Short G-rich oligonucleotides as a potential therapeutic for Huntington's Disease

Background  

Huntington's Disease (HD) is an inherited autosomal dominant genetic disorder in which neuronal tissue degenerates. The pathogenesis of the disease appears to center on the development of protein aggregates that arise initially from the misfolding of the mutant HD protein. Mutant huntingtin (Htt) is produced by HD genes that contain an increased number of glutamine codons within the first exon and this expansion leads to the production of a protein that misfolds. Recent studies suggest that mutant Htt can nucleate protein aggregation and interfere with a multitude of normal cellular functions.     

Altered neurological function in mice immunized with early endosome antigen 1

Background  

Autoantibodies directed against the 160 kDa endosome protein early endosome antigen 1 (EEA1) are seen in patients with neurological diseases. To determine if antibodies to EEA1 have a neuropathological effect, mice from three major histocompatability haplotype backgrounds (H2^q, H2^b and H2^d) were immunized with EEA1 (amino acids 82–1411) that was previously shown to contain the target EEA1 epitopes. The mice were then subjected to five neuro-behavioural tests: grid walking, forelimb strength, open field, reaching and rotarod.     

Nuclear localization of Annexin A7 during murine brain development

Background  

Annexin A7 is a member of the annexin protein family, which is characterized by its ability to interact with phospholipids in the presence of Ca²⁺-ions and which is thought to function in Ca²⁺-homeostasis. Results from mutant mice showed altered Ca²⁺-wave propagation in astrocytes. As the appearance and distribution of Annexin A7 during brain development has not been investigated so far, we focused on the distribution of Annexin A7 protein during mouse embryogenesis in the developing central nervous system and in the adult mouse brain.     

Inducible gene inactivation in neurons of the adult mouse forebrain

Background  

The analysis of the role of genes in important brain functions like learning, memory and synaptic plasticity requires gene inactivation at the adult stage to exclude developmental effects, adaptive changes or even lethality. In order to achieve temporally controlled somatic mutagenesis, the Cre/loxP-recombination system has been complemented with the tamoxifen-inducible fusion protein consisting of Cre recombinase and the mutated ligand binding domain of the human estrogen receptor (CreER^T2). To induce recombination of conditional alleles in neurons of the adult forebrain, we generated a bacterial artificial chromosome-derived transgene expressing the CreER^T2 fusion protein under control of the regulatory elements of the CaMKIIα gene (CaMKCreER^T2 transgene).     

Upregulation of synaptotagmin IV inhibits transmitter release in PC12 cells with targeted synaptotagmin I knockdown

Background  

The function of synaptotagmins (syt) in Ca²⁺-dependent transmitter release has been attributed primarily to Ca²⁺-dependent isoforms such as syt I. Recently, syt IV, an inducible Ca²⁺-independent isoform has been implicated in transmitter release. We postulated that the effects of syt IV on transmitter release are dependent on the expression of syt I.     

Ontogeny of ATP hydrolysis and isoform expression of the Plasma Membrane Ca2+-ATPase in mouse brain

Background  

Plasma membrane Ca²⁺-ATPases (PMCAs) are high affinity Ca²⁺ transporters actively involved in intracellular Ca²⁺ homeostasis. Considering the critical role of Ca²⁺ signalling in neuronal development and plasticity, we have analyzed PMCA-mediated Ca²⁺-ATPase activity and PMCA-isoform content in membranes from mouse cortex, hippocampus and cerebellum during postnatal development.     

An adeno-associated viral vector transduces the rat hypothalamus and amygdala more efficient than a lentiviral vector

Background  

This study compared the transduction efficiencies of an adeno-associated viral (AAV) vector, which was pseudotyped with an AAV1 capsid and encoded the green fluorescent protein (GFP), with a lentiviral (LV) vector, which was pseudotyped with a VSV-G envelop and encoded the discosoma red fluorescent protein (dsRed), to investigate which viral vector transduced the lateral hypothalamus or the amygdala more efficiently. The LV-dsRed and AAV1-GFP vector were mixed and injected into the lateral hypothalamus or into the amygdala of adult rats. The titers that were injected were 1 × 10⁸ or 1 × 10⁹ genomic copies of AAV1-GFP and 1 × 10⁵ transducing units of LV-dsRed.     

