Emotional states as distinct configurations of functional brain networks.

The conceptualization of emotional states as patterns of interactions between large-scale brain networks has recently gained support. Yet, few studies have directly examined the brain's network structure during emotional experiences. Here, we investigated the brain's functional network organization during experiences of sadness, amusement, and neutral states elicited by movies, in addition to a resting-state. We tested the effects of the experienced emotion on individual variability in the brain's functional connectome. Next, for each state, we defined a community structure of the brain and quantified its segregation and integration. We found that sadness, relative to amusement, was associated with higher modular integration and increased connectivity of cognitive control networks: the salience and fronto-parietal networks. Moreover, in both the functional connectome and the emotional report, the similarity between individuals was dependent on the sex. Our results suggest that the experience of emotion is linked to a reconfiguration of whole-brain distributed, not emotion-specific, functional networks and that the brain's topological structure carries information about the subjective emotional experience.

Coordinated Transcriptional Waves Define the Inflammatory Response of Primary Microglial Culture.

The primary role of microglia is to maintain homeostasis by effectively responding to various disturbances. Activation of transcriptional programs determines the microglia's response to external stimuli. In this study, we stimulated murine neonatal microglial cells with benzoyl ATP (bzATP) and lipopolysaccharide (LPS), and monitored their ability to release pro-inflammatory cytokines. When cells are exposed to bzATP, a purinergic receptor agonist, a short-lived wave of transcriptional changes, occurs. However, only combining bzATP and LPS led to a sustainable and robust response. The transcriptional profile is dominated by induced cytokines (e.g., IL-1α and IL-1ß), chemokines, and their membrane receptors. Several abundant long noncoding RNAs (lncRNAs) are induced by bzATP/LPS, including Ptgs2os2, Bc1, and Morrbid, that function in inflammation and cytokine production. Analyzing the observed changes through TNF (Tumor necrosis factor) and NF-κB (nuclear factor kappa light chain enhancer of activated B cells) pathways confirmed that neonatal glial cells exhibit a distinctive expression program in which inflammatory-related genes are upregulated by orders of magnitude. The observed capacity of the microglial culture to activate a robust inflammatory response is useful for studying neurons under stress, brain injury, and aging. We propose the use of a primary neonatal microglia culture as a responsive in vitro model for testing drugs that may interact with inflammatory signaling and the lncRNA regulatory network.

Trait-related changes in brain network topology in premenstrual dysphoric disorder.

The female predominance in the prevalence of depression is partially accounted by reactivity to hormonal fluctuations. Premenstrual dysphoric disorder (PMDD) is a reproductive subtype of depression characterized by cyclic emotional and somatic symptoms that recur before menstruation. Despite the growing understanding that most psychiatric disorders arise from dysfunctions in distributed brain circuits, the brain's functional connectome and its network properties of segregation and integration were not investigated in PMDD. To this end, we examined the brain's functional network organization in PMDD using graph theoretical analysis. 24 drug naïve women with PMDD and 27 controls without premenstrual symptoms underwent 2 resting-state fMRI scans, during the mid-follicular and late-luteal menstrual cycle phases. Functional connectivity MRI, graph theory metrics, and levels of sex hormones were computed during each menstrual phase. Altered network topology was found in PMDD across symptomatic and remitted stages in major graph metrics (characteristic path length, clustering coefficient, transitivity, local and global efficiency, centrality), indicating decreased functional network segregation and increased functional network integration. In addition, PMDD patients exhibited hypoconnectivity of the anterior temporal lobe and hyperconnectivity of the basal ganglia and thalamus, across menstrual phases. Furthermore, the relationship between difficulties in emotion regulation and PMDD was mediated by specific patterns of functional connectivity, including connections of the striatum, thalamus, and prefrontal cortex. The shifts in the functional connectome and its topology in PMDD may suggest trait vulnerability markers of the disorder.

Perinatal citalopram does not prevent the effect of prenatal stress on anxiety, depressive-like behaviour and serotonergic transmission in adult rat offspring.

