Binding of purified Reelin to ApoER2 and VLDLR mediates tyrosine phosphorylation of Disabled-1

Reelin, Disabled-1 (Dab1), apolipoprotein E receptor 2 (ApoER2), and very low density lipoprotein receptor (VLDLR) participate in a signaling pathway required for layer formation during mammalian brain development. Binding of Reelin to ApoER2 and VLDLR induces a rapid increase in tyrosine phosphorylation of Dab1, an adaptor protein that associates with the cytoplasmic domain of the receptors. However, Reelin has also been proposed to signal through integrin and protocadherin. Here we compare the roles of ApoER2 and VLDLR in Reelin signaling. We used layer-specific markers to identify the final positions of early- and late-born neurons in the cortices of mice lacking ApoER2, VLDLR, or both ApoER2 and VLDLR. Subtle alterations were observed in mice lacking VLDLR, whereas more severe abnormalities were detected in the absence of ApoER2, and major disruptions were obvious in mice lacking both receptors. Purified Reelin associated more readily with ApoER2 than with VLDLR and no synergy was observed in the presence of both receptors. Consistent with the binding data, the level of Reelin-induced Dab1 phosphorylation was more severely reduced in neurons lacking ApoER2 than in neurons lacking VLDLR. However, similarly low levels of Dab1 tyrosine phosphorylation were observed in ApoER2(-/-) and VLDLR(-/-) mice in vivo. Finally, there was a complete absence of Reelin-induced tyrosine phosphorylation of Dab1 in cortical neurons from mice lacking both ApoER2 and VLDLR. These findings demonstrate that ApoER2 and VLDLR are essential for Reelin signaling and that no other receptor molecules can compensate for their role in mediating tyrosine phosphorylation of Dab1.     

Malformation of the radial glial scaffold in the dentate gyrus of reeler mice,scrambler mice,and ApoER2/VLDLR-deficient mice

We studied the postnatal development of the radial glial scaffold in the dentate gyrus of reeler mice, lacking the extracellular matrix protein Reelin, in scrambler mice, deficient in the intracellular adaptor protein disabled1 (Dab1), which is required for the transmission of the Reelin signal into the cell, and in mutant mice lacking the Reelin receptors apolipoprotein receptor 2 (ApoER2) and/or the very low density lipoprotein receptor (VLDLR), known to transmit the Reelin signal via Dab1. By immunolabeling for the glial fibrillary acidic protein (GFAP), we show that a regular dentate radial glial scaffold fails to form in mutants deficient of Reelin, Dab1, and VLDLR and ApoER2. Mutant mice lacking only one of the Reelin receptors, VLDLR or ApoER2, display a gradual expression of the radial glial defects seen in mutants that lack both receptors. Our results suggest that Reelin signaling via ApoER2, VLDLR, and Dab1 is required for the formation of a regular radial glial scaffold in the dentate gyrus.     

Reelin receptors ApoER2 and VLDLR are expressed in distinct spatiotemporal patterns in developing mouse cerebral cortex

In mammalian developing brain, neuronal migration is regulated by a variety of signaling cascades, including Reelin signaling. Reelin is a glycoprotein that is mainly secreted by Cajal–Retzius neurons in the marginal zone, playing essential roles in the formation of the layered neocortex via its receptors, apolipoprotein E receptor 2 (ApoER2) and very low density lipoprotein receptor (VLDLR). However, the precise mechanisms by which Reelin signaling controls the neuronal migration process remain unclear. To gain insight into how Reelin signaling controls individual migrating neurons, we generated monoclonal antibodies against ApoER2 and VLDLR and examined the localization of Reelin receptors in the developing mouse cerebral cortex. Immunohistochemical analyses revealed that VLDLR is localized to the distal portion of leading processes in the marginal zone (MZ), whereas ApoER2 is mainly localized to neuronal processes and the cell membranes of multipolar cells in the multipolar cell accumulation zone (MAZ). These different expression patterns may contribute to the distinct actions of Reelin on migrating neurons during both the early and late migratory stages in the developing cerebral cortex. J. Comp. Neurol. 523:463–478, 2015. © 2014 Wiley Periodicals, Inc.     

