Learning abilities, NGF and BDNF brain levels in two lines of TNF-α transgenic mice, one characterized by neurological disorders, the other phenotypically normal

In this study we used two lines of transgenic mice overexpressing tumor necrosis factor alpha (TNF-α) in the central nervous system (CNS), one characterized by reactive gliosis, inflammatory demyelination and neurological deficits (Tg6074) the other showing no neurological or phenotypical alterations (TgK3) to investigate the effect of TNF-α on brain nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) levels and learning abilities. The results showed that the amount of NGF in the brain of Tg6074 and TgK3 transgenic mice is low in the hippocampus and in the spinal cord, increases in the hypothalamus of Tg6074 and showed no significant changes in the cortex. BDNF levels were low in the hippocampus and spinal cord of TgK3. BDNF increased in the hypothalamus of TgK3 and Tg6074 while in the cortex, BDNF increased only in Tg6074 mice. Transgenic mice also had memory impairments as revealed by the Morris Water Maze test. These findings indicate that TNF-α significantly influences BDNF and NGF synthesis, most probably in a dose-dependent manner. Learning abilities were also differently affected by overexpression of TNF-α, but were not associated with inflammatory activity. The possible functional implications of our findings are discussed.     

Learning performances, brain NGF distribution and NPY levels in transgenic mice expressing TNF-alpha

Tumor necrosis factor-alpha (TNF-alpha) is a cytokine involved in a variety of neurobiological activities including changing behavior and regulation of both neurotrophin and neuropeptide levels. In this study we used two lines of transgenic mice overexpressing brain TNF-alpha characterized by neurological deficits (line Tg6074) or phenotypically normal (line TgK3). We analyzed whether or not impairments in learning and memory processes due to TNF-alpha overexpression were associated with changes in endogenous brain NGF, NPY and beta-amyloid. The results indicate that full TNF-alpha transgene expression disrupted the learning capabilities of transgenic mice (both Tg6074 and TgK3). NGF decreased in the hippocampus of both transgenic mice whereas hippocampal NPY slightly potentiated in Tg6074. The decrease in NGF is correlated with deficits in spatial learning and memory whereas inflammation in the brain of Tg6074 could be responsible of the hippocampal increase in NPY. As a whole, these results show that transgenic mice overexpressing TNF-alpha in the brain represent a useful model for studying neuronal degeneration and brain inflammatory processes.     

Differential effect of short-term REM sleep deprivation on NGF and BDNF protein levels in the rat brain

It is well known that REM sleep is associated with memory consolidation, especially, procedural skill learning. Neurotrophic factors are known to be involved in synaptic plasticity. We therefore investigated the effects of selective REM sleep deprivation (RSD) on NGF and BDNF proteins in the hippocampus, cerebellum and brainstem in the rat. NGF and BDNF were detected by an ELISA. Our findings show that 6 h RSD affected the NGF and BDNF protein levels in different manner. In the cerebellum and brainstem, BDNF was significantly decreased, while NGF was not changed. Conversely, in the hippocampus, NGF was significantly decreased while BDNF was not changed. This study indicates that REM sleep may be associated with the secretion of neurotrophic factors and thus contribute to the memory functions.     

Thyroid hormone regulation of NGF, NT-3 and BDNF RNA in the adult rat brain.

The effects of peripherally administered thyroid hormone (TH; 500 micrograms/kg; i.p.; q.d.) on the relative abundances of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) RNA were determined by rtPCR in the cortex and hippocampus of young adult rats. Corresponding changes in choline acetyltransferase (ChAT) activity were measured since NGF and BDNF have been shown to enhance the expression of this marker enzyme of central cholinergic pathways. Abundance levels of NGF and NT-3, relative to cyclophilin (cycl), were increased significantly (+50%, P < 0.05) in the hippocampus following TH treatment. Despite enhanced abundance of NGF in the hippocampus, ChAT activity was unchanged, whereas ChAT activity was modestly increased by 28% in the cortex without corresponding changes in NGF, NT-3 or BDNF. These results demonstrate that TH administration is capable of inducing the accumulation of NT-3, in addition to NGF but that the induction levels of RNA cannot be directly correlated with responsivity of the cholinergic system as measured by ChAT activity.     