It is still not clear whether the selective serotonin reuptake inhibitors frequently prescribed to depressed pregnant women improve the behavioural outcome in their children. The current study investigated whether administration of citalopram to pregnant rats could prevent anxiety and depressive-like behaviour induced by gestational stress in their offspring, and restore the expression of serotonin 1A autoreceptors in GABAergic interneurons in the medial prefrontal cortex and dorsal raphe nuclei in males, and of corticotropin-releasing factor type 2 receptors in GABAergic interneurons in the dorsal raphe nuclei in females. Activation of these receptors modulates serotonergic transmission to target areas and is reduced in a sex-dependent manner by prenatal stress. Citalopram (10 mg/kg/day), administered orally from day 7 of gestation until 21 days postpartum, prevented the increase in anxiety in stressed mothers but did not reduce anxiety and depressive-like behaviour in their offspring and even induced depressive-like behaviour in the offspring of control mothers. Citalopram failed to restore the reduction in the expression of serotonin 1A autoreceptors in the prefrontal cortex of males and in corticotropin-releasing factor type 2 receptors in the dorsal raphe nuclei of females induced by prenatal stress. Prenatal citalopram did not prevent the behavioural changes or reduction in serotonergic transmission to target areas induced by prenatal stress. It had adverse behavioural effects in the offspring of control rats, which, together with the lack of any change in prenatally-stressed rats, may be due to inhibition of the foetal serotonin transporter thereby preventing normal development of the serotonin system.

Sex-Specific Effects of Prenatal Stress on Memory and Markers of Neuronal Activity in Juvenile Rats.

Stress during pregnancy can increase the incidence of emotional problems, learning and language difficulties in human infants and pre-adolescents. Most preclinical studies in rats that attempted to find experimental support for these observations were performed in adult male offspring, but the results are inconsistent. The aim of the current study was to examine the effect of prenatal stress on novel object recognition (NOR) and spatial learning and memory in the Morris water maze (MWM) of juvenile rats of both sexes. By the use of fluorescence immunohistochemistry and protein measurements by Western blot, we measured the expression of markers of neurogenesis (doublecortin, DCX) and neuronal activity that are important for synaptic plasticity and learning (c-fos, GluR1, nNOS). Since neuronal activity in the developing and adult brain can be regulated by astrocytes, we also measured the number of astrocytes and the expression of two astroglial proteins (GFAP and S100B) in the stress-responsive hippocampal dentate gyrus (DG). Experiments were performed on littermates of rats in which its effects on behavior were measured. We found for the first time that juvenile females performed better than males in the NOR and MWM tests. They also had higher densities of DCX and c-fos in the DG, together with the expression of nNOS and GluR1 in the subgranular zone (SGZ) of the DG. There were no sex differences in the expression of GFAP and S100B in astrocytes. Prenatal stress did not affect NOR in females, but improved it in males, together with an increase in DCX+ and c-fos, the number of GFAP-expressing astrocytes and the intensity of GFAP and S100B immunofluorescence in the DG. Staining intensity of GluR1 and nNOS in the hilus and SGZ of the DG, and protein expression in the whole DG, was unchanged in prenatally stressed males. Thus, prenatal stress changed the behavior and expression of key proteins in the DG to resemble that in females. A reduction in plasma testosterone, which although not attaining statistical significance was associated with that in anogenital distance, may contribute to the effect of prenatal stress in males. In females, prenatal stress had no effect on c-fos, DCX or the number of astrocytes but reduced the staining intensity of GluR1 and nNOS. Protein expression of nNOS was also significantly lower than that in prenatally stressed males. The differential effects of prenatal stress on hippocampal neuronal and glial markers may help to explain the sex-dependent effect on spatial learning in prepubertal rats.

Synthesis and in vitro evaluation of anti-inflammatory activity of ester and amine derivatives of indoline in RAW 264.7 and peritoneal macrophages.