Effects of NR1 splicing on NR1/NR3B-type excitatory glycine receptors

Background  

N-methyl-D-aspartate receptors (NMDARs) are the most complex of ionotropic glutamate receptors (iGluRs). Subunits of this subfamily assemble into heteromers, which – depending on the subunit combination – may display very different pharmacological and electrophysiological properties. The least studied members of the NMDAR family, the NR3 subunits, have been reported to assemble with NR1 to form excitatory glycine receptors in heterologous expression systems. The heterogeneity of NMDARs in vivo is in part conferred to the receptors by splicing of the NR1 subunit, especially with regard to proton sensitivity.     

Tyrosine phosphatases such as SHP-2 act in a balance with Src-family kinases in stabilization of postsynaptic clusters of acetylcholine receptors

Background  

Development of neural networks requires that synapses are formed, eliminated and stabilized. At the neuromuscular junction (NMJ), agrin/MuSK signaling, by triggering downstream pathways, causes clustering and phosphorylation of postsynaptic acetylcholine receptors (AChRs). Postnatally, AChR aggregates are stabilized by molecular pathways that are poorly characterized. Gain or loss of function of Src-family kinases (SFKs) disassembles AChR clusters at adult NMJs in vivo, whereas AChR aggregates disperse rapidly upon withdrawal of agrin from cultured src ^-/-;fyn ^-/- myotubes. This suggests that a balance between protein tyrosine phosphatases (PTPs) and protein tyrosine kinases (PTKs) such as those of the Src-family may be essential in stabilizing clusters of AChRs.     

Elevated intracellular chloride level in albino visual cortex neurons is mediated by Na-K-Cl co-transporter

Background  

During development the switch from a depolarizing to a hyperpolarizing action of GABA is a consequence of a decrease of the Na⁺-K⁺-2Cl^- co-transporter (NKCC1, Cl^--uptake) and increase of the K⁺-Cl^- co-transporter (KCC2, Cl^--extrusion) expression. However albino visual cortex neurons don't show a corresponding decrease in intracellular chloride concentration during development of the visual system as compared to pigmented animals.     

Prenatal cocaine exposure alters alpha2 receptor expression in adolescent rats

Background  

Prenatal cocaine exposure produces attentional deficits which to persist through early childhood. Given the role of norepinephrine (NE) in attentional processes, we examined the forebrain NE systems from prenatal cocaine exposed rats. Cocaine was administered during pregnancy via the clinically relevant intravenous route of administration. Specifically, we measured α₂-adrenergic receptor (α₂-AR) density in adolescent (35-days-old) rats, using [³H]RX821002 (5 nM).     

Serotonin-mediated modulation of Na<Superscript>+</Superscript>/K<Superscript>+</Superscript> pump current in rat hippocampal CA1 pyramidal neurons

Background  

The aim of this study was to investigate whether serotonin (5-hydroxytryptamine, 5-HT) can modulate Na⁺/K⁺ pump in rat hippocampal CA1 pyramidal neurons.     

A new approach for the large-scale generation of mature dendritic cells from adherent PBMC using roller bottle technology

Background  

Human monocyte-derived DC (mDC) loaded with peptides, protein, tumor cell lysates, or tumor cell RNA, are being tested as vaccines against multiple human malignancies and viral infection with great promise. One of the factors that has limited more widespread use of these vaccines is the need to generate mDC in large scale. Current methods for the large-scale cultivation of mDC in static culture vessels are labor- and time- intensive, and also require many culture vessels. Here, we describe a new method for the large-scale generation of human mDC from human PBMC from leukopheresis or buffy coat products using roller bottles, never attempted before for mDC generation. We have tested this technology using 850 cm² roller bottles compared to conventional T-175 flat-bottom static culture flasks.     

Equine estrogens differentially inhibit DNA fragmentation induced by glutamate in neuronal cells by modulation of regulatory proteins involved in programmed cell death

Background  

Recent data indicate that excitotoxicity of high levels of neurotransmitter glutamate may be mediated via programmed cell death (apoptosis) and that it can be prevented in HT22 mouse hippocampal cells by various equine estrogens with Δ⁸,17β-estradiol (Δ⁸,17β-E₂) being the most potent. In order to delineate the mechanism(s), glutamate-induced cell death of HT22 cells was assessed by measuring (a) DNA fragmentation in the presence or absence of 11 equine estrogens (components of the drug CEE); (b) cell death and (c) levels of anti-apoptotic (Bcl-2) and proapoptotic (Bax) proteins in the presence or absence of two equine estrogens, Δ⁸,17β-E₂ and 17β-estradiol (17β-E₂) by LDH release assay and Western blot analysis respectively.     