EGF and NGF injected into the brain of old mice enhance BDNF and ChAT in proliferating subventricular zone

The response of cells localized in the brain subventricular zone (SVZ) to growth factor stimulation has been largely described for development and adult life, whereas no information on their behavior during aging is available. To address the question of whether the cells in the SVZ of old mice respond to the intracerebroventricular administration of epidermal growth factor (EGF) and nerve growth factor (NGF), we studied the distribution of proliferating cells and the effects on ChAT and brain-derived neurotrophic factor (BDNF) synthesis in forebrain and SVZ. It was found that the conjoint administration of EGF + NGF produced a major increase in ChAT expression in both forebrain and SVZ. The ChAT mRNA levels and the number of ChAT positive cells localized in the ventricular border and in the parenchyma of SVZ area were also increased significantly in the mice receiving EGF + NGF. Enhanced numbers of SVZ cells expressing proliferative markers were also discovered in EGF + NGF treated mice and some of these cells expressed cholinergic markers, as demonstrated by double immunostaining. In addition, EGF and NGF treatments significantly upregulate BDNF protein and mRNA levels in this brain region. The present study demonstrates that cells localized in SVZ of aged mouse brain retain the capacity to respond to EGF and NGF and that after stimulation with these two growth factors, the synthesis of ChAT and BDNF also increases. The implication that cells of the SVZ remain a reservoir of cholinergic and BDNF-positive neurons in aged brain opens a new perspective for understanding the role of growth factors during neurodegenerative disorders associated with aging.     

Sodium butyrate and mood stabilizers block ouabain-induced hyperlocomotion and increase BDNF,NGF and GDNF levels in brain of Wistar rats

Bipolar Disorder (BD) is one of the most severe psychiatric disorders. Despite adequate treatment, patients continue to have recurrent mood episodes, residual symptoms, and functional impairment. Some preclinical studies have shown that histone deacetylase inhibitors may act on manic-like behaviors. Neurotrophins have been considered important mediators in the pathophysiology of BD. The present study aims to investigate the effects of lithium (Li), valproate (VPA), and sodium butyrate (SB), an HDAC inhibitor, on BDNF, NGF and GDNF in the brain of rats subjected to an animal model of mania induced by ouabain. Wistar rats received a single ICV injection of ouabain or artificial cerebrospinal fluid. From the day following ICV injection, the rats were treated for 6 days with intraperitoneal injections of saline, Li, VPA or SB twice a day. In the 7th day after ouabain injection, locomotor activity was measured using the open-field test. The BDNF, NGF and GDNF levels were measured in the hippocampus and frontal cortex by sandwich-ELISA. Li, VPA or SB treatments reversed ouabain-related manic-like behavior. Ouabain decreased BDNF, NGF and GDNF levels in hippocampus and frontal cortex of rats. The treatment with Li, VPA or SB reversed these impairment induced by ouabain. In addition, Li, VPA and SB per se increased NGF and GDNF levels in hippocampus of rats. Our data support the notion that neurotrophic factors play a role in BD and in the mechanisms of the action of Li, VPA and SB.     

