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The accumulation of amyloid-β (Aβ) is a characteristic event in the pathogenesis of Alzheimer’s disease (AD). Aquaporin 1 (AQP1) is a membrane water channel protein belonging to the AQP family. AQP1 levels are elevated in the cerebral cortex during the early stages of AD, but the role of AQP1 in AD pathogenesis is unclear. We first determined the expression and distribution of AQP1 in brain tissue samples of AD patients and two AD mouse models (3xTg-AD and 5xFAD). AQP1 accumulation was observed in vulnerable neurons in the cerebral cortex of AD patients, and in neurons affected by the Aβ or tau pathology in the 3xTg-AD and 5xFAD mice. AQP1 levels increased in neurons as aging progressed in the AD mouse models. Stress stimuli increased AQP1 in primary cortical neurons. In response to cellular stress, AQP1 appeared to translocate to endocytic compartments of β- and γ-secretase activities. Ectopic expression of AQP1 in human neuroblastoma cells overexpressing amyloid precussir protein (APP) with the Swedish mutations reduced β-secretase (BACE1)-mediated cleavage of APP and reduced Aβ production without altering the nonamyloidogenic pathway. Conversely, knockdown of AQP1 enhanced BACE1 activity and Aβ production. Immunoprecipitation experiments showed that AQP1 decreased the association of BACE1 with APP. Analysis of a human database showed that the amount of Aβ decreases as the expression of AQP1 increases. These results suggest that the upregulation of AQP1 is an adaptive response of neurons to stress that reduces Aβ production by inhibiting the binding between BACE1 and APP.

Multiple sclerosis (MS) is an inflammatory autoimmune disease of the central nervous system (CNS) involving demyelinating and neurodegenerative processes. Several of the major pathological CNS alterations and behavioral deficits of MS are recapitulated in the experimental autoimmune encephalitis (EAE) mouse model in which the disease process is induced by administration of myelin peptides. Development of EAE requires infiltration of inflammatory cytokine-generating monocytes and macrophages, and auto-reactive T cells, into the CNS. Very late antigen-4 (VLA-4, α4β1) is an integrin molecule that plays a role in inflammatory responses by facilitating the migration of leukocytes across the blood-brain barrier during inflammatory disease, and antibodies against VLA-4 exhibit therapeutic efficacy in mouse and monkey MS models. Here, we report that the tellurium compound AS101 (ammonium trichloro (dioxoethylene-o,o') tellurate) ameliorates EAE by inhibiting monocyte and T cell infiltration into the CNS. CD49d is an alpha subunit of the VLA-4 (α4β1) integrin. During the peak stage of EAE, AS101 treatment effectively ameliorated the disease process by reducing the number of CD49d(+) inflammatory monocyte/macrophage cells in the spinal cord. AS101 treatment markedly reduced the pro-inflammatory cytokine levels, while increasing anti-inflammatory cytokine levels. In contrast, AS101 treatment did not affect the peripheral populations of CD11b(+) monocytes and macrophages. AS101 treatment reduced the infiltration of CD4(+) and CD49(+)/VLA4 T cells. In addition, treatment of T cells from MS patients with AS101 resulted in apoptosis, while such treatment did not affect T cells from healthy donors. These results suggest that AS101 reduces accumulation of leukocytes in the CNS by inhibiting the activity of the VLA-4 integrin and provide a rationale for the potential use of Tellurium IV compounds for the treatment of MS.

Age-associated dysregulation of sleep can be worsened by Alzheimer’s disease (AD). AD and sleep restriction both impair cognition, yet it is unknown if mild chronic sleep restriction modifies the proteopathic processes involved in AD. The goal of this work was to test the hypothesis that sleep restriction worsens memory impairments, and amyloid β-peptide (Aβ) and pTau accumulations in the brain in a mouse model of AD, with a focus on a role for circulating glucocorticoids (GC). Male 3xTgAD mice were subjected to sleep restriction (SR) for 6 hrs/day for 6 weeks using the modified multiple platform technique, and behavioral (Morris water maze, fear conditioning, open field) and biochemical (immunoblot) outcomes were compared to mice undergoing daily cage transfers (large cage control; LCC) as well as control mice that remained in their home cage (control; CTL). At one week, both LCC and SR mice displayed significant elevations in plasma corticosterone compared to CTL (p

Rapid and effective wound healing requires a coordinated cellular response involving fibroblasts, keratinocytes and vascular endothelial cells (VECs). Impaired wound healing can result in multiple adverse health outcomes and, although antibiotics can forestall infection, treatments that accelerate wound healing are lacking. We now report that topical application of water soluble cerium oxide nanoparticles (Nanoceria) accelerates the healing of full-thickness dermal wounds in mice by a mechanism that involves enhancement of the proliferation and migration of fibroblasts, keratinocytes and VECs. The Nanoceria penetrated into the wound tissue and reduced oxidative damage to cellular membranes and proteins, suggesting a therapeutic potential for topical treatment of wounds with antioxidant nanoparticles.

Tumor necrosis factor-α (TNF) plays a prominent role in the brain damage and functional deficits that result from ischemic stroke. It was recently reported that the thalidomide analog 3,6'-dithiothalidomide (3,6'-DT) can selectively inhibit the synthesis of TNF in cultured cells. We therefore tested the therapeutic potential of 3,6'-DT in a mouse model of focal ischemic stroke. Administration of 3,6'-DT immediately prior to a stroke or within 3 hr after the stroke reduced infarct volume, neuronal death, and neurological deficits, whereas thalidomide was effective only when administered prior to stroke. Neuroprotection was accompanied by decreased inflammation; 3,6'-DT-treated mice exhibited reduced expression of TNF, interleukin-1β, and inducible nitric oxide synthase; reduced numbers of activated microglia/macrophages, astrocytes, and neutrophils; and reduced expression of intercellular adhesion molecule-1 in the ischemic brain tissue. 3,6'-DT treatment attenuated stroke-induced disruption of the blood-brain barrier by a mechanism that appears to involve suppression of matrix metalloproteinase-9 and preservation of occludin. Treatment with 3,6'-DT did not reduce ischemic brain damage in mice lacking TNF receptors, consistent with a critical role for suppression of TNF production and TNF signaling in the therapeutic action of 3,6'-DT. These findings suggest that anti-inflammatory mechanisms underlie the therapeutic actions of 3,6-DT in an animal model of stroke.

