Maternal Deprivation Enhances Behavioral Vulnerability to Stress Associated with miR-504 Expression in Nucleus Accumbens of Rats

Objective

In this study, the effect of maternal deprivation (MD) and chronic unpredictable stress (CUS) in inducing depressive behaviors and associated molecular mechanism were investigated in rats.

Methods

Maternal deprivation was established by separating pups from their mothers for 6 hours daily from postnatal day 1 to day 14. Chronic unpredictable stress was established by water deprivation, elevated open platform, food deprivation, restraint stress and electric foot shock. The depressive behaviors were determined by use of sucrose preference test and forced swim test.

Results

Rats in MD/CUS group exhibited lower sucrose preference rate, longer immobility time, and lighter body weights than rats in other groups (MD/control, non-MD/CUS and non-MD/control group). Meanwhile, higher miR-504 expression and lower dopamine receptor D1 (DRD1) and D2 (DRD2) expression were observed in the nucleus accumbens of rats in the MD/CUS group than in the other three groups. MiR-504 expression correlated negatively with DRD1 gene expression and sucrose preference rate in the sucrose preference test, but correlated positively with immobility time in forced swim test. Both DRD2 mRNA and protein expression correlated negatively with immobility time in forced swim test.

Conclusion

These results suggest that MD enhances behavioral vulnerability to stress during adulthood, which is associated with the upregulation of miR-504 and downregulation of DRD2 expression in the nucleus accumbens.     

Differential Response of Central Dopaminergic System in Acute and Chronic Unpredictable Stress Models in Rats

We aimed to evaluate the response of dopaminergic system in acute stress (AS) and chronic unpredictable stress (CUS) by measuring dopamine (DA) levels, its receptor densities in the frontal cortex, striatum, hippocampus, amygdala and orbito-frontal cortex regions of rat brain, and investigated the corresponding behavioral locomotor changes. Involvement of D₁ receptor was also examined during AS and CUS using A 68930, a D₁ selective agonist. Rats were exposed to AS (single immobilization for 150 min) and CUS (two different stressors for 7 days). AS significantly decreased the DA levels in the striatum and hippocampus, and A 68930 pretreatment significantly reverted these changes. However, in the frontal cortex significantly increased DA levels were remain unchanged following A 68930. CUS led to a decrease of DA levels in the frontal cortex, striatum and hippocampus, which were normalized by A 68930. Saturation radioligand binding assays revealed a significant decrease in the number of D₁-like receptors in the frontal cortex during CUS, which were further decreased by A 68930 pretreatment. However, in the striatum and hippocampus, A 68930 pretreatment reduced the CUS induced increase in the number of D₁-like receptors. No significant changes were observed in the amygdala and orbito-frontal cortex during AS and CUS, while D₂-like receptors were unchanged in all the brain regions studied. Locomotor activity was significantly decreased in both the stress models, A 68930 pretreatment significantly increased stereotypic counts and horizontal activity. Thus, present investigation provide insights into the differential regional response of dopaminergic system during AS and CUS. Further, neurochemical and behavioral effects of D₁ agonist pretreatment suggest specific modulatory role of D₁ receptor under such stressful episodes.     

NMDA Receptor in Nucleus Accumbens is Implicated in Morphine Withdrawal in Rats

The purpose of the present study is to elucidate whether ketamine, a non-competitive antagonist of the NMDA receptor, can suppress the morphine withdrawal syndrome in rats at a dose without affecting motor functions and to identify its site of action in the central nervous system. Rats were made dependent on morphine by multiple injections of morphine hydrochloride for 5 days. They were then given ketamine at the following doses and routes of administration: (a) intraperitoneal (i.p.) injections (2–16 mg/kg), (b) intracerebroventricular (i.c.v.) injections (4–100 g), and (c) intra-nucleus accumbens (NAc) or intra-amygdalar microinjections (0.4–10 g). Naloxone HCl (1 mg/kg, i.p.) was administered 3 h after the last ketamine injection to precipitate withdrawal syndrome, which was scored within a period of 30 min. Results showed that some of the precipitated withdrawal signs were dose-dependently suppressed by repeated injections of ketamine at 8 and 16 mg/kg, i.p. or 100 g, i.c.v. Dose-dependent suppression was observed by repeated microinjections (0.4–10 g) of ketamine to NAc, but not to amygdala. These results indicate that the NMDA receptor antagonist ketamine has the ability to suppress morphine withdrawal syndrome in experimental settings without motor interference, and NAc could be the critical CNS site mediating such effect.Special issue dedicated to Dr. Lawrence F. Eng.     

