging, and their function is impaired in AD.. Here, we compared the impact of aging on ionotropic glutamate receptor levels in the hippocampal formation of wild-type (WT) and Pdyn knock-out (KO) mice.. We observed a significant reduction in GluR1 and GluR2 AMPA receptor subunits in the hippocampal formation of 18- to 25-month-old WT mice in comparison with 6-month-old mice. Conversely, the GluR1 protein level was maintained in old Pdyn KO mice, and the NMDA NR2B subunit level was increased by 42% when compared to old WT animals.. These results suggest that elevated dynorphin expression occurring during aging and AD may mediate cognitive deficits by altering the glutamatergic system integrity.

Topics: Aging; Alzheimer Disease; Animals; Cognition Disorders; Disease Models, Animal; Dynorphins; Hippocampus; Mice; Mice, Inbred C57BL; Mice, Knockout; Receptors, AMPA

Electroacupuncture (EA) is used as a prescription to treat pruritus and atopic dermatitis. Whether EA affects experimental itch in rat models of immunologic or neuronal damages, however, is unknown.. The present study was designed to determine the therapeutic effects of high-frequency EA on atopic dermatitis-like lesions in rats.. Capsaicin (50mg/kg) was subcutaneously administered rat pups within 48h after birth. Rats then underwent 30min of EA at six acupoints (bilateral BL13, and unilateral LI11, ST36, SP10, SP6) every other day (EA group) for 3 weeks. Measurements of IgE, mast cells, scratching behavior, dynorphin release, skin thickness and dermatitis score were obtained.. Only the dermatitis score and dynorphin expression were decreased in the EA group compared with the control non-EA group.. We suggest that high-frequency EA alleviates pruritus of atopic dermatitis-like lesions in rats induced by capsaicin injection, via the release of dynorphin. These findings indicate a new potential therapeutic approach for the amelioration of symptoms of atopic dermatitis.

Topics: Animals; Animals, Newborn; Body Weight; Capsaicin; Dermatitis; Dermatitis, Atopic; Disease Models, Animal; Dynorphins; Electroacupuncture; Immunoglobulin E; Male; Mast Cells; Phenotype; Pruritus; Rats; Skin; Treatment Outcome

Interleukin-17 (IL-17) is involved in a wide range of inflammatory disorders and in recruitment of inflammatory cells to injury sites. A recent study of IL-17 knock-out mice revealed that IL-17 contributes to neuroinflammation and neuropathic pain after peripheral nerve injury. Surprisingly, little is known of micro-environment modulation by IL-17 in injured sites and in pathologically related neuroinflammation and chronic neuropathic pain. Therefore, we investigated nociceptive sensitization, immune cell infiltration, myeloperoxidase (MPO) activity, and expression of multiple cytokines and opioid peptides in damaged nerves of wild-type (IL-17(+/+)) and IL-17 knock-out (IL-17(-/-)) mice after partial sciatic nerve ligation. Our results demonstrated that the IL-17(-/-) mice had less behavioral hypersensitivity after partial sciatic nerve ligation, and inflammatory cell infiltration and pro-inflammatory cytokine (tumor necrosis factor-α, IL-6, and interferon-γ) levels in damaged nerves were significantly decreased, with the levels of anti-inflammatory cytokines IL-10 and IL-13, and expressions of enkephalin, β-endorphin, and dynorphin were also decreased compared to those in wild-type control mice. In conclusion, we provided evidence that IL-17 modulates the micro-environment at the level of the peripheral injured nerve site and regulates progression of behavioral hypersensitivity in a murine chronic neuropathic pain model. The attenuated behavioral hypersensitivity in IL-17(-/-) mice could be a result of decreased inflammatory cell infiltration to the injured site, resulting in modulation of the pro- and anti-inflammatory cytokine milieu within the injured nerve. Therefore, IL-17 may be a critical component for neuropathic pain pathogenesis and a novel target for therapeutic intervention for this and other chronic pain states.

Topics: Animals; Behavior, Animal; beta-Endorphin; Central Nervous System Sensitization; Cytokines; Disease Models, Animal; Dynorphins; Enkephalins; Hyperalgesia; Inflammation; Interleukin-10; Interleukin-13; Interleukin-17; Interleukin-1beta; Interleukin-2; Interleukin-6; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Neuralgia; Neutrophils; Nociception; Peripheral Nerve Injuries; Peroxidase; Sciatic Nerve; T-Lymphocytes; Tumor Necrosis Factor-alpha

Dynorphin A is an endogenous opioid peptide derived from the precursor prodynorphin. The proteolytic fragment dynorphin A (1-17) exhibits inhibitory effects via opioid receptors. Paradoxically, the activity of the dynorphin system increases with chronic pain and neuropathy is associated with the up-regulation of dynorphin biosynthesis. Dynorphin A (1-17) is cleaved in vivo to produce a non-opioid fragment, dynorphin A (2-17). Previously, a mechanism by which the non-opioid fragment promotes pain through agonist action at bradykinin receptors was revealed. Bradykinin receptor expression is up-regulated after nerve injury and both a truncated version of non-opioid fragment dynorphin A (2-17), referred to as 'Ligand 10', and novel bradykinin receptor antagonist 'Ligand 14', are known to bind to the bradykinin receptor. Here we show that Ligand 10 facilitates the response of wide dynamic range (WDR) neurons to innocuous and noxious mechanical stimuli in neuropathic, but not naïve, animals, while Ligand 14 exhibits inhibitory effects in neuropathic animals only. Furthermore, we reveal an inhibitory effect of Ligand 14 in naïve animals by pre-dosing with either Ligand 10 or a 5-HT3 receptor agonist to reflect activation of descending excitatory drives. Thus remarkably, by mimicking pro-excitatory pharmacological changes that occur after nerve injury in a naïve animal, we induce a state whereby the inhibitory actions of Ligand 14 are now effective. Ultimately our data support an increasing number of studies that suggest that blocking spinal bradykinin receptors may have a therapeutic potential in chronic pain states, here, in particular, in neuropathic pain.

Topics: Analgesics, Non-Narcotic; Animals; Bradykinin B2 Receptor Antagonists; Disease Models, Animal; Dynorphins; Male; Neuralgia; Neuronal Plasticity; Neurons; Nociception; Peptide Fragments; Physical Stimulation; Rats, Sprague-Dawley; Serotonin 5-HT3 Receptor Agonists; Spinal Cord; Spinal Nerves

Despite many advances, our understanding of the involvement of prodynorphin systems in the development of neuropathic pain is not fully understood. Recent studies suggest an important role of neuro-glial interactions in the dynorphin effects associated with neuropathic pain conditions. Our studies show that minocycline reduced prodynorphin mRNA levels that were previously elevated in the spinal and/or dorsal root ganglia (DRG) following sciatic nerve injury. The repeated intrathecal administration of minocycline enhanced the analgesic effects of low-dose dynorphin (0.15 nmol) and U50,488H (25-100 nmol) and prevented the development of flaccid paralysis following high-dose dynorphin administration (15 nmol), suggesting a neuroprotective effect. Minocycline reverts the expression of IL-1β and IL-6 mRNA within the spinal cord and IL-1β mRNA in DRG, which was elevated following intrathecal administration of dynorphin (15 nmol). These results suggest an important role of these proinflammatory cytokines in the development of the neurotoxic effects of dynorphin. Similar to minocycline, a selective inhibitor of MMP-9 (MMP-9 levels are reduced by minocycline) exerts an analgesic effect in behavioral studies, and its administration prevents the occurrence of flaccid paralysis caused by high-dose dynorphin administration (15 nmol). In conclusion, our results underline the importance of neuro-glial interactions as evidenced by the involvement of IL-1β and IL-6 and the minocycline effect in dynorphin-induced toxicity, which suggests that drugs that alter the prodynorphin system could be used to better control neuropathic pain.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analgesics, Non-Narcotic; Animals; Disease Models, Animal; Dynorphins; Ganglia, Spinal; Injections, Spinal; Interleukin-1beta; Interleukin-6; Male; Matrix Metalloproteinase 9; Minocycline; Neuralgia; Neuroprotective Agents; Paraplegia; Rats, Wistar; RNA, Messenger; Sciatic Neuropathy; Spinal Cord

The long term use of opioids for the treatment of pain leads to a group of maladaptations which includes opioid-induced hyperalgesia (OIH). OIH typically resolves within few days after cessation of morphine treatment in mice but is prolonged for weeks if histone deacetylase (HDAC) activity is inhibited during opioid treatment. The present work seeks to identify gene targets supporting the epigenetic effects responsible for OIH prolongation.. Mice were treated with morphine according to an ascending dose protocol. Some mice also received the selective HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) additionally. Chronic morphine treatment with simultaneous HDAC inhibition enhanced OIH, and several spinal cord genes were up-regulated. The expression of Bdnf (Brain-derived neurotrophic factor) and Pdyn (Prodynorphin) were most closely related to the observed behavioral changes. ChIP (Chromatin immuoprecipation) assays demonstrated that promoter regions of Pdyn and Bdnf were strongly associated with aceH3K9 (Acetylated histone H3 Lysine9) after morphine and SAHA treatment. Furthermore, morphine treatment caused an increase in spinal BDNF and dynorphin levels, and these levels were further increased in SAHA treated mice. The selective TrkB (tropomyosin-receptor-kinase) antagonist ANA-12 reduced OIH when given one or seven days after cessation of morphine. Treatment with the selective kappa opioid receptor antagonist nor-BNI also reduced established OIH. The co-administration of either receptor antagonist agent daily with morphine resulted in attenuation of hyperalgesia present one day after cessation of treatment. Additionally, repeated morphine exposure induced a rise in BDNF expression that was associated with an increased number of BDNF+ cells in the spinal cord dorsal horn, showing strong co-localization with aceH3K9 in neuronal cells. Lastly, spinal application of low dose BDNF or Dynorphin A after resolution of OIH produced mechanical hypersensitivity, with no effect in controls.. The present study identified two genes whose expression is regulated by epigenetic mechanisms during morphine exposure. Treatments aimed at preventing the acetylation of histones or blocking BDNF and dynorphin signaling may reduce OIH and improve long-term pain using opioids.

Topics: Analgesics, Opioid; Animals; Antineoplastic Agents; Azepines; Benzamides; Brain-Derived Neurotrophic Factor; Disease Models, Animal; Dynorphins; Epigenesis, Genetic; Gene Expression Regulation; Hydroxamic Acids; Hyperalgesia; Male; Mice; Mice, Inbred C57BL; Morphine; Naltrexone; Narcotic Antagonists; Pain Measurement; Spinal Cord; Vorinostat

A previous study of our group demonstrated that movement performance induced by dopamine agonist drugs in hemiparkinsonian rats unilaterally lesioned with 6-hydroxydopamine (6-OHDA), governs the occurrence of a sensitized motor response to a subsequent dopaminergic challenge (priming model). In the present study, we examined the influence of movement performance (rotational behavior) on the molecular events induced by priming in the striatum. To this end, unilaterally 6-OHDA-lesioned rats were primed with apomorphine (0.2 mg/kg) in immobilized or freely moving conditions (priming induction) and 3 days later the D1 receptor agonist SKF 38393 was administered (priming expression). Evaluation of striatal mRNA for enkephalin and dynorphin, markers of the indirect and direct striatonigral pathways, and of GAD67 showed an increase in dynorphin in primed SKF 38393-treated rats, no matter whether immobilized or freely moving during priming induction, whilst enkephalin and GAD67 did not show any changes. In contrast, evaluation of mRNA for the early gene zif-268 in the striatum showed a generalized increase after administration of SKF 38393, in both primed and unprimed rats. However, examination of zif-268 mRNA at the single-cell level, showed that only dynorphin(+) neurons of primed not immobilized rats displayed a significantly higher number of zif-268-positive silver grains in response to the SKF 38393 challenge. This selective activation of zif-268 in dynorphinergic striatonigral efferent neurons demonstrates that movement performance in response to dopaminergic drug administration under conditions of dopamine denervation is critical for the emergence of neurochemical modifications in selected striatal efferent neurons. Furthermore, these results may provide information on the first initial molecular events taking place in the complex processes that lead to dyskinetic movements in Parkinson's disease.

Topics: 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Adrenergic Agents; Analysis of Variance; Animals; Apomorphine; Corpus Striatum; Disease Models, Animal; Dopamine Agonists; Dynorphins; Early Growth Response Protein 1; Enkephalins; Functional Laterality; Gene Expression Regulation; Male; Motor Neurons; Movement; Oxidopamine; Parkinson Disease; Rats; Rats, Sprague-Dawley; RNA, Messenger

Parkinson's disease (PD) is an asymmetric neurodegenerative disorder, and secondary adaptive mechanisms of the less-affected side could potentially compensate for parkinsonian symptoms. Here, we analyzed gene expression changes on the healthy side of a unilateral PD rat model and correlated these changes with locomotor velocity, which is known to be decreased in PD. Four weeks after a unilateral 6-hydroxydopamine lesion, the spontaneous locomotor velocity of rats was recorded just prior to brain extraction. We then analyzed the gene expression levels of markers of the direct (dynorphin and D1-class dopamine receptors) and indirect (enkephalin and D2-class dopamine receptors) pathways in the contralateral healthy striatum by in situ hybridization histochemistry. In addition, we analyzed the expression of several striatal and cortical glutamatergic markers, as well as nigral tyrosine hydroxylase (TH) and nigral dopamine transporter (DAT). We found a significant positive correlation between the mRNA expression levels of contralateral D1-class dopamine receptors and the mean locomotor velocity, at 4 weeks after surgery in parkinsonian rats but not in controls. Moreover, we observed a significant increase in the level of dynorphin mRNA in the lateral part of the contralateral striatum of parkinsonian rats compared to the controls. In contrast, no contralateral changes were observed in the striatal indirect pathway. We also did not find any significant contralateral modifications of TH, DAT or glutamatergic markers in PD animals, indicating that changes in direct pathway genes are not due to nigrostriatal dopaminergic or corticostriatal glutamatergic innervation. In conclusion, our results suggest a role of the healthy striatal direct pathway in counteracting dopaminergic denervation effects on motor symptoms.

Topics: Adaptation, Physiological; Adrenergic Agents; Animals; Cerebral Cortex; Disease Models, Animal; Dopamine Plasma Membrane Transport Proteins; Dynorphins; Enkephalins; Female; Functional Laterality; Gene Expression Profiling; Locomotion; Neostriatum; Nerve Tissue Proteins; Neural Pathways; Oxidopamine; Parkinsonian Disorders; Random Allocation; Rats; Rats, Sprague-Dawley; Receptors, Dopamine D1; Receptors, Dopamine D2; RNA, Messenger; Tyrosine 3-Monooxygenase

In a previous study, steady-state methadone treatment was found to prevent associative cocaine learning, as well as related decreases in mRNA expression of preprohypocretin/preproorexin (ppHcrt) in the lateral hypothalamus (LH) and dopamine D2 receptor (DR2) in the caudate-putamen (CP), and increases in mu-opioid receptor in the ventral striatum of rats. To investigate whether the same regimen of methadone exposure could prevent the incubation of cocaine sensitization and related alterations in gene expression, male Sprague-Dawley rats received 45 mg/kg/day steady-dose "binge" cocaine administration (IP) for 14 days followed by mini-pumps releasing 30 mg/kg/day methadone (SC). After 14 days of methadone, and a subsequent 10-day drug-free period, all rats were tested for sensitization (cocaine test dose: 15 mg/kg) and brain tissue was collected to quantify mRNA expression. Rats exposed to cocaine displayed cocaine-induced stereotypy at test, as well as enhanced ppHcrt mRNA in the LH and reduced DR2 mRNA in the CP. Importantly, these alterations were significantly reduced in rats treated with methadone following cocaine. These results suggest that steady-state methadone can interfere with the incubation of neuroadaptations underlying changes in behavioral responses to cocaine and cocaine-associated stimuli, and that these effects can be observed even after withdrawal from methadone.

Topics: Analgesics, Opioid; Animals; Brain; Cocaine; Cocaine-Related Disorders; Disease Models, Animal; Dopamine Uptake Inhibitors; Drug Delivery Systems; Dynorphins; Gene Expression; Locomotion; Male; Methadone; Rats; Rats, Sprague-Dawley; Receptors, Dopamine D2; Receptors, Opioid, mu; RNA, Messenger; Stereotyped Behavior

The opiate withdrawal syndrome is a severe stressor that powerfully triggers addictive drug intake. However, no treatment yet exists that effectively relieves opiate withdrawal distress and spares stress-coping abilities. The corticotropin-releasing factor (CRF) system mediates the stress response, but its role in opiate withdrawal distress and bodily strategies aimed to cope with is unknown. CRF-like signaling is transmitted by two receptor pathways, termed CRF(1) and CRF(2). Here, we report that CRF(2) receptor-deficient (CRF(2)(-/-)) mice lack the dysphoria-like and the anhedonia-like states of opiate withdrawal. Moreover, in CRF(2)(-/-) mice opiate withdrawal does not increase the activity of brain dynorphin, CRF and periaqueductal gray circuitry, which are major substrates of opiate withdrawal distress. Nevertheless, CRF(2) receptor-deficiency does not impair brain, neuroendocrine and autonomic stress-coping responses to opiate withdrawal. The present findings point to the CRF(2) receptor pathway as a unique target to relieve opiate withdrawal distress without impairing stress-coping abilities.