Alzheimer's disease (AD) involves progressive accumulation of amyloid β-peptide (Aβ) and neurofibrillary pathologies, and glucose hypometabolism in brain regions critical for memory. The 3xTgAD mouse model was used to test the hypothesis that a ketone ester-based diet can ameliorate AD pathogenesis. Beginning at a presymptomatic age, 2 groups of male 3xTgAD mice were fed a diet containing a physiological enantiomeric precursor of ketone bodies (KET) or an isocaloric carbohydrate diet. The results of behavioral tests performed at 4 and 7 months after diet initiation revealed that KET-fed mice exhibited significantly less anxiety in 2 different tests. 3xTgAD mice on the KET diet also exhibited significant, albeit relatively subtle, improvements in performance on learning and memory tests. Immunohistochemical analyses revealed that KET-fed mice exhibited decreased Aβ deposition in the subiculum, CA1 and CA3 regions of the hippocampus, and the amygdala. KET-fed mice exhibited reduced levels of hyperphosphorylated tau deposition in the same regions of the hippocampus, amygdala, and cortex. Thus, a novel ketone ester can ameliorate proteopathic and behavioral deficits in a mouse AD model. © 2013 .

Endoplasmic reticulum (ER) stress has been implicated as an initiator or contributing factor in neurodegenerative diseases. The mechanisms that lead to ER stress and whereby ER stress contributes to the degenerative cascades remain unclear but their understanding is critical to devising effective therapies. Here we show that knockdown of Herp (Homocysteine-inducible ER stress protein), an ER stress-inducible protein with an ubiquitin-like (UBL) domain, aggravates ER stress-mediated cell death induced by mutant α-synuclein (αSyn) that causes an inherited form of Parkinson's disease (PD). Functionally, Herp plays a role in maintaining ER homeostasis by facilitating proteasome-mediated degradation of ER-resident Ca(2+) release channels. Deletion of the UBL domain or pharmacological inhibition of proteasomes abolishes the Herp-mediated stabilization of ER Ca(2+) homeostasis. Furthermore, knockdown or pharmacological inhibition of ER Ca(2+) release channels ameliorates ER stress, suggesting that impaired homeostatic regulation of Ca(2+) channels promotes a protracted ER stress with the consequent activation of ER stress-associated apoptotic pathways. Interestingly, sustained upregulation of ER stress markers and aberrant accumulation of ER Ca(2+) release channels were detected in transgenic mutant A53T-αSyn mice. Collectively, these data establish a causative link between impaired ER Ca(2+) homeostasis and chronic ER stress in the degenerative cascades induced by mutant αSyn and suggest that Herp is essential for the resolution of ER stress through maintenance of ER Ca(2+) homeostasis. Our findings suggest a therapeutic potential in PD for agents that increase Herp levels or its ER Ca(2+)-stabilizing action.

The cell fate determinant Numb exists in four alternatively spliced variants that differ in the length of their PTB (phosphotyrosine-binding domain, either lacking or containing an 11 amino acid insertion) and PRR (proline-rich region, either lacking or containing a 48 amino acid insertion). We previously reported that Numb switches from isoforms containing the PTB insertion to isoforms lacking this insertion in neural cultures subjected to stress induced by trophic factor withdrawal. The switch in Numb isoforms enhances the generation of amyloid-β peptide (Aβ), the principle component of senile plaques in Alzheimer's disease (AD). Here we examine the expression of the Numb isoforms in brains from AD patients and triple transgenic (3xTg) AD mice. We found that levels of the Numb isoforms lacking the PTB insertion are significantly elevated in the parietal cortex but not in the cerebellum of AD patients when compared to control subjects. Levels of Numb isoforms lacking the PTB insertion were also elevated in the cortex but not cerebellum of 12 month-old 3xTg AD mice with Aβ deposits compared to younger 3xTg-AD mice and to non-transgenic mice. Exposure of cultured neurons to Aβ resulted in an increase in the levels of Numb isoforms lacking the PTB domain, consistent with a role for Aβ in the aberrant expression of Numb in vulnerable brain regions of AD patients and mice. Collectively, the data show that altered expression of Numb isoforms in vulnerable neurons occurs during AD pathogenesis and suggest a role for Numb in the disease process.

Huntington's disease (HD) is an inherited neurodegenerative disorder caused by expanded polyglutamine repeats in the huntingtin (Htt) protein. Mutant Htt may damage and kill striatal neurons by a mechanism involving reduced production of brain-derived neurotrophic factor (BDNF) and increased oxidative and metabolic stress. Because electroconvulsive shock (ECS) can stimulate the production of BDNF and protect neurons against stress, we determined whether ECS treatment would modify the disease process and provide a therapeutic benefit in a mouse model of HD. ECS (50 mA for 0.2 s) or sham treatment was administered once weekly to male N171-82Q Htt mutant mice beginning at 2 months of age. Endpoints measured included motor function, striatal and cortical pathology, and levels of protein chaperones and BDNF. ECS treatment delayed the onset of motor symptoms and body weight loss and extended the survival of HD mice. Striatal neurodegeneration was attenuated and levels of protein chaperones (Hsp70 and Hsp40) and BDNF were elevated in striatal neurons of ECS-treated compared with sham-treated HD mice. Our findings demonstrate that ECS can increase the resistance of neurons to mutant Htt resulting in improved functional outcome and extended survival. The potential of ECS as an intervention in subjects that inherit the mutant Htt gene merits further consideration.

Engineered tissue strategies for central nervous system (CNS) repair have the potential for localizing treatment using a wide variety of cells or growth factors. However, these strategies are often limited by their ability to address only one aspect of the injury. Here we report the development of a novel alginate construct that acts as a multi-functional tissue scaffold for CNS repair, and as a localized growth factor delivery vehicle. We show that the surface of this alginate construct acts as an optimal growth environment for neural progenitor cell (NPC) attachment, survival, migration, and differentiation. Importantly, we show that tailor-made alginate constructs containing brain-derived neurotrophic factor or neurotrophin-3 differentially direct lineage fates of NPCs and may therefore be useful in treating a wide variety of injuries. It is this potential for directed differentiation of a scaffold prior to implantation at the injury site that we explore here.

Compromised cellular energy metabolism, cerebral hypoperfusion, and neuronal calcium dysregulation are involved in the pathological process of Alzheimer's disease (AD). ATP-sensitive potassium (KATP) channels in plasma membrane and inner mitochondrial membrane play important roles in modulating neuronal excitability, cell survival, and cerebral vascular tone. To investigate the therapeutic potential of drugs that activate KATP channels in AD, we first characterized the effects of the KATP channel opener diazoxide on cultured neurons, and then determined its ability to modify the disease process in the 3xTgAD mouse model of AD. Plasma and mitochondrial membrane potentials, cell excitability, intracellular Ca2+ levels and bioenergetics were measured in cultured cerebral cortical neurons exposed to diazoxide. Diazoxide hyperpolarized neurons, reduced the frequency of action potentials, attenuated Ca2+ influx through NMDA receptor channels, and reduced oxidative stress. 3xTgAD mice treated with diazoxide for 8 months exhibited improved performance in a learning and memory test, reduced levels of anxiety, decreased accumulation of Aβ oligomers and hyperphosphorylated tau in the cortex and hippocampus, and increased cerebral blood flow. Our findings show that diazoxide can ameliorate molecular, cytopathological, and behavioral alterations in a mouse model of AD suggesting a therapeutic potential for drugs that activate KATP channels in the treatment of AD.

