Increased vulnerability of hippocampal pyramidal neurons to the toxicity of kainic acid in OASIS-deficient mice

The endoplasmic reticulum (ER) stress response is a defense system for dealing with the accumulation of unfolded proteins in the ER lumen. Old astrocyte specifically induced substance (OASIS) is known to be expressed in astrocytes and involved in the ER stress response; however the function of OASIS in the injured brain has remained unclear. In this study, we examined the roles of OASIS in neuronal degeneration in the hippocampi of mice intraperitoneally injected with kainic acid (KA). OASIS mRNA was strongly induced in response to KA injection, with a similar time course to the induction of ER molecular chaperone immunoglobulin heavy chain binding protein mRNA. In situ hybridization showed that KA injection causes induction of immunoglobulin heavy chain binding protein mRNA in glial fibrillary acidic protein-positive astrocytes as well as in pyramidal neurons, although up-regulation of OASIS mRNA was only detected in glial fibrillary acidic protein-positive astrocytes. Primary cultured astrocytes, but not the neurons of OASIS −/− mice, revealed reduced vulnerability to ER stress. Furthermore, pyramidal neurons in the hippocampi of OASIS −/− mice were more susceptible to the toxicity induced by KA than those of wild-type mice. Taken together, these data suggest that OASIS expressed in astrocytes plays important roles in protection against the neuronal damage induced by KA.     

Deficiency of presenilin-1 increases calcium-dependent vulnerability of neurons to oxidative stress in vitro

We examined the function of presenilin-1 (PS1) on neuronal resistance to oxidative stress. CNS neurons cultured from PS1-deficient mice exhibited increased vulnerability to H2O2 treatment compared with those from wild-type mice. Antioxidants protected the cultured neurons against the oxidative stress. An intracellular calcium chelator, BAPTA AM, as well as an L-type voltage-dependent calcium channel blocker, nifedipine, rescued the neurons from H2O2-induced death, while an N-type voltage-dependent calcium channel blocker, omega-conotoxin, or calcium release blockers from ER stores, dantrolene and xestospongin C, failed to rescue them. Wild-type and PS1-deficient neurons showed comparable increases of cytoplasmic free calcium levels after exposure to H2O2. Taken together with the data that PS1-deficient neurons exhibited increased vulnerability to glutamate, these findings imply that PS1 confers resistance to oxidative stress on neurons in calcium-dependent manners.     

Developmental expression of wild-type and mutant presenilin-1 in hippocampal neurons from transgenic mice: evidence for novel species-specific properties of human presenilin-1.

Presenilins 1 (PS1) and 2 (PS2) are multispanning transmembrane proteins associated with familial Alzheimer disease (FAD). They are developmentally regulated, being expressed at highest levels during neuronal differentiation and are sustained at a lower level throughout life. We investigated the distribution and metabolism of endogenous murine PS1 as well as human wild-type (wtPS1) and the familial AD Met146Leu (M146L) mutant presenilins in dissociated cultures of hippocampal neurons derived from control and transgenic mice. We found that the PS1 endoproteolytic fragments and, to a lesser extent, the full-length protein, were expressed as early as day 3 post-plating. Both species increased until the cells were fully differentiated at day 12. Confocal microscopy revealed that presenilin is present in the Golgi and endoplasmic reticulum and, as in punctate, vesicle-like structures within developing neurites and growth cones. Using a human-specific PS1 antibody, we were able to independently examine the distribution of the transgenic protein which, although similar to the endogenous, showed some unique qualities. These included (i) some heterogeneity in the proteolytic fragments of human PS1; (ii) significantly reduced levels of full-length human PS1, possibly as a result of preferential processing; and (iii) a more discrete intracellular distribution of human PS1. Colocalization with organelle-specific proteins revealed that PS1 was located in a diffuse staining pattern in the MAP2-positive dendrites and in a punctate manner in GAP43-positive axons. PS1 showed considerable overlap with GAP43, particularly at the growth cones. Similar patterns of PS1 distribution were detected in cultures derived from transgenic animals expressing human wild-type or mutant presenilins. The studies demonstrate that mutant presenilins are not grossly different in their processing or distribution within cultured neurons, which may represent more physiological models as compared to transfection systems. Our data also suggest that the molecular pathology associated with PS1 mutations results from subtle alterations in presenilin function, which can be further investigated using these transgenic neuronal cell culture models.     