Differential Effect of Stress on In Vivo Dopamine Release in Striatum, Nucleus Accumbens, and Medial Frontal Cortex

Microdialysis was used to assess extracellular dopamine in striatum, nucleus accumbens, and medial frontal cortex of unanesthetized rats both under resting conditions and in response to intermittent tail-shock stress. The dopamine metabolites 3,4-dihydroxyphenylacetic acid and homovanillic acid also were measured. The resting extracellular concentration of dopamine was estimated to be approximately 10 nM in striatum, 11 nM in nucleus accumbens, and 3 nM in medial frontal cortex. In contrast, the resting extracellular levels of 3,4-dihydroxyphenylacetic acid and homovanillic acid were in the low micromolar range. Intermittent tail-shock stress increased extracellular dopamine relative to baseline by 25% in striatum, 39% in nucleus accumbens, and 95% in medial frontal cortex. 3,4-Dihydroxyphenylacetic acid and homovanillic acid also were generally increased by stress, although there was a great deal of variability in these responses. These data provide direct in vivo evidence for the global activation of dopaminergic systems by stress and support the concept that there exist regional variations in the regulation of dopamine release.     

Estrogen Regulation of Dopamine Release in the Nucleus Accumbens: Genomic-and Nongenomic-Mediated Effects

Abstract: The ability of estrogen to modulate mesolimbic dopamine (DA) was examined using in vivo voltammetry. Estrogen priming (5 μg, 48 h) of ovariectomized (ovx) female rats resulted in a slight decrease in K⁺-stimulated DA release measured in the nucleus accumbens: this decrease was accompanied by a significant increase in both DA reuptake and DA clearance times. Following estrogen priming nomifensine, a potent inhibitor of the DA uptake carrier, was still able to potentiate K⁺-stimulated DA release and alter the time course of DA availability, but the response was attenuated compared with ovx controls. Direct infusion of 17β-estradiol hemisuccinate (17β-E, 20–50 pg) into the nucleus accumbens resulted in a biphasic potentiation of K⁺-stimulated release. An initial increase in release was observed 2 min after 17β-E infusion; this increase, although reduced by 15 min, was still significantly higher than control values. A subsequent potentiation was observed 60 min after the initial 17β-E infusion; this response remained for at least an additional 60 min. Nomifensine did not significantly alter K⁺-stimulated DA release following 17β-E infusion, but was still able to potentiate the total time DA was available extracellularly. These data suggest that the mesolimbic A10 DA neurons that terminate in the nucleus accumbens can be modulated in vivo by estrogen and that this modulation may be mediated by both genomic (long term) and nongenomic (short term) mechanisms.     

Myocardial Creatine Levels Do Not Influence Response to Acute Oxidative Stress in Isolated Perfused Heart

Background

Multiple studies suggest creatine mediates anti-oxidant activity in addition to its established role in cellular energy metabolism. The functional significance for the heart has yet to be established, but antioxidant activity could contribute to the cardioprotective effect of creatine in ischaemia/reperfusion injury.

Objectives

To determine whether intracellular creatine levels influence responses to acute reactive oxygen species (ROS) exposure in the intact beating heart. We hypothesised that mice with elevated creatine due to over-expression of the creatine transporter (CrT-OE) would be relatively protected, while mice with creatine-deficiency (GAMT KO) would fare worse.