Topics: Adaptation, Psychological; Animals; Behavior, Addictive; Brain; Corticosterone; Corticotropin-Releasing Hormone; Disease Models, Animal; Dynorphins; Mice; Mice, Inbred C57BL; Mice, Knockout; Opioid-Related Disorders; Receptors, Corticotropin-Releasing Hormone; Stress, Psychological; Substance Withdrawal Syndrome; Tyrosine 3-Monooxygenase

Anhedonia is a core symptom of clinical depression. Two brain neuropeptides that have been implicated in anhedonia symptomology in preclinical depression models are dynorphin and orexin; which are concentrated along lateral hypothalamic dopamine reward pathways. These affect regulating neuropeptides modulate each other's function, implicating an interactive dysfunction between them in anhedonia symptomology. But whether their influences are modified or imbalanced within the hypothalamus or dopamine system in anhedonic preclinical depression models is not yet clear. We used radioimmunoassay to determine this in the rat social defeat model of depression; at a time that anhedonic sexual disinterest was expressed. In tissue samples of the medial prefrontal cortex (mPFC), ventral tegmental area (VTA) and nucleus accumbens, basal dynorphin levels were similar to normal animals. But orexin was reduced in the VTA and mPFC. Also, dynorphin and orexin were both diminished in the hypothalamus which is noteworthy since nearly all hypothalamic orexin cells co-express dynorphin. These findings suggest that orexin and dynorphin function may be imbalanced between the hypothalamus and mesocortical dopaminergic brain regions in depression.

Topics: Anhedonia; Animals; Brain; Depression; Disease Models, Animal; Dopamine; Dynorphins; Enzyme-Linked Immunosorbent Assay; Hypothalamus; Intracellular Signaling Peptides and Proteins; Male; Neuropeptides; Orexins; Prefrontal Cortex; Radioimmunoassay; Rats; Rats, Long-Evans; Ventral Tegmental Area

The aim of the present study was to evaluate whether eugenol, the main constituent of clove oil, has the capacity to provide analgesia in the monoiodoacetate-induced rat model of osteoarthritis. Animals (n = 6/group) received either eugenol (20 or 40 mg/kg) or a vehicle by gavage. Daily administrations were initiated 2 days post osteoarthritis induction and continued for the duration of the study (4 weeks). Gait analysis was performed using the CatWalk method and secondary mechanical allodynia was assessed with von Frey filaments. Selected spinal cord peptides (substance P, calcitonin gene-related peptide and dynorphin) were quantified by mass spectrometry. Significant changes were identified in dynamic gait parameters (swing speed, swing phase duration and duty cycle) of the affected limb following 40 mg/kg eugenol treatment compared with the vehicle (p < 0.05). Von Frey results revealed significant differences between the 40 mg/kg treatment and the vehicle group during the first and the third week of the study (p < 0.02). Spinal pain-related peptide analysis revealed a decreased content of substance P and CGRP accompanied by an increase of dynorphin in animals treated with 40 mg/kg eugenol. These results suggest a therapeutic potential of eugenol to alleviate osteoarthritis-related pain.

Topics: Analgesics; Animals; Calcitonin Gene-Related Peptide; Clove Oil; Disease Models, Animal; Dynorphins; Eugenol; Gait; Hyperalgesia; Iodoacetic Acid; Knee Joint; Male; Osteoarthritis; Rats; Rats, Sprague-Dawley; Spinal Cord; Substance P

Adenosine A(₂A) receptor antagonism provides a promising approach to developing nondopaminergic therapy for Parkinson's disease (PD). Clinical trials of A(₂A) antagonists have targeted PD patients with L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia (LID) in an effort to improve parkinsonian symptoms. The role of adenosine in the development of LID is little known, especially regarding its actions via A₁ receptors. We aimed to examine the effects of genetic deletion and pharmacological blockade of A₁ and/or A(₂A) receptors on the development of LID, on the induction of molecular markers of LID including striatal preprodynorphin and preproenkephalin (PPE), and on the integrity of dopaminergic nigrostriatal neurons in hemiparkinsonian mice. Following a unilateral 6-hydroxydopamine lesion A₁, A(₂A) and double A₁-A(₂A) knockout (KO) and wild-type littermate mice, and mice pretreated with caffeine (an antagonist of both A₁ and A(₂A) receptors) or saline were treated daily for 18-21 days with a low dose of L-DOPA. Total abnormal involuntary movements (AIMs, a measure of LID) were significantly attenuated (p<0.05) in A₁ and A(₂A) KOs, but not in A₁-A(₂A) KOs and caffeine-pretreated mice. An elevation of PPE mRNA ipsilateral to the lesion in WT mice was reduced in all KO mice. In addition, neuronal integrity assessed by striatal dopamine content was similar in all KOs and caffeine-pretreated mice following 6-hydroxydopamine lesioning. Our findings raise the possibility that A₁ or A(₂A) receptors blockade might also confer a disease-modifying benefit of reduced risk of disabling LID, whereas the effect of their combined inactivation is less clear.

Topics: Adrenergic Agents; Animals; Antiparkinson Agents; Caffeine; Corpus Striatum; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Gene Expression Regulation; Levodopa; Mice; Mice, Inbred C57BL; Mice, Knockout; Oxidopamine; Parkinson Disease; Protein Precursors; Purinergic P1 Receptor Antagonists; Receptor, Adenosine A1; Receptor, Adenosine A2A; RNA, Messenger; Statistics, Nonparametric; Time Factors

The kappa opioid receptor (KOR) system contributes to the prodepressive and aversive consequences of stress and is implicated in the facilitation of conditioned fear and anxiety in rodents. Here, we sought to identify neural circuits that mediate KOR system effects on fear and anxiety in rats.. We assessed whether fear conditioning induces plasticity in KOR or dynorphin (the endogenous KOR ligand) messenger RNA (mRNA) expression in the basolateral (BLA) and central (CeA) nuclei of the amygdala, hippocampus, or striatum. We then assessed whether microinfusions of the KOR antagonist JDTic (0-10 μg/side) into the BLA or CeA affect the expression of conditioned fear or anxiety. Finally, we examined whether fear extinction induces plasticity in KOR mRNA expression that relates to the quality of fear extinction.. Fear conditioning upregulated KOR mRNA in the BLA by 65% and downregulated it in the striatum by 22%, without affecting KOR levels in the CeA or hippocampus, or dynorphin levels in any region. KOR antagonism in either the BLA or CeA decreased conditioned fear in the fear-potentiated startle paradigm, whereas KOR antagonism in the BLA, but not the CeA, produced anxiolytic-like effects in the elevated plus maze. Effective fear extinction was associated with a 67% reduction in KOR mRNA in the BLA.. These findings suggest that fear conditioning and extinction dynamically regulate KOR expression in the BLA and provide evidence that the BLA and CeA are important neural substrates mediating the anxiolytic-like effects of KOR antagonists in models of fear and anxiety.

Topics: Amygdala; Animals; Anxiety; Conditioning, Psychological; Corpus Striatum; Disease Models, Animal; Dynorphins; Extinction, Psychological; Fear; Gene Expression Regulation; Hippocampus; Male; Maze Learning; Microinjections; Piperidines; Rats; Receptors, Opioid, kappa; Reflex, Startle; Signal Transduction; Tetrahydroisoquinolines

Dystonia is regarded as a basal ganglia disorder. In the dt(sz) hamster, a genetic animal model of paroxysmal dystonia, previous studies demonstrated a reduced density of striatal GABAergic interneurons which inhibit striatal GABAergic projection neurons. Although the disinhibition of striatal GABAergic projection neurons was evidenced in the dt(sz) hamster, alterations in their density have not been elucidated so far. Therefore, in the present study, the density of striatal methionin-(met-) enkephalin (ENK) immunoreactive GABAergic neurons, which project to the globus pallidus (indirect pathway), was determined in dt(sz) and control hamsters to clarify a possible role of an altered ratio between striatal interneurons and projection neurons. Furthermore, the immunoreactivity of dynorphin A (DYN), which is expressed in entopeduncular fibers of striatal neurons of the direct pathway, was verified by gray level measurements to illuminate the functional relevance of an enhanced striato-entopeduncular neuronal activity previously found in dt(sz) hamsters. While the density of striatal ENK immunoreactive (ENK(+) ) neurons did not significantly differ between mutant and control hamsters, there was a significantly enhanced ratio between the DYN immunoreactive area and the whole area of the EPN in dt(sz) hamsters compared to controls. These results support the hypothesis that a disbalance between a reduced density of striatal interneurons and an unchanged density of striatal projection neurons causes imbalances in the basal ganglia network. The consequentially enhanced striato-entopeduncular inhibition leads to an already evidenced reduced activity and an altered firing pattern of entopeduncular neurons in the dt(sz) hamster.

Topics: Animals; Animals, Genetically Modified; Basal Ganglia; Cricetinae; Disease Models, Animal; Dynorphins; Dystonia; Enkephalins; Female; Interneurons; Male; Mesocricetus; Neurons

The aim of this study was to investigate the underlying mechanism that dynorphin, an endogenous kappa opioid receptor (κ-OR) agonist, triggers antiapoptotic effect of postconditioning (Postcon). In addition to vehicle treatment, Sprague Dawley rats (n = 6) underwent a 30-minute left anterior descending occlusion followed by 2 hours of reperfusion with or without a Postcon stimulus. The selective κ-OR antagonist nor-binaltorphimine (Nor-BNI) was administered intravenously 5 minutes before reperfusion. Infarct size was determined by using 2,3,5-triphenyltetrazolium chloride staining. Blood plasma concentrations of creatine kinase (CK) and lactate dehydrogenase (LDH) and myocardial caspase-3 activity were analyzed spectrophotometrically. Myocardial apoptosis was analyzed by the detection of terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick-end labeling. Immunoreactive dynorphin in blood serum and myocardium was measured by means of an antigen-competitive enzyme-linked immunosorbent assay. Infarction size, caspase-3 activity, apoptotic index, and CK and LDH levels were significantly higher in the ischemic/reperfusion group than in the vehicle group (P < 0.01). Postcon significantly reduced infarction size, caspase-3 activity, apoptotic index, CK and LDH levels (P < 0.01 vs. ischemic/reperfusion). Dynorphin content significantly increased after Postcon (P < 0.01). All the effects described above were abolished by Nor-BNI, with the exception of dynorphin content. We found that cardiac protection and antiapoptotic effect of Postcon is mediated by the activation of κ-OR. Effect of Postcon is mediated, at least partially, by enhanced dynorphin expression.

Topics: Animals; Apoptosis; Disease Models, Animal; Dynorphins; Hemodynamics; In Situ Nick-End Labeling; Ischemic Postconditioning; Male; Myocardial Infarction; Myocardial Reperfusion Injury; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa

Opioid peptides are involved in various pathophysiological processes, including algesia, epilepsy, and drug dependence. A strong association between L-DOPA-induced dyskinesia (LID) and elevated prodynorphin mRNA levels has been established in both patients and in animal models of Parkinson's disease, but to date the endogenous prodynorphin peptide products have not been determined. Here, matrix-assisted laser desorption ionization (MALDI) imaging mass spectrometry (IMS) was used for characterization, localization, and relative quantification of striatal neuropeptides in a rat model of LID in Parkinson's disease. MALDI IMS has the unique advantage of high sensitivity and high molecular specificity, allowing comprehensive detection of multiple molecular species in a single tissue section. Indeed, several dynorphins and enkephalins could be detected in the present study, including dynorphin A(1-8), dynorphin B, α-neoendorphin, MetEnkRF, MetEnkRGL, PEnk (198-209, 219-229). IMS analysis revealed elevated levels of dynorphin B, α-neoendorphin, substance P, and PEnk (220-229) in the dorsolateral striatum of high-dyskinetic animals compared with low-dyskinetic and lesion-only control rats. Furthermore, the peak-intensities of the prodynorphin derived peptides, dynorphin B and α-neoendorphin, were strongly and positively correlated with LID severity. Interestingly, these LID associated dynorphin peptides are not those with high affinity to κ opioid receptors, but are known to bind and activate also μ- and Δ-opioid receptors. In addition, the peak intensities of a novel endogenous metabolite of α-neoendorphin lacking the N-terminal tyrosine correlated positively with dyskinesia severity. MALDI IMS of striatal sections from Pdyn knockout mice verified the identity of fully processed dynorphin peptides and the presence of endogenous des-tyrosine α-neoendorphin. Des-tyrosine dynorphins display reduced opioid receptor binding and this points to possible novel nonopioid receptor mediated changes in the striatum of dyskinetic rats. Because des-tyrosine dynorphins can only be detected by mass spectrometry, as no antibodies are available, these findings highlight the importance of MALDI IMS analysis for the study of molecular dynamics in neurological diseases.

Topics: Animals; Antiparkinson Agents; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Female; Humans; Levodopa; Mice; Neostriatum; Parkinson Disease; Protein Precursors; Rats; Rats, Sprague-Dawley; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Animal models are useful to evaluate pharmacological therapies to alleviate joint pain. The present study characterized central neuropeptides modulation in the monoiodoacetate (MIA) rat model. Animals receiving a single 3mg MIA injection were euthanized at 3, 7, 14, 21 and 28 days post injection. Spinal cords were analyzed by liquid chromatography ion trap mass spectrometry. Up-regulations of the calcitonin gene-related peptide and substance P were observed starting on days 7 and 28 respectively, whereas big dynorphin(₁₋₃₂) content decreased significantly on day 14 in comparison to control animals (P<0.05). Preclinical drug evaluations using this model should be conducted between 7 and 21 days post injection when the lesions resemble most to human osteoarthritis.

Topics: Animals; Calcitonin Gene-Related Peptide; Chromatography, Liquid; Disease Models, Animal; Dynorphins; Enzyme Inhibitors; Humans; Iodoacetic Acid; Knee Joint; Male; Neuropeptides; Osteoarthritis; Pain; Rats; Rats, Sprague-Dawley; Spinal Cord; Substance P; Tandem Mass Spectrometry; Time Factors

Dopamine denervation in Parkinson's disease and repeated Levodopa (L-DOPA) administration that induces dyskinesias are associated with an enhancement of basal ganglia neuropeptide transmission. Various adjunct non-dopaminergic treatments to Levodopa were shown to reduce and/or prevent dyskinesias. The aim of this study was to seek if non-dopaminergic drug treatments to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) lesioned monkeys combined with L-DOPA to prevent dyskinesia were associated with changes of striatal neuropeptides. Chronic treatment with Ro 61-8048 a kynurenine hydroxylase inhibitor, docosahexaenoic acid (DHA) a polyunsaturated fatty acid (omega-3), naltrexone an opioidergic antagonist and CI-1041 an N-methyl-D-aspartate (NMDA) glutamate receptor antagonist with L-DOPA prevented dyskinesias to various extents except naltrexone whereas all MPTP monkeys treated with L-DOPA alone developed dyskinesias. Striatal preproenkephalin (PPE), preprodynorphin (PPD) and preprotachykinin A (PPT-A) mRNA levels were measured by in situ hybridization. An increase of PPE and PPD mRNA levels was observed in anterior caudate nucleus of L-DOPA treated MPTP monkeys compared to controls and to Saline-treated MPTP monkeys whereas PPT-A mRNA levels were unchanged. Striatal PPE and PPD mRNA levels remained elevated in L-DOPA plus naltrexone-treated MPTP monkeys, while co-treatment with DHA, CI-1041 or Ro 61-8048 prevented their increase to various extents. Maximal dyskinesias scores of MPTP monkeys correlated significantly with striatal PPE and PPD mRNA levels but not with PPT-A mRNA levels. These results show that drugs displaying a wide range of pharmacological activities can modulate L-DOPA induced dyskinesias and this activity is correlated with striatal PPD and PPE mRNA levels suggesting a convergent mechanism.

Topics: Animals; Antiparkinson Agents; Benzoxazoles; Cocaine; Corpus Striatum; Disease Models, Animal; Docosahexaenoic Acids; Dopamine; Dopamine Uptake Inhibitors; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Female; Iodine Isotopes; Levodopa; Macaca fascicularis; Naltrexone; Neuropeptides; Ovariectomy; Parkinsonian Disorders; Piperidines; Protein Precursors; RNA, Messenger; Sulfonamides; Tachykinins; Thiazoles; Time Factors

The potential neuroprotective efficacy of dynorphin A antiserum on BBB dysfunction, edema formation and brain pathology was examined in a closed head injury (CHI) model in the rat. The CHI was produced by an impact of 0.224 N on the right parietal bone under anesthesia by dropping a weight of 114.6 g on the skull from a height of 20 cm through a guide tube. This concussive brain injury resulted in profound BBB disruption as evidenced by leakage of Evans blue and radioiodine in the brain. Edema formation and swelling at 5 h were most pronounced in the contralateral cerebral hemisphere. Pretreatment with dynorphin A antiserum (1:20, monoclonal) infused into the left lateral cerebral ventricle (30 microL in PBS) either 30 min before or 30 min after CHI significantly attenuated BBB dysfunction, brain edema formation, volume swelling and brain pathology. However, no reduction in brain edema, BBB permeability or improved brain pathology was seen when the antiserum was given 60 min post-CHI. These observations are the first to suggest that antiserum to dynorphin when administered into the CSF during early phase of CHI is neuroprotective. Our work further indicates that dynorphin is actively involved in the cellular and molecular mechanisms of edema formation and BBB breakdown in CHI.