Altered calcium homeostasis and mitochondrial dysfunction in cortical synaptic compartments of presenilin-1 mutant mice

Alzheimer's disease is characterized by amyloid beta-peptide deposition, synapse loss, and neuronal death, which are correlated with cognitive impairments. Mutations in the presenilin-1 gene on chromosome 14 are causally linked to many cases of early-onset inherited Alzheimer's disease. We report that synaptosomes prepared from transgenic mice harboring presenilin-1 mutations exhibit enhanced elevations of cytoplasmic calcium levels following exposure to depolarizing agents, amyloid beta-peptide, and a mitochondrial toxin compared with synaptosomes from nontransgenic mice and mice overexpressing wild-type presenilin-1. Mitochondrial dysfunction and caspase activation following exposures to amyloid beta-peptide and metabolic insults were exacerbated in synaptosomes from presenilin-1 mutant mice. Agents that buffer cytoplasmic calcium or that prevent calcium release from the endoplasmic reticulum protected synaptosomes against the adverse effect of presenilin-1 mutations on mitochondrial function. Abnormal synaptic calcium homeostasis and mitochondrial dysfunction may contribute to the pathogenic mechanism of presenilin-1 mutations.     

Increased fibrillar beta-amyloid in response to human clq injections into hippocampus and cortex of APP+PS1 transgenic mice

Human C1q when injected directly into hippocampus and cortex of doubly transgenic APP+PS1 mice results in the increase of Congo red-positive fibrillar deposits. Although there was no significant change in overall area stained for A total, qualitatively it appeared that there was less diffuse A in C1q-treated mice versus vehicle. There was no apparent change in astroglial or microglial activation caused by injection of C1q with respect to vehicle injections. These effects of C1q were only found in 50% BUB/BnJ mice, a strain with higher serum complement activity than other mouse lines. These in vivo data were consistent with the effects of C1q to increase fibrillogenesis of A in vitro. In conclusion, complement protein C1q, believed to be involved in the pathogenesis of Alzheimer's disease in humans, can cause increased fibrillogenesis in the APP+PS1 mouse model of amyloid deposition.     

Mutations in amyloid precursor protein and presenilin-1 genes increase the basal oxidative stress in murine neuronal cells and lead to increased sensitivity to oxidative stress mediated by amyloid beta-peptide (1-42), HO and kainic acid: implications for Alzheimer's disease

Oxidative stress is observed in Alzheimer's disease (AD) brain, including protein oxidation and lipid peroxidation. One of the major pathological hallmarks of AD is the brain deposition of amyloid beta-peptide (Abeta). This 42-mer peptide is derived from the beta-amyloid precursor protein (APP) and is associated with oxidative stress in vitro and in vivo. Mutations in the PS-1 and APP genes, which increase production of the highly amyloidogenic amyloid beta-peptide (Abeta42), are the major causes of early onset familial AD. Several lines of evidence suggest that enhanced oxidative stress, inflammation, and apoptosis play important roles in the pathogenesis of AD. In the present study, primary neuronal cultures from knock-in mice expressing mutant human PS-1 and APP were compared with those from wild-type mice, in the presence or absence of various oxidizing agents, viz, Abeta(1-42), H2O2 and kainic acid (KA). APP/PS-1 double mutant neurons displayed a significant basal increase in oxidative stress as measured by protein oxidation, lipid peroxidation, and 3-nitrotyrosine when compared with the wild-type neurons (p < 0.0005). Elevated levels of human APP, PS-1 and Abeta(1-42) were found in APP/PS-1 cultures compared with wild-type neurons. APP/PS-1 double mutant neuron cultures exhibited increased vulnerability to oxidative stress, mitochondrial dysfunction and apoptosis induced by Abeta(1-42), H2O2 and KA compared with wild-type neuronal cultures. The results are consonant with the hypothesis that Abeta(1-42)-associated oxidative stress and increased vulnerability to oxidative stress may contribute significantly to neuronal apoptosis and death in familial early onset AD.     