Reelin receptors in developing laminated brain structures of mouse and human

Reelin is an extracellular matrix protein secreted by a variety of cell types throughout the developing brain. The target cells for reelin express the cytoplasmic adapter protein Dab1, which binds to the reelin receptors VLDLR and ApoER2. In the present work, we have studied the localization of both receptors in developing mouse and human cortex, olfactory bulb and cerebellum. In mouse, some Cajal-Retzius cells express reelin and VLDLR; in humans, all the components of the signalling pathway (Reelin, Dab1, VLDLR and ApoER2) are present in subsets of Cajal-Retzius cells. In the mouse cortical plate, VLDLR and ApoER2 are present from E15 to postnatal stages; in human cortical plate they are most prominent at approximately 20 gestational weeks. In mice, cerebellar Purkinje cells only express VLDLR whereas in humans they express both VLDLR and ApoER2. Mitral cells of the mouse olfactory bulb are ApoER2-positive and VLDLR-negative. In sum, the receptor expression patterns are similar in the human and mouse cortical plate but differ in Cajal-Retzius and Purkinje cells, which in humans express additional components of the reelin-Dab1 pathway.     

Rescue of the reeler phenotype in the dentate gyrus by wild-type coculture is mediated by lipoprotein receptors for Reelin and Disabled 1

Reelin is a positional signal for the lamination of the dentate gyrus. In the reeler mutant lacking Reelin, granule cells are scattered all over the dentate gyrus. We have recently shown that the reeler phenotype of the dentate gyrus can be rescued in vitro by coculturing reeler hippocampal slices with slices from wild-type hippocampus. Here we studied whether Reelin from other brain regions can similarly induce this rescue effect and whether it is mediated via the Reelin receptors apolipoprotein E receptor 2 (ApoER2) and very-low-density lipoprotein receptor (VLDLR). We found that coculturing reeler hippocampal slices with slices from wild-type olfactory bulb, cerebellum, and neocortex rescued the reeler phenotype as seen before with hippocampal slices, provided that the Reelin-synthesizing cells of these regions were placed near the marginal zone of the reeler hippocampal slice. However, coculturing wild-type hippocampal slices with hippocampal slices from mutants deficient in ApoER2 and VLDLR did not rescue the reeler-like phenotype in these cultures. Similarly, no rescue of the reeler-like phenotype was observed in slices from mutants lacking Disabled 1 (Dab1), an adapter protein downstream of Reelin receptors. Conversely, reeler hippocampal slices were rescued by coculturing them with slices from Dab1(-/-) mutants or ApoER2(-/-)/VLDLR(-/-) mice. These findings show that Reelin from other brain regions can substitute for the loss of hippocampal Reelin and that rescue of the reeler phenotype observed in our coculture studies is mediated via lipoprotein receptors for Reelin and Dab1.     

ApoER2 function in the establishment and maintenance of retinal synaptic connectivity

The cellular and molecular mechanisms responsible for the development of inner retinal circuitry are poorly understood. Reelin and apolipoprotein E (apoE), ligands of apoE receptor 2 (ApoER2), are involved in retinal development and degeneration, respectively. Here we describe the function of ApoER2 in the developing and adult retina. ApoER2 expression was highest during postnatal inner retinal synaptic development and was considerably lower in the mature retina. Both during development and in the adult, ApoER2 was expressed by A-II amacrine cells. ApoER2 knock-out (KO) mice had rod bipolar morphogenic defects, altered A-II amacrine dendritic development, and impaired rod-driven retinal responses. The presence of an intact ApoER2 NPxY motif, necessary for binding Disabled-1 and transducing the Reelin signal, was also necessary for development of the rod bipolar pathway, while the alternatively spliced exon 19 was not. Mice deficient in another Reelin receptor, very low-density lipoprotein receptor (VLDLR), had normal rod bipolar morphology but altered A-II amacrine dendritic development. VLDLR KO mice also had reductions in oscillatory potentials and delayed synaptic response intervals. Interestingly, age-related reductions in rod and cone function were observed in both ApoER2 and VLDLR KOs. These results support a pivotal role for ApoER2 in the establishment and maintenance of normal retinal synaptic connectivity.     