Methods and Results

CrT-OE mice were pre-selected for creatine levels 20–100% above wild-type using in vivo ¹H^–MRS. Hearts were perfused in isovolumic Langendorff mode and cardiac function monitored throughout. After 20 min equilibration, hearts were perfused with either H₂O₂ 0.5 µM (30 min), or the anti-neoplastic drug doxorubicin 15 µM (100 min). Protein carbonylation, creatine kinase isoenzyme activities and phospho-PKCδ expression were quantified in perfused hearts as markers of oxidative damage and apoptotic signalling. Wild-type hearts responded to ROS challenge with a profound decline in contractile function that was ameliorated by co-administration of catalase or dexrazoxane as positive controls. In contrast, the functional deterioration in CrT-OE and GAMT KO hearts was indistinguishable from wild-type controls, as was the extent of oxidative damage and apoptosis. Exogenous creatine supplementation also failed to protect hearts from doxorubicin-induced dysfunction.

Conclusions

Intracellular creatine levels do not influence the response to acute ROS challenge in the intact beating heart, arguing against creatine exerting (patho-)physiologically relevant anti-oxidant activity.     

Blocking TRPV1 in Nucleus Accumbens Inhibits Persistent Morphine Conditioned Place Preference Expression in Rats

The function of TRPV1 (transient receptor potential vanilloid subfamily, member 1) in the central nervous system is gradually elucidated. It has been recently proved to be expressed in nucleus accumbens (NAc), a region playing an essential role in mediating opioid craving and taking behaviors. Based on the general role of TRPV1 antagonist in blocking neural over-excitability by both pre- and post-synaptic mechanisms, TRPV1 antagonist capsazepine (CPZ) was tested for its ability to prohibit persistent opioid craving in rats. In the present study, we assessed the expression of TRPV1 in nucleus accumbens and investigated the effect of CPZ in bilateral nucleus accumbens on persistent morphine conditioned place preference (mCPP) in rats. We also evaluated the side-effect of CPZ on activity by comparing cross-beam times between groups. We found that morphine conditioned place preference increased the TRPV1 expression and CPZ attenuated morphine conditioned place preference in a dose-dependent and target–specific manner after both short- and long-term spontaneous withdrawal, reflected by the reduction of the increased time in morphine-paired side. CPZ (10 nM) could induce prolonged and stable inhibition of morphine conditioned place preference expression. More importantly, CPZ did not cause dysfunction of activity in the subjects tested, which indicates the inhibitory effect was not obtained at the sacrifice of regular movement. Collectively, these results indicated that injection of TRPV1 antagonist in nucleus accumbens is capable of attenuating persistent morphine conditioned place preference without affecting normal activity. Thus, TRPV1 antagonist is one of the promising therapeutic drugs for the treatment of opioid addiction.     

Iron Deficiency Alters Discrete Proteins in Rat Caudate Nucleus and Nucleus Accumbens

Young rats (21 days old) made nutritionally iron deficient, by feeding them a semisynthetic diet containing skimmed milk for 5 weeks, had significantly lowered hemoglobin levels (5.2 +/- 4 g/100 ml). The nonheme iron content in caudate nucleus was decreased by 47%. The behavioral response of iron-deficient rats to apomorphine (2 mg/kg) and the density of 3,4-dihydroxyphenylethylamine (dopamine) D2 receptors, as measured by [3H]spiperone binding in caudate nucleus, were significantly reduced by 70 and 53%, respectively. The possibility that nutritional iron deficiency may affect protein content in brain was investigated by measuring the apparent concentration of proteins in caudate nucleus and nucleus accumbens from iron-deficient and control animals using two-dimensional gel electrophoresis. The data indicate that iron deficiency can affect content in these two brain regions. Significant changes in the content of 10 proteins were noted in the caudate nucleus and nucleus accumbens in iron-deficient rats. The albumin level was significantly increased in both regions studied, whereas the neuron-specific enolase level was increased in the nucleus accumbens and the glial fibrillary acidic protein level was reduced in the caudate nucleus. The significance of these protein content changes, as well as a reduction in content of a 94-kilodalton protein (a molecular size similar to that of the D2 dopamine receptor), remains to be established.     