Beta-secretase (BACE1), an enzyme responsible for the production of amyloid beta-peptide (Abeta), is increased by oxidative stress and is elevated in the brains of patients with sporadic Alzheimer's disease (AD). Here, we show that oxidative stress fails to induce BACE1 expression in presenilin-1 (gamma-secretase)-deficient cells and in normal cells treated with gamma-secretase inhibitors. Oxidative stress-induced beta-secretase activity and sAPPbeta levels were suppressed by gamma-secretase inhibitors. Levels of gamma- and beta-secretase activities were greater in brain tissue samples from AD patients compared to non-demented control subjects, and the elevated BACE1 level in the brains of 3xTgAD mice was reduced by treatment with a gamma-secretase inhibitor. Our findings suggest that gamma-secretase mediates oxidative stress-induced expression of BACE1 resulting in excessive Abeta production in AD.

Many phytochemicals function as noxious agents that protect plants against insects and other damaging organisms. However, at subtoxic doses, the same phytochemicals may activate adaptive cellular stress response pathways that can protect cells against a variety of adverse conditions. We screened a panel of botanical pesticides using cultured human and rodent neuronal cell models, and identified plumbagin as a novel potent activator of the nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway. In vitro, plumbagin increases nuclear localization and transcriptional activity of Nrf2, and induces the expression of the Nrf2/ARE-dependent genes, such as heme oxygenase 1 in human neuroblastoma cells. Plumbagin specifically activates the Nrf2/ARE pathway in primary mixed cultures from ARE-human placental alkaline phosphatase reporter mice. Exposure of neuroblastoma cells and primary cortical neurons to plumbagin provides protection against subsequent oxidative and metabolic insults. The neuroprotective effects of plumbagin are abolished by RNA interference-mediated knockdown of Nrf2 expression. In vivo, administration of plumbagin significantly reduces the amount of brain damage and ameliorates-associated neurological deficits in a mouse model of focal ischemic stroke. Our findings establish precedence for the identification and characterization of neuroprotective phytochemicals based upon their ability to activate adaptive cellular stress response pathways.

beta1 integrin is a cell surface molecule that is critical for endothelial cell adhesion, migration and survival during angiogenesis. In the present study we employed in vivo and in vitro models to elucidate the role of beta1 integrin in vascular remodelling and stroke outcomes. At 24 h after cerebral ischemia and reperfusion (I/R), the ischemic cortex (ipsilateral area) exhibited modest beta1 integrin immunoreactivity and a robust increase was observed at 72 h. Double-label immunohistochemical analysis for beta1 integrin with neuronal (NeuN), microglial (Iba-1), astrocyte (GFAP), progenitor cell (Ng2) and blood vessel (collagen 4) markers showed that beta1 integrin expression only localized to blood vessels. In vitro studies using cultured endothelial cells and a beta1 integrin blocking antibody confirmed that beta1 integrin is required for endothelial cell migration, proliferation and blood vessel formation. In vivo studies in the cerebral I/R model using the beta1 integrin blocking antibody further confirmed that beta1 integrin signaling is involved in vascular formation and recovery following ischemic stroke. Finally, we found that beta1 integrin is critically involved in functional deficits and survival after a stroke. These results suggest that beta1 integrin plays important roles in neurovascular remodelling and functional outcomes following stroke, and that targeting the beta1 integrin signalling may provide a novel strategy for modulating angiogenesis in ischemic stroke and other pathological conditions.

Mutations of the presenilin 1 (PS1) gene are the most common cause of early onset familial Alzheimer disease (FAD). PS1 mutations alter the activity of the gamma-secretase on the beta-amyloid precursor protein (APP), leading to selective overproduction of beta-amyloid (Abeta) 42 peptides, the species that forms oligomers that may exert toxic effects on neurons. Here we show that PS1 mutations, expressed both transiently and stably, in non-neuronal and neuronal cell lines increase the expression and the activity of the beta-secretase (BACE1), the rate-limiting step of Abeta production. Also, BACE1 expression and activity are elevated in brains of PS1 mutant knock-in mice compared with wild type littermates as well as in cerebral cortex of FAD cases bearing various PS1 mutations compared with in sporadic AD cases and controls. The up-regulation of BACE1 by PS1 mutations requires the gamma-secretase cleavage of APP and is proportional to the amount of secreted Abeta42. Abeta42, and not AICD (APP intracellular domain), is indeed the APP derivative that mediates the overexpression of BACE1. The effect of PS1 mutations on BACE1 may contribute to determine the wide clinical and pathological phenotype of early onset FAD.

Emerging data suggest that, much like epithelial cells, the polarized growth of neurons requires both the secretory and endocytic pathways. The clathrin assembly proteins AP180 and CALM (clathrin assembly lymphoid myeloid protein) are known to be involved in clathrin-mediated endocytosis, but their roles in mammalian neurons and, in particular, in developmental processes before synaptogenesis are unknown. Here we provide evidence that AP180 and CALM play critical roles in establishing the polarity and controlling the growth of axons and dendrites in embryonic hippocampal neurons. Knockdown of AP180 primarily impairs axonal development, whereas reducing CALM levels results in dendritic dystrophy. Conversely, neurons that overexpress AP180 or CALM generate multiple axons. Ultrastructural analysis shows that CALM affiliates with a wider range of intracellular trafficking organelles than does AP180. Functional analysis shows that endocytosis is reduced in both AP180-deficient and CALM-deficient neurons. Additionally, CALM-deficient neurons show disrupted secretory transport. Our data demonstrate previously unknown functions for AP180 and CALM in intracellular trafficking that are essential in the growth of neurons.

The innate immune system senses the invasion of pathogenic microorganisms and tissue injury through Toll-like receptors (TLR), a mechanism thought to be limited to immune cells. We recently found that neurons express several TLRs, and that the levels of TLR2 and TLR4 are increased in neurons in response to energy deprivation. Here we report that TLR4 expression increases in neurons when exposed to amyloid beta-peptide (Abeta1-42) or the lipid peroxidation product 4-hydroxynonenal (HNE). Neuronal apoptosis triggered by Abeta and HNE was mediated by jun N-terminal kinase (JNK); neurons from TLR4 mutant mice exhibited reduced JNK and caspase-3 activation and were protected against apoptosis induced by Abeta and HNE. Levels of TLR4 were decreased in inferior parietal cortex tissue specimens from end-stage AD patients compared to aged-matched control subjects, possibly as the result of loss of neurons expressing TLR4. Our findings suggest that TLR4 signaling increases the vulnerability of neurons to Abeta and oxidative stress in AD, and identify TLR4 as a potential therapeutic target for AD.

