Beatriz G. Pérez-Nievas

Stress-Induced Oxidative Changes in Brain

Numerous systems and organs are affected by stress. In this review we will focus on the effects in brain. Some of the most impressive effects of the stress in brain are the atrophy of hippocampal dendrites or even the reduction of the hippocampal size observed in brains from subjects exposed to severe or chronic stress. Obviously, before reaching this point of damage there are many other processes taking place in the stressed CNS. The release of glucocorticoids is one of the first features of the stress response. Glucocorticoids can result in neurotoxicity through different mechanisms, including modifications in the energy metabolism or via an increase in excitatory amino acids such as glutamate in the extracellular space. Glutamate can induce neuronal excitotoxicity. This sequence of events leads to the activation of TNFalpha convertase (TACE) and TNFalpha release in brain of rats subjected to restraint stress. One of the multiple effects exerted by this cytokine is to initiate the translocation of the transcription factor NFkappaB to neuronal nuclei. NFkappaB activation results in the induction of iNOS and COX2, two enzymes responsible for a great portion of the neurological damage produced in models of stress.

Effects of peroxisome proliferator-activated receptor gamma agonists on brain glucose and glutamate transporters after stress in rats.

Repeated stress causes an energy-compromised status in the brain, with a decrease in glucose utilization by the brain cells, which might account for excitotoxicity processes seen in this condition. In fact, brain glucose metabolism mechanisms are impaired in some neurodegenerative disorders, including stress-related neuropsychopathologies. More recently, it has been demonstrated that some synthetic peroxisome proliferator-activated receptor gamma (PPARγ) agonists increase glucose utilization in rat cortical slices and astrocytes, as well as inhibit brain oxidative damage after repeated stress, which add support for considering these drugs as potential neuroprotective agents. To assess if stress causes glucose utilization impairment in the brain and to study the mechanisms by which this effect is achieved, young-adult male Wistar rats (control and immobilized for 6 h during 7 or 14 consecutive days, S7, S14) were i.p. injected with the natural ligand 15-deoxy-Δ-12,14-prostaglandin J2 (PGJ2, 120 μg/kg) or the high-affinity ligand rosiglitazone (RG, 3 mg/kg) at the onset of stress. Repeated immobilization during 1 or 2 weeks produces a decrease in brain cortical synaptosomal glucose uptake, and this effect was prevented by treatment with both natural and synthetic PPARγ ligands by restoring protein expression of the neuronal glucose transporter, GLUT-3 in membrane fractions. On the other hand, treatment with PPARγ ligands prevents stress-induced ATP loss in rat brain. Finally, repeated immobilization stress also produces a decrease in brain cortical synaptosomal glutamate uptake, and this effect was prevented by treatment with PPARγ ligands by restoring synaptosomal protein expression of the glial glutamate transporter, EAAT2. In summary, our results demonstrate that 15d-PGJ2 and the thiazolidinedione rosiglitazone increase neuronal glucose metabolism, restore brain ATP levels and prevent the impairment in glutamate uptake mechanisms induced by exposure to stress, suggesting that this class of drugs may be therapeutically useful in conditions in which brain glucose levels or availability are limited after exposure to stress.

Depressive-like States Heighten the Aversion to Painful Stimuli in a Rat Model of Comorbid Chronic Pain and Depression