The Role of Nucleus Accumbens Core/Shell in Sleep-Wake Regulation and their Involvement in Modafinil-Induced Arousal

Background

We have previously shown that modafinil promotes wakefulness via dopamine receptor D₁ and D₂ receptors; however, the locus where dopamine acts has not been identified. We proposed that the nucleus accumbens (NAc) that receives the ventral tegmental area dopamine inputs play an important role not only in reward and addiction but also in sleep-wake cycle and in mediating modafinil-induced arousal.

Methodology/Principal Findings

In the present study, we further explored the role of NAc in sleep-wake cycle and sleep homeostasis by ablating the NAc core and shell, respectively, and examined arousal response following modafinil administration. We found that discrete NAc core and shell lesions produced 26.5% and 17.4% increase in total wakefulness per day, respectively, with sleep fragmentation and a reduced sleep rebound after a 6-hr sleep deprivation compared to control. Finally, NAc core but not shell lesions eliminated arousal effects of modafinil.

Conclusions/Significance

These results indicate that the NAc regulates sleep-wake behavior and mediates arousal effects of the midbrain dopamine system and stimulant modafinil.     

Restraint Stress Potentiated Morphine Sensitization: Involvement of Dopamine Receptors within the Nucleus Accumbens

Sensitization to psychostimulant drugs, as well as morphine, subjected to cross-sensitization with stress. The development of morphine sensitization is associated with enhancements in dopamine overflow in the Nucleus accumbens (NAc). This study aimed to examine the role of accumbal D1/D2-like dopamine receptors in restraint stress (RS) induced sensitization to morphine antinociceptive effects. Adult male Wistar rats weighing 220–250 g underwent stereotaxic surgery. Two stainless steel guide cannulae were bilaterally implanted, 1 mm above the NAc injection site. Different solutions of SCH-23390, as a D1-like receptor antagonist or sulpiride, as a D2-like receptor antagonist, were microinjected into the NAc five min before exposure to RS. Restraint stress lasted for 3 h, 10 min after RS termination; animals received a subcutaneous injection of morphine (1 mg/kg) for 3 consecutive days. The procedure was followed by a 5-day drug and/or stress-free period. After that, on the 9th day, the nociceptive response was evaluated by the tail-flick test. The results revealed that intra-NAc administration of D1/D2-like dopamine receptor antagonists, SCH-23390 or sulpiride, respectively, blocked morphine sensitization-induced by RS and morphine co-administration in rats for three consecutive days. This work provides new insight into the determinant role of accumbal dopamine receptors in morphine sensitization produced by RS-morphine co-administration.

     

Differential Response of Hippocampal Subregions to Stress and Learning

The hippocampus has two functionally distinct subregions–the dorsal portion, primarily associated with spatial navigation, and the ventral portion, primarily associated with anxiety. In a prior study of chronic unpredictable stress (CUS) in rodents, we found that it selectively enhanced cellular plasticity in the dorsal hippocampal subregion while negatively impacting it in the ventral. In the present study, we determined whether this adaptive plasticity in the dorsal subregion would confer CUS rats an advantage in a spatial task–the radial arm water maze (RAWM). RAWM exposure is both stressful and requires spatial navigation, and therefore places demands simultaneously upon both hippocampal subregions. Therefore, we used Western blotting to investigate differential expression of plasticity-associated proteins (brain derived neurotrophic factor [BDNF], proBDNF and postsynaptic density-95 [PSD-95]) in the dorsal and ventral subregions following RAWM exposure. Lastly, we used unbiased stereology to compare the effects of CUS on proliferation, survival and neuronal differentiation of cells in the dorsal and ventral hippocampal subregions. We found that CUS and exposure to the RAWM both increased corticosterone, indicating that both are stressful; nevertheless, CUS animals had significantly better long-term spatial memory. We also observed a subregion-specific pattern of protein expression following RAWM, with proBDNF increased in the dorsal and decreased in the ventral subregion, while PSD-95 was selectively upregulated in the ventral. Finally, consistent with our previous study, we found that CUS most negatively affected neurogenesis in the ventral (compared to the dorsal) subregion. Taken together, our data support a dual role for the hippocampus in stressful experiences, with the more resilient dorsal portion undergoing adaptive plasticity (perhaps to facilitate escape from or neutralization of the stressor), and the ventral portion involved in affective responses.     