Neuroticism, other personality variables, and serum lipid levels in patients with anxiety disorders and normal controls

OBJECTIVE: This study evaluates the association between certain personality variables, especially neuroticism, and serum lipid levels in patients suffering from anxiety disorders as opposed to normal control individuals. It addresses the question whether in anxiety disorder patients neuroticism is associated with elevated serum cholesterol and serum LDL cholesterol levels. METHOD: Thirty (15 male, 15 female) patients with anxiety disorders as defined by DSM-IV and thirty normal matched controls were enrolled in this study. The patients were either admitted for inpatient treatment or referred to the behavioral therapy outpatient clinic at the Department of Psychiatry of the University Hospital Eppendorf (Hamburg/Germany). Control individuals were selected on a voluntary basis and recruited from the authors' colleagues and peers. Matching included Body-Mass-Index, age, and gender. RESULTS: This study showed a significant positive association of neuroticism and serum cholesterol (r = .29) and serum LDL cholesterol (r = .35) within the entire cohort of participants. An even higher significant positive relation (r = .43) could be detected between neuroticism and serum LDL cholesterol within the psychiatric patient group. As far as differences of personality variables between patients and normal controls are concerned, patients seemed to be significantly less satisfied with life, more generally inhibited, more irritable, more stressed, and more emotionally labile than controls. Anxiety disorder patients also tended to complain significantly more often about physical problems. CONCLUSIONS: Our data suggest that high scores of neuroticism in anxiety disorders patients may be involved in elevated serum LDL cholesterol levels and, thereby, may lead to an increased risk of coronary heart disease.     

Cortico-hippocampal APP and NGF levels are dynamically altered by cholinergic muscarinic antagonist or M1 agonist treatment in normal mice

To determine whether altered cholinergic neurotransmission can modify the long-term secretion of amyloid precursor protein (APP), endogenous levels of APP and nerve growth factor (NGF), we administered a selective M1 muscarinic receptor agonist (RS86) or the muscarinic antagonist, atropine, for 7 days in vivo into young adult mice (C57BL/6j). The levels of NGF and total APP in the hippocampus, frontal cortex, striatum, parietal cortex and cerebrospinal fluid (CSF) were examined by ELISA and Western blot. We found that this repeated i.m. administration of M1 receptor agonist resulted in decreased total APP levels in the hippocampus, frontal cortex and parietal cortex, and increased secreted alpha-APPs levels in the CSF. M1 agonist treatment also resulted in decreased NGF levels in the hippocampus and CSF. These effects of the M1 muscarinic agonist could be blocked by atropine, which by itself elevated tissue levels of total APP. Interestingly, we found that the decrease of total APP in the hippocampus and striatum after M1 agonist treatment inversely correlated with the change in NGF levels. These data suggest that a sustained increased cholinergic, M1-mediated neurotransmission will enhance secretion of alpha-APPs in CSF and adaptively reduce the levels of total APP and NGF in the corticohippocampal regions of normal mice. The dynamic and adaptive regulation linking total APP and NGF levels in normal adult mice is relevant for understanding the pathophysiology of conditions with cholinergic and APP related pathologies, like Alzheimer's disease and Down's syndrome.     

Abeta levels in serum,CSF and brain,and cognitive deficits in APP + PS1 transgenic mice

We compared beta-amyloid peptide (Abeta) levels in the serum, CSF and brain (hippocampus) and correlated these with spatial learning in APP+PS1 transgenic mice. Compared with non-transgenic littermates, male 14-month-old APP + PS1 mice were impaired in spatial learning in the water maze. Among the APP + PS1 mice, only the hippocampal insoluble Abeta42 level correlated with spatial memory (r = -0.44). The levels of insoluble Abeta40 and Abeta42 were highly correlated (r = 0.92), and also correlated with soluble hippocampal Abeta42 (r = 0.64/0.69), which further correlated with the CSF Abeta42 (r = 0.52). None of these parameters correlated with serum Abeta40 levels. These findings support the role of insoluble Abeta42 in memory dysfunction and suggest a model with several pools (insoluble, extracellular soluble, CSF) of Abeta being in partial equilibrium with each other.     