Background: Chronic pain and depression are two complex states with sensory/somatic and emotional components, and they may mutually exacerbate one another in conditions of comorbidity, leading to a poorer prognosis. Methods: The authors have evaluated the sensory and emotional components in a rat model combining chronic constriction injury (CCI, a model of chronic neuropathic pain) with unpredictable chronic mild stress (CMS, an experimental model of depression). In addition, the phosphorylation/activation of the extracellular signal-regulated kinases 1 and 2 and neuronal density was also evaluated in the anterior cingulate cortex. Four groups were tested: sham-control, sham-CMS, CCI-control, and CCI-CMS. Results: CMS selectively heightens aversion to painful experiences in animals subjected to CCI, as measured in the place escape/avoidance test at 20, 25, and 30 min (CCI-CMS (mean ± SEM): 75.68 ± 3.32, 66.75 ± 4.70, 77.54 ± 3.60 vs. CCI-control: 44.66 ± 6.07, 43.17 ± 6.92, 52.83 ± 5.92, respectively), in conjunction with an increase in the accumulation of phosphorylation/activation of the extracellular signal-regulated kinases (CCI-CMS: 4.17 ± 0.52 vs. sham-control: 0.96 ± 0.05) and a decrease in neuronal density in the anterior cingulate cortex. In contrast, chronic pain did not exacerbate the characteristic profile of depression (anhedonia and behavioral despair) in rats subjected to CMS. Furthermore, depression enhances the perception of some specific modalities of sensorial pain such as cold allodynia but has no influence on mechanical threshold. Conclusions: These findings support the theory that depression leads to emotional dysfunction in the interpretation of pain in patients suffering chronic pain. In addition, combined animal models of pain-depression may provide a valuable tool to study the comorbidity of pain and depression.

The anti-inflammatory prostaglandin 15d-PGJ2 and its nuclear receptor PPARgamma are decreased in schizophrenia

A number of findings suggest that inflammation plays a role in the pathophysiology of schizophrenia. Taking into account a physiological balance between pro- and anti-inflammatory mediators, we measured the plasma levels of cyclooxygenase-derived mediators and other key pro- and anti-inflammatory transcription factors in peripheral blood mononuclear cells (PBMC). Forty healthy subjects and 46 treated chronic schizophrenic patients with an acutely exacerbated condition who met DSM-IV criteria were included. COX by-products prostaglandin E2 (PGE2) and 15d-prostaglandin J2 (15d-PGJ2) plasma levels were measured by EIA. Peroxisome proliferator-activated receptor gamma (PPARγ) as well as nuclear factor kappaB (NFκB) activity in nuclear extracts from PBMC and expression of its inhibitory subunit IκBα in cytosolic extracts were determined using ELISA-based kits. Schizophrenic patients showed higher plasma levels of pro-inflammatory PGE2 than age-matched controls (p=0.043). On the contrary, levels of anti-inflammatory 15-d-PGJ2 were lower (p=0.004), correlating with a lower expression of its nuclear target, PPARγ in nuclear extracts from PBMC (p=0.001). Although no changes in NFκB activity were observed between patients and healthy controls, the expression of its inhibitory protein IκBα was lower in the patients compared to the controls (p=0.027). These findings suggest that schizophrenia is associated with a systemic imbalance in the plasma levels of pro-inflammatory/anti-inflammatory prostaglandins in favor of the former. Furthermore, the expression and activity of anti-inflammatory PPARγ are diminished in PBMC, which indicates a state of inflammation and blunted anti-inflammatory counterbalancing mechanisms at systemic level in these patients.

Corticosterone as a marker of susceptibility to oxidative/nitrosative cerebral damage after stress exposure in rats

There are important individual differences in susceptibility to stress-induced diseases, most of them associated to the hypothalamic-pituitary and sympatho-medullo-adrenal axis functioning. Characterization of individual differences in animals may help to find the origin of this susceptibility. In order to study differences in oxidative and neuroinflammatory consequences in brain after stress exposure, we used an adult, male, outbred (Wistar:Hannover) population of 60 rats. Animals were subjected to 6h of immobilisation stress. Basal (1 week before stress) and post-stress (immediately after stress) plasma corticosterone (CC) was measured for each animal from the tail vein (basal: 239.74+/-19.44 ng/ml at 1500 h). Group H was assigned to animals with 33% higher levels of CC (>279.53 ng/ml) and group L to animals with 33% lower levels of CC (<199.09 ng/ml). After stress, animals with higher plasma CC levels in basal conditions showed higher adrenal response (higher post-stress CC levels) than rats with lower levels of basal CC. Furthermore, rats from H group are more vulnerable to accumulation of oxidative/nitrosative mediators in brain (higher calcium-independent nitric oxide activity and higher lipid peroxidation, by malondialdehyde determination, MDA) and also to the accumulation of proinflammatory mediators (higher PGE(2) levels) whereas showing less antiinflammatory protection (less 15-deoxy-PGJ(2) levels). Statistical analysis, by using ROC curves revealed cut-off values of basal plasma CC predicting animals with higher post-stress MDA and PGE(2) and lower PGJ(2) levels in brain. These data indicate that plasma basal levels of CC are an easily detectable and reproducible parameter for predicting the response of the individuals after an acute stress, providing further support for studies on individual differences.