Beneficial effects of dietary restriction on cerebral cortical synaptic terminals: preservation of glucose and glutamate transport and mitochondrial function after exposure to amyloid beta-peptide, iron, and 3-nitropropionic acid

Recent studies have shown that rats and mice maintained on a dietary restriction (DR) regimen exhibit increased resistance of neurons to excitotoxic, oxidative, and metabolic insults in experimental models of Alzheimer's, Parkinson's, and Huntington's diseases and stroke. Because synaptic terminals are sites where the neurodegenerative process may begin in such neurodegenerative disorders, we determined the effects of DR on synaptic homeostasis and vulnerability to oxidative and metabolic insults. Basal levels of glucose uptake were similar in cerebral cortical synaptosomes from rats maintained on DR for 3 months compared with synaptosomes from rats fed ad libitum. Exposure of synaptosomes to oxidative insults (amyloid beta-peptide and Fe(2+)) and a metabolic insult (the mitochondrial toxin 3-nitropropionic acid) resulted in decreased levels of glucose uptake. Impairment of glucose uptake following oxidative and metabolic insults was significantly attenuated in synaptosomes from rats maintained on DR. DR was also effective in protecting synaptosomes against oxidative and metabolic impairment of glutamate uptake. Loss of mitochondrial function caused by oxidative and metabolic insults, as indicated by increased levels of reactive oxygen species and decreased transmembrane potential, was significantly attenuated in synaptosomes from rats maintained on DR. Levels of the stress proteins HSP-70 and GRP-78 were increased in synaptosomes from DR rats, consistent with previous data suggesting that the neuroprotective mechanism of DR involves a "preconditioning" effect. Collectively, our data provide the first evidence that DR can alter synaptic homeostasis in a manner that enhances the ability of synapses to withstand adversity.     

Interleukin-1 beta up-regulates TACE to enhance alpha-cleavage of APP in neurons: resulting decrease in Abeta production

The proinflammatory cytokine interleukin (IL)-1β is up-regulated in microglial cells surrounding amyloid plaques, leading to the hypothesis that IL-1β is a risk factor for Alzheimer's disease. However, we unexpectedly found that IL-1β significantly enhanced α-cleavage, indicated by increases in sAPPα and C83, but reduced β-cleavage, indicated by decreases in sAPPβ and Aβ40/42, in human neuroblastoma SK-N-SH cells. IL-1β did not significantly alter the mRNA levels of BACE1, ADAM-9, and ADAM-10, but up-regulated that of TACE by threefold. The proform and mature form of TACE protein were also significantly up-regulated. A TACE inhibitor (TAPI-2) concomitantly reversed the IL-1β-dependent increase in sAPPα and decrease in sAPPβ, suggesting that APP consumption in the α-cleavage pathway reduced its consumption in the β-cleavage pathway. IL-1Ra, a physiological antagonist for the IL-1 receptor, reversed the effects of IL-1β, suggesting that the IL-1β-dependent up-regulation of α-cleavage is mediated by the IL-1 receptor. IL-1β also induced this concomitant increase in α-cleavage and decrease in β-cleavage in mouse primary cultured neurons. Taken together we conclude that IL-1β is an anti-amyloidogenic factor, and that enhancement of its signaling or inhibition of IL-1Ra activity could represent potential therapeutic strategies against Alzheimer's disease.     