Abnormalities of cell packing density and dendritic complexity in the MeCP2 A140V mouse model of Rett syndrome/X-linked mental retardation

Background  

Rett syndrome (RTT), a common cause of mental retardation in girls, is associated with mutations in the MECP2 gene. Most human cases of MECP2 mutation in girls result in classical or variant forms of RTT. When these same mutations occur in males, they often present as severe neonatal encephalopathy. However, some MECP2 mutations can also lead to diseases characterized as mental retardation syndromes, particularly in boys. One of these mutations, A140V, is a common, recurring missense mutation accounting for about 0.6% of all MeCP2 mutations and ranking 21^st by frequency. It has been described in familial X-linked mental retardation (XLMR), PPM- X syndrome (Parkinsonism, Pyramidal signs, Macroorchidism, X-linked mental retardation) and in other neuropsychiatric syndromes. Interestingly, this mutation has been reported to preserve the methyl-CpG binding function of the MeCP2 protein while compromising its ability to bind to the mental retardation associated protein ATRX.     

Calcium-sensitive regulation of monoamine oxidase-A contributes to the production of peroxyradicals in hippocampal cultures: implications for Alzheimer disease-related pathology

Background  

Calcium (Ca²⁺) has recently been shown to selectively increase the activity of monoamine oxidase-A (MAO-A), a mitochondria-bound enzyme that generates peroxyradicals as a natural by-product of the deamination of neurotransmitters such as serotonin. It has also been suggested that increased intracellular free Ca²⁺ levels as well as MAO-A may be contributing to the oxidative stress associated with Alzheimer disease (AD).     

Synthesis of novel 4H-pyrimido[1,6-a]pyrimidines via a one-pot three-component condensation

Abstract  

Highly substituted novel 4H-pyrimido[1,6-a] pyrimidines were prepared by a trifluoromethanesulfonic acid catalyzed one-pot three-component condensation of 4-aminopyrimidines, aldehydes, and β-ketoesters. A preliminary feasibility study was undertaken on these compounds, to assess the potential production of a library of further diversified compounds by nucleophilic replacement of Cl (R¹) or by reaction of electrophiles with the NH₂ (R²) group.     

Functional distribution of Ca<Superscript>2+</Superscript>-coupled P2 purinergic receptors among adrenergic and noradrenergic bovine adrenal chromaffin cells

Background  

Adrenal chromaffin cells mediate acute responses to stress through the release of epinephrine. Chromaffin cell function is regulated by several receptors, present both in adrenergic (AD) and noradrenergic (NA) cells. Extracellular ATP exerts excitatory and inhibitory actions on chromaffin cells via ionotropic (P2X) and metabotropic (P2Y) receptors. We have taken advantage of the actions of the purinergic agonists ATP and UTP on cytosolic free Ca²⁺ concentration ([Ca²⁺]_i) to determine whether P2X and P2Y receptors might be asymmetrically distributed among AD and NA chromaffin cells.     

Protection of cortical cells by equine estrogens against glutamate-induced excitotoxicity is mediated through a calcium independent mechanism

Background  

High concentrations of glutamate can accumulate in the brain and may be involved in the pathogenesis of neurodegenerative disorders such as Alzheimer's disease. This form of neurotoxicity involves changes in the regulation of cellular calcium (Ca²⁺) and generation of free radicals such as peroxynitrite (ONOO^-). Estrogen may protect against glutamate-induced cell death by reducing the excitotoxic Ca²⁺ influx associated with glutamate excitotoxicity. In this study, the inhibition of N-methyl-D-aspartate (NMDA) receptor and nitric oxide synthase (NOS) along with the effect of 17β-estradiol (17β-E₂) and a more potent antioxidant Δ⁸, 17β-estradiol (Δ⁸, 17β-E₂) on cell viability and intracellular Ca²⁺ ([Ca²⁺]_i), following treatment of rat cortical cells with glutamate, was investigated.