Chronic immobilisation stress ameliorates clinical score and neuroinflammation in a MOG-induced EAE in Dark Agouti rats: mechanisms implicated.

BackgroundMultiple sclerosis (MS) is the endpoint of a complex and still poorly understood process which results in inflammation, demyelination and axonal and neuronal degeneration. Since the first description of MS, psychological stress has been suggested to be one of the trigger factors in the onset and/or relapse of symptoms. However, data from animal models of MS, such as experimental autoimmune encephalomyelitis (EAE) are inconsistent and the effect of stress on EAE onset and severity depends on duration and time of application of the stress protocol and the underlying mechanisms.MethodsDark Agouti rats were inoculated with MOG/CFA to induce EAE, and an immobilisation stress protocol with two different durations (12 and 21 days, starting at the moment of MOG-inoculation) was applied in order to analyse the effect of stress on disease onset and neuroinflammation.ResultsTwelve days of stress exposure increased EAE clinical score in Dark Agouti rats. In addition, these animals presented higher levels of MMP-9 and proinflammatory PGE2 in spinal cord. In contrast, animals chronically exposed to stress (21 days) showed a significantly lower incidence of EAE clinical signs and reduced myelin loss, leukocyte infiltration and accumulation of inflammatory/oxidative mediators in spinal cord. Interestingly, chronically stressed animals showed a parallel increase in levels of the anti-inflammatory prostaglandin 15d-PGJ2, the main endogenous agonist of PPARγ.ConclusionsOur results demonstrate that, depending on duration, stress exposure elicits opposite effects on PGE2/15d-PGJ2 ratios in spinal cord of EAE-induced Dark Agouti rats. Further studies are needed to elucidate if these changes in prostaglandin balance are sufficient to mediate the differences in clinical score and inflammation here reported, and to establish the potential utility of pharmacological intervention in MS directed toward anti-inflammatory pathways.

Cannabinoid/agonist WIN 55,212-2 reduces cardiac ischaemia–reperfusion injury in Zucker diabetic fatty rats: role of CB2 receptors and iNOS/eNOS.

Diabetes increases cardiac damage after myocardial ischaemia. Cannabinoids can protect against myocardial ischaemia/reperfusion injury. The aim of this study was to examine the cardioprotective effect of the cannabinoid agonist WIN 55,212‐2 (WIN) against ischaemia/reperfusion injury in an experimental model of type 2 diabetes. We performed these experiments in the Zucker diabetic fatty rat, and focused on the role of cannabinoid receptors in modulation of cardiac inducible nitric oxide synthase (iNOS)/endothelial‐type nitric oxide synthase (eNOS) expression.

Endogenous cannabinoid system regulates intestinal barrier function in vivo through cannabinoid type 1 receptor activation