Regulation of Neuropeptides in Adult Rat Forebrain by the Neurotrophins BDNF and NGF

The expression of neuropeptides and neurotrophic factors is altered in the hippocampus after seizure induction in rats. Because the increase in brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) mRNAs precede changes in neuropeptide expression after seizure, it is possible that BDNF and NGF mediate subsequent alterations in peptide expression. To test this hypothesis directly, BDNF or NGF was infused into the hippocampus and cortex of adult rats. To ascertain the regional specificity of any observed effects of neurotrophin administration on neuropeptide expression, infusions into the striatum were also studied. To control for specificity, vehicle was also infused into the same sites. Peptide and mRNA alterations were assessed by Northern analysis, immunohistochemistry and radioimmunoassay. BDNF produced elevations of peptide and mRNA for neuropeptide Y and cholecystokinin in hippocampus and cortex, and somatostatin in cortex. BDNF increased mRNAs for neuropeptide Y, cholecystokinin, substance P and dynorphin in striatum. In contrast, BDNF decreased dynorphin peptide and mRNA in hippocampus. NGF's effects were limited to small mRNA increases, without corresponding changes in peptide levels, for neuropeptide Y in hippocampus and striatum, substance P in cortex and cholecystokinin in striatum. The distinct and limited effects of NGF infusion on neuropeptide expression demonstrate that BDNF's effects are not non-specific results of protein infusion into the brain. These findings indicate that BDNF may play a regionally specific role in modulating neuropeptide expression in the normal brain as well as in various pathophysiological states.     

Human olfactory epithelium in normal aging, Alzheimer's disease, and other neurodegenerative disorders.

By use of immunohistochemistry, we characterized the molecular phenotype of human olfactory epithelial (OE) cells and assessed the nature of the dystrophic olfactory neurites described initially in Alzheimer's disease (AD). Keratin 8 was present in all classes of OE cells. Sustentacular cells lacked other cell type specific polypeptides and were distinguished from neurons and basal cells because the latter two classes of OE cells expressed neural cell adhesion molecules (N-CAMs) and microtubule associated proteins (MAPs), i.e., MAP5. Basal cells expressed nerve growth factor receptors (NGFRs), which distinguished them from olfactory neurons. Unlike their perikarya, olfactory axons expressed vimentin and GAP-43, but not peripherin or neurofilament (NF) proteins. Olfactory nerves were distinguished from other axons because the latter were positive for all three NF subunits and peripherin, in addition to vimentin and GAP-43. Dystrophic neurites in the OE were GAP-43 positive, but they also expressed proteins that were not detected in normal olfactory nerves (i.e., synaptophysin, MAP2, tau, peripherin, NF proteins). Further, rare NF positive olfactory neurons gave rise to NF positive dystrophic neurites. These neurites were present in all 11 AD cases, 11 of 14 subjects with other neurodegenerative diseases, and 6 of 8 neurologically normal adult controls, but no dystrophic neurites were seen in 9 fetal and neonatal cases. We conclude that the molecular phenotype of different human OE cells is distinct and that dystrophic olfactory neurites occur very frequently in neurologically normal adults. The relevance of these neurites to aging or specific disease processes remains speculative.     

PCR-mediated search for herpes simplex virus DNA in sections of brain from patients with multiple sclerosis and other neurological disorders.

Herpes simplex virus (HSV) has been shown to cause central nervous system demyelination in experimental animals and several studies have implicated HSV in the aetiology of multiple sclerosis (MS). We have used the polymerase chain reaction to look for DNA of both type 1 HSV (HSV-1) and type 2 HSV (HSV-2) in formalin-fixed paraffin-embedded brain tissues from patients with MS and other neurological diseases. Primers which amplify a fragment of the normal cellular gene c-myc were included in the reactions to assess the preservation of DNA in the tissue samples. 77 plaques of demyelination from 23 patients with MS were examined. HSV-1 DNA was amplified from only one plaque. This plaque involved the trigeminal root entry zone in the pons and it is suggested that the presence of viral DNA was related to the site examined rather than to the demyelination per se. HSV-2 DNA was amplified from none of the plaques. As expected, HSV-1 DNA was detected in the brains of 6 patients who died of HSV-1 encephalitis and HSV-2 DNA was amplified from the brain of a neonate with congenital HSV-2 infection. In sections of brain from 39 patients with a wide range of other neurological diseases HSV-1 DNA was detected in the pons of only 1 patient, who had AIDS associated with cytomegalovirus ventriculitis; subsequent investigation confirmed the presence of concomitant HSV-1 brain stem infection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Maternal infection regulates BDNF and NGF expression in fetal and neonatal brain and maternal-fetal unit of the rat