Topics: Analysis of Variance; Animals; Antibodies; Blood-Brain Barrier; Brain Edema; Disease Models, Animal; Dynorphins; Head Injuries, Closed; Male; Rats

Stress exposure increases the risk of addictive drug use in human and animal models of drug addiction by mechanisms that are not completely understood. Mice subjected to repeated forced swim stress (FSS) before cocaine develop significantly greater conditioned place preference (CPP) for the drug-paired chamber than unstressed mice. Analysis of the dose dependency showed that FSS increased both the maximal CPP response and sensitivity to cocaine. To determine whether FSS potentiated CPP by enhancing associative learning mechanisms, mice were conditioned with cocaine in the absence of stress, then challenged after association was complete with the kappa-opioid receptor (KOR) agonist U50,488 or repeated FSS, before preference testing. Mice challenged with U50,488 60 min before CPP preference testing expressed significantly greater cocaine-CPP than saline-challenged mice. Potentiation by U50,488 was dose and time dependent and blocked by the KOR antagonist norbinaltorphimine (norBNI). Similarly, mice subjected to repeated FSS before the final preference test expressed significantly greater cocaine-CPP than unstressed controls, and FSS-induced potentiation was blocked by norBNI. Novel object recognition (NOR) performance was not affected by U50,488 given 60 min before assay, but was impaired when given 15 min before NOR assay, suggesting that KOR activation did not potentiate CPP by facilitating memory retrieval or expression. The results from this study show that the potentiation of cocaine-CPP by KOR activation does not result from an enhancement of associative learning mechanisms and that stress may instead enhance the rewarding valence of cocaine-associated cues by a dynorphin-dependent mechanism.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analgesics, Non-Narcotic; Analysis of Variance; Anesthetics, Local; Animals; Association Learning; Behavior, Animal; Cocaine; Conditioning, Operant; Cues; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Schedule; Drug Interactions; Dynorphins; Exploratory Behavior; Male; Mental Recall; Mice; Mice, Inbred C57BL; Naltrexone; Narcotic Antagonists; Receptors, Opioid, kappa; Reward; Stress, Psychological; Swimming

To observe the analgesic effects of moxibustion in rats with chronic visceral hyperalgesia and its influence on the concentration of dynorphin (Dyn) and endomorphin (EM) in spinal cord.. The rat model of chronic visceral hyperalgesia was established by colorectal distention (CRD). In moxibustion (MX) group, moxibustion was applied once daily for 7 d; in sham moxibustion (SM) group, moxibustion was given to the same acupoints but with the non-smoldered end of the moxa stick. Model control (MC) group and normal control group were also studied. The scoring system of abdominal withdrawal reflex was used to evaluate visceral pain for behavioral assessment. Enzyme linked immunosorbent assay was performed to determine the concentrations of Dyn and EM in spinal cord.. Moxibustion significantly decreased visceral pain to CRD in this rat model, and no significant difference was detected between the SM group and the MC group. In MX group, moxibustion also increased the concentrations of Dyn and EM in spinal cord, and no significant difference was found between the SM group and the MC group.. Moxibustion therapy can significantly enhance the pain threshold of rats with chronic visceral hyperalgesia, and the effect may be closely related to the increased concentration of Dyn and EM in spinal cord.

Topics: Analgesics, Opioid; Animals; Animals, Newborn; Disease Models, Animal; Dynorphins; Humans; Hyperalgesia; Male; Moxibustion; Oligopeptides; Pain Threshold; Rats; Rats, Sprague-Dawley; Reflex, Abdominal; Viscera

Opioids/opiates are commonly administered to alleviate pain, unload the heart, or decrease breathlessness in patients with advanced heart failure. As such, it is important to evaluate whether the myocardial opioidergic system is altered in cardiac disease. A hamster model of spontaneous hypertension was investigated before the development of hypertension (1 mo of age) and in the hypertensive state (10 mo of age) to evaluate the effect of prolonged hypertension on myocardial opioidergic activity. Plasma beta-endorphin was decreased before the development of hypertension and in the hypertensive state (P < 0.05). There was no change in cardiac beta-endorphin content at either time point. No differences were detected in cardiac or plasma dynorphin A, Met-enkephalin, or Leu-enkephalin, or in cardiac peptide expression of kappa- or delta-opioid receptors. mu-Opioid receptor was not detected in either model. To determine how hypertension affects myocardial opioid signaling, the ex vivo work-performing heart was used to assess the cardiac response to opioid administration in healthy hearts and those subjected to chronic hypertension. Agonists selective for the kappa- and delta-opioid receptors, but not mu-opioid receptors, induced a concentration-dependent decrease in cardiac function. The decrease in left ventricular systolic pressure on administration of the kappa-opioid receptor-selective agonist, U50488H, was attenuated in hearts from hamsters subjected to chronic, untreated hypertension (P < 0.05) compared with control. These results show that peripheral and myocardial opioid expression and signaling are altered in hypertension.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Antihypertensive Agents; Benzamides; beta-Endorphin; Blood Pressure; Cricetinae; Cyclic AMP; Disease Models, Animal; Dynorphins; Enkephalin, Leucine; Enkephalin, Methionine; Hypertension; Hypertrophy, Left Ventricular; Myocardial Contraction; Myocardium; Piperazines; Receptors, Opioid, delta; Receptors, Opioid, kappa; Systole; Ventricular Remodeling

The clinically important opioid fentanyl, administered acutely, enhances mechanical hypersensitivity in a model of surgical pain induced by plantar incision. Activity of neurokinin-1 (NK-1) receptor-expressing ascending spinal neurons, descending pathways originating in the rostral ventromedial medulla (RVM), and spinal dynorphin are necessary for the development and maintenance of hyperalgesia during sustained morphine exposure, suggesting that these mechanisms may also be important in opioid enhancement of surgical pain. Therefore, we examined the roles of these mechanisms in sensory hypersensitivity produced by acute fentanyl administration in rats not undergoing surgical incision and in rats undergoing plantar incision. In non-operated rats, fentanyl induced analgesia followed by immediate and long-lasting sensory hypersensitivity, as previously described. Fentanyl also enhanced pain sensitivity induced by plantar incision. Ablation of NK-1-expressing spinal neurons by pre-treatment with substance P-Saporin reduced sensory hypersensitivity in fentanyl-treated rats and, to a lesser extent, in fentanyl-treated rats with a surgical incision. Microinjection of lidocaine into the RVM completely reversed fentanyl-induced sensory hypersensitivity and fentanyl enhancement of incision-induced sensory hypersensitivity. RVM lidocaine injection resulted in a slight reduction of incision-induced sensory hypersensitivity in the absence of fentanyl pre-treatment. Spinal dynorphin content increased by 30 +/- 7% and 66 +/- 17% in fentanyl- and fentanyl/incision-treated rats. Spinal administration of antiserum to dynorphin attenuated sensory hypersensitivity in fentanyl-treated rats. These data support a partial role of NK-1 receptor-containing ascending pathways and a crucial role of descending facilitatory pathways in fentanyl-induced hyperalgesia and in the enhanced hyperalgesia produced by fentanyl treatment following surgical incision.

Topics: Analgesics, Opioid; Anesthetics, Local; Animals; Disease Models, Animal; Dynorphins; Fentanyl; Immunohistochemistry; Lidocaine; Male; Medulla Oblongata; Neural Pathways; Neurons; Pain Threshold; Pain, Postoperative; Rats; Rats, Sprague-Dawley; Receptors, Neurokinin-1; Spinal Cord

Rosiglitazone is a commonly prescribed insulin-sensitizing drug with a selective agonistic activity on the peroxisome proliferator-activated receptor-gamma (PPAR-gamma). PPAR-gamma can modulate inflammatory responses in the brain, and agonists might be beneficial in neurodegenerative diseases. In the present study we used a chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine plus probenecid (MPTPp) mouse model of progressive Parkinson's disease (PD) to assess the therapeutic efficacy of rosiglitazone on behavioural impairment, neurodegeneration and inflammation. Mice chronically treated with MPTPp displayed typical features of PD, including impairment of motor and olfactory functions associated with partial loss of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra pars compacta (SNc), decrease of dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) content and dynorphin (Dyn) mRNA levels in the caudate-putamen (CPu), intense microglial and astroglial response in the SNc and CPu. Chronic rosiglitazone, administered in association with MPTPp, completely prevented motor and olfactory dysfunctions and loss of TH-positive cells in the SNc. In the CPu, loss of striatal DA was partially prevented, whereas decreases in DOPAC content and Dyn were fully counteracted. Moreover, rosiglitazone completely inhibited microglia reactivity in SNc and CPu, as measured by CD11b immunostaining, and partially inhibited astroglial response assessed by glial fibrillary acidic protein immunoreactivity. Measurement of striatal MPP+ levels 2, 4, 6 h and 3 days after chronic treatment indicated that MPTP metabolism was not altered by rosiglitazone. The results support the use of PPAR-gamma agonists as a putative anti-inflammatory therapy aimed at arresting PD progression, and suggest that assessment in PD clinical trials is warranted.

Topics: 3,4-Dihydroxyphenylacetic Acid; Animals; Catalepsy; CD11b Antigen; Chromatography, High Pressure Liquid; Chronic Disease; Disease Models, Animal; Dopamine; Drug Interactions; Dynorphins; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Globus Pallidus; Locomotion; Male; Mice; Mice, Inbred C57BL; Motor Activity; MPTP Poisoning; PPAR gamma; Psychomotor Performance; RNA, Messenger; Rosiglitazone; Smell; Tandem Mass Spectrometry; Thiazolidinediones; Tyrosine 3-Monooxygenase

Experimental evidence suggests that ethanol alters the activity of the endogenous opioid peptide systems in a dose and brain-region dependent manner. These alterations may influence the processes of ethanol reward and reinforcement. Thus, it was the objective of this study to investigate the response of the 3 major opioid peptide systems (endorphins, enkephalins, and dynorphins) to acute ethanol administration, at the level of the midbrain including the ventral tegmental area (midbrain/VTA), a region important for drug, including ethanol reinforcement.. Using the in vivo microdialysis technique coupled with specific solid-phase radioimmunoassay for beta-endorphin, met-enkephalin, and dynorphin A(1-8,) changes in the extracellular concentration of theses peptides at the level of midbrain/VTA were determined at distinct time points following the administration of 0.0 (saline), 0.8, 1.2, 1.6, 2.0, and 2.4 g ethanol/kg B.Wt.. A biphasic effect of ethanol on beta-endorphin release was found, with low to medium (1.2, 1.6, and 2.0 g) but not high (2.4 g) doses of ethanol, inducing a significant increase in the dialysate content of beta-endorphin. A late increase in the dialysate content of dynorphin A(1-8) was observed in response to the 1.2 g ethanol dose. However, none of the ethanol doses tested significantly altered the content of met-enkephalin in the dialysate.. The present findings suggest that the ethanol-induced increase of beta-endorphin release at the level of midbrain/VTA may influence alcohol reinforcement.

Topics: Alcoholism; Animals; beta-Endorphin; Central Nervous System Depressants; Disease Models, Animal; Dose-Response Relationship, Drug; Dynorphins; Enkephalin, Methionine; Ethanol; Injections, Intraperitoneal; Male; Mesencephalon; Opioid Peptides; Peptide Fragments; Rats; Rats, Sprague-Dawley; Time Factors; Ventral Tegmental Area

Pharmacologic studies have implicated dopamine D1-like receptors in the development of dopamine precursor molecule 3,4-dihydroxyphenyl-L-alanine (L-DOPA)-induced dyskinesias and associated molecular changes in hemiparkinsonian mice. However, pharmacologic agents for D1 or D2 receptors also recognize other receptor family members. Genetic inactivation of the dopamine D1 or D2 receptor was used to define the involvement of these receptor subtypes.. During a 3-week period of daily L-DOPA treatment (25 mg/kg), mice were examined for development of contralateral turning behavior and dyskinesias. L-DOPA-induced changes in expression of signaling molecules and other proteins in the lesioned striatum were examined immunohistochemically.. Chronic L-DOPA treatment gradually induced rotational behavior and dyskinesia in wildtype hemiparkinsonian mice. Dyskinetic symptoms were associated with increased FosB and dynorphin expression, phosphorylation of extracellular signal-regulated kinase, and phosphoacetylation of histone 3 (H3) in the lesioned striatum. These molecular changes were restricted to striatal areas with complete dopaminergic denervation and occurred only in dynorphin-containing neurons of the direct pathway. D1 receptor inactivation abolished L-DOPA-induced dyskinesias and associated molecular changes. Inactivation of the D2 receptor had no significant effect on the behavioral or molecular response to chronic L-DOPA.. Our results demonstrate that the dopamine D1 receptor is critical for the development of L-DOPA-induced dyskinesias in mice and in the underlying molecular changes in the denervated striatum and that the D2 receptor has little or no involvement. In addition, we demonstrate that H3 phosphoacetylation is blocked by D1 receptor inactivation, suggesting that inhibitors of H3 acetylation and/or phosphorylation may be useful in preventing or reversing dyskinesia.

Topics: Acetylation; Analysis of Variance; Animals; Antiparkinson Agents; Behavior, Animal; Corpus Striatum; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Extracellular Signal-Regulated MAP Kinases; Functional Laterality; Gene Expression Regulation; Histones; Levodopa; Mice; Mice, Knockout; Motor Activity; Neurons; Oxidopamine; Parkinsonian Disorders; Phosphorylation; Proto-Oncogene Proteins c-fos; Receptors, Dopamine D1; Receptors, Dopamine D2; Statistics as Topic; Tyrosine 3-Monooxygenase

Subthalamic nucleus high frequency stimulation (STN-HFS) efficiently alleviates L-DOPA-sensitive parkinsonian motor symptoms, but has no direct beneficial action on L-DOPA-induced dyskinesias (LID). Here, we provide evidence that anti-akinetic STN-HFS or dyskinesiogenic L-DOPA similarly reversed the dopamine lesion-induced increases in gene expression of cytochrome oxidase subunit I (CoI), a metabolic marker of neuronal activity, in the globus pallidus, STN and substantia nigra pars reticulata (SNr) in rats. In contrast, in entopeduncular nucleus (EP), STN-HFS did not modify the lesion-induced increase in CoI mRNA levels, whereas L-DOPA induced a marked decrease versus control. Combining the two treatments did not reveal significant interaction. Interestingly, CoI gene expression in EP but not in SNr was inversely correlated with striatal preprodynorphin mRNA level, a LID marker. This work suggests the existence of two functional basal ganglia subcircuits: the one, including STN and SNr, involved in antiparkinsonian action, and the other, including EP, preferentially involved in LID.

Topics: Animals; Antiparkinson Agents; Basal Ganglia; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Electric Stimulation; Electrodes, Implanted; Electron Transport Complex IV; Gene Expression Regulation; Levodopa; Male; Nerve Net; Oxidopamine; Parkinsonian Disorders; Protein Precursors; Protein Subunits; Rats; Rats, Wistar; RNA, Messenger

Administration of amphetamine overstimulates medium spiny neurons (MSNs) by releasing dopamine and glutamate from afferents in the striatum. However, these afferents also release brain-derived neurotrophic factor (BDNF) that protects striatal MSNs from overstimulation. Intriguingly, all three neurochemicals increase opioid gene expression in MSNs. In contrast, striatal opioid expression is less in naive BDNF heterozygous (BDNF(+/-)) vs. wild-type (WT) mice. This study was designed to determine whether partial genetic depletion of BDNF influences the behavioral and molecular response to an acute amphetamine injection. An acute injection of amphetamine [5 mg/kg, intraperitoneal (i.p.)] or saline was administered to WT and BDNF(+/-) mice. WT and BDNF(+/-) mice exhibited similar locomotor activity during habituation, whereas BDNF(+/-) mice exhibited more prolonged locomotor activation during the third hour after injection of amphetamine. Three hours after amphetamine injection, there was an increase of preprodynorphin mRNA in the caudate putamen and nucleus accumbens (Acb) and dopamine D(3) receptor mRNA levels were increased in the Acb of BDNF(+/-) and WT mice. Striatal/cortical trkB and BDNF, and mesencephalic tyrosine hydroxylase mRNA levels were only increased in WT mice. These results indicate that BDNF modifies the locomotor responses of mice to acute amphetamine and differentially regulates amphetamine-induced gene expression.

Topics: Adrenergic Uptake Inhibitors; Amphetamine; Amphetamine-Related Disorders; Animals; Brain Chemistry; Brain-Derived Neurotrophic Factor; Disease Models, Animal; Down-Regulation; Dynorphins; Gene Expression Regulation; Habituation, Psychophysiologic; Heterozygote; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Motor Activity; Neostriatum; Neurons; Nucleus Accumbens; Protein Precursors; Receptor, trkB; Receptors, Dopamine D3; Time Factors

Central neuropathic pain is refractory to conventional treatment and thus remains a therapeutic challenge. In this work, we used a well-recognized model of central neuropathic pain to evaluate time-dependent expression of preprodynorphin (ppD), protein kinase C gamma (PKCgamma) and NMDA receptor (NMDAR) subunits NR1, NR2A and NR2B, all critical players in nociceptive processing at the spinal level. Male Sprague-Dawley rats were subjected to spinal hemisection at T13 level and sham-operated rats were included as control animals. The development of hindpaw mechanical allodynia was assessed using the von Frey filaments test. Real time RT-PCR was employed to determine the relative mRNA levels of NMDAR subunits, ppD and PKCgamma in the dorsal spinal cord 1, 14 and 28 days after injury. Our results show that, coincident with the allodynic phase after injury, there was a strong up-regulation of the mRNAs coding for ppD, PKCgamma and NMDAR subunits in the dorsal spinal cord caudal to the injury site. The present study provides further evidence that these molecules are involved in the development/maintenance of central neuropathic pain and thus could be the target of therapeutic approaches.

Topics: Analysis of Variance; Animals; Disease Models, Animal; Dynorphins; Functional Laterality; Hyperalgesia; Male; Pain Measurement; Pain Threshold; Protein Kinase C; Protein Precursors; Protein Subunits; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Spinal Cord Injuries; Time Factors

Alterations in the opioidergic system have been found in cerebral ischemia. Neuroprotection studies have demonstrated the involvement of the opioidergic system in cerebral ischemia/reperfusion (I/R). However, the neuroprotective mechanisms remain largely unclear. This study was conducted to investigate whether intracerebroventricular administration of opioidergic agonists has a neuroprotective effect against cerebral ischemia in rats and, if this proved to be the case, to determine the potential neuroprotective mechanisms. Using a focal cerebral I/R rat model, we demonstrated that the opioidergic agents, BW373U86 (delta agonist) and Dynorphin A 1-13 (kappa agonist), but not TAPP (mu agonist), attenuated cerebral ischemic injury as manifested in the reduction of cerebral infarction and preservation of neurons. The antagonism assay showed that the neuroprotective effect of Dynorphin A was attenuated by nor-Binaltorphimine (kappa antagonist). Surprisingly, BW373U86-induced neuroprotection was not changed by Naltrindole (delta antagonist). These findings indicate that BW373U86 and Dynorphin A exerted distinct neuroprotection against ischemia via opioid-independent and -dependent mechanisms, respectively. The post-ischemic protection in beneficial treatments was accompanied by alleviations in brain edema, inflammatory cell infiltration, and pro-inflammatory cytokine interleukin 6 (IL-6) expression. In vitro cell study further demonstrated that the opioidergic agonists, delta and kappa, but not mu, attenuated IL-6 production from stimulated glial cells. Our findings indicate that opioidergic agents have a role in post-ischemic progression through both opioid-dependent and -independent mechanisms. In spite of the distinct-involved action mechanism, the potential neuroprotective effect of opioidergic compounds was associated with immune suppression. Taken together, these findings suggest a potential role for opioidergic agents in the therapeutic consideration of neuroinflammatory diseases. However, a better understanding of the mechanisms involved is necessary before this therapeutic potential can be realized.

