Patterns of activity correlate with symptom severity in major depressive disorder patients.

Objective measures, such as activity monitoring, can potentially complement clinical assessment for psychiatric patients. Alterations in rest-activity patterns are commonly encountered in patients with major depressive disorder. The aim of this study was to investigate whether features of activity patterns correlate with severity of depression symptoms (evaluated by Montgomery-Åsberg Rating Scale (MADRS) for depression). We used actigraphy recordings collected during ongoing major depressive episodes from patients not undergoing any antidepressant treatment. The recordings were acquired from two independent studies using different actigraphy systems. Data was quality-controlled and pre-processed for feature extraction following uniform procedures. We trained multiple regression models to predict MADRS score from features of activity patterns using brute-force and semi-supervised machine learning algorithms. The models were filtered based on the precision and the accuracy of fitting on training dataset before undergoing external validation on an independent dataset. The features enriched in the models surviving external validation point to high depressive symptom severity being associated with less complex activity patterns and stronger coupling to external circadian entrainers. Our results bring proof-of-concept evidence that activity patterns correlate with severity of depressive symptoms and suggest that actigraphy recordings may be a useful tool for individual evaluation of patients with major depressive disorder.

Methylmercury interferes with glucocorticoid receptor: Potential role in the mediation of developmental neurotoxicity.

Methylmercury (MeHg) is a widespread environmental contaminant with established developmental neurotoxic effects. Computational models have identified glucocorticoid receptor (GR) signaling to be a key mediator behind the birth defects induced by Hg, but the mechanisms were not elucidated. Using molecular dynamics simulations, we found that MeHg can bind to the GR protein at Cys736 (located close to the ligand binding site) and distort the conformation of the ligand binging site. To assess the functional consequences of MeHg interaction with GR, we used a human cell line expressing a luciferase reporter system (HeLa AZ-GR). We found that 100 nM MeHg does not have any significant effect on GR activity alone, but the transactivation of gene expression by GR upon Dex (a synthetic GR agonist) administration was reduced in cells pre-treated with MeHg. Similar effects were found in transgenic zebrafish larvae expressing a GR reporter system (SR4G). Next we asked whether the effects of developmental exposure to MeHg are mediated by the effects on GR. Using a mutant zebrafish line carrying a loss-of-function mutation in the GR (grS357) we could show that the effects of developmental exposure to 2.5 nM MeHg are mitigated in absence of functional GR signaling. Taken together, our data indicate that inhibition of GR signaling may have a role in the developmental neurotoxic effects of MeHg.

Effects of developmental exposure to perfluorooctanoic acid (PFOA) on long bone morphology and bone cell differentiation.

Perfluorooctanoic acid (PFOA) is a ubiquitous and persistent environmental chemical, which has been used extensively due to its stability and surface tension-lowering properties. Toxicological effects include induction of neonatal mortality and reproductive toxicity. In this study, pregnant C57BL/6 mice were exposed orally to 0.3mg PFOA/kg/day throughout pregnancy, and female offspring were studied at the age of 13 or 17months. Morphometrical and biomechanical properties of femurs and tibias were analyzed with micro-computed tomography and 3-point bending, and bone PFOA concentrations were determined by mass spectrometry. The effects of PFOA on bone cell differentiation were studied in osteoclasts from C57BL/6 mice and in the MC3T3 pre-osteoblast cell line. PFOA exposed mice showed increased femoral periosteal area as well as decreased mineral density of tibias. Biomechanical properties of these bones were not affected. Bone PFOA concentrations were clearly elevated even at the age of 17months. In osteoblasts, low concentrations of PFOA increased osteocalcin (OCN) expression and calcium secretion, but at PFOA concentrations of 100µM and above osteocalcin (OCN) expression and calcium secretion were decreased. The number of osteoclasts was increased at all PFOA concentrations tested and resorption activity dose-dependently increased from 0.1-1.0µM, but decreased at higher concentrations. The results show that PFOA accumulates in bone and is present in bones until the old age. PFOA has the potential to influence bone turnover over a long period of time. Therefore bone is a target tissue for PFOA, and altered bone geometry and mineral density seem to persist throughout the life of the animal.

Alterations in circadian entrainment precede the onset of depression-like behavior that does not respond to fluoxetine.