Stroke is among the three leading causes of death worldwide and the most frequent cause of permanent disability. Brain ischemia induces an inflammatory response involving activated complement fragments. Here we show that i.v. Ig (IVIG) treatment, which scavenges complement fragments, protects brain cells against the deleterious effects of experimental ischemia and reperfusion (I/R) and prevents I/R-induced mortality in mice. Animals administered IVIG either 30 min before ischemia or after 3 h of reperfusion exhibited a 50–60% reduction of brain infarct size and a 2- to 3-fold improvement of the functional outcome. Even a single low dose of IVIG given after stroke was effective. IVIG was protective in the nonreperfusion model of murine stroke as well and did not exert any peripheral effects. Human IgG as well as intrinsic murine C3 levels were significantly higher in the infarcted brain region compared with the noninjured side, and their physical association was demonstrated by immuno-coprecipitation. C5-deficient mice were significantly protected from I/R injury compared with their wild-type littermates. Exposure of cultured neurons to oxygen/glucose deprivation resulted in increased levels of C3 associated with activation of caspase 3, a marker of apoptosis; both signals were attenuated with IVIG treatment. Our data suggest a major role for complement-mediated cell death in ischemic brain injury and the prospect of using IVIG in relatively low doses as an interventional therapy for stroke.

The mechanisms by which neural and glial progenitor cells in the adult brain respond to tissue injury are unknown. We studied the responses of these cells to stab wound injury in rats and in two transgenic mouse models in which Y/GFP is driven either by Sox2 (a neural stem cell marker) or by Talpha-1 (which marks newly born neurons). The response of neural progenitors was low in all nonneurogenic regions, and no neurogenesis occurred at the injury site. Glial progenitors expressing Olig2 and NG2 showed the greatest response. The appearance of these progenitors preceded the appearance of reactive astrocytes. Surprisingly, we found evidence of the translocation of the transcription factor Olig2 into cytoplasm in the first week after injury, a mechanism that is known to mediate the differentiation of astrocytes during brain development. Translocation of Olig2, down-regulation of NG2, and increased glial fibrillary acidic protein expression were recapitulated in vitro after exposure of glial progenitors to serum components or bone morphogentic protein by up-regulation of Notch-1. The glial differentiation and Olig2 translocation could be blocked by inhibition of Notch-1 with the gamma-secretase inhibitor DAPT. Together, these data indicate that the prompt maturation of numerous Olig2(+) glial progenitors to astrocytes underlies the repair process after a traumatic injury. In contrast, neural stem cells and neuronal progenitor cells appear to play only a minor role in the injured adult CNS.

Neuroinflammation in the central nervous system is detrimental for learning and memory, as evident form epidemiological studies linking developmental defects and maternal exposure to harmful pathogens. Postnatal infections can also induce neuroinflammatory responses with long-term consequences. These inflammatory responses can lead to motor deficits and/or behavioral disabilities. Toll like receptors (TLRs) are a family of innate immune receptors best known as sensors of microbial-associated molecular patterns, and are the first responders to infection. TLR2 forms heterodimers with either TLR1 or TLR6, is activated in response to gram-positive bacterial infections, and is expressed in the brain during embryonic development. We hypothesized that early postnatal TLR2-mediated neuroinflammation would adversely affect cognitive behavior in the adult. Our data indicate that postnatal TLR2 activation affects learning and memory in adult mice in a heterodimer-dependent manner. TLR2/6 activation improved motor function and fear learning, while TLR2/1 activation impaired spatial learning and enhanced fear learning. Moreover, developmental TLR2 deficiency significantly impairs spatial learning and enhances fear learning, stressing the involvement of the TLR2 pathway in learning and memory. Analysis of the transcriptional effects of TLR2 activation reveals both common and unique transcriptional programs following heterodimer-specific TLR2 activation. These results imply that adult cognitive behavior could be influenced in part, by activation or alterations in the TLR2 pathway at birth.

Tomosyn, a syntaxin-binding protein, is known to inhibit vesicle priming and synaptic transmission via interference with the formation of SNARE complexes. Using a lentiviral vector, we specifically overexpressed tomosyn1 in hippocampal dentate gyrus neurons in adult mice. Mice were then subjected to spatial learning and memory tasks and electrophysiological measurements from hippocampal slices. Tomosyn1-overexpression significantly impaired hippocampus-dependent spatial memory while tested in the Morris water maze. Further, tomosyn1-overexpressing mice utilize swimming strategies of lesser cognitive ability in the Morris water maze compared with control mice. Electrophysiological measurements at mossy fiber-CA3 synapses revealed impaired paired-pulse facilitation in the mossy fiber of tomosyn1-overexpressing mice. This study provides evidence for novel roles for tomosyn1 in hippocampus-dependent spatial learning and memory, potentially via decreased synaptic transmission in mossy fiber-CA3 synapses. Moreover, it provides new insight regarding the role of the hippocampal dentate gyrus and mossy fiber-CA3 synapses in swimming strategy preference, and in learning and memory.

Toll-like receptors (TLRs) play essential roles in innate immunity and increasing evidence indicates that these receptors are expressed in neurons, astrocytes and microglia in the brain where they mediate responses to infection, stress and injury. Very little is known about the roles of TLRs in cognition. To test the hypothesis that TLR4 has a role in hippocampus-dependent spatial learning and memory, we used mice deficient for TLR4 and mice receiving chronic TLR4 antagonist infusion to the lateral ventricles in the brain. We found that developmental TLR4 deficiency enhances spatial reference memory acquisition and memory retention, impairs contextual fear-learning and enhances motor functions, traits that were correlated with CREB up-regulation in the hippocampus. TLR4 antagonist infusion into the cerebral ventricles of adult mice did not affect cognitive behavior, but instead affected anxiety responses. Our findings indicate a developmental role for TLR4 in shaping spatial reference memory, and fear learning and memory. Moreover, we show that central TLR4 inhibition using a TLR4 antagonist has no discernible physiological role in regulating spatial and contextual hippocampus-dependent cognitive behavior.

Preclinical evaluation of drugs for neurological disorders is usually performed on overfed rodents, without consideration of how metabolic state might affect drug efficacy. Using a widely employed mouse model of focal ischemic stroke, we found that that the NMDA receptor antagonist dizocilpine (MK-801) reduces brain damage and improves functional outcome in mice on the usual ad libitum diet, but exhibits little or no therapeutic efficacy in mice maintained on an energy-restricted diet. Thus, NMDA receptor activation plays a central role in the mechanism by which a high dietary energy intake exacerbates ischemic brain injury. These findings suggest that inclusion of subjects with a wide range of energy intakes in clinical trials for stroke may mask a drug benefit in the overfed/obese subpopulation of subjects.

Pregabalin, a Ca(2+) channel α(2)δ-subunit antagonist with analgesic and antiepileptic activity, reduced neuronal loss and improved functional outcome in a mouse model of focal ischemic stroke. Pregabalin administration (5-10mg/kg, i.p.) 30-90 min after transient middle cerebral artery occlusion/reperfusion reduced infarct volume, neuronal death in the ischemic penumbra and neurological deficits at 24h post-stroke. Pregabalin significantly decreased the amount of Ca(2+)/calpain-mediated α-spectrin proteolysis in the cerebral cortex measured at 6h post-stroke. Together with the extensive clinical experience with pregabalin for other neurological indications, our findings suggest the potential for a therapeutic benefit of pregabalin in stroke patients.