Topics: Amidines; Analgesics, Opioid; Animals; Benzamides; Brain; Brain Ischemia; Cytoprotection; Disease Models, Animal; Disease Progression; Dynorphins; Encephalitis; Immune Tolerance; Interleukin-6; Male; Narcotic Antagonists; Neuroprotective Agents; Peptide Fragments; Piperazines; Rats; Rats, Sprague-Dawley; Receptors, Opioid; Stroke

We recently identified brain-derived neurotrophic factor (BDNF) in the dorsal striatum to be a major component of a homeostatic pathway controlling ethanol consumption. We hypothesized that ethanol-mediated activation of the BDNF signaling cascade is required for the ethanol-related function of the neurotrophic factor. Here, we demonstrate that exposure of striatal neurons to ethanol results in the activation of the BDNF receptor TrkB, leading to the activation of the mitogen-activated protein kinase (MAP kinase) signaling pathway and the subsequent increase in the expression of preprodynorphin (Pdyn) via BDNF. Finally, we show that activation of the dynorphin receptor, the kappa opioid receptor (KOR), is required for the BDNF-mediated decrease in ethanol intake, illustrating a function of dynorphin in BDNF's homeostatic control of ethanol consumption. Taken together, these results demonstrate that BDNF regulates ethanol intake by initiation of MAP kinase signaling and the ensuing production of downstream gene products, including Pdyn.

Topics: Alcohol Drinking; Animals; Brain-Derived Neurotrophic Factor; Cell Culture Techniques; Corpus Striatum; Disease Models, Animal; Dynorphins; Extracellular Signal-Regulated MAP Kinases; Gene Expression Regulation; Homeostasis; Humans; Mice; Mice, Inbred C57BL; Neurons; Rats; Rats, Long-Evans; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction

The Fischer 344 (F344) rat strain differs from the Lewis strain in the response to neuropathic pain. Recently, we found that F344 rats totally recover from mechanical allodynia induced by chronic constriction injury (CCI) of the sciatic nerve 28 days after surgery whereas Lewis rats are initiating their recovery at this time point. Thus, the use of this neuropathic pain model in these different rat strains constitutes a good strategy to identify possible target genes involved in the development of neuropathic pain. Since differences between Lewis and F344 rats in their response to pain stimuli in acute pain models have been related to differences in the endogenous opioid and noradrenergic systems, we aimed to determine the levels of expression of key genes of both systems in the spinal cord and dorsal root ganglia (DRG) of both strains 28 days after CCI surgery. Real time RT-PCR revealed minimal changes in gene expression in the spinal cord after CCI despite the strain considered, but marked changes in DRG were observed. A significant upregulation of prodynorphin gene expression occurred only in injured DRG of F344 rats, the most resistant strain to neuropathic pain. In addition, we found a significant downregulation of tyrosine hydroxylase and proenkephalin gene expression levels in both strains whereas delta-opioid receptor was found to be significantly downregulated only in injured DRG of Lewis rats although the same trend was observed in F344 rats. The data strongly suggest that dynorphins could be involved in strain differences concerning CCI resistance.

Topics: Animals; Chronic Disease; Denervation; Disease Models, Animal; Down-Regulation; Dynorphins; Enkephalins; Ganglia, Spinal; Gene Expression Regulation; Hyperalgesia; Ligation; Male; Neurons, Afferent; Norepinephrine; Peripheral Nerve Injuries; Peripheral Nerves; Peripheral Nervous System Diseases; Protein Precursors; Rats; Rats, Inbred F344; Rats, Inbred Lew; Receptors, Opioid, delta; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Species Specificity; Spinal Cord; Tyrosine 3-Monooxygenase

Escape from the extremely stressful opiate withdrawal syndrome may motivate opiate seeking and taking. The corticotropin-releasing factor receptor-1 (CRF1) pathway mediates behavioral and endocrine responses to stress. Here, we report that genetic inactivation (CRF1-/-) as well as pharmacological antagonism of the CRF/CRF1 receptor pathway increased and prolonged the somatic expression of opiate withdrawal. Opiate-withdrawn CRF1-/- mice also showed aberrant CRF and dynorphin expression in the paraventricular nucleus of the hypothalamus (PVN) and the striatum, indicating profound impairments in stress-responsive brain circuitry. Intake of nonstressful amounts of corticosterone effectively reduced the exaggerated somatic reactions of CRF1-/- mice to opiate withdrawal. Exogenous corticosterone also restored "wild-type-like" patterns of CRF and dynorphin gene expression in the PVN and the striatum of opiate-withdrawn CRF1-/- mice, respectively. The present findings unravel a key role for the hypothalamus-pituitary-adrenal (HPA) system and brain extra-hypothalamic CRF/CRF1 receptor circuitry in somatic, molecular, and endocrine alterations induced by opiate withdrawal.

Topics: Analysis of Variance; Animals; Behavior, Animal; Corticosterone; Corticotropin-Releasing Hormone; Disease Models, Animal; Dynorphins; Female; Mice; Mice, Knockout; Morphine; Paraventricular Hypothalamic Nucleus; Receptors, Corticotropin-Releasing Hormone; Signal Transduction; Stress, Psychological; Substance Withdrawal Syndrome; Time Factors

L-DOPA-induced dyskinesia (LID) is one of the main limitations of long term L-DOPA use in Parkinson's disease (PD) patients. We show that chronic L-DOPA treatment induces novel dyskinetic behaviors in aphakia mouse with selective nigrostriatal deficit mimicking PD. The stereotypical abnormal involuntary movements were induced by dopamine receptor agonists and attenuated by antidyskinetic agents. The development of LID was accompanied by preprodynorphin and preproenkephalin expression changes in the denervated dorsal striatum. Increased FosB-expression was also noted in the dorsal striatum. In addition, FosB expression was noted in the pedunculopontine nucleus and the zona incerta, structures previously not examined in the setting of LID. The aphakia mouse is a novel genetic model with behavioral and biochemical characteristics consistent with those of PD dyskinesia and provides a more consistent, convenient, and physiologic model than toxic lesion models to study the mechanism of LID and to test therapeutic approaches for LID.

Topics: Afferent Pathways; Animals; Antiparkinson Agents; Aphakia; Corpus Striatum; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Levodopa; Mice; Mice, Inbred C57BL; Mice, Neurologic Mutants; Parkinsonian Disorders; Protein Precursors; Proto-Oncogene Proteins c-fos; Substantia Nigra

Current therapies to treat skeletal fracture pain are extremely limited. Some non-steroidal anti-inflammatory drugs have been shown to inhibit bone healing and opiates induce cognitive dysfunction and respiratory depression which are especially problematic in the elderly suffering from osteoporotic fractures. In the present report, we developed a closed femur fracture pain model in the mouse where skeletal pain behaviors such as flinching and guarding of the fractured limb are reversed by 10mg/kg morphine. Using this model we showed that the administration of a monoclonal antibody against nerve growth factor (anti-NGF) reduced fracture-induced pain-related behaviors by over 50%. Treatment with anti-NGF reduced c-Fos and dynorphin up-regulation in the spinal cord at day 2 post-fracture. However, anti-NGF treatment did not reduce p-ERK and c-Fos expression at 20 and 90 min, respectively, following fracture. This suggests NGF is involved in maintenance but not the acute generation of fracture pain. Anti-NGF therapy did not inhibit bone healing as measured by callus formation, bridging of the fracture site or mechanical strength of the bone. As the anti-NGF antibody does not appreciably cross the blood-brain barrier, the present data suggest that the anti-hyperalgesic action of anti-NGF therapy results from blockade of activation and/or sensitization of the CGRP/trkA positive fibers that normally constitute the majority of sensory fibers that innervate the bone. These results demonstrate that NGF plays a significant role in driving fracture pain and that NGF sequestering therapies may be efficacious in attenuating this pain.

Topics: Animals; Antibodies; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Disease Models, Animal; Dynorphins; Exodeoxyribonucleases; Femoral Fractures; Male; Mice; Mice, Inbred C3H; Nerve Fibers; Nerve Growth Factors; Pain; Proto-Oncogene Proteins c-fos; Radiography; Time Factors; Trans-Activators; Tyrosine 3-Monooxygenase; Up-Regulation

Physical activity has documented beneficial effect in treatment of depression. Recently, we found an antidepressant-like effect of running in an animal model of depression, the Flinders Sensitive Line (FSL) and demonstrated that it was associated with increased hippocampal cell proliferation. In this study, we analyzed levels of mRNAs encoding the neuropeptide Y (NPY) and the opioid peptides dynorphin and enkephalin in hippocampus and correlated these to cell proliferation in the FSL and in the 'nondepressed' Flinders Resistant Line (FRL) strain, with/without access to running wheels. Running increased NPY mRNA in dentate gyrus and the CA4 region in FSL, but not in FRL rats. NPY mRNA increase was correlated to increased cell proliferation in the subgranular zone of dentate gyrus. Baseline dynorphin and enkephalin mRNA levels in the dentate gyrus were lower in the FSL compared to the FRL strain. Running had no effect on dynorphin and enkephalin mRNAs in the FSL strain but it decreased dynorphin mRNA, and there was a trend to increased enkephalin mRNA in the FRL rats. Thus, it would appear that the CNS effects of running are different in 'depressed' and control animals; modification of NPY, a peptide associated with depression and anxiety, in depressed animals, vs effects on opioids, associated with the reward systems, in healthy controls. Our data support the hypothesis that NPY neurotransmission in hippocampus is malfunctioning in depression and that antidepressive treatment, in this case wheel running, will normalize it. In addition, we also show that the increased NPY after running is correlated to increased cell proliferation, which is associated with an antidepressive-like effect.

Topics: Analysis of Variance; Animals; Bromodeoxyuridine; Cell Count; Cell Proliferation; Depression; Disease Models, Animal; Dynorphins; Enkephalins; Exercise Therapy; Hippocampus; Immunohistochemistry; In Situ Hybridization; Male; Neuropeptide Y; Rats; Rats, Inbred Strains; Running

Clinically, it has been reported that chronic pain induces depression, anxiety, and reduced quality of life. The endogenous opioid system has been implicated in nociception, anxiety, and stress. The present study was undertaken to investigate whether chronic pain could induce anxiogenic effects and changes in the opioidergic function in the amygdala in mice. We found that either injection of complete Freund's adjuvant (CFA) or neuropathic pain induced by sciatic nerve ligation produced a significant anxiogenic effect at 4 weeks after the injection or surgery. Under these conditions, the selective mu-opioid receptor agonist [D-Ala2,N-MePhe4,Gly5-ol]-enkephalin (DAMGO)- and the selective delta-opioid receptor agonist (+)-4-[(alphaR)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80)-stimulated [35S]GTPgammaS binding in membranes of the amygdala was significantly suppressed by CFA injection or nerve ligation. CFA injection was associated with a significant increase in the kappa-opioid receptor agonist 2-(3,4-dichlorophenyl)-N-methyl-N-[(1S)-1-phenyl-2-(1-pyrrolidinyl)ethyl]acetamide hydrochloride (ICI199,441)-stimulated [35S]GTPgammaS binding in membranes of the amygdala. The intracerebroventricular administration and microinjection of a selective mu-opioid receptor antagonist, a selective delta-opioid receptor antagonist, and the endogenous kappa-opioid receptor ligand dynorphin A caused a significant anxiogenic effect in mice. We also found that thermal hyperalgesia induced by sciatic nerve ligation was reversed at 8 weeks after surgery. In the light-dark test, the time spent in the lit compartment was not changed at 8 weeks after surgery. Collectively, the present data constitute the first evidence that chronic pain has an anxiogenic effect in mice. This phenomenon may be associated with changes in opioidergic function in the amygdala.

Topics: Amygdala; Analgesics, Opioid; Analysis of Variance; Animals; Anxiety; Behavior, Animal; Benzamides; Chronic Disease; Diazepam; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Dynorphins; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Freund's Adjuvant; Guanosine 5'-O-(3-Thiotriphosphate); Injections, Intraventricular; Male; Maze Learning; Mice; Mice, Inbred C57BL; Naltrexone; Narcotic Antagonists; Narcotics; Pain; Pain Measurement; Piperazines; Protein Binding; Pyrrolidines; Rats; Rats, Sprague-Dawley; Reaction Time; Sciatica; Somatostatin; Sulfur Isotopes; Time Factors; Tranquilizing Agents

Opiates are commonly used to treat moderate to severe pain and can be used over prolonged periods in states of chronic pain such as those associated with cancer. In addition, to analgesic actions, studies show that opiate administration can paradoxically induce hyperalgesia. At the pre-clinical level, such hyperalgesia is associated with numerous pronociceptive neuroplastic changes within the primary afferent fibers and the spinal cord. In rodents, sustained opiate administration also induces antinociceptive tolerance. The mechanisms by which prolonged opiate exposure induces hyperalgesia and the relationship of this state to antinociceptive tolerance remain unclear. The present study was aimed at determining whether sustained opiate-induced hyperalgesia, associated neuroplasticity and antinociceptive tolerance are the result of specific opiate interaction at opiate receptors. Enantiomers of oxymorphone, a mu opioid receptor agonist, were administered to rats by spinal infusion across 7 days. Sustained spinal administration of (-)-oxymorphone, but not its inactive enantiomer (+)-oxymorphone or vehicle, upregulated spinal dynorphin content, produced thermal and tactile hypersensitivity, and produced antinociceptive tolerance. These results indicate that these pronociceptive actions of sustained opiate administration require specific interaction with opiate receptors and are unlikely to be the result of accumulation of potentially excitatory metabolic products. While the precise mechanisms, which may account for these pronociceptive changes remain to be unraveled, the present data point to plasticity initiated by opiate receptor interaction.

Topics: Afferent Pathways; Animals; Central Nervous System; Disease Models, Animal; Drug Administration Schedule; Drug Tolerance; Dynorphins; Hyperalgesia; Isomerism; Male; Narcotics; Neuronal Plasticity; Nociceptors; Oxymorphone; Pain; Pain Measurement; Pain Threshold; Rats; Rats, Sprague-Dawley; Receptors, Opioid; Receptors, Opioid, mu; Spinal Nerve Roots; Up-Regulation

Psychostimulant-induced molecular changes in cortico-basal ganglia-cortical circuits play a critical role in addiction and dependence. These changes include alterations in gene regulation particularly in projection neurons of the sensorimotor striatum. We previously showed that cocaine-induced gene regulation in such neurons is dependent on the behavior performed during drug action. Rats trained on a running wheel under the influence of cocaine for 4 days subsequently displayed greater c-fos induction by cocaine than untrained controls. This effect was selective for the sensorimotor striatum, which is known to mediate forms of motor learning. In the present study, we investigated whether this enhanced cellular responsiveness was associated with learning of wheel running or with prolonged running (exercising), by assessing c-fos inducibility after 1, 2, or 8 days of training. Wheel training was performed after injection of cocaine (25 mg/kg) or vehicle, and c-fos induction by a cocaine challenge was measured 24 h later. Rats that trained under cocaine (but not vehicle) showed a greater c-fos response in the striatum compared to locked-wheel controls. This effect was present after the 1-day training, peaked after 2 days, and dissipated by 8 days of training. Similar effects were found for substance P, but not enkephalin, expression. These changes in striatal gene regulation paralleled improvement in wheel running, which was facilitated by cocaine. Thus, these training-induced molecular changes do not appear to represent exercising effects, but may reflect motor learning-associated neuronal changes altered by cocaine. Such cocaine effects may contribute to aberrant motor learning implicated in psychostimulant addiction.

Topics: Animals; Cocaine; Cocaine-Related Disorders; Corpus Striatum; Disease Models, Animal; Dopamine; Dynorphins; Enkephalins; Gene Expression Regulation; Learning; Male; Motor Activity; Motor Skills; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Substance P; Up-Regulation

Epilepsy remains a major medical problem of unknown aetiology. Potentially, viruses can be environmental triggers for development of seizures in genetically vulnerable individuals. An estimated half of encephalitis patients experience seizures and approximately 4% develop status epilepticus. Epilepsy vulnerability has been associated with a dynorphin promoter region polymorphism or low dynorphin expression genotype, in man. In animals, the dynorphin system in the hippocampus is known to regulate excitability. The present study was designed to test the hypothesis that reduced dynorphin expression in the dentate gyrus of hippocampus due to periadolescent virus exposure leads to epileptic responses. Encephalitis produced by the neurotropic Borna disease virus in the rat caused epileptic responses and dynorphin to disappear via dentate granule cell loss, failed neurogenesis and poor survival of new neurons. Kappa opioid (dynorphin) agonists prevented the behavioural and electroencephalographic seizures produced by convulsant compounds, and these effects were associated with an absence of dynorphin from the dentate gyrus granule cell layer and upregulation of enkephalin in CA1 interneurons, thus reproducing a neurochemical marker of epilepsy, namely low dynorphin tone. A key role for kappa opioids in anticonvulsant protection provides a framework for exploration of viral and other insults that increase seizure vulnerability and may provide insights into potential interventions for treatment of epilepsy.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Blotting, Northern; Borna Disease; Cell Survival; Disease Models, Animal; Dynorphins; Electroencephalography; Encephalitis, Viral; Enkephalins; Hippocampus; Male; Naloxone; Narcotic Antagonists; Neurons; Rats; Rats, Inbred Lew; Receptors, Opioid, kappa; Seizures

The aim of this study was to determine whether inhibition of the extracellular-regulated kinase signaling pathway decreases acute amphetamine-induced behavioral activity and neuropeptide gene expression in the rat striatum. Western blotting revealed that extracellular-regulated kinase1/2 phosphorylation was highly induced in the rat striatum 15 min after an acute amphetamine (2.5 mg/kg, i.p.) injection without altering the total amount of extracellular-regulated kinase protein. In a separate experiment, the systemic injection of SL327, a selective inhibitor of extracellular regulated kinase kinase that crosses the blood-brain barrier, 1 h prior to amphetamine administration decreased amphetamine-induced vertical and horizontal activity. Quantitative in situ hybridization histochemistry showed that SL327 abolished the high levels of preproenkephalin and preprodynorphin mRNA induced by amphetamine in the striatum with no alteration of their basal levels. In another set of experiments, the hyperlocomotor activity induced by amphetamine was reduced by pretreatment with intra-striatal infusion of U0126. U0126 also blocked the amphetamine-induced increases in phospho-extracellular-regulated kinase and preproenkephalin and preprodynorphin gene expression in the striatum. These data indicate that activation of the extracellular-regulated kinase cascade contributes to the behavioral effects and changes in striatal neuropeptide gene expression induced by acute amphetamine.