High affinity binding of Dab1 to Reelin receptors promotes normal positioning of upper layer cortical plate neurons

The positions of neurons in the neocortex, hippocampus, cerebellum and various other laminated brain regions are regulated by a signaling pathway initiated by the secreted protein Reelin and requiring the intracellular adaptor protein Dab1. Dab1 and the Reelin receptors VLDLR and ApoER2 are expressed by neurons whose migrations are coordinated by Reelin. In vitro, Dab1 binds with high affinity to the cytoplasmic tails of VLDLR and ApoER2 via its PTB domain. To test the importance of Dab1 binding to VLDLR and ApoER2, we replaced the Dab1 gene with a cDNA cassette encoding a point mutant allele, Dab1(F158V). This mutation strongly decreases Dab1 binding in vitro to peptides containing the ApoER2 or VLDLR cytoplasmic regions. Surprisingly, Dab1(F158V/F158V) homozygotes have no discernable phenotype. However, Dab1(F158V/-) hemizygous animals have a subtle phenotype in which late-generated cortical plate neurons migrate excessively into the marginal zone. Early cortical plate neurons, subplate neurons, hippocampal pyramidal cells and cerebellar Purkinje cells are positioned normally. Thus Dab(F158V) is a weak loss-of-function (hypomorphic) allele that has no detectable effect when homozygous. The phenotype of Dab1(F158V/-) hemizygotes shows that late cortical plate neurons of layers 2-3 require efficient Reelin-Dab1 signaling to prevent them entering the marginal zone. The Dab1(F158V) allele adds to a series of Dab1 alleles that demonstrates cell type-specific variation in the Reelin-Dab1 pathway.     

Decreased expression of reelin receptor VLDLR in peripheral lymphocytes of drug-naive schizophrenic patients

Reelin, a secretory protease that plays major roles in neurodevelopment and synaptic plasticity, may also play a role in the pathogenesis of schizophrenia. The present study was undertaken to examine whether the expression of two receptors for reelin, very low-density lipoprotein receptor (VLDLR) and apolipoprotein E receptor type 2 (ApoER2), were abnormal in peripheral blood lymphocytes of schizophrenic patients. In this study, we measured the mRNA levels of VLDLR and ApoER2 in blood lymphocytes from patients with schizophrenia (drug-naive patients (n=20) and medicated patients (n=20)) and age-and gender-matched healthy controls (n=40) using quantitative real-time RT-PCR. Furthermore, we examined the correlation between mRNA levels and clinical variables in patients. Levels of VLDLR mRNA in drug-naive, unmedicated patients with schizophrenia were significantly lower than those of controls. In contrast, levels of ApoER2 mRNA in drug-naive patients did not differ from those of controls, although the levels of ApoER2 mRNA in medicated patients were significantly lower than those of controls. Interestingly, levels of VLDLR mRNA in drug-naive patients showed significant increases with respect to baseline after six months of antipsychotic treatment, whereas levels of ApoER2 mRNA were significantly lower than baseline after six months of treatment. In all patients, there was a negative correlation between VLDLR mRNA levels and the severity of clinical symptoms. Our findings suggest that peripheral VLDLR mRNA levels may serve as a reliable peripheral biological marker of schizophrenia, and that the reelin-VLDLR/ApoER2 signaling pathway plays a role in the pathophysiology of schizophrenia.     