Maternal infection during pregnancy is associated with increased risk for neurodevelopmental disorders. Lipopolysaccharide (LPS) or saline was administered to rats to model maternal infection, and levels of brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in maternal plasma, placenta, amniotic fluid, fetal liver/spleen, fetal brain, and cerebral cortex after birth were determined by ELISA or semiquantitative Western blot analysis. BDNF expression was significantly increased in the fetal brain (p=0.039); NGF expression was significantly increased in neonatal cortex (p=0.0009). Neurotrophic factor expression was also altered in other tissues of the maternal-fetal unit. Abnormal expression of neurotrophic factors represents a potential mechanism through which maternal infection increases risk for neurodevelopmental disorders.     

Maternal micronutrient imbalance alters gene expression of BDNF,NGF, TrkB and CREB in the offspring brain at an adult age

Micronutrients like folate, vitamin B₁₂, and fatty acids which are interlinked in the one carbon cycle play a vital role in mediating epigenetic processes leading to an increased risk for neurodevelopmental disorders in the offspring. Our earlier study demonstrates that a micronutrient imbalanced diet adversely affects docosahexaenoic acid (DHA) and protein levels of neurotrophins like brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in the brain and cognition in the offspring by 3 months of age. In this study we attempt to analyze if these effects are a consequence of a change in gene expression of these molecules. Further, we also examined the effect of either a postnatal control diet or a prenatal omega-3 fatty acid supplementation on gene expression in the cortex of the offspring. Pregnant rats were divided into control and five treatment groups at two levels of folic acid (normal and excess folate) in the presence and absence of vitamin B₁₂. Omega-3 fatty acid (eicosapentaenoic acid – EPA + DHA) supplementation was given to vitamin B₁₂ deficient groups. Following delivery, 8 dams from each group were shifted to control diet and remaining continued on the same treatment diet. Our results demonstrate that the imbalanced diet caused a marked reduction in the mRNA levels of BDNF, NGF, TrkB, and cAMP response element-binding protein (CREB). Prenatal omega-3 fatty acid supplementation to the maternal imbalanced diet was able to normalize the mRNA levels of all the above genes. This study demonstrates that a maternal diet imbalanced in micronutrients (folic acid, vitamin B₁₂) influences gene expression of neurotrophins and their signalling molecules and thereby adversely affects the brain of the offspring.     

Effects of repeated minimal electroshock seizures on NGF,BDNF and FGF-2 protein in the rat brain during postnatal development

Repeated brief seizures, such as those induced by electroconvulsive therapy (ECT), markedly elevate neurotrophic factor levels in the adult rat brain, but it is not known whether a similar response to seizures occurs in immature animals. To address this question, we evoked brief seizures with electroconvulsive shock (ECS) in rat pups at different stages of postnatal development and examined basic fibroblast growth factor (FGF-2), nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF) proteins in selected brain regions in which these trophic factors are known to increase in the adult rat following ECS-induced seizures.     

Atorvastatin induction of VEGF and BDNF promotes brain plasticity after stroke in mice.