Topics: Aminoacetonitrile; Amphetamine; Amphetamine-Related Disorders; Animals; Butadienes; Corpus Striatum; Disease Models, Animal; Dynorphins; Enkephalins; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Gene Expression Regulation; Male; MAP Kinase Signaling System; Motor Activity; Neuropeptides; Nitriles; Phosphorylation; Protein Precursors; Rats; Rats, Sprague-Dawley; RNA, Messenger; Up-Regulation

Up to 89% of patients with herpes simplex virus type-1 (HSV-1) encephalitis can have seizures. Possibly, viruses are environmental triggers for seizures in genetically vulnerable individuals. Inherited dynorphin promoter polymorphisms are associated with temporal lobe epilepsy and febrile seizures in man. In animals, the dynorphin system in the hippocampus regulates excitability. The hypothesis that reduced dynorphin expression in dentate gyrus of hippocampus due to HSV-1 infection leads to epileptic responses was tested in a rat model of HSV-1 encephalitis using EEG recording, histopathological and neuropharmacologic probes. HSV-1 infection causes loss of dynorphin A-like immunoreactivity in hippocampus, an effect independent of direct viral interference and cell loss. A kappa opioid receptor agonist U50488 effectively blocks ictal activity, linking absence of dynorphin to propensity for epileptic activity. These findings show a vulnerability of hippocampal dynorphin during infection, suggesting a neurochemical basis for seizures that may be generalizable to other encephalitic viruses.

Topics: Action Potentials; Analgesics, Opioid; Animals; Dentate Gyrus; Disease Models, Animal; Down-Regulation; Dynorphins; Electroencephalography; Encephalitis, Herpes Simplex; Epilepsy; Genetic Predisposition to Disease; Herpesvirus 1, Human; Male; Rats; Rats, Inbred Lew; Receptors, Opioid, kappa; Risk Factors

The in vivo biotransformation of dynorphin A(1-17) (Dyn A) was studied in the striatum of hemiparkinsonian rats by using microdialysis in combination with nanoflow reversed-phase liquid chromatography/electrospray time-of-flight mass spectrometry. The microdialysis probes were implanted into both hemispheres of unilaterally 6-hydroxydopamine (6-OHDA) lesioned rats. Dyn A (10 pmol microl(-1)) was infused through the probes at 0.4 microl min(-1) for 2 h. Samples were collected every 30 min and analyzed by mass spectrometry. The results showed for the first time that there was a difference in the Dyn A biotransformation when comparing the two corresponding sides of the brain. Dyn A metabolites 1-8, 1-16, 5-17, 10-17, 7-10 and 8-10 were detected in the dopamine-depleted striatum but not in the untreated striatum. Dyn A biotransformed fragments found in both hemispheres were N-terminal fragments 1-4, 1-5, 1-6, 1-11, 1-12 and 1-13, C-terminal fragments 2-17, 3-17, 4-17, 7-17 and 8-17 and internal fragments 2-5, 2-10, 2-11, 2-12, and 8-15. The relative levels of these fragments were lower in the dopamine-depleted striatum. The results imply that the extracellular in vivo processing of the dynorphin system is being disturbed in the 6-OHDA-lesion animal model of Parkinson's disease.

Topics: Amino Acid Sequence; Animals; Apomorphine; Behavior, Animal; Brain Chemistry; Chromatography, Liquid; Corpus Striatum; Disease Models, Animal; Dynorphins; Mass Spectrometry; Microdialysis; Molecular Sequence Data; Oxidopamine; Parkinson Disease; Peptide Fragments; Rats; Rats, Sprague-Dawley; Spectrometry, Mass, Electrospray Ionization

A common trait of antidepressant drugs, electroconvulsive treatment and physical exercise is that they relieve depression and up-regulate neurotrophic factors as well as cell proliferation and neurogenesis in the hippocampus. In order to identify possible biological underpinnings of depression and the antidepressant effect of running, we analysed cell proliferation, the level of the neurotrophic factor BDNF in hippocampus and dynorphin in striatum/accumbens in 'depressed' Flinders Sensitive Line rats (FSL) and Flinders Resistant Line (FRL) rats with and without access to running-wheels. The FRL strain exhibited a higher daily running activity than the FSL strain. Wheel-running had an antidepressant effect in the 'depressed' FSL rats, as indicated by the forced swim test. In the hippocampus, cell proliferation was lower in the 'depressed' rats compared to the control FRL rats but there was no difference in BDNF or dynorphin levels in striatum/accumbens. After 5 wk of running, cell proliferation increased in FSL but not in FRL rats. BDNF and dynorphin mRNA levels were increased in FRL but not to the same extent in the in FSL rats; thus, increased BDNF and dynorphin levels were correlated to the running activity but not to the antidepressant effect of running. The only parameter that was associated to basal level of 'depression' and to the antidepressant effect was cell proliferation in the hippocampus. Thus, suppression of cell proliferation in the hippocampus could constitute one of the mechanisms that underlie depression, and physical activity might be an efficient antidepressant.

Topics: Analysis of Variance; Animals; Antidepressive Agents; Behavior, Animal; Body Weight; Brain-Derived Neurotrophic Factor; Bromodeoxyuridine; Cell Count; Cell Proliferation; Depression; Disease Models, Animal; Dynorphins; Enkephalins; Freezing Reaction, Cataleptic; Hippocampus; Immunohistochemistry; In Situ Hybridization; Male; Rats; Rats, Inbred Strains; RNA, Messenger; Running; Substance P

Marked fluctuation of dopamine concentration in the striatum following long-term L-DOPA administration contributes to the development of L-DOPA-induced motor complications including L-DOPA-induced dyskinesias and wearing-off in patients with Parkinson's disease. We have shown that pretreatment with 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), a 5-HT1A (5-hydroxytryptamine) receptor agonist, alleviates fluctuation of dopamine levels in the dopamine-denervated striatum of 6-hydroxydopamine-lesioned (hemiparkinsonian) rats after L-DOPA treatment. To determine whether co-administration of 8-OH-DPAT with L-DOPA prevents L-DOPA-induced motor complications, we examined rotation behavior and levels of messenger RNAs coding for dynorphin and glutamic acid decarboxylase in the striatum of 6-hydroxydopamine-lesioned rats treated with L-DOPA alone or L-DOPA + 8-OH-DPAT, twice daily, for 2 weeks. Co-administration of 8-OH-DPAT inhibited an increase of rotation behavior to L-DOPA and L-DOPA-induced increases in levels of messenger RNAs coding for dynorphin and glutamic acid decarboxylase in the dopamine-denervated striatum, both of which are established indices of L-DOPA-induced motor complications. These results suggest that pharmaceutical products that stimulate 5-HT1A receptors could prove useful in prevention of the development of L-DOPA-induced motor complications in patients with Parkinson's disease.

Topics: 8-Hydroxy-2-(di-n-propylamino)tetralin; Animals; Antiparkinson Agents; Behavior, Animal; Corpus Striatum; Disease Models, Animal; Drug Therapy, Combination; Dynorphins; Dyskinesia, Drug-Induced; Female; Glutamate Decarboxylase; In Situ Hybridization; Levodopa; Parkinson Disease; Rats; Rats, Wistar; Serotonin Receptor Agonists

Motor complications induced through repeated L-DOPA treatment in patients with Parkinson's disease are thought to be the consequence of molecular adaptations that occur in response to repeated dopamine receptors stimulation. Here, we studied the molecular changes taking place in the denervated striatum of unilaterally 6-OHDA-lesioned rats repeatedly treated with L-DOPA alone or combined to the D1 receptor antagonist SCH23390. We looked at the territorial patterns of expression of neurotensin (NT), dynorphin (DYN), enkephalin (ENK) and Nur77 (also known as NGFI-B) mRNA expression in the striatum and contrasted these with markers of glutamatergic transport and dopaminergic receptor functions. The denervation process induced NT and Nur77 mRNA expression in ENK-positive cells. Subsequent repeated L-DOPA treatment led to a sensitization of L-DOPA-induced rotational response and produced a second surge of NT induction, this time limited to DYN-positive cells and preferentially restricted to the lateral striatum. In this specific territory, the number of Nur77-positive cells was decreased, in response to L-DOPA, when compared to the medial part of the lesioned striatum. L-DOPA treatment increased dopamine D3 receptor and glutamate transporter 1 (GLT1) mRNA expression in the lesioned striatum and that, specifically in an area overlapping one of Nur77 decrease and of NT/DYN induction. The concomitant administration of SCH23390 with repeated L-DOPA treatment blocked the development of behavioral sensitization and the appearance of all L-DOPA-induced molecular reorganizations reported above. Our results showed that repeated L-DOPA treatment produces, in a denervated striatum, a complex pattern of genes regulation in both the direct and the indirect striatal output pathways. This phenomenon is located preferentially in a striatal area receiving converging inputs from the thalamus and sensorimotor cortex and is dependent upon D1 receptor stimulation.

Topics: Animals; Corpus Striatum; Denervation; Disease Models, Animal; DNA-Binding Proteins; Dopamine; Dopamine Antagonists; Dynorphins; Enkephalins; Excitatory Amino Acid Transporter 2; Gene Expression Regulation; Levodopa; Male; Neurons; Neurotensin; Nuclear Receptor Subfamily 4, Group A, Member 1; Parkinson Disease; Phenotype; Rats; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; Receptors, Dopamine D1; Receptors, Dopamine D3; Receptors, Steroid; RNA, Messenger; Transcription Factors; Tyrosine 3-Monooxygenase

It is generally accepted that neuropathic pain is resistant to amelioration by morphine in clinical studies and insensitivity to intrathecal (i.t.) administered morphine in experimental models of neuropathic pain has been demonstrated. This study is to determine if endogenous dynorphin, substance P or cholecystokinin is involved in the lack of anti-allodynia of morphine in a partial sciatic nerve ligation (PSL) model of CD-1 mice. Mice exhibited tactile allodynia in the ipsilateral hind paw 1 day after PSL, and reached its maximal allodynic effect at 2 days and remained allodynic for 7 days. Morphine (3.0 nmol) given i.t. did not alter the tactile allodynic threshold in ipsilateral paw of mice pretreated i.t. with normal rabbit serum 2 days after PSL. However, the same dose of morphine (3.0 nmol) given i.t. reduced markedly allodynia in mice pretreated for 2h with antiserum against dynorphin A(1-17) (200 microg); the morphine-produced anti-allodynia developed slowly, reached its peak effect at 30 min and returned to an allodynic state in 60 min. Similarly, i.t. injection of morphine reduced the allodynia in PSL mice pretreated with antiserum against substance P (10 microg) or cholecystokinin (200 microg) for 2h. Intrathecal pretreatment with antiserum against dynorphin A(1-17), substance P or cholecystokinin for 2h injected alone did not affect the baseline mechanical tactile threshold in ipsilateral paw 2 days after PSL. The results indicate that endogenous dynorphin A(1-17), substance P and cholecystokinin are involved in PSL-induced neuropathic allodynia to attenuate the anti-allodynic effect of morphine.

Topics: Analgesics, Opioid; Animals; Antibodies; Cholecystokinin; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Resistance; Drug Synergism; Dynorphins; Immune Sera; Injections, Spinal; Ligation; Male; Mice; Morphine; Neuralgia; Peripheral Nervous System Diseases; Sciatic Nerve; Sciatic Neuropathy; Substance P

Increased endogenous opioid activity has been implicated in cholestatic pruritus. In the present study, we have further defined the involvement of opioids in cholestasis. Rats underwent either bile duct ligation or a sham operation. Five days after surgery, brains were removed and agonist-stimulated [35S]GTPgammaS binding was measured in ten brain regions. Serum endomorphin-2, leu-enkephalin and dynorphin A levels were measured using ELISA on day five. Microdialysis to the dorsal hypothalamic area was conducted in the same animal before and after cholestasis. Dialysate endomorphin-1, leu-enkephalin and dynorphin A levels also were measured. Delta- and kappa-stimulated binding was significantly decreased in cholestasic animals compared to controls in the dorsal hypothalamic area. The serum dynorphin A level was lower in the cholestasic group than in controls (2.56+/-0.09 and 3.29+/-0.22 ng/ml, respectively, P<0.01). We propose that pruritus in cholestasis may result from an impaired balance between mu- and kappa-opioid systems.

Topics: Animals; Binding, Competitive; Brain; Cholestasis; Dialysis Solutions; Disease Models, Animal; Dynorphins; Enkephalin, Leucine; Guanosine 5'-O-(3-Thiotriphosphate); Hypothalamus; Male; Microdialysis; Oligopeptides; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Sulfur Radioisotopes

Although oral administration of L-Dopa remains the best therapy for Parkinson disease, its long-term administration causes the appearance of abnormal involuntary movements such as dyskinesia. Although persistent striatal induction of some genes has already been associated with such pathologic profiles in hemiparkinsonian rats, molecular and cellular mechanisms underlying such long-term adaptations remain to be elucidated. In this study, using a rat model of L-Dopa-induced dyskinesia, we report that activity regulated cytoskeletal (Arc)-associated protein is strongly upregulated in the lesioned striatum and that the extent of its induction further varies according to the occurrence or absence of locomotor sensitization. Moreover, Arc is preferentially induced, along with FosB, nur77, and homer-1a, in striatonigral neurons, which express mRNA encoding the precursor of dynorphin. Given the likely importance of Arc in the regulation of cytoskeleton during synaptic plasticity, its upregulation supports the hypothesis that a relationship exists between cytoskeletal modifications and the longlasting action of chronically administrated L-Dopa.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Adrenergic Agents; AIDS-Related Complex; Amphetamine; Analgesics, Non-Narcotic; Animals; Antiparkinson Agents; Behavior, Animal; Carrier Proteins; Central Nervous System Stimulants; Corpus Striatum; Disease Models, Animal; Drug Interactions; Dynorphins; Dyskinesia, Drug-Induced; Functional Laterality; Homer Scaffolding Proteins; Immunohistochemistry; In Situ Hybridization; Levodopa; Male; Motor Activity; Naloxone; Naltrexone; Narcotic Antagonists; Neurons; Oxidopamine; Protein Precursors; Proto-Oncogene Proteins c-fos; Rats; Rats, Wistar; Stereotyped Behavior; Substantia Nigra; Time Factors; Tyrosine 3-Monooxygenase; Up-Regulation

The aging process is associated with various morphological and biochemical changes in the nervous system that may affect the processing of noxious inputs. This study showed greater hyperalgesia and up-regulation of spinal dynorphin (DYN) expression in aging than in young adult rats during CFA-induced peripheral inflammation. These data indicate that nociception is regulated differently in aging individuals, a fact that should be considered when selecting treatment strategies for aging populations with persistent pain.

Topics: Aging; Animals; Chronic Disease; Disease Models, Animal; Dynorphins; Freund's Adjuvant; Functional Laterality; Hyperalgesia; Immunohistochemistry; Inflammation; Male; Peripheral Nervous System Diseases; Physical Stimulation; Posterior Horn Cells; Protein Precursors; Rats; Rats, Sprague-Dawley; RNA, Messenger; Up-Regulation

Release of endogenous dynorphin opioids within the spinal cord after partial sciatic nerve ligation (pSNL) is known to contribute to the neuropathic pain processes. Using a phosphoselective antibody [kappa opioid receptor (KOR-P)] able to detect the serine 369 phosphorylated form of the KOR, we determined possible sites of dynorphin action within the spinal cord after pSNL. KOR-P immunoreactivity (IR) was markedly increased in the L4-L5 spinal dorsal horn of wild-type C57BL/6 mice (7-21 d) after lesion, but not in mice pretreated with the KOR antagonist nor-binaltorphimine (norBNI). In addition, knock-out mice lacking prodynorphin, KOR, or G-protein receptor kinase 3 (GRK3) did not show significant increases in KOR-P IR after pSNL. KOR-P IR was colocalized in both GABAergic neurons and GFAP-positive astrocytes in both ipsilateral and contralateral spinal dorsal horn. Consistent with sustained opioid release, KOR knock-out mice developed significantly increased tactile allodynia and thermal hyperalgesia in both the early (first week) and late (third week) interval after lesion. Similarly, mice pretreated with norBNI showed enhanced hyperalgesia and allodynia during the 3 weeks after pSNL. Because sustained activation of opioid receptors might induce tolerance, we measured the antinociceptive effect of the kappa agonist U50,488 using radiant heat applied to the ipsilateral hindpaw, and we found that agonist potency was significantly decreased 7 d after pSNL. In contrast, neither prodynorphin nor GRK3 knock-out mice showed U50,488 tolerance after pSNL. These findings suggest that pSNL induced a sustained release of endogenous prodynorphin-derived opioid peptides that activated an anti-nociceptive KOR system in mouse spinal cord. Thus, endogenous dynorphin had both pronociceptive and antinociceptive actions after nerve injury and induced GRK3-mediated opioid tolerance.