UNLABELLED: A prolonged increase in pro-inflammatory cytokines, TNF-α and IL-6 occurs in inflammatory diseases. Although existing therapies like steroids and TNF-α antagonists are effective they may cause serious adverse effects. We describe the preparation and evaluation for anti-inflammatory activity of 11 novel derivatives of indoline carbamates with a propionic ester, 2-aminoethyl, 3-aminopropyl 2-(dimethylamino)ethyl or 3-(dimethylamino)propyl group in positions 3 or 1. Compounds 25, 26 and 29 were previously shown to inhibit acetylcholinesterase with IC50s ranging from 0.4 to 55µM and to prevent cytotoxicity induced by reactive oxygen species in a concentration range of 100pM-1µM. Compounds 25, 26, 29, 9, 10, 17 and 18, reduced NO, TNF-α and IL-6 at concentrations of 1-10pM in LPS-activated RAW-264.7 and mouse peritoneal macrophages. The reduction in cytokines by compound 25 was associated with an increase in IκBα degradation and a decrease in the phosphorylation of p38 but not that of ERK. CONCLUSION: Indoline derivatives substituted at position 3 with chains carrying ester or amino groups may have potential for the treatment of chronic inflammatory and neurodegenerative diseases.

Ladostigil Reduces the Adenoside Triphosphate/Lipopolysaccharide-Induced Secretion of Pro-Inflammatory Cytokines from Microglia and Modulate-Immune Regulators, TNFAIP3, and EGR1.

Treatment of aging rats for 6 months with ladostigil (1 mg/kg/day) prevented a decline in recognition and spatial memory and suppressed the overexpression of gene-encoding pro-inflammatory cytokines, TNFα, IL1ß, and IL6 in the brain and microglial cultures. Primary cultures of mouse microglia stimulated by lipopolysaccharides (LPS, 0.75 µg/mL) and benzoyl ATPs (BzATP) were used to determine the concentration of ladostigil that reduces the secretion of these cytokine proteins. Ladostigil (1 × 10-11 M), a concentration compatible with the blood of aging rats in, prevented memory decline and reduced secretion of IL1ß and IL6 by ≈50%. RNA sequencing analysis showed that BzATP/LPS upregulated 25 genes, including early-growth response protein 1, (Egr1) which increased in the brain of subjects with neurodegenerative diseases. Ladostigil significantly decreased Egr1 gene expression and levels of the protein in the nucleus and increased TNF alpha-induced protein 3 (TNFaIP3), which suppresses cytokine release, in the microglial cytoplasm. Restoration of the aberrant signaling of these proteins in ATP/LPS-activated microglia in vivo might explain the prevention by ladostigil of the morphological and inflammatory changes in the brain of aging rats.

AN1284 attenuates steatosis, lipogenesis, and fibrosis in mice with pre-existing non-alcoholic steatohepatitis and directly affects aryl hydrocarbon receptor in a hepatic cell line.

Non-alcoholic steatohepatitis (NASH) is an aggressive form of fatty liver disease with hepatic inflammation and fibrosis for which there is currently no drug treatment. This study determined whether an indoline derivative, AN1284, which significantly reduced damage in a model of acute liver disease, can reverse steatosis and fibrosis in mice with pre-existing NASH and explore its mechanism of action. The mouse model of dietary-induced NASH reproduces most of the liver pathology seen in human subjects. This was confirmed by RNA-sequencing analysis. The Western diet, given for 4 months, caused steatosis, inflammation, and liver fibrosis. AN1284 (1 mg or 5 mg/kg/day) was administered for the last 2 months of the diet by micro-osmotic-pumps (mps). Both doses significantly decreased hepatic damage, liver weight, hepatic fat content, triglyceride, serum alanine transaminase, and fibrosis. AN1284 (1 mg/kg/day) given by mps or in the drinking fluid significantly reduced fibrosis produced by carbon tetrachloride injections. In human HUH7 hepatoma cells incubated with palmitic acid, AN1284 (2.1 and 6.3 ng/ml), concentrations compatible with those in the liver of mice treated with AN1284, decreased lipid formation by causing nuclear translocation of the aryl hydrocarbon receptor (AhR). AN1284 downregulated fatty acid synthase (FASN) and sterol regulatory element-binding protein 1c (SREBP-1c) and upregulated Acyl-CoA Oxidase 1 and Cytochrome P450-a1, genes involved in lipid metabolism. In conclusion, chronic treatment with AN1284 (1mg/kg/day) reduced pre-existing steatosis and fibrosis through AhR, which affects several contributors to the development of fatty liver disease. Additional pathways are also influenced by AN1284 treatment.

Prenatal stress diminishes gender differences in behavior and in expression of hippocampal synaptic genes and proteins in rats.