Differential Regulation of CuZnSOD Expression in Rat Brain by Acute and/or Chronic Stress

Neuroendocrine stress (NES) causes increase of glucocorticoids and alters physiological levels of reactive oxygen species production in cells, which might involve modifications in the antioxidant defense system. We investigated the hypothesis that acute, chronic, or combined stress alters copper–zinc superoxide dismutase (CuZnSOD) expression pattern at both, mRNA and subcellular protein level in the cerebral cortex and hippocampus of rats and that there may be a relationship between stress-induced corticosterone and CuZnSOD expression. The most effective stress model which led to the most pronounced changes in CuZnSOD expression patterns was also investigated. Our results demonstrated that acute stress immobilization up-regulates mRNA expression of hippocampal CuZnSOD, while cytosolic protein expression of this enzyme was increased in both brain structures. Chronic stress isolation had no effect on either mRNA and protein expression level and caused a lack of significant up-regulation to a novel acute stressors. The presence of this protein in nuclear fractions of both brain structures was also confirmed. The elevated cytosolic CuZnSOD protein levels following acute immobilization might reflect on the defense system against oxidative stress. Chronic isolation compromises CuZnSOD protein expression, which may lead to the inefficient defense against reactive oxygen species (ROS). The stress-triggered CuZnSOD protein expression was not correlated by the corresponding mRNA. The results suggest that different stress models exert a different degree of influence on mRNA and protein level of CuZnSOD in both brain structures as well as serum corticosterone.     

Non-NMDA Glutamate Receptor Antagonist Injected into the Hypothalamic Paraventricular Nucleus Inhibits the Prolactin Response to Formalin Stress of Male Rats

The aim of the present investigations was to test the involvement of the glutamatergic innervation of the hypothalamic paraventricular nucleus in the prolactin response to stress. A non-NMDA (6-cyano-7-nitroquinoxaline-2,3-dione disodium, CNQX) or an NMDA glutamate receptor antagonist (dizocilpine hydrogen malate, MK-801) was injected bilaterally into the paraventricular nucleus of freely moving male rats and 15 min later the animals were exposed to formalin stress. Blood samples for prolactin and corticosterone were taken at different time points before and after administration of formalin. CNQX, when injected into the paraventricular nucleus, inhibited the formalin-induced rise in plasma prolactin and not significantly the increase in corticosterone. A similar effect was not observed if MK-801 was administered into the paraventricular nuclei or CNQX was injected outside the cell group. The findings indicate that the glutamatergic innervation of the paraventricular nucleus is involved in the mediation of the formalin-induced prolactin release.Special Issue Dedicated to Miklós Palkovits.     