Aims Glucagon-like peptide 1 (GLP-1) is an incretin hormone released from the gut in response to food intake. Whereas GLP-1 acts in the periphery to inhibit glucagon secretion and stimulate insulin release, it also acts in the central nervous system to mediate autonomic control of feeding, body temperature, and cardiovascular function. Because of its role as an incretin hormone, GLP-1 receptor analogs are used as a treatment for type 2 diabetes. Central or peripheral administration of GLP-1 increases blood pressure and heart rate, possibly by activating brainstem autonomic nuclei and increasing vagus nerve activity. However, the mechanism(s) by which GLP-1 receptor stimulation affects cardiovascular function are unknown. We used the long-lasting GLP-1 receptor agonist Exendin-4 (Ex-4) to test the hypothesis that GLP-1 signalling modulates central parasympathetic control of heart rate. Methods and results Using a telemetry system, we assessed heart rate in mice during central Ex-4 administration. Heart rate was increased by both acute and chronic central Ex-4 administration. Spectral analysis indicated that the high frequency and low frequency powers of heart rate variability were diminished by Ex-4 treatment. Finally, Ex-4 decreased both excitatory glutamatergic and inhibitory glycinergic neurotransmission to preganglionic parasympathetic cardiac vagal neurons. Conclusion These data suggest that central GLP-1 receptor stimulation diminishes parasympathetic modulation of the heart thereby increasing heart rate.

Toll-like receptors (TLRs) are innate immune receptors that have recently emerged as regulators of neuronal survival and developmental neuroplasticity. Adult TLR3-deficient mice exhibited enhanced hippocampus-dependent working memory in the Morris water maze, novel object recognition, and contextual fear-conditioning tasks. In contrast, TLR3-deficient mice demonstrated impaired amygdala-related behavior and anxiety in the cued fear-conditioning, open field, and elevated plus maze tasks. Further, TLR3-deficient mice exhibited increased hippocampal CA1 and dentate gyrus volumes, increased hippocampal neurogenesis, and elevated levels of the AMPA receptor subunit GluR1 in the CA1 region of the hippocampus. In addition, levels of activated forms of the kinase ERK and the transcription factor CREB were elevated in the hippocampus of TLR3-deficient mice, suggesting that constitutive TLR3 signaling negatively regulates pathways known to play important roles in hippocampal plasticity. Direct activation of TLR3 by intracerebroventricular infusion of a TLR3 ligand impaired working memory, but not reference memory. Our findings reveal previously undescribed roles for TLR3 as a suppressor of hippocampal cellular plasticity and memory retention.

Clathrin-coated vesicles are known to play diverse and pivotal roles in cells. The proper formation of clathrin-coated vesicles is dependent on, and highly regulated by, a large number of clathrin assembly proteins. These assembly proteins likely determine the functional specificity of clathrin-coated vesicles, and together they control a multitude of intracellular trafficking pathways—including those involved in embryonic development. In this study, we focus on two closely related clathrin assembly proteins, AP180 and CALM (clathrin assembly lymphoid myeloid leukemia protein), in the developing embryonic rat brain. We find that AP180 begins to be expressed at embryonic day 14 (E14), but only in postmitotic cells that have acquired a neuronal fate. CALM, on the other hand, is expressed as early as E12, by both neural stem cells and postmitotic neurons. In vitro loss-of-function studies using RNA interference (RNAi) indicate that AP180 and CALM are dispensable for some aspects of embryonic neurogenesis but are required for the growth of postmitotic neurons. These results identify the developmental staging of AP180 and CALM expression and suggest that each protein has distinct functions in neural development.

The subependymal zone (SEZ) of the lateral ventricles is one of the areas of the adult brain where new neurons are continuously generated from neural stem cells (NSCs), via rapidly dividing precursors. This neurogenic niche is a complex cellular and extracellular microenvironment, highly vascularized compared to non-neurogenic periventricular areas, within which NSCs and precursors exhibit distinct behavior. Here, we investigate the possible mechanisms by which extracellular matrix molecules and their receptors might regulate this differential behavior. We show that NSCs and precursors proceed through mitosis in the same domains within the SEZ of adult male mice-albeit with NSCs nearer ependymal cells-and that distance from the ventricle is a stronger limiting factor for neurogenic activity than distance from blood vessels. Furthermore, we show that NSCs and precursors are embedded in a laminin-rich extracellular matrix, to which they can both contribute. Importantly, they express differential levels of extracellular matrix receptors, withNSCsexpressing low levels of alpha 6 beta 1 integrin, syndecan-1, and lutheran, and in vivo blocking of beta 1 integrin selectively induced the proliferation and ectopic migration of precursors. Finally, when NSCs are activated to reconstitute the niche after depletion of precursors, expression of laminin receptors is upregulated. These results indicate that the distinct behavior of adult NSCs and precursors is not necessarily regulated via exposure to differential extracellular signals, but rather via intrinsic regulation of their interaction with their microenvironment.

Activity-dependent modulation of neuronal gene expression promotes neuronal survival and plasticity, and neuronal network activity is perturbed in aging and Alzheimer's disease (AD). Here we show that cerebral cortical neurons respond to chronic suppression of excitability by downregulating the expression of genes and their encoded proteins involved in inhibitory transmission (GABAergic and somatostatin) and Ca(2+) signaling; alterations in pathways involved in lipid metabolism and energy management are also features of silenced neuronal networks. A molecular fingerprint strikingly similar to that of diminished network activity occurs in the human brain during aging and in AD, and opposite changes occur in response to activation of N-methyl-D-aspartate (NMDA) and brain-derived neurotrophic factor (BDNF) receptors in cultured cortical neurons and in mice in response to an enriched environment or electroconvulsive shock. Our findings suggest that reduced inhibitory neurotransmission during aging and in AD may be the result of compensatory responses that, paradoxically, render the neurons vulnerable to Ca(2+)-mediated degeneration.

Toll-like receptors (TLRs) play essential roles in innate immunity, and increasing evidence indicates that these receptors are expressed in neurons, astrocytes, and microglia in the brain, where they mediate responses to infection, stress, and injury. To address the possibility that TLR2 heterodimer activation could affect progenitor cells in the developing brain, we analyzed the expression of TLR2 throughout mouse cortical development, and assessed the role of TLR2 heterodimer activation in neuronal progenitor cell (NPC) proliferation. TLR2 mRNA and protein was expressed in the cortex in embryonic and early postnatal stages of development, and in cultured cortical NPC. While NPC from TLR2-deficient and wild type embryos had the same proliferative capacity, TLR2 activation by the synthetic bacterial lipopeptides Pam(3)CSK(4) and FSL1, or low molecular weight hyaluronan, an endogenous ligand for TLR2, inhibited neurosphere formation in vitro. Intracerebral in utero administration of TLR2 ligands resulted in ventricular dysgenesis characterized by increased ventricle size, reduced proliferative area around the ventricles, increased cell density, an increase in phospho-histone 3 cells, and a decrease in BrdU(+) cells in the sub-ventricular zone. Our findings indicate that loss of TLR2 does not result in defects in cerebral development. However, TLR2 is expressed and functional in the developing telencephalon from early embryonic stages and infectious agent-related activation of TLR2 inhibits NPC proliferation. TLR2-mediated inhibition of NPC proliferation may therefore be a mechanism by which infection, ischemia, and inflammation adversely affect brain development.