Components of the reelin signaling pathway are expressed in the spinal cord

The Reelin signaling pathway in the brain involves the binding of Reelin to very-low-density lipoprotein receptors (VLDLR) and apolipoprotein E receptor 2 (ApoER2). After Reelin binds the lipoprotein receptors on migrating neurons, the intracellular adaptor protein Disabled-1 (Dab1) becomes phosphorylated, ultimately resulting in the proper positioning of cortical neurons. Previous work showed that Reelin also affects the positioning of sympathetic preganglionic neurons (SPN) in the spinal cord (Yip et al. [2000] Proc Natl Acad Sci USA 97:8612-8616). We asked in the present study whether components of the Reelin signaling pathway in the brain also function to control SPN migration in developing spinal cord. Results showed that Reelin and reelin mRNA are found adjacent to migrating SPN. In addition, dab1 mRNA and protein are expressed by migrating SPN, and dab1-null mice show abnormal SPN migration similar to that seen in reeler. Finally, vldlr and apoER2 are also expressed in migrating SPN, and mice lacking both vldlr and apoER2 show aberrant SPN location that is identical to that of reeler and dab1-null mice. Because molecules known to be involved in Reelin signaling in the brain are present in the developing spinal cord, it is likely that the Reelin signaling pathways in the brain and spinal cord function similarly. The relative simplicity of the organization of the spinal cord makes it a potentially useful model system with which to study the molecular and cellular function of the Reelin signaling pathway in control of neuronal migration.     

Tyrosine phosphorylation of Disabled-1 is essential for Reelin-stimulated activation of Akt and Src family kinases

Reelin is a large secreted signaling protein that is essential for proper positioning of migratory neurons during mammalian brain development. The Reelin signal is transduced into the cell by the lipoprotein receptors VLDLR and ApoER2, leading to tyrosine phosphorylation of the associated intracellular adaptor protein Disabled-1 (Dab1). Tyrosine phosphorylation of Dab1 is essential for responding to Reelin, as knock-in mice expressing a form of Dab1 that cannot be phosphorylated on tyrosine are indistinguishable from mice lacking Reelin, Reelin-receptors or Dab1. Molecular events dependent on Dab1 tyrosine phosphorylation are unknown. However, Reelin has recently been shown to activate the phosphoinositide-3-kinase (PI 3-K)-dependent kinase, Akt, as well as Src family kinases in wild type but not Dab1-/- primary embryonic neuronal cultures. Using pharmacological inhibitors and mice harboring mutant alleles of Dab1, we show here that tyrosine phosphorylation, but not the carboxyl-terminal region, of Dab1 is required for Reelin-induced activation of Akt and Src family kinases. Additionally, although Fyn is an important regulator of Dab1, Fyn deficiency does not prevent acute Reelin-induced Akt activation. Finally, whereas a number of growth factors propagate signals simultaneously through PI 3-K and mitogen-activated protein kinase (MAPK) cascades, we find Reelin does not engage the canonical MAPK cascade. These results define the first molecular events strictly dependent on Reelin-induced Dab1 tyrosine phosphorylation, and suggest that propagation of the Reelin signal is mediated by Akt, substrates of Src family kinases and/or unidentified molecules that share with these a common molecular link to phosphorylated Dab1.     

Reelin and disabled-1 expression in developing and mature human cortical neurons

In developing mammalian (mouse) brain, Reelin (Reln) is secreted by the Cajal-Retzius (CR) neurons in the marginal zone, binds apolipoprotein E receptor 2 (ApoER2) and very low density lipoprotein receptor (Vldlr), and induces the phosphorylation of the downstream cytoplasmic molecule disabled-1 (Dab1) in cortical plate neurons. Although this is a well-characterized signaling pathway in mice, it has not been well defined in human brain. In this paper we examined the expression of RELN, APOER2, VLDLR, and DAB1 in the developing human brain by RT-PCR. We further determined the cellular expression of the proteins RELN and DAB1 in 50 human brains ranging in age from 10 gestational weeks (GW) to 62 years using immunochemistry. We found that the pattern of expression of RELN and DAB1 in the human brain isnot identical to that observed in the mouse brain. In particular, we report the novel finding that human DAB1and RELN are coexpressed in CR neurons during cortical development and in cortical pyramidal neurons after neuronal migration is complete. Thus, in the human brain, the whole RELN signaling pathway is present within selected populations of cortical neurons throughout life. We speculate that RELN and DAB1 coexpression in these neurons is necessary for both normal cortical development and mature function.     