Molecular mechanisms underlying the role of statins in the induction of brain plasticity and subsequent improvement of neurologic outcome after treatment of stroke have not been adequately investigated. Here, we use both in vivo and in vitro studies to investigate the potential roles of two prominent factors, vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF), in mediating brain plasticity after treatment of stroke with atorvastatin. Treatment of stroke in adult mice with atorvastatin daily for 14 days, starting at 24 hours after MCAO, shows significant improvement in functional recovery compared with control animals. Atorvastatin increases VEGF, VEGFR2 and BDNF expression in the ischemic border. Numbers of migrating neurons, developmental neurons and synaptophysin-positive cells as well as indices of angiogenesis were significantly increased in the atorvastatin treatment group, compared with controls. In addition, atorvastatin significantly increased brain subventricular zone (SVZ) explant cell migration in vitro. Anti-BDNF antibody significantly inhibited atorvastatin-induced SVZ explant cell migration, indicating a prominent role for BDNF in progenitor cell migration. Mouse brain endothelial cell culture expression of BDNF and VEGFR2 was significantly increased in atorvastatin-treated cells compared with control cells. Inhibition of VEGFR2 significantly decreased expression of BDNF in brain endothelial cells. These data indicate that atorvastatin promotes angiogenesis, brain plasticity and enhances functional recovery after stroke. In addition, VEGF, VEGFR2 and BDNF likely contribute to these restorative processes.     

Compartmentalized tau hyperphosphorylation and increased levels of kinases in transgenic mice.

The formation of neurofibrillary tangles in Alzheimer's disease is preceded by a pretangle stage of hyperphosphorylated tau. To characterize pretangle tau in vivo, we correlated, in human tau transgenic mice, levels of kinases known to phosphorylate tau in vitro with the phosphorylation of tau at specific epitopes. Levels of cyclin-dependent kinase-5 were increased in axons of CA1 pyramidal neurons, where tau was phosphorylated specifically at the AD2 epitope Ser396/Ser404. The 12E8 epitope serine262/serine356 and the AT180 epitope threonine231/serine235 were phosphorylated in dendrites, and colocalized with increased levels of glycogen synthase kinase-3. CA1 neurons phosphorylated tau at more epitopes than dentate gyrus neurons, suggesting that tau phosphorylation is cell type-specific, a possible explanation for the spatial distribution of neurofibrillary tangles.     

Prolonged prenatal exposure to low-level ozone affects aggressive behaviour as well as NGF and BDNF levels in the central nervous system of CD-1 mice

The long-term effects on isolation-induced aggressive behaviour and central NGF and BDNF levels of gestational exposures to ozone (O(3)) were evaluated in adult CD-1 mice. Females were exposed to O(3), at the dose of 0.0, 0.3 or 0.6 ppm from 30 days prior the formation of breeding pairs until gestational day 17. Litters were fostered at birth to untreated dams and, at adulthood, male offspring underwent five successive daily encounters (15 min each) with a standard opponent of the same strain, sex, weight and age. The encounters on day 1, 3 and 5 were videotaped and agonistic and non-agonistic behavioural items finely scored. O(3)-exposed mice showed a significant increase in freezing and defensive postures, a decrease in nose-sniffing behaviour and reduced progressively the aggressive behavioural profile displayed on day 1. Reduced NGF levels in the hippocampus and increased BDNF in the striatum were also found upon O(3) exposure.     

Differential effects of lovastatin treatment on brain cholesterol levels in normal and apoE-deficient mice

Growing evidence indicates that membrane cholesterol is involved in the development of Alzheimer's disease. Therefore, the availability of pharmacological strategies to modify brain cholesterol is of increasing importance. Accordingly, we investigated the effects of the HMG-CoA reductase inhibitor lovastatin on brain cholesterol levels in vivo. Brain cholesterol was significantly decreased by lovastatin treatment (100 mg/kg/day) in 1- and 12-month-old C57BL/6J mice. Reduced brain cholesterol was associated with decreased pyrene-excimer fluorescence, indicating altered membrane function. Lovastatin had no effect on brain cholesterol ApoE-/- mice. Peripheral cholesterol levels were not affected by lovastatin in all three groups of mice. We demonstrate for the first time that lovastatin represents a valid pharmacological tool to significantly modulate brain cholesterol levels.