Reelin Secreted by GABAergic Neurons Regulates Glutamate Receptor Homeostasis

BACKGROUND: Reelin is a large secreted protein of the extracellular matrix that has been proposed to participate to the etiology of schizophrenia. During development, reelin is crucial for the correct cytoarchitecture of laminated brain structures and is produced by a subset of neurons named Cajal-Retzius. After birth, most of these cells degenerate and reelin expression persists in postnatal and adult brain. The phenotype of neurons that bind secreted reelin and whether the continuous secretion of reelin is required for physiological functions at postnatal stages remain unknown. METHODOLOGY/PRINCIPAL FINDINGS: Combining immunocytochemical and pharmacological approaches, we first report that two distinct patterns of reelin expression are present in cultured hippocampal neurons. We show that in hippocampal cultures, reelin is secreted by GABAergic neurons displaying an intense reelin immunoreactivity (IR). We demonstrate that secreted reelin binds to receptors of the lipoprotein family on neurons with a punctate reelin IR. Secondly, using calcium imaging techniques, we examined the physiological consequences of reelin secretion blockade. Blocking protein secretion rapidly and reversibly changes the subunit composition of N-methyl-D-aspartate glutamate receptors (NMDARs) to a predominance of NR2B-containing NMDARs. Addition of recombinant or endogenously secreted reelin rescues the effects of protein secretion blockade and reverts the fraction of NR2B-containing NMDARs to control levels. Therefore, the continuous secretion of reelin is necessary to control the subunit composition of NMDARs in hippocampal neurons. CONCLUSIONS/SIGNIFICANCE: Our data show that the heterogeneity of reelin immunoreactivity correlates with distinct functional populations: neurons synthesizing and secreting reelin and/or neurons binding reelin. Furthermore, we show that continuous reelin secretion is a strict requirement to maintain the composition of NMDARs. We propose that reelin is a trans-neuronal messenger secreted by GABAergic neurons that regulates NMDARs homeostasis in postnatal hippocampus. Defects in reelin secretion could play a major role in the development of neuropsychiatric disorders, particularly those associated with deregulation of NMDARs such as schizophrenia

HIV Testing of At Risk Patients in a Large Integrated Health Care System

OBJECTIVE: Early identification of HIV infection is critical for patients to receive life-prolonging treatment and risk-reduction counseling. Understanding HIV screening practices and barriers to HIV testing is an important prelude to designing successful HIV screening programs. Our objective was to evaluate current practice patterns for identification of HIV. METHODS: We used a retrospective cohort analysis of 13,991 at-risk patients seen at 4 large Department of Veterans Affairs (VA) health-care systems. We also reviewed 1,100 medical records of tested patients. We assessed HIV testing rates among at-risk patients, the rationale for HIV testing, and predictors of HIV testing and of HIV infection. RESULTS: Of the 13,991 patients at risk for HIV, only 36% had been HIV-tested. The prevalence of HIV ranged from 1% to 20% among tested patients at the 4 sites. Approximately 90% of patients who were tested had a documented reason for testing. CONCLUSION: One-half to two-thirds of patients at risk for HIV had not been tested within our selected VA sites. Among tested patients, the rationale for HIV testing was well documented. Further testing of at-risk patients could clearly benefit patients who have unidentified HIV infection by providing earlier access to life-prolonging therapy

Fragile X Related Protein 1 Clusters with Ribosomes and Messenger RNAs at a Subset of Dendritic Spines in the Mouse Hippocampus

The formation and storage of memories in neuronal networks relies on new protein synthesis, which can occur locally at synapses using translational machinery present in dendrites and at spines. These new proteins support long-lasting changes in synapse strength and size in response to high levels of synaptic activity. To ensure that proteins are made at the appropriate time and location to enable these synaptic changes, messenger RNA (mRNA) translation is tightly controlled by dendritic RNA-binding proteins. Fragile X Related Protein 1 (FXR1P) is an RNA-binding protein with high homology to Fragile X Mental Retardation Protein (FMRP) and is known to repress and activate mRNA translation in non-neuronal cells. However, unlike FMRP, very little is known about the role of FXR1P in the central nervous system. To understand if FXR1P is positioned to regulate local mRNA translation in dendrites and at synapses, we investigated the expression and targeting of FXR1P in developing hippocampal neurons in vivo and in vitro. We found that FXR1P was highly expressed during hippocampal development and co-localized with ribosomes and mRNAs in the dendrite and at a subset of spines in mouse hippocampal neurons. Our data indicate that FXR1P is properly positioned to control local protein synthesis in the dendrite and at synapses in the central nervous system

Involvement of the Glycogen Synthase Kinase-3 Signaling Pathway in TBI Pathology and Neurocognitive Outcome