Because the olfactory system plays a major role in food consumption, and because “food addiction” and associated morbidities have reached epidemic proportions, we tested the hypothesis that dietary energy restriction can modify adverse effects of cocaine on behavior and olfactory cellular and molecular plasticity. Mice maintained on an alternate day fasting (ADF) diet exhibited increased baseline locomotion and increased cocaine-sensitized locomotion during cocaine conditioning, despite no change in cocaine conditioned place preference, compared to mice fed ad libitum. Levels of dopamine and its metabolites in the olfactory bulb (OB) were suppressed in mice on the ADF diet compared to mice on the control diet, independent of acute or chronic cocaine treatment. The expression of several enzymes involved in dopamine metabolism including tyrosine hydroxylase, monoamine oxidases A and B (MAOA), and catechol-O-methyltransferase were significantly reduced in OBs of mice on the ADF diet. Both acute and chronic administration of cocaine suppressed the production of new OB cells, and this effect of cocaine was attenuated in mice on the ADF diet. Cocaine administration to mice on the control diet resulted in up-regulation of OB genes involved in mitochondrial energy metabolism, synaptic plasticity, cellular stress responses, and calcium- and cyclic AMP-mediated signaling, whereas multiple olfactory receptor genes were down-regulated by cocaine treatment. ADF abolished many of the effects of cocaine on OB gene expression. Our findings reveal that dietary energy intake modifies the neural substrates underlying some of the behavioral and physiological responses to repeated cocaine treatment, and also suggest novel roles for the olfactory system in addiction. The data further suggest that modification of dietary energy intake could provide a novel potential approach to addiction treatments.

Wound healing is a complex process involving intrinsic dermal and epidermal cells, and infiltrating macrophages and leukocytes. Excessive oxidative stress and associated inflammatory processes can impair wound healing, and antioxidants have been reported to improve wound healing in animal models and human subjects. Uric acid (UA) is an efficient free radical scavenger, but has a very low solubility and poor tissue penetrability. We recently developed novel UA analogs with increased solubility and excellent free radical-scavenging properties and demonstrated their ability to protect neural cells against oxidative damage. Here we show that the uric acid analog (6, 8 dithio-UA, but not equimolar concentrations of UA or 1, 7 dimethyl-UA) modified the behaviors of cultured vascular endothelial cells, keratinocytes and fibroblasts in ways consistent with enhancement of the wound healing functions of all three cell types. We further show that 6, 8 dithio-UA significantly accelerates the wound healing process when applied topically (once daily) to full-thickness wounds in mice. Levels of Cu/Zn superoxide dismutase were increased in wound tissue from mice treated with 6, 8 dithio-UA compared to vehicle-treated mice, suggesting that the UA analog enhances endogenous cellular antioxidant defenses. These results support an adverse role for oxidative stress in wound healing and tissue repair, and provide a rationale for the development of UA analogs in the treatment of wounds and for modulation of angiogenesis in other pathological conditions.

Membrane-associated oxidative stress has been implicated in the synaptic dysfunction and neuronal degeneration that occurs in Alzheimer's disease (AD), but the underlying mechanisms are unknown. Enzymes of the plasma membrane redox system (PMRS) provide electrons for energy metabolism and recycling of antioxidants. Here, we show that activities of several PMRS enzymes are selectively decreased in plasma membranes from the hippocampus and cerebral cortex of 3xTgAD mice, an animal model of AD. Our results that indicate the decreased PMRS enzyme activities are associated with decreased levels of coenzyme Q(10) and increased levels of oxidative stress markers. Neurons overexpressing the PMRS enzymes (NQO1 or cytochrome b5 reductase) exhibit increased resistance to amyloid β-peptide (Aβ). If and to what extent Aβ is the cause of the impaired PMRS enzymes in the 3xTgAD mice is unknown. Because these mice also express mutant tau and presenilin-1, it is possible that one or more of the PMRS could be adversely affected by these mutations. Nevertheless, the results of our cell culture studies clearly show that exposure of neurons to Aβ1-42 is sufficient to impair PMRS enzymes. The impairment of the PMRS in an animal model of AD, and the ability of PMRS enzyme activities to protect neurons against Aβ-toxicity, suggest enhancement PMRS function as a novel approach for protecting neurons against oxidative damage in AD and related disorders.

Claudin proteins are frequently overexpressed in various tumors such as breast, prostate and ovarian cancer. While their functions in cancer have not been completely elucidated, roles in survival, adhesion, and invasion have been suggested. In order to clarify the roles of claudins in ovarian cancer, we have performed gene expression profiling of ovarian surface epithelial cells overexpressing claudin-4 and compared the expression patterns to the parental, non-expressing cells. Claudin-4 expression leads to the differential expression of several genes, including many that have previously been implicated in angiogenesis. In particular, angiogenic cytokines, such as IL-8, were found elevated while genes of the angiostatic interferon pathway were found down-regulated. In vitro assays show that claudin-4-expressing cells produce factors that can stimulate angiogenesis as measured by tube formation and migration in HUVEC cells. In addition, an in vivo mouse dorsal skinfold assay confirms that cells expressing claudin-4 secrete factors that can mediate angiogenesis in the dorsal skin of mice. Our data suggest a novel function for claudin-4 in cancer and provide an additional rationale for its common overexpression in human tumors.