The deleterious effects of stress on the gastrointestinal tract seem to be mainly mediated by the induction of intestinal barrier dysfunction and subsequent subtle mucosal inflammation. Cannabinoid 1 receptor (CB1R) is expressed in the mammalian gut under physiological circumstances. The aim of this investigation is to study the possible role of CB1R in the maintenance of mucosal homeostasis after stress exposure. CB1R knockout mice (CB1R(-/-)) and their wild-type (WT) counterparts were exposed to immobilization and acoustic (IA) stress for 2 h per day during 4 consecutive days. Colonic protein expression of the inducible forms of the nitric oxide synthase and cyclooxygenase (NOS2 and COX2), IgA production, permeability to (51)Cr-EDTA, and bacterial translocation to mesenteric lymph nodes were evaluated. Stress exposure induced greater expression of proinflammatory enzymes NOS2 and COX2 in colonic mucosa of CB1R(-/-) mice when compared with WT animals. These changes were related with a greater degree of colonic barrier dysfunction in CB1R(-/-) animals determined by 1) a significantly lower IgA secretion, 2) higher paracellular permeability to (51)Cr-EDTA, and 3) higher bacterial translocation, both under basal conditions and after IA stress exposure. Pharmacological antagonism with rimonabant reproduced stress-induced increase of proinflammatory enzymes in the colon described in CB1R(-/-) mice. In conclusion, CB1R exerts a protective role in the colon in vivo through the regulation of intestinal secretion of IgA and paracellular permeability. Pharmacological modulation of cannabinoid system within the gastrointestinal tract might be therapeutically useful in conditions on which intestinal inflammation and barrier dysfunction takes place after exposure to stress.

Increased vulnerability to 6-hydroxydopamine lesion and reduced development of dyskinesias in mice lacking CB1 cannabinoid receptors

Motor impairment, dopamine (DA) neuronal activity and proenkephalin (PENK) gene expression in the caudate-putamen (CPu) were measured in 6-OHDA-lesioned and treated (L-DOPA+benserazide) CB1 KO and WT mice. A lesion induced by 6-OHDA produced more severe motor deterioration in CB1 KO mice accompanied by more loss of DA neurons and increased PENK gene expression in the CPu. Oxidative/nitrosative and neuroinflammatory parameters were estimated in the CPu and cingulate cortex (Cg). CB1 KO mice exhibited higher MDA levels and iNOS protein expression in the CPu and Cg compared to WT mice. Treatment with L-DOPA+benserazide (12 weeks) resulted in less severe dyskinesias in CB1 KO than in WT mice. The results revealed that the lack of cannabinoid CB1 receptors increased the severity of motor impairment and DA lesion, and reduced L-DOPA-induced dyskinesias. These results suggest that activation of CB1 receptors offers neuroprotection against dopaminergic lesion and the development of L-DOPA-induced dyskinesias.

Is there a role for the nuclear receptor PPARγ in neuropsychiatric diseases

The aetiology of psychiatric diseases such as depression or schizophrenia remains largely unknown, even though multiple theories have been proposed. Although monoamine theory is the cornerstone of available pharmacological therapies, relapses, incomplete control of symptoms or failure in treatment occur frequently. From an inflammatory/immune point of view, both entities share several common hallmarks in their pathophysiology, e.g. neuroendocrine/immune alterations, structural/functional abnormalities in particular brain areas, and cognitive deficits, suggesting a dysregulated inflammatory-related component of these diseases that better explains the myriad of symptoms presented by affected individuals. In this review we aimed to explore the role and relevance of inflammatory related lipids (prostanoids) derived from arachidonic acid metabolism by identification of new inflammatory markers and possible pharmacological/dietary modulation of these compounds, with the aim of improving some of the symptoms developed by individuals affected with psychiatric diseases (a critical review of basic and clinical studies about inflammatory-related arachidonic acid metabolism on neuropsychiatric diseases is included). As a specific candidate, one of these immunoregulatory lipids, the anti-inflammatory prostaglandin 15d-PGJ₂ and its nuclear receptor peroxisome proliferator-activated nuclear receptor (PPARγ) could be used as a biological marker for psychiatric diseases. In addition, its pharmacological activation can be considered as a multi-faceted therapeutic target due to its anti-inflammatory/antioxidant/anti-excitotoxic/pro-energetic profile, reported in some inflammatory-related scenarios (neurological and stress-related diseases). PPARs are activated by a great variety of compounds, the most relevant being the currently prescribed group of anti-diabetic drugs thiazolidinediones, and some cannabinoids (both endocannabinoids, phytocannabinoids or synthetic), as possible novel therapeutical strategy.