Topics: Animals; Astrocytes; Disease Models, Animal; Disease Progression; Drug Tolerance; Dynorphins; Enkephalins; G-Protein-Coupled Receptor Kinase 3; Hyperalgesia; Lumbosacral Region; Mice; Mice, Inbred C57BL; Mice, Knockout; Narcotic Antagonists; Narcotics; Neuralgia; Neurons; Protein Precursors; Protein Serine-Threonine Kinases; Receptors, Opioid; Receptors, Opioid, kappa; Sciatic Neuropathy; Spinal Cord

We studied a mouse model of the haploinsufficiency form of Rubinstein-Taybi syndrome (RTS), an inheritable disorder caused by mutations in the gene encoding the CREB binding protein (CBP) and characterized by mental retardation and skeletal abnormalities. In these mice, chromatin acetylation, some forms of long-term memory, and the late phase of hippocampal long-term potentiation (L-LTP) were impaired. We ameliorated the L-LTP deficit in two ways: (1) by enhancing the expression of CREB-dependent genes, and (2) by inhibiting histone deacetyltransferase activity (HDAC), the molecular counterpart of the histone acetylation function of CBP. Inhibition of HDAC also reversed the memory defect observed in fear conditioning. These findings suggest that some of the cognitive and physiological deficits observed on RTS are not simply due to the reduction of CBP during development but may also result from the continued requirement throughout life for both the CREB co-activation and the histone acetylation function of CBP.

Topics: Acetylation; Analysis of Variance; Animals; Blotting, Western; Brain-Derived Neurotrophic Factor; Cell Line; Chromatin; Conditioning, Psychological; CREB-Binding Protein; Disease Models, Animal; Dynorphins; Electrophysiology; Embryo, Mammalian; Excitatory Postsynaptic Potentials; Fear; Female; Gene Expression; Heterozygote; Hippocampus; Humans; Immunohistochemistry; In Situ Hybridization; In Vitro Techniques; Kidney; Long-Term Potentiation; Male; Maze Learning; Memory; Mice; Mice, Inbred C57BL; Mice, Transgenic; Motor Activity; Neural Inhibition; Neurons; Nuclear Proteins; Phosphodiesterase Inhibitors; Proto-Oncogene Proteins c-fos; Psychomotor Performance; Reaction Time; Recognition, Psychology; Rolipram; Rubinstein-Taybi Syndrome; Synaptophysin; Time Factors; Trans-Activators; Transfection

Tumors including sarcomas and breast, prostate, and lung carcinomas frequently grow in or metastasize to the skeleton where they can induce significant bone remodeling and cancer pain. To define products that are released from tumors that are involved in the generation and maintenance of bone cancer pain, we focus here on endothelin-1 (ET-1) and endothelin receptors as several tumors including human prostate and breast have been shown to express high levels of ETs and the application of ETs to peripheral nerves can induce pain. Here we show that in a murine osteolytic 2472 sarcoma model of bone cancer pain, the 2472 sarcoma cells express high levels of ET-1, but express low or undetectable levels of endothelin A (ETAR) or B (ETBR) receptors whereas a subpopulation of sensory neurons express the ETAR and non-myelinating Schwann cells express the ETBR. Acute (10 mg/kg, i.p.) or chronic (10 mg/kg/day, p.o.) administration of the ETAR selective antagonist ABT-627 significantly attenuated ongoing and movement-evoked bone cancer pain and chronic administration of ABT-627 reduced several neurochemical indices of peripheral and central sensitization without influencing tumor growth or bone destruction. In contrast, acute treatment (30 mg/kg, i.p.) with the ETBR selective antagonist, A-192621 increased several measures of ongoing and movement evoked pain. As tumor expression and release of ET-1 has been shown to be regulated by the local environment, location specific expression and release of ET-1 by tumor cells may provide insight into the mechanisms that underlie the heterogeneity of bone cancer pain that is frequently observed in humans with multiple skeletal metastases.

Topics: Analysis of Variance; Animals; Atrasentan; Behavior, Animal; Bone Neoplasms; Calcitonin Gene-Related Peptide; Disease Models, Animal; Dynorphins; Endothelin Receptor Antagonists; Endothelin-1; Ganglia, Spinal; Gene Expression Regulation, Neoplastic; Glial Fibrillary Acidic Protein; Immunohistochemistry; Male; Mice; Mice, Inbred Strains; Pain; Pain Measurement; Pyrrolidines; Receptors, Endothelin; Sarcoma; Sciatic Nerve; Time Factors

Rats exposed to learned helplessness (LH), an animal model of depression, showed a recovery following an intracerebroventricular injection of nor-binaltorphimine dihydrochloride (norBNI; a kappa-opioid antagonist). To investigate the potential role of dynorphin A and dynorphin B, we examined the effects of different stress/depression models on dynorphin A and dynorphin B immunoreactivity in hippocampus and nucleus accumbens (NAc). Immobilization stress (3 h) caused an increase in levels of dynorphin A and dynorphin B immunoreactivity in the hippocampus and the NAc. Forced swim stress also temporally increased dynorphin A levels in the hippocampus. Furthermore, exposure to LH produced a similar increase in dynorphin A and dynorphin B in the hippocampus and NAc. Infusions of norBNI into the dentate gyrus or CA3 regions of hippocampus and into the shell or core regions of NAc produced antidepressant-like effects in the LH paradigm. The degrees of norBNI's effects were stronger in the CA3 region and NAc shell and less effective in the dentate gyrus of hippocampus and NAc core. These results indicate that both dynorphin A and dynorphin B contribute to the effects of stress, and suggest that blockade of kappa-opioid receptors may have therapeutic potential for the treatment of depression.

Topics: Animals; Behavior, Animal; Cell Count; Disease Models, Animal; Dose-Response Relationship, Drug; Dynorphins; Endorphins; Escape Reaction; Helplessness, Learned; Immobilization; Immunohistochemistry; Limbic System; Male; Naltrexone; Rats; Rats, Sprague-Dawley; Stress, Physiological

Previous studies suggest that cholecystokinin (CCK) is implicated in the modulation of pain sensitivity and the development of neuropathic pain. We used CCK(2) receptor deficient (CCK(2) (-/-)) mice and assessed their mechanical sensitivity using Von Frey filaments, as well as the development and time course of mechanical hyperalgesia in a model of neuropathic pain. We found that CCK(2) (-/-) mice displayed mechanical hyposensitivity, which was reversed to the level of wild-type animals after administration of naloxone (0.1-10 mg/kg). On the other hand, injection of L-365260 (0.01-1 mg/kg), an antagonist of CCK(2) receptors, decreased dose-dependently, mechanical sensitivity in wild-type mice. The mechanism of reduced mechanical sensitivity in CCK(2) (-/-) mice may be explained by changes in interactions between CCK and opioid systems. Indeed, CCK(2) (-/-) mice natively expressed higher levels of lumbar CCK(1), opioid delta and kappa receptors. Next, we found that CCK(2) (-/-) mice did not develop mechanical hyperalgesia in the Bennett's neuropathic pain model. Induction of neuropathy resulted in decrease of lumbar pro-opiomelanocortin (POMC) gene expression in wild-type mice, but increase of POMC expression in CCK(2) (-/-) mice. In addition, induction of neuropathy resulted in further increase of opioid delta receptor in CCK(2) (-/-) mice. Gene expression results indicate up-regulation of opioid system in CCK(2) (-/-) mice, which apparently result in decreased neuropathy score. Our study suggests that not only pain sensitivity, but also mechanical sensitivity and the development of neuropathic pain are regulated by antagonistic interactions between CCK and opioid systems.

Topics: Animals; Benzodiazepinones; Disease Models, Animal; Dose-Response Relationship, Drug; Dynorphins; Enkephalins; Gene Expression; Hyperalgesia; Inflammation; Ligation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Naloxone; Narcotic Antagonists; Pain Measurement; Pain Threshold; Phenylurea Compounds; Pro-Opiomelanocortin; Protein Precursors; Receptor, Cholecystokinin B; Receptors, Cholecystokinin; Receptors, Opioid; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sciatic Neuropathy; Time Factors

The transcription factor DeltaFosB accumulates in substance P-dynorphin-containing striatal neurons with repeated cocaine use. Here, we show that inducible transgenic DeltaFosB overexpression in this same striatal cell type facilitates acquisition of cocaine self-administration at low-threshold doses, consistent with increased sensitivity to the pharmacological effects of the drug. Importantly, DeltaFosB also enhances the degree of effort mice will exert to maintain self-administration of higher doses on a progressive ratio schedule of reinforcement, whereas levels of cocaine intake are not altered on less demanding fixed-ratio schedules. Acquisition and extinction of behavior reinforced by food pellets is not altered in DeltaFosB-overexpressing mice, indicating that DeltaFosB does not alter the capacity to learn an instrumental response or cause response perseveration in the absence of reinforcement. These data suggest that accumulation of DeltaFosB contributes to drug addiction by increasing the incentive properties of cocaine, an effect that could increase the risk for relapse long after cocaine use ceases.

Topics: Animals; Behavior, Animal; Cocaine; Cocaine-Related Disorders; Corpus Striatum; Disease Models, Animal; Dose-Response Relationship, Drug; Doxycycline; Drug Administration Schedule; Dynorphins; Gene Expression; Mice; Mice, Transgenic; Motivation; Neurons; Organ Specificity; Proto-Oncogene Proteins c-fos; Reinforcement, Psychology; Self Administration; Substance P; Transgenes

Topically administered capsaicin produces thermal allodynia, and this effect has been used to investigate pain transduction and its pharmacological modulation. This study investigated the parameters of topical capsaicin-induced thermal allodynia in unanesthetized rhesus monkeys and its pharmacological modulation by centrally acting compounds [a kappa-opioid agonist: (5alpha,7alpha,8beta)-(+)-N-methyl-N-(7-[1-pyrrolidinyl]-1-oxaspiro [4.5]dec-8-yl)-benzeneacetamide (U69,593); and noncompetitive N-methyl-d-aspartate (NMDA) antagonists: ketamine and MK-801 (dizocilpine)]. Rhesus monkeys (n = 4) were studied within the warm water tail withdrawal assay (20-s maximum latency), using thermal stimuli that are normally not noxious (38 and 42 degrees C). Capsaicin was applied topically on the tail (0.0013 and 0.004 M capsaicin solution on a 1-cm2 patch; 15-min contact). Topical capsaicin produced concentration-dependent thermal allodynia in both temperatures, robustly detected 15 to 90 min after topical capsaicin removal. A similar allodynic profile was observed with topical administration of the "endovanilloid" N-arachidonoyl-dopamine. The kappa-agonist U69,593 (0.01-0.1 mg/kg, s.c.) dose dependently prevented capsaicin (0.004 M)-induced allodynia in 38 and 42 degrees C, and the largest U69,593 dose also reversed ongoing allodynia within this model. Two NMDA antagonists, ketamine and MK-801 (0.32-1.8 and 0.032-0.056 mg/kg, respectively), also prevented capsaicin-induced allodynia in 38 degrees C, but only variably in 42 degrees C, at doses that did not cause robust thermal antinociceptive effects. At the largest doses studied, ketamine but not MK-801 also briefly reversed ongoing capsaicin-induced allodynia. The present model of topical capsaicin administration may be used to study antiallodynic effects of opioid and nonopioid compounds, as well as their ability to prevent and reverse allodynia, in unanesthetized nonhuman primates in the absence of tissue disruption.

Topics: Aminobutyrates; Analgesics; Animals; Benzeneacetamides; Capsaicin; Disease Models, Animal; Dizocilpine Maleate; Dynorphins; Female; Ketamine; Macaca mulatta; Pain; Pain Measurement; Peptide Fragments; Pyrrolidines

It has been suggested that kainic acid enhances opioid peptide release. However, no direct evidence exists to support this hypothesis. The main aim of the present study was to determine whether such release occurs in the hippocampus of the rat after status epilepticus induced by kainic acid. Microdialysis experiments revealed significant opioid peptide release in the hippocampus 90-150 min (100%) and 270-300 min (50%) after kainic acid-induced status epilepticus. The peptides released were identified by high-performance liquid chromatography linked to radioimmunoassay as Met-enkephalin, Leu-enkephalin, Dynorphin-A (1-6), and Dynorphin-A (1-8). Reduced extracellular opioid peptide immunoreactivity was detected 28 days after status epilepticus (38% compared with control situation). The present results indicate an important activation of opioid peptide systems by kainic acid-induced status epilepticus. In addition, the reduced hippocampal extracellular opioid peptide levels long-term after kainic acid administration could have important implications for the progressive nature of epileptogenesis.

Topics: Animals; Disease Models, Animal; Disease Progression; Down-Regulation; Dynorphins; Enkephalins; Epilepsy, Temporal Lobe; Excitatory Amino Acid Agonists; Glutamic Acid; Hippocampus; Immunohistochemistry; Kainic Acid; Male; Opioid Peptides; Rats; Rats, Wistar; Reaction Time; Status Epilepticus

Nerve injury-induced afferent discharge is thought to elicit spinal sensitization and consequent abnormal pain. Experimental neuropathic pain, however, also depends on central changes, including descending facilitation arising from the rostral ventromedial medulla (RVM) and upregulation of spinal dynorphin. A possible intersection of these influences at the spinal level was explored by measuring evoked, excitatory transmitter release in tissues taken from nerve-injured animals with or without previous manipulation of descending modulatory systems. Spinal nerve ligation (SNL) produced expected tactile and thermal hyperesthesias. Capsaicin-evoked calcitonin gene-related peptide (CGRP) release was markedly enhanced in lumbar spinal tissue from SNL rats when compared with sham-operated controls. Enhanced, evoked CGRP release from SNL rats was blocked by anti-dynorphin A(1-13) antiserum; this treatment did not alter evoked release in tissues from sham-operated rats. Dorsolateral funiculus lesion (DLF) or destruction of RVM neurons expressing mu-opioid receptors with dermorphin-saporin, blocked tactile and thermal hypersensitivity, as well as SNL-induced upregulation of spinal dynorphin. Spinal tissues from these DLF-lesioned or dermorphin-saporin-treated SNL rats did not exhibit enhanced capsaicin-evoked CGRP-IR release. These data demonstrate exaggerated release of excitatory transmitter from primary afferents after injury to peripheral nerves, supporting the likely importance of increased afferent input as a driving force of neuropathic pain. The data also show that modulatory influences of descending facilitation are required for enhanced evoked transmitter release after nerve injury. Thus, convergence of descending modulation, spinal plasticity, and afferent drive in the nerve-injured state reveals a mechanism by which some aspects of nerve injury-induced hyperesthesias may occur.

Topics: Afferent Pathways; Analgesics, Opioid; Animals; Calcitonin Gene-Related Peptide; Capsaicin; Disease Models, Animal; Dynorphins; Enkephalins; Hyperesthesia; Ligation; Lumbosacral Region; Male; Medulla Oblongata; Microinjections; N-Glycosyl Hydrolases; Nerve Compression Syndromes; Neuralgia; Neuronal Plasticity; Neurotransmitter Agents; Oligopeptides; Opioid Peptides; Pain Measurement; Protein Precursors; Rats; Rats, Sprague-Dawley; Receptors, Opioid, mu; Recombinant Fusion Proteins; Ribosome Inactivating Proteins, Type 1; Saporins; Sensory Thresholds; Spinal Cord; Spinal Nerves

To explore changes and role of dynorphinA(1 - 13) on the experimental treatment of hypoxic-ischemic brain injury (HIBI) by intracerebral transplantation of genetically modified myoblasts expressing and secreting brain-derived neurotrophic factor (BDNF) in neonatal rats.. Seven-day-old Sprague-Dawley rats were randomly divided into HIBI + BDNF group (A), HIBI + BDNF + U50, 488H group (B), HIBI group (C) and sham-operated group (D). Pups were intracerebroparenchymally transplanted with either genetically modified myoblasts producing and secreting BDNF (A, B) or their parent cells (C) at 0.8 microl (4 x 10(4)/microl) followed by a cerebroventricular microinjection of opioid kappa receptor agonist U50, 488H (0.5 microg, B) or vehicles (A, C) shortly after HIBI undergone by a permanent ligation of left common carotid artery followed by a 2.5 h inhalation of humidified 8%O(2) + 92%N(2) at 37 degrees C. Water contents of the brain, levels of malondiadehye (MDA) and cell apoptosis were investigated 1 d after the procedure. Contents of dynorphinA(1 - 13)-like immunoreactivity (ir-DynA(1 - 13)) at the left and right cortex and hippocampus were tested by radioimmunoassay 0, 1, 3 d postinjury.. Levels of ir-DynA(1 - 13) in left cortex were markedly increased in group A, B or C when compared to group D different times after the procedure and markedly decreased in group A or B vs group C 1, 3 d after the procedure. Levels of ir-DynA(1 - 13) in left hippocampus were also markedly increased in group A, B and C (0 d), B, C (1 d) and C (3 d) when compared to group D and markedly decreased in group A (1 d) and B (3 d) vs group C, respectively. Water contents, MDA levels and percentage of cell apoptosis were significantly higher in group A, B or C than those in group D and these parameters were obviously lowered in group A compared to group C in the left brain 1 d after the procedure. There were no significant differences of water contents and levels of MDA between group B and C but significantly lowered percentage of cell apoptosis in group B was seen when compared to group C in the left brain.. It is suggested that dynorphinA(1 - 13) participates in the pathophysiological process of HIBI. One of the pathways of the beneficial effects of intracerebral transplantation of genetically modified myoblasts producing BDNF on HIBI lies on the inhibition of the function of dynorphin A(1 - 13) through the binding of kappa opioid receptor.