Acute Thermal and Stress Response in Moose to Chemical Immobilization

Management and research of moose (Alces alces) in Alaska, USA, often require chemical immobilization; however, moose may be prone to capture-induced hyperthermia while immobilized. We chemically immobilized moose with carfentanil citrate and xylazine hydrochloride to measure rump fat depth, collect blood and fecal samples, and to deploy modified vaginal implant transmitters and global positioning system (GPS)-collars for recording body temperature and movement during and after the chemical immobilization. We predicted wild moose pursued and captured from a helicopter would have elevated body temperature at time of capture, whereas body temperature would remain stable in hand-raised captive moose not pursued and only hand-injected for immobilization. Additionally, we expected post-capture body temperature would be a function of activity, time immobilized, and ambient temperature. As predicted, body temperature of wild moose was elevated 1 hour after capture (38.9°C, 95% CI = 38.7–39.1°C) but returned to baseline levels within 3 hours (38.0°C, 95% CI = 37.9–38.1°C); however, body temperatures then rose above baseline levels and remained elevated 12–48 hours post-capture when movement rates were also elevated. Body temperatures in captive moose were not elevated 1-hour post-immobilization (37.9°C, 95% CI = 37.8–38.0°C). Body temperatures of wild moose were positively related to cortisol levels at time of capture. Two moose that died after immobilization had initial body temperatures similar to other immobilized moose; however, their body temperature began to rise at 17 hours and 40 hours post-immobilization. Our study provides evidence that chemical immobilization affects body temperature and movement of wild moose up to 48 hours after capture, possibly as a result of renarcotization from carfentanil citrate. With advancements in technology, we recommend fine-scale GPS data (<1-hr fix rates) and continuous body temperature be evaluated to detect evidence of renarcotization during and after opioid-based captures of northern ungulates. © 2020 The Wildlife Society.     

The Adenosine A2A Receptor Activation in Nucleus Accumbens Suppress Cue-Induced Reinstatement of Propofol Self-administration in Rats

Neurochemical Research - Propofol has shown strong addictive properties in rats and humans. Adenosine A2A receptors (A2AR) in the nucleus accumbens (NAc) modulate dopamine signal and addictive...     

乙烯调控植物耐盐性的研究进展

乙烯具有复杂的生物学功能,它调节着植物生长发育和许多的生理生化过程。乙烯也被认为是一种胁迫应答激素,直到近几年关于乙烯生物合成及信号转导途径与植物盐胁迫的关系才逐渐被挖掘出来。乙烯在不同水平、层次参与盐胁迫反应,包括乙烯合成关键酶(ACS)和乙烯受体,细胞质中CTR1和EIN2以及细胞核中EIN3传导、响应盐信号。但是乙烯合成和信号转导途径在植物盐胁迫响应过程中仍然存在许多未解决的问题。主要介绍乙烯合成及信号转导途径的各组分与盐胁迫关系的最新研究进展,并讨论其存在的主要问题。     

S-Nitroso-Proteome in Poplar Leaves in Response to Acute Ozone Stress

Protein S-nitrosylation, the covalent binding of nitric oxide (NO) to protein cysteine residues, is one of the main mechanisms of NO signaling in plant and animal cells. Using a combination of the biotin switch assay and label-free LC-MS/MS analysis, we revealed the S-nitroso-proteome of the woody model plant Populus x canescens. Under normal conditions, constitutively S-nitrosylated proteins in poplar leaves and calli comprise all aspects of primary and secondary metabolism. Acute ozone fumigation was applied to elicit ROS-mediated changes of the S-nitroso-proteome. This treatment changed the total nitrite and nitrosothiol contents of poplar leaves and affected the homeostasis of 32 S-nitrosylated proteins. Multivariate data analysis revealed that ozone exposure negatively affected the S-nitrosylation status of leaf proteins: 23 proteins were de-nitrosylated and 9 proteins had increased S-nitrosylation content compared to the control. Phenylalanine ammonia-lyase 2 (log2[ozone/control] = −3.6) and caffeic acid O-methyltransferase (−3.4), key enzymes catalyzing important steps in the phenylpropanoid and subsequent lignin biosynthetic pathways, respectively, were de-nitrosylated upon ozone stress. Measuring the in vivo and in vitro phenylalanine ammonia-lyase activity indicated that the increase of the phenylalanine ammonia-lyase activity in response to acute ozone is partly regulated by de-nitrosylation, which might favor a higher metabolic flux through the phenylpropanoid pathway within minutes after ozone exposure.