The study determined whether there were gender differences in the expression of hippocampal genes in adult rats in association with dissimilarity in their behavior, and how these were affected by prenatal stress. Pregnant Wistar rats were subjected to varied stress once daily on days 14-20 of gestation. Adult female offspring of control rats showed significantly less anxiogenic behavior in the elevated plus maze and better discrimination between a novel and familiar object than males in the object recognition test. These gender differences in behavior were markedly attenuated by prenatal stress. Using Affymetrix DNA chip technology on hippocampal extracts prepared from littermates of the offspring used for behavioral tests, we found that 1,680 genes were differentially expressed in control males and females. The gender difference in gene expression was decreased to 11% (191 genes) by prenatal stress. In both sexes, processes like the translational machinery, mitochondrial activity, and cation transport were downregulated compared to controls, but there was a greater suppression of genes involved in vesicle trafficking, regulation of synaptic plasticity, and neurogenesis in females than in males. This was compensated by a higher expression of other components of vesicle trafficking, microtubule-based processes, and neurite development. Prenatal stress decreased the expression of 19 Rab proteins in females and five Rabs in males, but a compensatory increase of Rab partner proteins and effectors only occurred in females. Exposure to stress decreased the expression of synaptic proteins, synaptophysin, and synaptopodin in prenatally stressed males and females and increased those of PSD-95 and NR1 subunit of the N-methyl-D-aspartic acid (NMDA) glutamate receptor only in females. The study provides an unbiased view of key genes and proteins that act as gender dependent molecular sensors. The disruption of their expression by adverse early life stress may explain the alterations that occur in behavior.

Comparison of the tissue distribution and metabolism of AN1284, a potent anti-inflammatory agent, after subcutaneous and oral administration in mice.

This study is to compare the tissue distribution and metabolism of AN1284 after subcutaneous and oral administration at doses causing maximal reductions in IL-6 in plasma and tissues of mice. Anti-inflammatory activity of AN1284 and its metabolites was detected in lipopolysaccharide (LPS) activated RAW 264.7 macrophages. Mice were given AN1284 by injection or gavage, 15 min before LPS. IL-6 protein levels were measured after 4 h. Using a liquid chromatography/mass spectrometry method we developed, we showed that AN1284 is rapidly metabolized to the indole (AN1422), a 7-OH derivative (AN1280) and its glucuronide. AN1422 has weaker anti-inflammatory activity than AN1284 in LPS-activated macrophages and in mice. AN1284 (0.5 mg/kg) caused maximal reductions in IL-6 in the plasma, brain, and liver when injected subcutaneously and after gavage only in the liver. Similar reductions in the plasma and brain required a dose of 2.5 mg/kg, which resulted in 5.5-fold higher hepatic levels than after injection of 0.5 mg/kg, but 7, 11, and 19-fold lower ones in the plasma, brain, and kidneys, respectively. Hepatic concentrations produced by AN1284 were 2.5 mg/kg/day given by subcutaneously implanted mini-pumps that were only 12% of the peak levels seen after acute injection of 0.5 mg/kg. Similar hepatic concentrations were obtained by (1 mg/kg/day), administered in the drinking fluid. These were sufficient to decrease hepatocellular damage and liver triglycerides in previous experiments in diabetic mice. AN1284 can be given orally by a method of continuous release to treat chronic liver disease, and its preferential concentration in the liver should limit any adverse effects.

Effect of ladostigil treatment of aging rats on gene expression in four brain areas associated with regulation of memory.

Episodic and spatial memory decline in aging and are controlled by the hippocampus, perirhinal, frontal and parietal cortices and the connections between them. Ladostigil, a drug with antioxidant and anti-inflammatory activity, was shown to prevent the loss of episodic and spatial memory in aging rats. To better understand the molecular effects of aging and ladostigil on these brain regions we characterized the changes in gene expression using RNA-sequencing technology in rats aged 6 and 22 months. We found that the changes induced by aging and chronic ladostigil treatment were brain region specific. In the hippocampus, frontal and perirhinal cortex, ladostigil decreased the overexpression of genes regulating calcium homeostasis, ion channels and those adversely affecting synaptic function. In the parietal cortex, ladostigil increased the expression of several genes that provide neurotrophic support, while reducing that of pro-apoptotic genes and those encoding pro-inflammatory cytokines and their receptors. Ladostigil also decreased the expression of axonal growth inhibitors and those impairing mitochondrial function. Together, these actions could explain the protection by ladostigil against age-related memory decline.