Reduced expression of apolipoprotein E receptor type 2 in peripheral blood lymphocytes from patients with major depressive disorder

We measured the mRNA levels of apolipoprotein E receptor type 2 (ApoER2) and very low-density lipoprotein receptor (VLDLR) in peripheral blood lymphocytes from 43 patients with major depressive disorder (27 drug-free patients and 16 medicated patients) and 43 age-matched healthy controls using a quantitative real-time RT-PCR method. The correlations between mRNA levels of both receptors and clinical variables in patients were also examined. The expression of ApoER2 mRNA, but not VLDLR, was significantly lower in patients as compared to controls, irrespective of the medication status. There was no statistically significant correlation between the reduction of ApoER2 mRNA levels and any of the clinical variables measured in patients. Results from this preliminary study suggest that the expression of ApoER2 may serve as a trait marker for major depressive disorder.     

Dentate granule cells in reeler mutants and VLDLR and ApoER2 knockout mice

We have studied the organization and cellular differentiation of dentate granule cells and their axons, the mossy fibers, in reeler mutant mice lacking reelin and in mutants lacking the reelin receptors very low density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2). We show that granule cells in reeler mice do not form a densely packed granular layer, but are loosely distributed throughout the hilar region. Immunolabeling for calbindin and calretinin revealed that the sharp border between dentate granule cells and hilar mossy cells is completely lost in reeler mice. ApoER2/VLDLR double-knockout mice copy the reeler phenotype. Mice deficient only in VLDLR showed minor alterations of dentate organization; migration defects were more prominent in ApoER2 knockout mice. Tracing of the mossy fibers with Phaseolus vulgaris leukoagglutinin and calbindin immunolabeling revealed an irregular broad projection in reeler mice and ApoER2/VLDLR double knockouts, likely caused by the irregular wide distribution of granule cell somata. Mutants lacking only one of the lipoprotein receptors showed only minor changes in the mossy fiber projection. In all mutants, mossy fibers respected the CA3-CA1 border. Retrograde labeling with DiI showed that malpositioned granule cells also projected as normal to the CA3 region. These results indicate that ( 1 ) reelin signaling via ApoER2 and VLDLR is required for the normal positioning of dentate granule cells and (2) the reelin signaling pathway is not involved in pathfinding and target recognition of granule cell axons.     

Disabled-1 is expressed in type AII amacrine cells in the mouse retina

The organization of several laminated structures in the brain is controlled by a signaling pathway activated by Reelin, a large glycoprotein secreted by pioneer neurons in the developing brain. Reelin binds to transmembrane receptors, including VLDLR and ApoER2, and stimulates tyrosine phosphorylation of Disabled-1 (Dab1), which associates with an NPxY motif present in the cytoplasmic domain of the receptors. Disruption of reelin, dab1, or both the vldr and apoer2 genes results in similar cell positioning defects in laminated brain regions including the cerebellum, hippocampus, and cerebral cortex. Although retinal ganglion cells express reelin during development, there is no obvious disruption of cell positioning in the retina of reeler mice. Here, we examine the expression pattern of Dab1 as a first step toward understanding the function of the Reelin signaling pathway in neural retina. Immunohistochemical analysis of the adult retina revealed that Dab1 is expressed in a specific type of amacrine cell. These cells display a narrow dendritic field and they project to two distinct sublaminae within the inner plexiform layer. Dab1 co-localizes with the high-affinity glycine transporter, indicating that these amacrine cells are glycinergic. Cells that express Dab1 are surrounded by dopaminergic fibers originating from wide-field amacrine cells. These features are characteristic of type AII amacrine cells described in other mammalian species. Analysis of the retina at several stages of development revealed that Dab1 is expressed shortly after birth during the time at which AII amacrine cells extend neurites and form synaptic connections in the inner retina. This raises the possibility that the Reelin/Dab1 signaling pathway contributes to formation of intraretinal circuitry in the neural retina.     