IInteractions between laminins and integrin receptors hold neural stem cells in place at the ventricular surface of embryonic brain. Transient disruption leads to abnormal stem cell divisions and permanent cortical malformation., During embryogenesis, the neural stem cells (NSC) of the developing cerebral cortex are located in the ventricular zone (VZ) lining the cerebral ventricles. They exhibit apical and basal processes that contact the ventricular surface and the pial basement membrane, respectively. This unique architecture is important for VZ physical integrity and fate determination of NSC daughter cells. In addition, the shorter apical process is critical for interkinetic nuclear migration (INM), which enables VZ cell mitoses at the ventricular surface. Despite their importance, the mechanisms required for NSC adhesion to the ventricle are poorly understood. We have shown previously that one class of candidate adhesion molecules, laminins, are present in the ventricular region and that their integrin receptors are expressed by NSC. However, prior studies only demonstrate a role for their interaction in the attachment of the basal process to the overlying pial basement membrane. Here we use antibody-blocking and genetic experiments to reveal an additional and novel requirement for laminin/integrin interactions in apical process adhesion and NSC regulation. Transient abrogation of integrin binding and signalling using blocking antibodies to specifically target the ventricular region in utero results in abnormal INM and alterations in the orientation of NSC divisions. We found that these defects were also observed in laminin α2 deficient mice. More detailed analyses using a multidisciplinary approach to analyse stem cell behaviour by expression of fluorescent transgenes and multiphoton time-lapse imaging revealed that the transient embryonic disruption of laminin/integrin signalling at the VZ surface resulted in apical process detachment from the ventricular surface, dystrophic radial glia fibers, and substantial layering defects in the postnatal neocortex. Collectively, these data reveal novel roles for the laminin/integrin interaction in anchoring embryonic NSCs to the ventricular surface and maintaining the physical integrity of the neocortical niche, with even transient perturbations resulting in long-lasting cortical defects., Author Summary The developing cerebral cortex contains bipolar neural stem cells that span the cortical layers between the inner ventricular surface and the outer pial surface of the embryonic brain. The nuclei of these cells remain near the ventricular cavity, a microenvironment or niche thought to provide vital signals. It is not known how this inner end of the bipolar stem cell is held in place, or what would happen if its attachment to the inner surface were lost. Genetic manipulation can be used to disrupt candidate molecules involved in this adhesion, but this will affect all adhesion points and complicate the results. We have therefore developed an approach to target the stem cell attachments specifically at the ventricular surface by placing blocking antibodies directly into the ventricles of mouse embryos and then expressing fluorescent markers in the stem cells to see the effects of losing this attachment in living tissue. We examined laminins and integrins, whose expression and properties make them excellent candidates. Blocking integrin signalling by antibody application caused the inner end of the stem cells to rapidly detach and then undergo aberrant cell division. We also showed that a transient block of integrins (for

Brain iron insufficiency has been implicated in several neurological disorders. The dopamine system is consistently altered in studies of iron deficiency in rodent models. Changes in striatal dopamine D(2) receptors are directly proportional to the degree of iron deficiency. In light of the unknown mechanism for the iron deficiency-dopamine connection and because of the known interplay between adenosinergic and dopaminergic systems in the striatum we examined the effects of iron deficiency on the adenosine system. We first attempted to assess whether there is a functional change in the levels of adenosine receptors in response to this low iron. Mice made iron-deficient by diet had an increase in the density of striatal adenosine A(2A) (A(2A)R) but not A(1) receptor (A(1)R) compared to mice on a normal diet. Between two inbred murine strains, which had 2-fold differences in their striatal iron concentrations under normal dietary conditions, the strain with the lower striatal iron had the highest striatal A(2A)R density. Treatment of SH-SY5Y (human neuroblastoma) cells with an iron chelator resulted in increased density of A(2A)R. In these cells, A(2A)R agonist-induced cyclic AMP production was enhanced in response to iron chelation, also demonstrating a functional upregulation of A(2A)R. A significant correlation (r(2)=0.79) was found between a primary marker of cellular iron status (transferrin receptor (TfR)) and A(2A)R protein density. In conclusion, the A(2A)R is increased across different iron-insufficient conditions. The relation between A(2A)R and cellular iron status may be an important pathway by which adenosine may alter the function of the dopaminergic system.

Regular exercise can counteract the adverse effects of aging on the musculoskeletal and cardiovascular systems. In males the normal aging process is associated with reductions in testosterone production’(Matsumoto 2002); (synder 2001) and impaired spermatogenesis, but the underlying mechanisms and their potential modification by exercise are unknown. Here we report that lifelong regular exercise (running) protects the testes against maturation as well as age related changes, and that this effect of running is associated with decreased amounts of oxidative damage to proteins, lipids and DNA in spermatogenic and Leydig cells. Six month-old male mice were divided into a sedentary group and a group which ran an average of 1.75 km/day, until the mice reached the age of 20 months. Seminiferous tubules of runners exhibited a full complement of cells at different stages of the spermatogenic process and a clear central lumen with large numbers of spermatozoa, in contrast to sedentary mice which exhibited disorganized spermatogenic cells and lacked spermatocytes in a central lumen. Levels of protein carbonyls, nitrotyrosine, lipid peroxidation products and oxidatively modified DNA were significantly greater in spermatogenic and Leydig cells of sedentary mice compared to runners. These findings suggest that lifelong regular exercise suppresses aging of the testes by a mechanism that involves reduced oxidative damage to spermatogenic and Leydig cells.

Epidemiological studies indicate that women have a higher risk of Alzheimer's disease (AD) even after adjustment for age. Though transgenic mouse models of AD develop AD-related amyloid beta (Abeta) and/or tau pathology, gender differences have not been well documented in these models. In this study, we found that female 3xTg-AD transgenic mice expressing mutant APP, presenilin-1 and tau have significantly more aggressive Abeta pathology. We also found an increase in beta-secretase activity and a reduction of neprilysin in female mice compared to males; this suggests that a combination of increased Abeta production and decreased Abeta degradation may contribute to higher risk of AD in females. In contrast to significantly more aggressive Abeta pathology in females, gender did not affect the levels of phosphorylated tau in 3xTg-AD mice. These results point to the involvement of Abeta pathways in the higher risk of AD in women. In addition to comparison of pathology between genders at 9, 16 and 23 months of age, we examined the progression of Abeta pathology at additional age points; i.e., brain Abeta load, intraneuronal oligomeric Abeta distribution and plaque load, in male 3xTg-AD mice at 3, 6, 9, 12, 16, 20 and 23 months of age. These findings confirm progressive Abeta pathology in 3xTg-AD transgenic mice, and provide guidance for their use in therapeutic research.

The Notch signaling pathway is critically involved in cell fate decisions during development of many tissues and organs. In the present study we employed in vivo and cell culture models to elucidate the role of Notch signaling in wound healing. The healing of full-thickness dermal wounds was significantly delayed in Notch antisense transgenic mice and in normal mice treated with gamma-secretase inhibitors that block proteolytic cleavage and activation of Notch. In contrast, mice treated with a Notch ligand Jagged peptide showed significantly enhanced wound healing compared to controls. Activation or inhibition of Notch signaling altered the behaviors of cultured vascular endothelial cells, keratinocytes and fibroblasts in a scratch wound healing model in ways consistent with roles for Notch signaling in wound healing functions all three cell types. These results suggest that Notch signaling plays important roles in wound healing and tissue repair, and that targeting the Notch pathway might provide a novel strategy for treatment of wounds and for modulation of angiogenesis in other pathological conditions.