A Novel Indoline Derivative Ameliorates Diabesity-Induced Chronic Kidney Disease by Reducing Metabolic Abnormalities.

Both diabetes and obesity (diabesity) contribute significantly to the development of chronic kidney disease (CKD). In search of new remedies to reverse or arrest the progression of CKD, we examined the therapeutic potential of a novel compound, AN1284, in a mouse model of CKD induced by type 2 diabetes with obesity. Six-week-old BKS Cg-Dock 7m+/+ Leprdb/J mice with type 2 diabetes and obesity were treated with AN1284 (2.5 or 5 mg kg-1 per day) via micro-osmotic pumps implanted subcutaneously for 3 months. Measures included renal, pancreatic, and liver assessment as well as energy utilization. AN1284 improved kidney function in BSK-db/db animals by reducing albumin and creatinine and preventing renal inflammation and morphological changes. The treatment was associated with weight loss, decreased body fat mass, increased utilization of body fat toward energy, preservation of insulin sensitivity and pancreatic ß cell mass, and reduction of dyslipidemia, hepatic steatosis, and liver injury. This indoline derivative protected the kidney from the deleterious effects of hyperglycemia by ameliorating the metabolic abnormalities of diabetes. It could have therapeutic potential for preventing CKD in human subjects with diabesity.

Age-Induced Spatial Memory Deficits in Rats Are Correlated with Specific Brain Region Alterations in Microglial Morphology and Gene Expression.

Effect of age and ladostigil treatment (1 mg/kg/day), given for 6 months to 16 month old rats, was investigated on microglial morphology in brain regions associated with control of spatial learning. This was assessed in the Morris water maze (MWM). Microglial morphology was assessed with diaminobenzidine and fluorescent staining with Iba1 and CD11b in these brain regions. Aging did not change the number of microglia in the parietal cortex (PC) or hippocampal CA1 region (CA1-HC), but decreased microglial process tips in the CA1-HC, increased the area fraction stained by CD11b and number of bulbs on processes in PC and CA1-HC and thickness of microglial processes in corpus callosum (CC) and fornix (Fx). Performance in MWM (distance swam to escape platform) was negatively correlated with number of bulbs in PC and thickness of process in CC, and positively correlated with number of process tips in CA1-HC. Aging increased expression of MHC class II genes and others associated with motility and membrane adhesion in the PC and hippocampus, but Adora2a (Adenosine A2a receptor), only in hippocampus. Age-related increase in the number of bulbs and expression of inflammatory genes was prevented by ladostigil in PC. In the CA1-HC, ladostigil increased the number of process tips and prevented the increase in expression of Adora2a and genes regulating ion channels. Ladostigil also decreased thickening of the processes in CC and Fx. The data show brain region-specific relations induced by age in spatial learning, microglial morphology and associated genes and their response to ladostigil treatment. Graphical Abstract.

Low-dose ladostigil for mild cognitive impairment: A phase 2 placebo-controlled clinical trial.