Reduced blood‐brain barrier expression of fatty acid‐binding protein 5 is associated with increased vulnerability of APP/PS1 mice to cognitive deficits from low omega‐3 fatty acid diets

Lower levels of the cognitively beneficial docosahexaenoic acid (DHA) are often observed in Alzheimer's disease (AD) brains. Brain DHA levels are regulated by the blood‐brain barrier (BBB) transport of plasma‐derived DHA, a process facilitated by fatty acid‐binding protein 5 (FABP5). This study reports a 42.1 ± 12.6% decrease in the BBB transport of ¹⁴C‐DHA in 8‐month‐old AD transgenic mice (APPswe,PSEN1?E9) relative to wild‐type mice, associated with a 34.5 ± 6.7% reduction in FABP5 expression in isolated brain capillaries of AD mice. Furthermore, short‐term spatial and recognition memory deficits were observed in AD mice on a 6‐month n‐3 fatty acid‐depleted diet, but not in AD mice on control diet. This intervention led to a dramatic reduction (41.5 ± 11.9%) of brain DHA levels in AD mice. This study demonstrates FABP5 deficiency and impaired DHA transport at the BBB are associated with increased vulnerability to cognitive deficits in mice fed an n‐3 fatty acid‐depleted diet, in line with our previous studies demonstrating a crucial role of FABP5 in BBB transport of DHA and cognitive function.

     

Endogenous GLP-1 acts on paraventricular nucleus to suppress feeding: Projection from nucleus tractus solitarius and activation of corticotropin-releasing hormone,nesfatin-1 and oxytocin neurons

Glucagon-like peptide-1 (GLP-1) receptor agonists have been used to treat type 2 diabetic patients and shown to reduce food intake and body weight. The anorexigenic effects of GLP-1 and GLP-1 receptor agonists are thought to be mediated primarily via the hypothalamic paraventricular nucleus (PVN). GLP-1, an intestinal hormone, is also localized in the nucleus tractus solitarius (NTS) of the brain stem. However, the role of endogenous GLP-1, particularly that in the NTS neurons, in feeding regulation remains to be established. The present study examined whether the NTS GLP-1 neurons project to PVN and whether the endogenous GLP-1 acts on PVN to restrict feeding. Intra-PVN injection of GLP-1 receptor antagonist exendin (9–39) increased food intake. Injection of retrograde tracer into PVN combined with immunohistochemistry for GLP-1 in NTS revealed direct projection of NTS GLP-1 neurons to PVN. Moreover, GLP-1 evoked Ca²⁺ signaling in single neurons isolated from PVN. The majority of GLP-1-responsive neurons were immunoreactive predominantly to corticotropin-releasing hormone (CRH) and nesfatin-1, and less frequently to oxytocin. These results indicate that endogenous GLP-1 targets PVN to restrict feeding behavior, in which the projection from NTS GLP-1 neurons and activation of CRH and nesfatin-1 neurons might be implicated. This study reveals a neuronal basis for the anorexigenic effect of endogenous GLP-1 in the brain.     

Susceptibility of cerebellar granule neurons from GM2/GD2 synthase-null mice to apoptosis induced by glutamate excitotoxicity and elevated KCl: Rescue by GM1 and LIGA20

Our previous study showed an impaired regulation of Ca(2+) homeostasis in cultured cerebellar granule neurons (CGN) from neonatal mice lacking GM2, GD2 and all gangliotetraose gangliosides, due to disruption of the GM2/GD2 synthase (GalNAc-T) gene. In the presence of depolarizing concentration (55 mM) K(+), these cells showed persistent elevation of intracellular Ca(2+) ([Ca(2+)]( i )) leading to apoptosis and cell destruction. This was in contrast to CGN from normal littermates whose survival was enhanced by high K(+). In this study we demonstrate that glutamate has the same effect as K(+) on CGN from these ganglioside-deficient knockout (KO) mice and that apoptosis in both cases is averted by exogenous GM1. Even more effective rescue was obtained with LIGA20, a semi-synthetic derivative of GM1. LC(50) of glutamate in the KO cells was 3.1 microM, compared to 46 microM in normal CGN. [Ca(2+)]( i ) measurement with fura-2 revealed no difference in glutamate-stimulated Ca(2+) influx between the 2 cell types. However, reduction of [Ca(2+)]( i ) following application of Mg(2+) was significantly impaired in the mutant CGN. The rescuing effects of exogenous GM1 and LIGA20 corresponded to their ability to restore Ca(2+) homeostasis. The greater potency of LIGA20 is attributed to its greater membrane permeability with resultant ability to insert into both plasma and nuclear membranes at low concentration (相似文献