The extracellular matrix (ECM) provides both a physical framework and a microenvironment that supplies instructive signals from the earliest stages of multicellular development. As a first step toward understanding the role of the ECM in regulating the behavior of neural stem cells (NSCs), here we show the localization of laminins, a heterotrimeric family of ECM molecules expressed in many different stem cell microenvironments, and their corresponding receptors in the embryonic murine ventricular zone (VZ) within which the NSCs undergo symmetrical and asymmetrical divisions required for cortical development. In addition to the presence of laminins containing both the α2 and α4 chains, we find distinct patterns of ECM receptor expression in the VZ and in the overlying cortex. Neural stem cells derived from the VZ express high levels of the integrin laminin receptor α6β1. At developmental stages at which NSCs undergo asymmetrical divisions, integrin β1 was unevenly distributed in some mitotic pairs at the ventricular wall. These results suggest a significant role in the regulation of NSC fate for laminin/integrin signaling within the microenvironment of the VZ and provide a framework for future molecular and cellular analyses of the role of the ECM in neural development. J. Comp. Neurol. 505:630–643, 2007. © 2007 Wiley-Liss, Inc.

The innate immune system senses the invasion of pathogenic microorganisms and tissue injury through Toll-like receptors (TLR), a mechanism thought to be limited to immune cells. We now report that neurons express several TLRs, and that the levels of TLR2 and -4 are increased in neurons in response to IFN-γ stimulation and energy deprivation. Neurons from both TLR2 knockout and -4 mutant mice were protected against energy deprivation-induced cell death, which was associated with decreased activation of a proapoptotic signaling cascade involving jun N-terminal kinase and the transcription factor AP-1. TLR2 and -4 expression was increased in cerebral cortical neurons in response to ischemia/reperfusion injury, and the amount of brain damage and neurological deficits caused by a stroke were significantly less in mice deficient in TLR2 or -4 compared with WT control mice. Our findings establish a proapoptotic signaling pathway for TLR2 and -4 in neurons that may render them vulnerable to ischemic death.

Histidine is a naturally occurring amino acid with antioxidant properties, which is present in low amounts in tissues throughout the body. We recently synthesized and characterized histidine analogues related to the natural dipeptide carnosine, which selectively scavenge the toxic lipid peroxidation product 4-hydroxynonenal (HNE). We now report that the histidine analogue histidyl hydrazide is effective in reducing brain damage and improving functional outcome in a mouse model of focal ischemic stroke when administered intravenously at a dose of 20 mg/kg, either 30 min before or 60 min and 3 h after the onset of middle cerebral artery occlusion. The histidine analogue also protected cultured rat primary neurons against death induced by HNE, chemical hypoxia, glucose deprivation, and combined oxygen and glucose deprivation. The histidine analogue prevented neuronal apoptosis as indicated by decreased production of cleaved caspase-3 protein. These findings suggest a therapeutic potential for HNE-scavenging histidine analogues in the treatment of stroke and related neurodegenerative conditions.

Mice transgenic for antisense Notch and normal mice treated with inhibitors of the Notch-activating enzyme gamma-secretase showed reduced damage to brain cells and improved functional outcome in a model of focal ischemic stroke. Notch endangers neurons by modulating pathways that increase their vulnerability to apoptosis, and by activating microglial cells and stimulating the infiltration of proinflammatory leukocytes. These findings suggest that Notch signaling may be a therapeutic target for treatment of stroke and related neurodegenerative conditions.

Huntington's disease (HD) is associated with profound autonomic dysfunction including dysregulation of cardiovascular control often preceding cognitive or motor symptoms. Brain-derived neurotrophic factor (BDNF) levels are decreased in the brains of HD patients and HD mouse models, and restoring BDNF levels prevents neuronal loss and extends survival in HD mice. We reasoned that heart rate changes in HD may be associated with altered BDNF signaling in cardiovascular control nuclei in the brainstem. Here we show that heart rate is elevated in HD (N171-82Q) mice at presymptomatic and early disease stages, and heart rate responses to restraint stress are attenuated. BDNF levels were significantly reduced in brainstem regions containing cardiovascular nuclei in HD mice and human HD patients. Central administration of BDNF restored the heart rate to control levels. Our findings establish a link between diminished BDNF expression in brainstem cardiovascular nuclei and abnormal heart rates in HD mice, and suggest a novel therapeutic target for correcting cardiovascular dysfunction in HD.

Chronic stress may be a risk factor for developing Alzheimer's disease (AD), but most studies of the effects of stress in models of AD utilize acute adverse stressors of questionable clinical relevance. The goal of this work was to determine how chronic psychosocial stress affects behavioral and pathological outcomes in an animal model of AD, and to elucidate underlying mechanisms. A triple-transgenic mouse model of AD (3xTgAD mice) and nontransgenic control mice were used to test for an affect of chronic mild social stress on blood glucose, plasma glucocorticoids, plasma insulin, anxiety, and hippocampal amyloid β-particle (Aβ), phosphorylated tau (ptau), and brain-derived neurotrophic factor (BDNF) levels. Despite the fact that both control and 3xTgAD mice experienced rises in corticosterone during episodes of mild social stress, at the end of the 6-week stress period 3xTgAD mice displayed increased anxiety, elevated levels of Aβ oligomers and intraneuronal Aβ, and decreased brain-derived neurotrophic factor levels, whereas control mice did not. Findings suggest 3xTgAD mice are more vulnerable than control mice to chronic psychosocial stress, and that such chronic stress exacerbates Aβ accumulation and impairs neurotrophic signaling.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by a progressive loss of lower motor neurons in the spinal cord. The incretin hormone, glucagon-like peptide-1 (GLP-1), facilitates insulin signaling, and the long acting GLP-1 receptor agonist exendin-4 (Ex-4) is currently used as an anti-diabetic drug. GLP-1 receptors are widely expressed in the brain and spinal cord, and our prior studies have shown that Ex-4 is neuroprotective in several neurodegenerative disease rodent models, including stroke, Parkinson's disease and Alzheimer's disease. Here we hypothesized that Ex-4 may provide neuroprotective activity in ALS, and hence characterized Ex-4 actions in both cell culture (NSC-19 neuroblastoma cells) and in vivo (SOD1 G93A mutant mice) models of ALS. Ex-4 proved to be neurotrophic in NSC-19 cells, elevating choline acetyltransferase (ChAT) activity, as well as neuroprotective, protecting cells from hydrogen peroxide-induced oxidative stress and staurosporine-induced apoptosis. Additionally, in both wild-type SOD1 and mutant SOD1 (G37R) stably transfected NSC-19 cell lines, Ex-4 protected against trophic factor withdrawal-induced toxicity. To assess in vivo translation, SOD1 mutant mice were administered vehicle or Ex-4 at 6-weeks of age onwards to end-stage disease via subcutaneous osmotic pump to provide steady-state infusion. ALS mice treated with Ex-4 showed improved glucose tolerance and normalization of behavior, as assessed by running wheel, compared to control ALS mice. Furthermore, Ex-4 treatment attenuated neuronal cell death in the lumbar spinal cord; immunohistochemical analysis demonstrated the rescue of neuronal markers, such as ChAT, associated with motor neurons. Together, our results suggest that GLP-1 receptor agonists warrant further evaluation to assess whether their neuroprotective potential is of therapeutic relevance in ALS.