Growing evidence links adverse prenatal conditions to mood disorders. We investigated the long-term behavioral alterations induced by prenatal exposure to excess glucocorticoids (dexamethasone--DEX). At 12 months, but not earlier, DEX-exposed mice displayed depression-like behavior and impaired hippocampal neurogenesis, not reversible by the antidepressant fluoxetine (FLX). Concomitantly, we observed arrhythmic glucocorticoid secretion and absent circadian oscillations in hippocampal clock gene expression. Analysis of spontaneous activity showed progressive alterations in circadian entrainment preceding depression. Circadian oscillations in clock gene expression (measured by means of quantitative PCR) were also attenuated in skin fibroblasts before the appearance of depression. Interestingly, circadian entrainment is not altered in a model of depression (induced by methylmercury prenatal exposure) that responds to FLX. Altogether, our results suggest that alterations in circadian entrainment of spontaneous activity, and possibly clock gene expression in fibroblasts, may predict the onset of depression and the response to FLX in patients.

Tet3 mediates stable glucocorticoid-induced alterations in DNA methylation and Dnmt3a/Dkk1 expression in neural progenitors.

Developmental exposure to excess glucocorticoids (GCs) has harmful neurodevelopmental effects, which include persistent alterations in the differentiation potential of embryonic neural stem cells (NSCs). The mechanisms, however, are largely unknown. Here, we investigated the effects of dexamethasone (Dex, a synthetic GC analog) by MeDIP-like genome-wide analysis of differentially methylated DNA regions (DMRs) in NSCs isolated from embryonic rat cortices. We found that Dex-induced genome-wide DNA hypomethylation in the NSCs in vitro. Similarly, in utero exposure to Dex resulted in global DNA hypomethylation in the cerebral cortex of 3-day-old mouse pups. Dex-exposed NSCs displayed stable changes in the expression of the DNA methyltransferase Dnmt3a, and Dkk1, an essential factor for neuronal differentiation. These alterations were dependent on Tet3 upregulation. In conclusion, we propose that GCs elicit strong and persistent effects on DNA methylation in NSCs with Tet3 playing an essential role in the regulation of Dnmt3a and Dkk1. Noteworthy is the occurrence of similar changes in Dnmt3a and Dkk1 gene expression after exposure to excess GC in vivo.

Long-lasting neurotoxic effects of exposure to methylmercury during development.

Amongst environmental chemical contaminants, methylmercury (MeHg) remains a major concern because of its detrimental effects on developing organisms, which appear to be particularly susceptible to its toxicity. Here, we investigated the effects of low MeHg levels on the development of the nervous system using both in vitro and in vivo experimental models. In neural stem cells (NSCs), MeHg decreased proliferation and neuronal differentiation and induced cellular senescence associated with impairment in mitochondrial function and a concomitant decrease in global DNA methylation. Interestingly, the effects were heritable and could be observed in daughter NSCs never directly exposed to MeHg. By chronically exposing pregnant/lactating mice to MeHg, we found persistent behavioural changes in the male offspring, which exhibited depression-like behaviour that could be reversed by chronic treatment with the antidepressant fluoxetine. The behavioural alterations were associated with a decreased number of proliferating cells and lower expression of brain-derived neurotrophic factor (Bdnf) mRNA in the hippocampal dentate gyrus. MeHg exposure also induced long-lasting DNA hypermethylation, increased histone H3-K27 tri-methylation and decreased H3 acetylation at the Bdnf promoter IV, indicating that epigenetic mechanisms play a critical role in mediating the long-lasting effects of perinatal exposure to MeHg. Fluoxetine treatment restored the Bdnf mRNA expression levels, as well as the number of proliferating cells in the granule cell layer of the dentate gyrus, which further supports the hypothesis that links depression to impaired neurogenesis. Altogether, our findings have shown that low concentrations of MeHg induce long-lasting effects in NSCs that can potentially predispose individuals to depression, which we have reported earlier to occur in experimental animals exposed to MeHg during prenatal and early postnatal development.

IL-1/IL-1ra balance in the brain revisited - evidence from transgenic mouse models.

The interleukin-1 (IL-1) family is unique in its including an endogenous antagonist of the IL-1 receptor (IL-1ra). IL-1ra has been shown to antagonise IL-1 signalling so effectively, that it came into clinical use within a few years from its discovery. Although barely detectable in the normal brain, IL-1 is dramatically upregulated during neuroinflammation, and also displays peaks of expression in the brain during development, as well as following the induction of long-term potentiation. IL-1 has been ascribed a central role in neuroinflammation accompanying ageing and age-related neurodegenerative conditions. Several experimental models based on genetically modified mice have been used in order to address the role of IL-1 in neurodegeneration and neuroprotection. Most of the findings here are based on the experiments involving a transgenic mouse strain with brain-directed overexpression of human IL-1ra, in which the balance between IL-1 and IL-1ra is permanently tipped towards inhibiting IL-1 signalling. The developmental effects of IL-1 are evident in the altered brain morphology in adult transgenic mice. In addition, IL-1 appears to be central in regulating the elasticity of the brain response to injury. Thus, a number of lines of evidence support the essential role played by IL-1 in development, plasticity, and physiological brain function.