Localization of ApoER2, VLDLR and Dab1 in radial glia: groundwork for a new model of reelin action during cortical development

The reelin signaling pathway regulates laminar positioning of radially migrating neurons during cortical development. It has been suggested that reelin secreted by Cajal-Retzius cells in the marginal zone could provide either a stop or an attractant signal for migratory neurons expressing reelin receptors, but the proposed models fail to explain recent experimental findings. Here we provide evidence that the reelin receptor machinery, including the lipoprotein receptors ApoER2 and VLDLR along with the cytoplasmic adaptor protein Dab1, is located in radial glia precursors whose processes span the entire cortical wall from the ventricular zone to the pial surface. Moreover, in reeler mice, defective in reelin, decreased levels of Dab1 in the ventricular zone correspond to an accumulation of the protein in radial end-feet beneath the pia matter. Our results support that neural stem cells receive a functional reelin signal. They are also consistent with a working model of reelin action, according to which reelin signaling on the newborn neuron-inherited radial process regulates perikaryal translocation and positioning.     

Postnatal ethanol exposure blunts upregulation of GABAA receptor currents in Purkinje neurons.

Recently, we found that early postnatal ethanol exposure inhibits the maturation of GABAA receptors (GABAARs) in developing medial septum/diagonal band (MS/DB) neurons, suggesting that these receptors may represent a target for ethanol related to fetal alcohol syndrome (FAS). To determine whether GABAARs on other neurons are also sensitive to a postnatal ethanol insult, postnatal day (PD) 4-9, rat pups were artificially reared and exposed to ethanol (4.5 g kg-1 day-1, 10.2% v/v). The pharmacological profile of acutely dissociated cerebellar Purkinje cell GABAARs from untreated, artificially reared controls and ethanol-treated animals was examined with conventional whole-cell patch clamp recordings during PD 12-16 (juveniles) and PD 25-35 (young adults). For untreated animals, GABA (0.3-100 microM) consistently induced inward Cl- currents in a concentration-dependent manner showing an age-related increase in maximum response without change in EC50 or slope value. Acute ethanol (100 mM) consistently inhibited 3 microM GABA currents (10-20%); positive modulators, pentobarbital (10 microM), midazolam (1 microM) and loreclezole (10 microM), consistently potentiated; the negative modulator, Zn2+ (30 microM), inhibited GABA currents across both juvenile and young adult groups. Loreclezole potentiation increased while Zn2+ inhibition decreased with age in untreated Purkinje neurons. Postnatal ethanol exposure (PD 4-9) decreased GABAAR maximum current density in young adult Purkinje cells but not in juvenile neurons. However, sensitivity to allosteric modulators did not change after ethanol. These data are consistent with the hypothesis that postnatal ethanol exposure during the brain growth spurt can disturb GABAAR development across the brain, although the mechanism(s) underlying this action remains to be determined.     

A genetic interaction between the APP and Dab1 genes influences brain development

The Dab1 docking protein is required for the proper organization of brain laminae and for a signal transduction pathway initiated by Reelin binding to the ApoER2 and VLDLR receptors on the cell surface of neurons. Dab1 physically interacts with APP; however, it is not known whether the APP gene influences Dab1 function. Here we demonstrate a genetic interaction between Dab1 and APP. Dab1-hypomorphic animals have neuronal ectopias in the neocortex and reduced cerebellar volume, possibly a consequence of Purkinje cell misplacement. These phenotypes are exacerbated in transgenic animals overexpressing a mutant form of APP, APP(swe), which is characterized by increased processing at the beta-secretase site. The Dab1-hypomorphic phenotype is improved in the cerebellum of animals that are deficient for APP. Together this suggests that APP expression constrains the consequences of Dab1 activity during brain development.     