BACKGROUND: Traumatic brain injury (TBI) sets in motion cascades of biochemical changes that result in delayed cell death and altered neuronal architecture. Studies have demonstrated that inhibition of glycogen synthase kinase-3 (GSK-3) effectively reduces apoptosis following a number of stimuli. The Wnt family of proteins, and growth factors are two major factors that regulate GSK-3 activity. In the absence of stimuli, GSK-3 is constitutively active and is complexed with Axin, adenomatous polyposis coli (APC), and casein kinase Iα (CK1α) and phosphorylates ß-Catenin leading to its degradation. Binding of Wnt to Frizzled receptors causes the translocation of GSK-3 to the plasma membrane, where it phosphorylates and inactivates the Frizzled co-receptor lipoprotein-related protein 6 (LRP6). Furthermore, the translocation of GSK-3 reduces ß-Catenin phosphorylation and degradation, leading to ß-Catenin accumulation and gene expression. Growth factors activate Akt, which in turn inhibits GSK-3 activity by direct phosphorylation, leading to a reduction in apoptosis. METHODOLOGY/PRINCIPAL FINDINGS: Using a rodent model, we found that TBI caused a rapid, but transient, increase in LRP6 phosphorylation that is followed by a modest decrease in ß-Catenin phosphorylation. Phospho-GSK-3β immunoreactivity was found to increase three days post injury, a time point at which increased Akt activity following TBI has been observed. Lithium influences several neurochemical cascades, including inhibiting GSK-3. When the efficacy of daily lithium was assessed, reduced hippocampal neuronal cell loss and learning and memory improvements were observed. These influences were partially mimicked by administration of the GSK-3-selective inhibitor SB-216763, as this drug resulted in improved motor function, but only a modest improvement in memory retention and no overt neuroprotection. CONCLUSION/SIGNIFICANCE: Taken together, our findings suggest that selective inhibition of GSK-3 may offer partial cognitive improvement. As a broad spectrum inhibitor of GSK-3, lithium offers neuroprotection and robust cognitive improvement, supporting its clinical testing as a treatment for TBI

Newly Developed Mg2+–Selective Fluorescent Probe Enables Visualization of Mg2+ Dynamics in Mitochondria

Mg2+ plays important roles in numerous cellular functions. Mitochondria take part in intracellular Mg2+ regulation and the Mg2+ concentration in mitochondria affects the synthesis of ATP. However, there are few methods to observe Mg2+ in mitochondria in intact cells. Here, we have developed a novel Mg2+–selective fluorescent probe, KMG-301, that is functional in mitochondria. This probe changes its fluorescence properties solely depending on the Mg2+ concentration in mitochondria under physiologically normal conditions. Simultaneous measurements using this probe together with a probe for cytosolic Mg2+, KMG-104, enabled us to compare the dynamics of Mg2+ in the cytosol and in mitochondria. With this method, carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP)–induced Mg2+ mobilization from mitochondria to the cytosol was visualized. Although a FCCP–induced decrease in the Mg2+ concentration in mitochondria and an increase in the cytosol were observed both in differentiated PC12 cells and in hippocampal neurons, the time-courses of concentration changes varied with cell type. Moreover, the relationship between mitochondrial Mg2+ and Parkinson's disease was analyzed in a cellular model of Parkinson's disease by using the 1-methyl-4-phenylpyridinium ion (MPP+). A gradual decrease in the Mg2+ concentration in mitochondria was observed in response to MPP+ in differentiated PC12 cells. These results indicate that KMG-301 is useful for investigating Mg2+ dynamics in mitochondria. All animal procedures to obtain neurons from Wistar rats were approved by the ethical committee of Keio University (permit number is 09106-(1))

Inhibition of Soluble Tumor Necrosis Factor Ameliorates Synaptic Alterations and Ca2+ Dysregulation in Aged Rats

The role of tumor necrosis factor α (TNF) in neural function has been investigated extensively in several neurodegenerative conditions, but rarely in brain aging, where cognitive and physiologic changes are milder and more variable. Here, we show that protein levels for TNF receptor 1 (TNFR1) are significantly elevated in the hippocampus relative to TNF receptor 2 (TNFR2) in aged (22 months) but not young adult (6 months) Fischer 344 rats. To determine if altered TNF/TNFR1 interactions contribute to key brain aging biomarkers, aged rats received chronic (4–6 week) intracranial infusions of XPro1595: a soluble dominant negative TNF that preferentially inhibits TNFR1 signaling. Aged rats treated with XPro1595 showed improved Morris Water Maze performance, reduced microglial activation, reduced susceptibility to hippocampal long-term depression, increased protein levels for the GluR1 type glutamate receptor, and lower L-type voltage sensitive Ca2+ channel (VSCC) activity in hippocampal CA1 neurons. The results suggest that diverse functional changes associated with brain aging may arise, in part, from selective alterations in TNF signaling