Studies on brain volume, Alzheimer-related proteins and cytokines in mice with chronic overexpression of IL-1 receptor antagonist.

Inflammation is associated with both acute and chronic neurological disorders, including stroke and Alzheimer's disease (AD). Cytokines such as interleukin (IL)-1 have several activities in the brain both under physiological and pathophysiological conditions. The objective of this study was to evaluate consequences of the central blockade of IL-1 transmission in a previously developed transgenic mouse strain with brain-directed overexpression of human soluble IL-1 receptor antagonist (Tg hsIL-1ra). Effects on brain morphology and brain levels of the AD-related proteins beta-amyloid precursor protein (APP) and presenilin 1(PS1), as well as the levels of IL-1beta, IL-6 and tumour necrosis factor-alpha (TNF-alpha) were analysed in homozygotic and heterozygotic mice and wild type (WT) controls, of both genders and of young (30-40 days) and adult (13-14 months) age. A marked reduction in brain volume was observed in transgenic mice as determined by volumetry. Western blot analysis showed higher levels of APP, but lower levels of PS1, in adult animals than in young ones. In the cerebellum, heterozygotic (Tg hsIL-1ra(+/-)) mice had lower levels of APP and PS1 than WT mice. With one exception, there were no genotypic differences in the levels of IL-1beta, IL-6 and TNF-alpha. The cytokine levels were generally higher in adult than in young mice. In conclusion, the chronic blockade of IL-1 signalling in the brain was associated with an atrophic phenotype of the brain, and with modified levels of APP and PS1. Brain-directed overexpression of hsIL-1ra was not followed by major compensatory changes in the levels of pro-inflammatory cytokines.

The relationship between respiratory sinus arrhythmia and heart rate during anesthesia in rat.

During inspiration the heart rate (HR) increases and during expiration it decreases. Contribution of respiratory sinus arrhythmia (RSA) to spontaneous heart rate variability (HRV) can be measured as the high frequency (HF) component of variation in consecutive R-R intervals on ECG. In conscious rats, slowing of HR is associated with an increase in HF. The aim of this study was to investigate whether this relationship between HF and HR is preserved during anesthesia in rat. A 15 minutes long ECG signal was recorded from rats (N=15) under moderate chloral hydrate (CHL) anesthesia. Recordings were extended with 45 minutes to investigate the effect of atropine (N=3), against controls (N=3). Short term HRV was investigated in 30 seconds long epochs. HF was considered the frequency band between 0.8 and 1.6 Hz. RSA was quantified as the relative spectral power of the HF. Respiratory frequency (RF) was quantified as the mean spectral frequency within the HF band. One minute estimates of HR, RSA and HF were calculated by averaging 3 epochs of 30 seconds overlapped 50%. The average HR was 427 +/- 3 bpm. The magnitude of RSA was 45 +/- 1% at a RF of 71 +/- 1 rpm. We found that: (1) the decrease in HR that occurs during CHL anesthesia in rat correlates with an increase in RSA; (2) atropine reduces RSA and the time-dependent decrease in HR; (3) the time-dependent increase in RSA is preserved after atropine. We conclude that the correlation between RSA and HR reflects the cardio-pulmonary coupling under parasympathetic control.

Oxidative damage following cerebral ischemia depends on reperfusion - a biochemical study in rat.

The extent of brain injury during reperfusion appears to depend on the experimental pattern of ischemia/reperfusion. The goals of this study were: first, to identify the rate of free radicals generation and the antioxidant activity during ischemia and reperfusion by means of biochemical measurement of lipid peroxidation (LPO) and both enzymatic (superoxide dismutase - SOD, catalase - CAT, glutathione peroxidase - GPx) and non-enzymatic antioxidants activity (glutathione - GSH); and second, to try to find out how the pattern of reperfusion may influence the balance between free radical production and clearance. Wistar male rats were subject of four-vessel occlusion model (Pulsinelly & Brierley) cerebral blood flow being controlled by means of two atraumatic arterial microclamps placed on carotid arteries. The level of free radicals and the antioxidant activity were measured in ischemic rat brain tissue homogenate using spectrophotometrical techniques. All groups subjected to ischemia shown an increase of LPO and a reduction of the activity of enzymatic antioxidative systems (CAT, GPx, SOD) and non-enzymatic systems (GSH). For both groups subjected to ischemia and reperfusion, results shown an important increase of LPO but less significant than the levels found in the group with ischemia only. Statistically relevant differences (p<0.01) between continuous reperfusion and fragmented reperfusion were observed concerning the LPO, CAT, SOD and GSH levels, oxidative aggression during fragmented reperfusion being more important.