Topics: Animals; Brain Ischemia; Brain-Derived Neurotrophic Factor; Disease Models, Animal; Dynorphins; Female; Hypoxia-Ischemia, Brain; Male; Malondialdehyde; Oxygen; Peptide Fragments; Rats; Rats, Sprague-Dawley

In a previous study, we have shown in unilaterally dopamine-depleted rats that increased behavioral responsiveness to the dopamine D1-receptor agonist SKF-38393, which was induced by pretreatment with L-DOPA, is paralleled by specific alterations in striatal neuropeptide mRNA levels. The behavioral 'priming' effect of L-DOPA is prevented if L-DOPA is preceded by the NMDA-receptor antagonist MK-801. In the present study, the question is addressed whether blockade of the increased behavioral responsiveness with MK-801 also prevents the observed changes in striatal neuropeptide mRNA levels. After a challenge with SKF-38393 (3 mg/kg, s.c.), the striatal levels of preprodynorphin, preprotachykinin, and preproenkephalin mRNA were compared between unilaterally dopamine-depleted rats that were either primed with a single administration of L-DOPA (50 mg/kg, i.p.) or with L-DOPA preceded by MK-801 (0.1 mg/kg, i.p.). Priming with L-DOPA enhanced the increase in dynorphin mRNA levels in the dorsolateral part of the dopamine-depleted striatum that occurred after SKF-38393. On the other hand, it had no significant effect on substance P or enkephalin mRNA levels. MK-801 prior to L-DOPA prevented the increased responsiveness of dynorphin regulation. However, it induced a decreased response to dopamine D1-receptor stimulation in the substance P mRNA levels in dorsal regions of the dopamine-depleted striatum. The levels of enkephalin mRNA after challenge with SKF-38393 were not affected by the MK-801 administration. These results demonstrate that the increased behavioral responsiveness to the D1-receptor agonist SKF-38393 after priming with L-DOPA is primarily related to the upregulation of dynorphin mRNA levels in the dopamine-depleted striatum.

Topics: 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Animals; Disease Models, Animal; Dizocilpine Maleate; Dopamine Agents; Dopamine Agonists; Drug Interactions; Dynorphins; Dyskinesia, Drug-Induced; Enkephalins; Excitatory Amino Acid Antagonists; Immunohistochemistry; Levodopa; Male; Motor Activity; Neostriatum; Neurons; Neuropeptides; Oxidopamine; Parkinsonian Disorders; Protein Precursors; Rats; Rats, Wistar; Receptors, Dopamine D1; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Tachykinins; Tyrosine 3-Monooxygenase

The influence of chronic arthritic pain on two endogenous opioid peptides, dynorphin B and [Met5]enkephalin-Arg6-Phe7, and multiple opioid receptors in discrete brain, lumbar spinal cord and pituitary pools was investigated. Using radioimmunoassay and receptor binding assay, we examined the changes in regional opioid peptide levels and opioid receptor activity due to chronic inflammation in adjuvant arthritic rats. At 4 weeks post-inoculation, increased levels of immunoreactive dynorphin B and [Met5]enkephalin-Arg6-Phe7 were measured in tissues of arthritic rats compared with controls. No significant changes in mu-, delta- or kappa-opioid receptors were seen after chronic inflammation. Taken together, these results indicate that in chronic arthritis, opioid receptor changes do not follow the peptide alterations of pro-dynorphin and pro-enkephalin systems. Thus, dynamic modification and modulation of nociceptive information takes place during chronic inflammation. This supports the key role of the central nervous system in chronic inflammatory pain conditions.

Topics: Animals; Brain; Chronic Disease; Disease Models, Animal; Dynorphins; Endorphins; Enkephalin, Methionine; Female; Inflammation; Opioid Peptides; Pain; Pain Measurement; Rats; Rats, Inbred Lew; Receptors, Opioid; Spinal Cord

Whereas tissue injury increases spinal dynorphin expression, the functional relevance of this upregulation to persistent pain is unknown. Here, mice lacking the prodynorphin gene were studied for sensitivity to non-noxious and noxious stimuli, before and after induction of experimental neuropathic pain. Prodynorphin knock-out (KO) mice had normal responses to acute non-noxious stimuli and a mild increased sensitivity to some noxious stimuli. After spinal nerve ligation (SNL), both wild-type (WT) and KO mice demonstrated decreased thresholds to innocuous mechanical and to noxious thermal stimuli, indicating that dynorphin is not required for initiation of neuropathic pain. However, whereas neuropathic pain was sustained in WT mice, KO mice showed a return to baselines by post-SNL day 10. In WT mice, SNL upregulated lumbar dynorphin content on day 10, but not day 2, after injury. Intrathecal dynorphin antiserum reversed neuropathic pain in WT mice at post-SNL day 10 (when dynorphin was upregulated) but not on post-SNL day 2; intrathecal MK-801 reversed SNL-pain at both times. Opioid (mu, delta, and kappa) receptor density and G-protein activation were not different between WT and KO mice and were unchanged by SNL injury. The observations suggest (1) an early, dynorphin-independent phase of neuropathic pain and a later dynorphin-dependent stage, (2) that upregulated spinal dynorphin is pronociceptive and required for the maintenance of persistent neuropathic pain, and (3) that processes required for the initiation and the maintenance of the neuropathic pain state are distinct. Identification of mechanisms that maintain neuropathic pain appears important for strategies to treat neuropathic pain.

Topics: Animals; Chronic Disease; Disease Models, Animal; Dizocilpine Maleate; Dynorphins; Excitatory Amino Acid Antagonists; Hyperesthesia; Immune Sera; Injections, Spinal; Ligation; Lumbosacral Region; Male; Mice; Mice, Knockout; Neuralgia; Pain Measurement; Pain Threshold; Physical Stimulation; Reaction Time; Receptors, Opioid; Spinal Cord; Spinal Nerves

Previous studies in piglets show that opioid-induced pial artery dilation was impaired following fluid percussion brain injury (FPI). This study was designed to determine the role of the newly described opioid nociceptin orphanin FQ (NOC/oFQ) in such impaired dilation to other opioids after FPI. CSF NOC/oFQ concentration was elevated from 70+/-6 to 444+/-56 pg/ml ( approximately 10(-10) M) within 1 h of FPI. Coadministration of NOC/oFQ (10(-10) M) with methionine enkephalin (10(-10), 10(-8), 10(-6) M) attenuated pial dilation induced by this opioid (7+/-1, 13+/-2, and 19+/-2 vs. 2+/-1, 6+/-1, and 7+/-2%) under non-brain injury conditions. Similar inhibition by NOC/oFQ was observed for leucine enkephalin and dynorphin. Methionine enkephalin (10(-10), 10(-8), 10(-6) M)-induced pial artery dilation was also inhibited within 1 h of FPI, but such responses were partially restored in animals pretreated with the NOC/oFQ receptor antagonist [F/G] NOC/oFQ (1-13) NH(2) (10(-6) M) (8+/-1, 14+/-1, and 21+/-1 vs. 1+/-1, 3+/-1, and 4+/-1 vs. 7+/-1, 11+/-1, and 17+/-1% for sham control, FPI and FPI pretreated with the NOC/oFQ receptor antagonist). Leucine enkephalin and dynorphin-induced pial artery dilation were similarly altered by FPI and partially restored by [F/G] NOC/oFQ (1-13) NH(2). These data indicate that the NOC/oFQ released by FPI contributes to impaired dilation to other opioids observed following this insult.

Topics: Animals; Brain Injuries; Cerebral Arteries; Disease Models, Animal; Dynorphins; Enkephalin, Leucine; Enkephalin, Methionine; Female; Male; Nociceptin; Opioid Peptides; Pia Mater; Swine; Vasodilation

The aim of this investigation was to determine whether murine models of inflammatory, neuropathic and cancer pain are each characterized by a unique set of neurochemical changes in the spinal cord and sensory neurons. All models were generated in C3H/HeJ mice and hyperalgesia and allodynia behaviorally characterized. A variety of neurochemical markers that have been implicated in the generation and maintenance of chronic pain were then examined in spinal cord and primary afferent neurons.Three days after injection of complete Freund's adjuvant into the hindpaw (a model of persistent inflammatory pain) increases in substance P, calcitonin gene-related peptide, protein kinase C gamma, and substance P receptor were observed in the spinal cord. Following sciatic nerve transection or L5 spinal nerve ligation (a model of persistent neuropathic pain) significant decreases in substance P and calcitonin gene-related peptide and increases in galanin and neuropeptide Y were observed in both primary afferent neurons and the spinal cord. In contrast, in a model of cancer pain induced by injection of osteolytic sarcoma cells into the femur, there were no detectable changes in any of these markers in either primary afferent neurons or the spinal cord. However, in this cancer-pain model, changes including massive astrocyte hypertrophy without neuronal loss, increase in the neuronal expression of c-Fos, and increase in the number of dynorphin-immunoreactive neurons were observed in the spinal cord, ipsilateral to the limb with cancer. These results indicate that a unique set of neurochemical changes occur with inflammatory, neuropathic and cancer pain in C3H/HeJ mice and further suggest that cancer induces a unique persistent pain state. Determining whether these neurochemical changes are involved in the generation and maintenance of each type of persistent pain may provide insight into the mechanisms that underlie each of these pain states.

Topics: Animals; Astrocytes; Axotomy; Behavior, Animal; Disease Models, Animal; Dynorphins; Fluorescent Antibody Technique; Freund's Adjuvant; Ganglia, Spinal; Male; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Neuralgia; Neuritis; Neurons, Afferent; Osteolysis; Pain; Palpation; Physical Stimulation; Proto-Oncogene Proteins c-fos; Receptors, Neurokinin-1; Sarcoma, Experimental; Sciatic Nerve; Spinal Cord; Spinal Nerves; Tumor Cells, Cultured

Chronic treatment of humans or experimental animals with classical neuroleptic drugs can lead to abnormal, tardive movements that persist long after the drugs are withdrawn. A role in these neuroleptic-induced dyskinesias may be played by a structural change in the shell of the nucleus accumbens where the opioid peptide dynorphin is upregulated in treated rats that show vacuous chewing movements (VCMs). The shell of the nucleus accumbens normally contains a dense plexus of dynorphinergic fibers especially in its caudomedial part. After 27 weeks of haloperidol administration and 18 weeks of withdrawal, the immunoreactive labeling of this plexus is intensified when compared with that after vehicle treatment. In addition, medium spiny neurons here show a significant increase in spine density, dendritic branching, and numbers of terminal segments. In the VCM-positive animals, the dendritic surface area is reduced, and dynorphin-positive terminals contact more spines and form more asymmetrical specializations than do those in animals without the syndrome (VCM-negative and vehicle-treated groups). Persistent, neuroleptic-induced oral dyskinesias could therefore be caused by incontrovertible alterations, involving terminal remodeling or sprouting, to the synaptic connectivity of the accumbal shell.

Topics: Animals; Antipsychotic Agents; Behavior, Animal; Dendrites; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Haloperidol; Male; Mastication; Microscopy, Electron; Neurons; Nucleus Accumbens; Presynaptic Terminals; Rats; Rats, Sprague-Dawley; Rats, Wistar; Synapses

In a model of self-sustaining status epilepticus induced in rats by 30 min intermittent stimulation of the perforant path through chronically implanted electrodes, a decrease in dynorphin-like immunoreactivity in the dentate gyrus and CA3 was observed 3 h and 24 h after the induction of status epilepticus. Enkephalin-like immunoreactivity decreased 3 h but not 24 h after perforant path stimulation. Injection into the hilus of the dentate gyrus 10 min prior to stimulation of the kappa-receptor agonist dynorphin-A(1-13), the delta-receptor antagonists ICI-174864 and naltrindole, as well as i.p. injection of naloxone prevented the development of status epilepticus. Perihilar administration of the delta-agonist [D-Ser2]Leu-enkephalin-Thr6 or the kappa-antagonist nor-Binaltorphimine, but not of the mu-agonist [D-Ala2,N-Me-Phe4,Gly-ol5]-Enkephalin, facilitated the establishment of self-sustaining status epilepticus. Injection into the hilus of dynorphin-A(1-13) after the end of perforant path stimulation, stopped established status epilepticus, while administration of naloxone, naltrindole and ICI-174864 were ineffective. We conclude that kappa-opioids in the hippocampus counteract initiation and maintenance of status epilepticus, while delta-opioids promote initiation, but not maintenance of seizure activity. These data are important for the understanding the mechanisms which underlie initiation and maintenance of status epilepticus and for the development of new approaches for its effective management.

Topics: Action Potentials; Analgesics; Analgesics, Opioid; Animals; Disease Models, Animal; Dynorphins; Electric Stimulation; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, Leucine; Enkephalins; Immunohistochemistry; Male; Naloxone; Naltrexone; Narcotic Antagonists; Neurons; Opioid Peptides; Peptide Fragments; Perforant Pathway; Rats; Rats, Wistar; Receptors, Opioid, delta; Receptors, Opioid, kappa; Status Epilepticus

While the morphometry of classical transmitter systems has been extensively studied, relatively little quantitative information is available on the subcellular distribution of peptidergic dense core vesicles (DCVs) within axonal arbors and terminals, and how distribution patterns change in response to neural activity. This study used correlated quantitative light and electron microscopic immunohistochemistry to examine dynorphin B-like immunoreactivity (dyn B-LI) in the rat hippocampal mossy fiber pathway before and after seizures. Forty-eight hours after seizures induced by two pentylenetetrazol injections, light microscopic dyn B-LI was decreased dorsally and increased ventrally. Ultrastructural examination indicated that, in the hilus of the dentate gyrus, these alterations resulted from changes that were almost entirely restricted to the profiles of the large mossy-like terminals formed by mossy fiber collaterals (which primarily contact spines), compared to the profiles of the smaller, less-convoluted terminals found on the same collaterals (which primarily contact aspiny dendritic shafts). Dorsally, mossy terminal profile labeled DCV (/DCV) density dropped substantially, while ventrally, both mossy terminal profile perimeter and /DCV density increased. In all terminal profile examined, /DCVs also were closely associated with the plasma membrane. Following seizures, there was a reorientation of /DCVs along the inner surface of mossy terminal profile membranes, in relation to the types of profiles adjacent to the membrane: in both the dorsal and ventral hilus, significantly fewer /DCVs were observed at sites apposed to dendrites, and significantly more were observed at sites apposed to spines. Thus, after seizures, changes specific to: (1) the dorsoventral level of the hippocampal formation, (2) the type of terminal, and (3) the type of profile in apposition to the portion of the terminal membrane examined were all observed. An explanation of these complex, interdependent alterations will probably require evoking multiple interrelated mechanisms, including selective prodynorphin synthesis, transport, and release.

Topics: Animals; Disease Models, Animal; Dynorphins; Endorphins; Immunohistochemistry; Linear Models; Male; Microscopy, Electron; Mossy Fibers, Hippocampal; Neuropeptides; Neurotransmitter Agents; Rats; Rats, Sprague-Dawley; Seizures

The effects of the extremely selective mu-opioid receptor agonist, [D-Arg2,Lys4]-dermorphin-(1-4)-amide (DALDA), the mu-opioid receptor agonist morphine, the mu/delta agonist D-Ala2, Leu5, Arg6-enkephalin (dalargin), the kappa-opioid receptor agonist spiradoline, and the sigma1-receptor antagonist DuP 734 on ventricular fibrillation threshold (VFT) was investigated in an experimental post-infarction cardiosclerosis model and an immobilization stress-induced model in rats. Both models produced a significant decrease in VFT. The postinfarction cardiosclerosis-induced decrease in VFT was significantly reversed by intravenous administration of dalargin (0.1 mg/kg), DALDA (0.1 mg/kg), or morphine HCl (1.5 mg/kg). Pretreatment with naloxone (0.2 mg/kg) completely eliminated the increase in cardiac electrical stability produced by DALDA. Both spiradoline (8 mg/kg, i.p.) and DuP 734 (1 mg/kg, i.p.) produced a significant increase in VFT in rats with post-infarction cardiosclerosis. This effect of spiradoline was blocked by nor-binaltorphimine. The immobilization stress-induced decrease in VFT was significantly reversed by administration of either DALDA, spiradoline or DuP 734. In conclusion, activation of either mu- or kappa1-opioid receptors or blockade of sigma1-receptors reversed the decrease in VFT in both cardiac compromised models. Since DALDA and dalargin essentially do not cross blood brain barriers, their effects on VFT may be mediated through peripheral mu-opioid receptors.

Topics: Animals; Anti-Arrhythmia Agents; beta-Endorphin; Disease Models, Animal; Dynorphins; Enkephalin, Leucine-2-Alanine; Heart; Immobilization; Ligands; Morphine; Myocardial Infarction; Myocardium; Naloxone; Naltrexone; Narcotic Antagonists; Oligopeptides; Piperidines; Pyrrolidines; Rats; Receptors, Opioid; Receptors, Opioid, delta; Stress, Physiological; Ventricular Fibrillation

The effects of N-methyl-D-aspartate(NMDA) receptor antagonist, Mk-801, on the expression of spinal dynorphin (DYN) mRNA and the hyperalgesia induced by peripheral inflammation were studied by Northern analysis and behavioral test. Following an unilateral injection of complete Freund's adjuvant (CFA) into the rat hindpaw, there appeared a significant hyperalgesia of inflamed hindpaw and up-regulation of ipsilateral spinal DYN mRNA; while the pre-emptive and continuous intrathecal administration of Mk-801 (10 microg/microl per h) could significantly suppress both the hyperalgesia and the up-regulation of spinal DYN mRNA induced by peripheral inflammation. The results suggest that NMDA receptor activation may contribute to the development and maintenance of the thermal hyperalgesia that is associated with the up-regulation of DYN expression in spinal dorsal horn.