OBJECTIVE: Ladostigil reduces oxidative stress and microglial activation in aging rats. We assessed its safety and potential efficacy in a 3-year, randomized, double-blind, placebo-controlled phase 2 clinical trial in patients with mild cognitive impairment (MCI) and medial temporal lobe atrophy. METHODS: Patients 55 to 85 years of age with MCI, Clinical Dementia Rating (CDR) score of 0.5, Mini-Mental State Examination (MMSE) score >24, Wechsler Memory Scale-Revised Verbal Paired Associates I score ≤18, and Medial Temporal Lobe Atrophy Scale score >1 were stratified by APOE ε4 genotype and randomly assigned (1:1) to ladostigil 10 mg/d or placebo. Primary outcomes were safety and onset of Alzheimer disease dementia. Secondary endpoints were Neuropsychological Test Battery (NTB) composite, Disability Assessment in Dementia (DAD), and Geriatric Depression Scale (GDS) scores. Exploratory outcomes were NTB component, CDR, and MMSE scores. Biomarkers included MRI-derived whole-brain, hippocampus, and entorhinal cortex volumes. RESULTS: Two hundred ten patients from 15 sites in Austria, Germany, and Israel were randomly allocated to placebo (107 patients) or ladostigil (103 patients). After 36 months, 21 of 103 patients on placebo and 14 of 99 patients receiving ladostigil progressed to Alzheimer disease (log-rank test p = 0.162). There were no significant effects on the NTB composite, DAD, or GDS score. Whole-brain and hippocampus volumes decreased more in the placebo than in the ladostigil group (whole brain, p = 0.025, Cohen d = 0.43; hippocampus, p = 0.043, d = 0.43). Serious adverse events were reported by 28 of 107 patients treated with placebo and 26 of 103 with ladostigil. CONCLUSION: Ladostigil was safe and well tolerated but did not delay progression to dementia. Its association with reduced brain and hippocampus volume loss suggests a potential effect on atrophy. CLINICALTRIALSGOV IDENTIFIER: NCT01429623. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that for patients with MCI and medial temporal lobe atrophy, ladostigil did not significantly decrease the risk of the development of Alzheimer disease.

Sex differences during emotion processing are dependent on the menstrual cycle phase.

Sex differences in the neural processing of emotion are of special interest considering that mood and anxiety disorders predominant in females. However, these sex-related differences were typically studied without considering the hormonal status of female subjects, although emotion processing in the brain was shown to differ between phases of the menstrual cycle. In this functional MRI study, we demonstrated the influence of the menstrual cycle phase on sex differences in brain activity and functional connectivity during negative and positive emotions, using two different paradigms: emotion perception and emotion experience. Twenty naturally cycling healthy women without premenstrual symptoms were scanned twice: during the mid-follicular and late-luteal menstrual phases, and compared to a matched group of twenty healthy men. During negative emotion perception, men showed increased neural activity in the right hippocampal formation relative to women in the mid-follicular phase, and increased activity in the right cerebellum relative to women in the late-luteal phase. During experience of amusement, reduced putamen-ventrolateral prefrontal cortex and putamen-dorsomedial prefrontal cortex functional connectivity were observed for women in the late-luteal phase relative to men and associated with levels of sex hormones. These neural and hormonal findings were complemented by behavioral reports of reduced amusement and increased sadness in late-luteal women. Our results demonstrate menstrual phase-dependent sex differences in emotion perception and experience and may suggest a biological tendency for a deficient experience of pleasure and reward during the late-luteal phase. These findings may further shed light on the underlying pathophysiology of premenstrual dysphoric disorder.

The long-term behavioural consequences of prenatal stress.

UNLABELLED: Maternal distress during pregnancy increases plasma levels of cortisol and corticotrophin releasing hormone in the mother and foetus. These may contribute to insulin resistance and behaviour disorders in their offspring that include attention and learning deficits, generalized anxiety and depression. The changes in behaviour, with or independent of alterations in the function of the hypothalamic pituitary adrenal (HPA) axis, can be induced by prenatal stress in laboratory rodents and non-human primates. The appearance of such changes depends on the timing of the maternal stress, its intensity and duration, gender of the offspring and is associated with structural changes in the hippocampus, frontal cortex, amygdala and nucleus accumbens. The dysregulation of the HPA axis and behaviour changes can be prevented by maternal adrenalectomy. However, only the increased anxiety and alterations in HPA axis are re-instated by maternal injection of corticosterone. CONCLUSION: Excess circulating maternal stress hormones alter the programming of foetal neurons, and together with genetic factors, the postnatal environment and quality of maternal attention, determine the behaviour of the offspring.

Suppression of neuroinflammation and immunomodulation by the acetylcholinesterase inhibitor rivastigmine.

In this study we determined the influence of cholinergic up-regulation by rivastigmine, an acetylcholinesterase inhibitor, on central nervous system inflammation. Neuroinflammation was induced in experimental autoimmune encephalomyelitis (EAE). Rivastigmine markedly ameliorated clinical symptoms of EAE and the spatial memory deficits induced by EAE. It also reduced demyelination, microglia activation and axonal damage. Rivastigmine decreased the reactivity of encephalitogenic T-cells and the production of pro-inflammatory cytokines (TNF-alpha, IFN-gamma and IL-17) without affecting IL-10 production. These effects were abolished by alpha7 nicotinic acetylcholine receptor antagonists. Antigen presentation was also affected by this treatment. Thus, rivastigmine treatment had immunomodulatory activity in EAE.