Functional role of lipoprotein receptors in Alzheimer's disease

The LDL receptor gene family constitutes a class of structurally closely related cell surface receptors fulfilling diverse functions in different organs, tissues, and cell types. The LDL receptor is the prototype of this family, which also includes the VLDLR, ApoER2/LRP8, LRP1 and LRP1B, as well as Megalin/GP330, SorLA/LR11, LRP5, LRP6 and MEGF7. Recently several lines of evidence have positioned the LDL receptor gene family as one of the key players in Alzheimer's disease (AD) research. Initially this receptor family was of high interest due to its key function in cholesterol/apolipoprotein E (ApoE) uptake, with the epsilon4 allele of ApoE as the strongest genetic risk factor for late-onset AD. It has been established that the cholesterol metabolism of the cell has a strong impact on the production of Abeta, the major component of the plaques found in the brain of AD-patients. The original report that soluble amyloid precursor protein (APP) containing the kunitz proteinase inhibitor (KPI) domain might act as a ligand for LRP1 led to a complex investigation of the interaction of both proteins and their potential function in AD development. Meanwhile, it has been demonstrated that LRP1 might bind to APP independent of the KPI domain in APP. This APP - LRP1 interaction is facilitated through a trimeric complex of APP-FE65-LRP1, which has a functional role in APP processing. Along with LRP1, APP is transported from the early secretory compartments to the cell surface and subsequently internalised into the endosomal / lysosomal compartments. Recent investigations indicate that ApoER2 and SorLA fulfil a similar role in shifting APP localisation in the cell, which affects APP processing and the production of the APP derived amyloid beta-peptide (Abeta). In addition to the effect of lipoprotein receptors on APP processing and Abeta production, LRP1 has been shown to bind Abeta directly or indirectly through Abeta-lactoferrin, Abeta-alpha2M and Abeta-ApoE complexes in vitro and in vivo. Based on these observations two LRP1 mediated clearance mechanisms of Abeta are proposed to play a crucial role in the prevention of AD: either Abeta-uptake into a cell with its subsequent degradation or its transport out of the brain over the blood brain barrier into the periphery. Following this export Abeta is degraded in the liver, where LRP1 potentially conducts the removal of Abeta from the blood stream. Although the involvement of LDLR family members in AD is not yet fully understood it becomes clear that they can directly affect APP production, Abeta-clearance and Abeta-transport over the blood brain barrier.     

Developmental expression of NMDA and AMPA receptor subunits in vestibular nuclear neurons that encode gravity-related horizontal orientations

We examined the expression profile of subunits of ionotropic glutamate receptors [N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionate (AMPA)] during postnatal development of connectivity in the rat vestibular nucleus. Vestibular nuclear neurons were functionally activated by constant velocity off-vertical axis rotation, a strategy to stimulate otolith organs in the inner ear. These neurons indicated Fos expression as a result. By immunodetection for Fos, otolith-related neurons that expressed NMDA/AMPA receptor subunits were identified as early as P7, and these neurons were found to increase progressively up to adulthood. Although there was developmental invariance in the percentage of Fos-immunoreactive neurons expressing the NR1, NR2A, GluR1, or GluR2/3 subunits, those expressing the NR2B subunit decreased from P14 onward, and those expressing the GluR4 subunit decreased in adults. These double-immunohistochemical data were corroborated by combined immuno-/hybridization histochemical data obtained from Fos-immunoreactive neurons expressing NR2B mRNA or GluR4 mRNA. The staining of both NR2B and GluR4 in the cytoplasm of these neurons decreased upon maturation. The percentage of Fos-immunoreactive neurons expressing the other ionotropic glutamate receptor subunits (viz. NR1, NR2A, GluR1, and GluR2/3) remained relatively constant throughout postnatal maturation. Triple immunofluorescence further demonstrated coexpression of NR1 and NR2 subunits in Fos-immunoreactive neurons. Coexpression of NR1 subunit with each of the GluR subunits was also observed among the Fos-immunoreactive neurons. Taken together, the different expression profiles of ionotropic glutamate receptor subunits constitute the histological basis for glutamatergic neurotransmission in the maturation of central vestibular connectivity for the coding of gravity-related horizontal head movements.