Topics: Animals; Binding, Competitive; Chronic Disease; Disease Models, Animal; Dizocilpine Maleate; Dynorphins; Freund's Adjuvant; Hindlimb; Hyperalgesia; Inflammation; Infusion Pumps, Implantable; Injections, Spinal; Male; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Spinal Cord; Up-Regulation

Dynorphin, an endogenous opioid, may contribute to secondary nervous tissue damage following spinal cord injury. The temporal and spatial distribution of preprodynorphin (PPD) mRNA expression in the injured rat spinal cord was examined by in situ hybridization. Rats were subjected to traumatic spinal cord injury at the T13 spinal segment using the weight-drop method. Motor function of these rats was evaluated by their ability to maintain their position on an inclined plane. Two double-labeling experiments revealed that increased PPD mRNA and dynorphin peptide expression were found exclusively in dorsal horn neurons. Neurons exhibiting an increase in the level of PPD mRNA were concentrated in the superficial laminae and the neck of dorsal horn within several spinal segments from the epicenter of the injury at 24 and 48 h after injury. A number of neurons showing increased PPD mRNA were found in gray matter adjacent to the injury areas. Segments caudal to the injury site exhibited a long-lasting elevation of PPD mRNA in neurons, compared to the rostral segments. The number of neurons expressing PPD mRNA in each rat was significantly positively correlated with its motor dysfunction. These findings suggest that increased expression of dynorphin mRNA and peptide in dorsal horn neurons occurs after traumatic spinal cord injury. This also supports the hypothesis that the dynorphin has a pathological role in secondary tissue damage and neurological dysfunction after spinal cord injury.

Topics: Animals; Autoradiography; Disease Models, Animal; Disease Progression; Dynorphins; Gene Expression; In Situ Hybridization; Male; Microscopy, Confocal; Neurons; Photomicrography; Protein Precursors; Rats; Rats, Sprague-Dawley; RNA, Messenger; Spinal Cord; Spinal Cord Injuries; Time Factors

We have used a partial sciatic nerve ligation model to examine the time course for changes in the expression of mRNA for three peptides related to pain transmission at spinal sites (dynorphin, enkephalin and substance P), during the development of allodynia. Enhanced expression of mRNA for dynorphin and substance P was observed in the dorsal horn on the same side as the partial nerve ligation. Increased expression of dynorphin mRNA was biphasic. The initial increases in expression of dynorphin mRNA occurred at 3 h, and a secondary peak was observed 1-3 days after surgery. The secondary increases coincided roughly with increased substance P mRNA expression. However, both dynorphin and substance P mRNA returned to control values after 1 week despite continuing allodynia. No significant changes in expression of mRNA for enkephalin were observed. The elevation of substance P mRNA in intrinsic spinal cord neurons may be secondary to changes in immediate early genes c-fos and jun-B, whereas the expression of dynorphin and enkephalin mRNA is differently regulated. The results also suggest that changes in the expression of the three neuropeptides are not critically involved in the development and maintenance of chronic pain or allodynia.

Topics: Animals; Behavior, Animal; Chronic Disease; Disease Models, Animal; Dynorphins; Enkephalins; Gene Expression; Male; Neuropeptides; Pain; Rats; Rats, Sprague-Dawley; RNA, Messenger; Spinal Cord; Substance P; Time Factors

Several studies have demonstrated that cocaine increases preprodynorphin, c-fos, and zif/268 mRNAs in rat dorsal striatum. Multiple, closely spaced exposures to cocaine appear to elicit the greatest increases in dynorphin. However, the response of preproenkephalin, c-fos and zif/268 mRNAs to such a dosing regimen is unknown. Therefore, we used a 'binge' paradigm to evaluate changes in mRNA for preprodynorphin, preproenkephalin, c-fos and zif/268. Male Wistar rats received three hourly i.p. injections of saline or 10 or 20 mg/kg cocaine for 1, 5, or 10 days. Although cocaine-induced locomotor and stereotypical behaviors were significantly increased as compared to saline on days 1, 5 and 10, these behaviors were significantly less on day 10 than on days 1 and 5. One hour after the last injection on days 1, 5, or 10, the rats were anesthetized and decapitated for quantitative in situ hybridization histochemistry. C-fos mRNA was undetectable in all treatment groups whereas zif/268 mRNA in the dorsal striatum was increased in a dose-dependent manner (20 mg/kg > 10 mg/kg) but the intensity of hybridization signal decreased over time (1 day >> 5 days > 10 days) as compared to that in saline-treated controls. In contrast, 10 mg/kg cocaine binges caused an increase in preprodynorphin, but not preproenkephalin, mRNA in the dorsal, but not ventral, striatum in a time-dependent manner (day 10 >> day 5 > day 1) whereas 20 mg/kg cocaine binges caused an increase in striatal preprodynorphin that was greater on day 1 and day 5 than on day 10. These data indicate that (1) c-fos, zif/268 and preprodynorphin mRNAs are differentially regulated in dorsal striatum, (2) behavioral tolerance results from chronic binges with 10 and 20 mg/kg cocaine and (3) the preprodynorphin genomic response exhibits tolerance to chronic high dose, but not low dose, cocaine binges.

Topics: Analysis of Variance; Animals; Behavior, Animal; Cocaine; Corpus Striatum; Disease Models, Animal; DNA-Binding Proteins; Drug Administration Schedule; Dynorphins; Early Growth Response Protein 1; Enkephalins; Immediate-Early Proteins; Male; Nerve Tissue Proteins; Protein Precursors; Proto-Oncogene Proteins c-fos; Rats; Rats, Wistar; RNA, Messenger; Transcription Factors

Dynorphin A reduced lumbosacral blood flow, elevated cerebrospinal fluid lactic acid concentrations and caused flaccid hindlimb paralysis and striking neuropathological changes after its injection into the spinal subarachnoid space in rats. Coadministration of the vasodilator hydralazine substantially eliminated the paralytic, anaerobic metabolic and neuropathological responses to dynorphin A. In contrast, in concentrations up to 1 mM, dynorphin A did not alter the viability of cultured rat spinal cord neurons. Thus, it appears that this peptide lacks direct neurotoxic effects and that neuronal injuries in vivo result primarily from ischemia associated with dynorphin A-induced blood flow reductions. NMDA receptor antagonists significantly improved recovery from dynorphin A-induced hindlimb paralysis, and substantially eliminated neuropathological changes without attenuating the acute blood flow reductions or lactic acid elevations. Additionally, glutamate and aspartate concentrations were increased significantly in spinal cord cerebrospinal fluid samples removed during the time that peptide-induced spinal cord blood flow reductions were observed. In contrast, neither amino acid concentration was elevated in media removed after 1-hr exposure of spinal cord neuronal cell cultures to 100 microM concentrations of dynorphin A. These results indicate that the paralysis and spinal cord injuries produced in rats after spinal subarachnoid injection of dynorphin A result predominantly from spinal cord ischemia, and further identify excitatory amino acids and N-methyl-D-aspartate receptor mechanisms as important mediators in this injury model.

Topics: Amino Acids; Animals; Cells, Cultured; Disease Models, Animal; Dynorphins; Hindlimb; Ischemia; Male; Neurons; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Spinal Cord Injuries

The effects of dynorphin A-(1-13), an endogenous kappa opioid agonist, on basal forebrain (BF)-lesion-induced amnesia in rats were investigated using step-through-type passive avoidance task. The BF was lesioned by injecting the cholinergic neurotoxin ibotenic acid (6 micrograms/side). The number of rats achieving the cut-off time (600 s) of step-through latency (STL) in BF-lesioned group significantly decreased as compared with that in sham-operated group. Dynorphin A-(1-13) (0.3 micrograms) significantly increased the number of rats achieving the cut-off time of STL in BF-lesioned rats. These results suggest that dynorphins play an improving role in the impairment of memory processes in BF-lesioned rats.

Topics: Alzheimer Disease; Amnesia; Animals; Avoidance Learning; Cognition; Disease Models, Animal; Dynorphins; Ibotenic Acid; Male; Peptide Fragments; Prosencephalon; Rats; Rats, Wistar; Receptors, Opioid, kappa

Tardive dyskinesia has been connected with regional reductions of GABA functions in the basal ganglia. In view of the possibility that peptides are involved in neuroleptic-induced dyskinesias substance P and dynorphin A levels were measured in the basal ganglia of the Cebus apella model for tardive dyskinesia. In addition, regional glutamate decarboxylase activities, dopamine, homovanillic acid and dihydroxyphenylacetic acid levels were monitored. A significant dyskinesia-related decrease in glutamate decarboxylase activity was found in the subthalamic nucleus, the medial segment of globus pallidus and the rostral part of substantia nigra in accordance with earlier findings. Cebus monkeys with an intact GABA system (neuroleptic-treated controls without dyskinesia) showed increased levels of substance P and homovanillic acid in the caudate nucleus. The changes were confined to the caudal part of the body of the caudate and the nucleus accumbens. On the other hand, the dyskinetic monkeys, with a defective GABA system, did not demonstrate a similar substance P rise in the caudate or nucleus accumbens, but showed a depression of homovanillic acid levels in the caudal part of the body of the caudate nucleus. Dynorphin A, dopamine and dihydroxyphenylacetic acid showed no dyskinesia-related changes. In conclusion, the difference in glutamate decarboxylase activity between animals developing dyskinetic symptoms vs those who did not, was reflected by regional changes in substance P and homovanillic acid levels.

Topics: 3,4-Dihydroxyphenylacetic Acid; Animals; Behavior, Animal; Brain Chemistry; Cebus; Corpus Striatum; Disease Models, Animal; Dynorphins; Dyskinesia, Drug-Induced; Female; Globus Pallidus; Glutamate Decarboxylase; Homovanillic Acid; Male; Neuropeptides; Substance P; Substantia Nigra

In this study the possible role of hippocampal dynorphin in the development of hypertension in spontaneously hypertensive rats (SHR) was investigated by determining dynorphin A (1-8) (DN A (1-8] levels in hippocampus in 16 week old SRH, Wistar Kyoto (WKY) controls and SHR treated with antihypertensive drugs as well as DOCA-salt hypertensive Sprague Dawley (SD) rats, using radioimmunoassay (RIA). We found that DN A (1-8) was decreased significantly in both dorsal (-68%) and ventral (-58%) hippocampus in SHR compared with WKY rats. Treatment with hydralazine and guanethidine (25 mg/kg/24 hr of each drug in drinking water) for 8 weeks to prevent the development of hypertension in young SHR had no effect on this low hippocampal dynorphin level. We failed to find significant changes in hippocampal DN A (1-8) level in DOCA-salt hypertensive rats. The low level of hippocampal dynorphin existed before the development of hypertension in 6 day neonatal SHR (-73%). Hippocampal Met-enkephalin was unchanged in all experimental groups except for a slight decrease in neonatal SHR. The results establish a genetic difference in the hippocampal dynorphin system of SHR compared with WKY, the significance of which, for the development of hypertension, remains to be investigated.

Topics: Animals; Animals, Newborn; Antihypertensive Agents; Desoxycorticosterone; Disease Models, Animal; Dynorphins; Enkephalin, Methionine; Hippocampus; Hypertension; Peptide Fragments; Radioimmunoassay; Rats; Rats, Inbred SHR; Rats, Inbred Strains; Rats, Inbred WKY; Sodium Chloride

Dynorphin, an opioid peptide, is thought to play an important role in the modulation of nociceptive neural circuits at the level of the spinal cord. In a model of peripheral inflammation and hyperalgesia, an oligodeoxyribonucleotide probe complementary to a portion of preprodynorphin mRNA and antisera to dynorphin A-(1-8) were used to localize changes in dynorphin mRNA and peptide to individual spinal cord neurons. Intraplantar injection in rats of complete Freund's adjuvant resulted in edema and hyperalgesia to radiant heat stimulation of the injected hind paw that reached a peak at 4 days. At the same time, in situ hybridization histochemistry and immunocytochemistry identified an increase in transcription of preprodynorphin mRNA that was paralleled by an increase in dynorphin peptide. These changes were seen in spinal neurons in the medial two-thirds of laminae I and II and in laminae V and VI of lumbar segments receiving innervation from the inflamed paw. Since neurons demonstrating the increase in dynorphin biosynthesis are located in both the superficial and deep dorsal horn laminae, our data provide evidence for opioid modulation of nociceptive neural circuits in these two distinct spinal locations.

Topics: Animals; Disease Models, Animal; Dynorphins; Hyperalgesia; Hyperesthesia; Inflammation; Male; Neurons; Nucleic Acid Hybridization; Rats; Rats, Inbred Strains; RNA, Messenger; Spinal Cord

Inoculation of the right hind paw with Mycobacterium butyricum rapidly led to swelling and inflammation. The afflicted limb showed an enhanced sensitivity to noxious pressure (hyperalgesia) and a reduced sensitivity to noxious heat 24 h following treatment. Both naloxone and MR 2266 (which has greater activity at kappa-opioid receptors) further increased the sensitivity to pressure (that is, potentiated the hyperalgesia) but did not affect the response to heat. They did not affect the response of the uninflamed paw. At 1 week, only MR 2266 was effective. At both 24 h and 1 week, the inflamed paw showed pronounced supersensitivity to the antinociceptive action of morphine against noxious pressure. At both 24 h and (to a greater extent) 1 week, a rise in levels of immunoreactive (ir)-dynorphin (DYN) was seen in the ipsilateral dorsal horn of the lumbar spinal cord. There was no alteration in the contralateral dorsal horn or in either ventral horn. Furthermore, levels of ir-met-enkephalin (ME) and ir-leu-enkephalin (LE) were unaffected. There was no difference in the density of mu-, delta- or kappa-binding sites in any part of the lumbar cord, at either 24 h or 1 week, between ipsilateral and contralateral tissue. By 3 and 5 weeks postinoculation, the symptoms had spread to the contralateral hind limb and ir-DYN was elevated in the contralateral dorsal horn and the ipsilateral ventral horn. At 5 weeks, levels of ir-ME and ir-LE also were increased in the ipsilateral and contralateral dorsal horns, but not in the contralateral ventral horn. Furthermore, levels of ir-DYN were increased in the cervico-thoracic spinal cord, and rats displayed adrenal hypertrophy and a rise in plasma levels of ir-beta-endorphin (beta-EP). These data indicate: (1) Peripheral inflammation localized to a single limb selectively modifies levels of ir-DYN in ipsilateral dorsal horn. The effect is specific to DYN as compared to ME and LE. The density of mu-, delta-, or kappa-receptors in the lumbar spinal cord is unmodified. (2) The altered response to opioid agonists and antagonists shown by rats with an inflamed limb may be selective to the injured tissue. (3) Alterations in opioid systems associated with unilateral hind limb inflammation may not be exclusively chronic in nature: they appear very rapidly (within 24 h) of the induction of pain. With time, the contralateral limb becomes affected and, eventually, the effects resemble those seen with generalized polyarthritis.

Topics: Animals; Benzomorphans; Disease Models, Animal; Dynorphins; Endorphins; Hindlimb; Inflammation; Male; Naloxone; Pain; Rats; Rats, Inbred Strains; Receptors, Opioid; Spinal Cord

Chronic arthritic pain was induced by intradermally inoculating rats at the tail-base with Mycobacterium butyricum, which results in swelling, inflammation, and hyperalgesia of the joints. These symptoms peak at 3 weeks after inoculation and disappear by 10 weeks. The following changes were seen at 3 weeks. Immunoreactive dynorphin (ir-Dyn) and ir-alpha-neo-endorphin (alpha-NE) manifested comparable patterns of change. Their levels were increased in the anterior, but not neurointermediate, pituitary. The thalamus showed a rise in ir-Dyn and ir-alpha-NE, but no alterations were seen in other brain regions. In each case, cervical, thoracic, and lumbosacral sections of the spinal cord showed a rise in ir-Dyn and ir-alpha-NE: This was most pronounced in the lumbosacral region, where the magnitude of these shifts correlated with the intensity of arthritic symptoms. In addition, a moderate elevation in ir-methionine-enkephalin (ME) was seen in lumbosacral spinal cord. In brain, ir was not changed. The level of ir-beta-endorphin (beta-EP) was elevated both in the plasma and the anterior, but not the neurointermediate, pituitary. In addition, the content of messenger RNA encoding the beta-EP precursor, proopiomelanocortin (POMC), was enhanced in the anterior lobe. Thus, there was a selective activation of synthesis of beta-EP in, and its secretion from, the anterior lobe. In no brain tissue did levels of ir-beta-EP change. At 10 weeks postinoculation, the above changes were no longer apparent, indicating their reversibility.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Arthritis; Arthritis, Experimental; beta-Endorphin; Brain; Brain Chemistry; Chronic Disease; Diprenorphine; Disease Models, Animal; Dynorphins; Endorphins; Hypothalamus; Male; Mesencephalon; Pain; Rats; Rats, Inbred Strains; Receptors, Opioid; Spinal Cord; Thalamus

We previously reported that the opioid peptide dynorphin1-13 improves survival chances in stroked cats. Some evidence also suggests that changes in dopamine and gamma-aminobutyric acid (GABA) uptake may be associated with stroke. In the present study, therefore, we determined binding of the opiate [3H]ethylketocyclazocine (EKC), as well as dopamine and GABA uptake in various brain regions of control, stroked and dynorphin1-13-treated stroked cats. Cats were stroked by middle cerebral artery occlusion. In the EKC binding study, the Kd of the high-affinity site of the occluded cortex was significantly increased, relative to that of both the unoccluded side and control cortex. Dynorphin1-13 treatment reversed this effect, lowering the Kd to control level. In the dopamine uptake study, the Km was decreased and Vmax was increased significantly in unoccluded cortex, compared with that in the occluded cortex or in control cortex. Again, dynorphin1-13 reversed these effects, raising the Km and lowering the Vmax. However, the Km of occluded cortex was also increased so that it became significantly higher than that of control cortex. The Km of unoccluded subcortex in stroked cats treated with dynorphin1-13 was significantly reduced compared with control. In the GABA uptake study, there was no significant change in any parameter. The change in opioid binding observed here and its reversal by dynorphin1-13 are consistent with the notion that the peptide's beneficial effect on stroke is mediated through opiate receptors. Since opioid systems in the brain are known to have association with dopaminergic ones, the change in dopamine uptake could also be the result of an opioid effect.

Topics: Animals; Brain; Brain Ischemia; Cats; Cyclazocine; Disease Models, Animal; Dopamine; Dynorphins; Ethylketocyclazocine; gamma-Aminobutyric Acid; Kinetics; Male; Peptide Fragments; Receptors, Opioid

Topics: Animals; Disease Models, Animal; Drug Tolerance; Dynorphins; Humans; In Vitro Techniques; Narcotics; Opioid-Related Disorders; Receptors, Opioid