Synthesis and Biological Evaluation of Derivatives of Indoline as Highly Potent Antioxidant and Anti-inflammatory Agents.

We describe the preparation and evaluation of novel indoline derivatives with potent antioxidant and anti-inflammatory activities for the treatment of pathological conditions associated with chronic inflammation. The indolines are substituted at position 1 with chains carrying amino, ester, amide, or alcohol groups, and some have additional substituents, Cl, MeO, Me, F, HO, or BnO, on the benzo ring. Concentrations of 1 pM to 1 nM of several compounds protected RAW264.7 macrophages against H2O2 induced cytotoxicity and LPS induced elevation of NO, TNF-α, and IL-6. Several derivatives had anti-inflammatory activity at 1/100th of the concentration of unsubstituted indoline. Four compounds with ester, amine, amide, or alcohol side chains injected subcutaneously in mice at a dose of 1 µmol/kg or less, like dexamethasone (5.6 µmol/kg) prevented LPS-induced cytokine elevation in the brain and peripheral tissues. Subcutaneous injection of 100 µmol/kg of these compounds caused no noticeable adverse effects in mice during 3 days of observation.

Ladostigil prevents gliosis, oxidative-nitrative stress and memory deficits induced by intracerebroventricular injection of streptozotocin in rats.

Glial activation and oxidative-nitrative stress occur at an early stage in Alzheimer's disease (AD). In a rat model of AD, deficits in cerebral glucose utilization and memory were seen 3-4 weeks after intracerebroventricular (icv) injection of streptozotocin (STZ). This study examined whether icv STZ induced glial activation and oxidative-nitrative stress preceded the memory deficits and whether they could be prevented by ladostigil a novel drug, a cholinesterase and monoamine oxidase inhibitor with neuroprotective activity. One week after STZ injection activated microglia and astrocytes were seen in the cortex, around the cannula penetration area, in the hippocampal CA1 region, corpus callosum, medial and lateral septum. The activated astrocytes showed a significant increase in nitrotyrosine immunoreactivity, a measure of oxidative-nitrative stress. Only 3 weeks later were deficits in episodic (object recognition test) and spatial memory (place recognition) seen in STZ-injected rats. Daily oral administrations of ladostigil (1mg/kg) for 1 week, before and after STZ prevented the glial changes, increase in nitrotyrosine immunoreactivity and memory deficits. Taken together the data support the role of glial activation and oxidative-nitrative stress in discrete brain areas in the aetiology of memory deficits and indicate a potential mechanism for their prevention by drug treatment.

Effect of varied gestational stress on acquisition of spatial memory, hippocampal LTP and synaptic proteins in juvenile male rats.

Some but not other forms of prenatal stress have been shown to impair spatial memory in adult male offspring. It is not clear if this is because of the intensity of the stress, age of rats, or the way in which learning is assessed. We examined the effect of daily varied prenatal stress consisting of 30 min restraint, saline injections and 15 min forced swim on day 17-21 of gestation on spatial learning, synaptic plasticity and the expression of key proteins of the post synaptic density (PSD) in the hippocampus of males aged 4-5 weeks. Prenatal stress impaired spatial learning in the Morris water maze and induced a significant decrease in long-term potentiation (LTP) in hippocampal slices. There was no change in the paired pulse facilitation ratio but there was a significant reduction in the expression of the NR2B subunit of the glutamate type NMDA receptor and the GluR1 subunit of the AMPA receptor, both of which are important modulators of LTP. These changes were accompanied by a remarkable increase in the scaffolding protein PSD95, which interacts with the intracellular carboxy terminal domains of the NR2 subunits. The high levels of PSD95 may have contributed to the impairment of LTP by disrupting the clustering of NMDA receptors in CA1 synapses. The alteration by prenatal stress in the relative amounts of scaffolding proteins and those which compose glutamate receptors could explain the depression of LTP and impairment in the acquisition of spatial learning.