scholarly journals Multiplex expression cloning of blood-brain barrier membrane proteins

PROTEOMICS ◽  
2009 ◽  
Vol 9 (4) ◽  
pp. 1099-1108 ◽  
Author(s):  
Nitin Agarwal ◽  
Eric V. Shusta
Keyword(s):  
Membrane Proteins ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Expression Cloning ◽  
Barrier Membrane ◽  
Blood Brain

scholarly journals Novel Molecular Biomarkers at the Blood-Brain Barrier in ALS

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Danijela Bataveljic ◽  
Milena Milosevic ◽  
Lidija Radenovic ◽  
Pavle Andjus
Keyword(s):  
Membrane Proteins ◽  
Potassium Channel ◽  
Blood Brain Barrier ◽  
Disease Onset ◽  
Water Channel ◽  
Aquaporin 4 ◽  
Brain Barrier ◽  
Inwardly Rectifying Potassium Channel ◽  
Blood Brain ◽  
Inwardly Rectifying

Recently neuroinflammation has gained a particular focus as a key mechanism of ALS. Several studiesin vivoas well asin vitrohave nominated immunoglobulin G (IgG) isolated from ALS patients as an active contributor to disease onset and progression. We have shown that ALS IgG affects astroglial Ca2+excitability and induces downstream activation of phosphatidylinositol 3-kinase. These studies were hampered by a lack of knowledge of the pathway of entry of immune factors in the CNS. Our MRI data revealed the blood-brain barrier BBB leakage and T cell infiltration into brain parenchyma in ALS G93A rats. Since astrocyte ensheathes blood vessel wall contributing to BBB stability and plays an important role in ALS pathogenesis, we have studied astrocytic membrane proteins water channel aquaporin-4 and the inwardly rectifying potassium channel. In this review, we will summarize data related to BBB disruption with particular emphasis on impaired function of astrocytes in ALS. We will discuss implication of membrane proteins expressed on astrocytic endfeet, aquaporin-4, and inwardly rectifying potassium channel in the pathology of ALS. In addition to ALS-specific IgGs, these membrane proteins are proposed as novel biomarkers of the disease.

Neurothelin: molecular characteristics and developmental regulation in the chick CNS

Development ◽  
1991 ◽  
Vol 113 (1) ◽  
pp. 129-140
Author(s):  
B. Schlosshauer
Keyword(s):  
Endothelial Cells ◽  
Membrane Proteins ◽  
Molecular Mass ◽  
Blood Brain Barrier ◽  
Adult Brain ◽  
Brain Barrier ◽  
Relative Molecular Mass ◽  
Brain Neurons ◽  
Blood Brain

Neurothelin has recently been identified as a cell surface protein specific for chick endothelial cells forming the blood-brain barrier. Neurons of the adult brain are essentially devoid of neurothelin. In contrast, neurons of the chick retina, which lack blood vessels and accessory astrocytes, express neurothelin. Here we demonstrate that during chick brain development initially neurothelin is expressed probably in all neuroblasts. With proceeding cytodifferentiation, such as vascularization and gliogenesis, brain neurons become neurothelin negative. Coincidentally the endothelial cells forming the blood-brain barrier start to synthesize neurothelin. In contrast to brain neurons, in retina neurons, neurothelin expression increases by one order of magnitude during the course of histogenesis. Coculturing of chick retinal cells with purified rat astrocytes in vitro results in reduction of neural neurothelin expression as quantified by ELISA. Conversely, disruption of the glia-neuron interactions by culturing brain neurons as individualized cells in vitro leads to a reexpression of neurothelin. This is consistent with the hypothesis that astrocytes inhibit neurothelin expression in neurons. Biochemical characterization classifies neurothelin as an integral membrane protein. Temperature-induced-detergent phase separation, phospholipase C digestion and sodium carbonate treatment were employed to distinguish between integral membrane proteins, lipid-anchored proteins and peripheral membrane proteins. Two-dimensional gel electrophoresis reveals an isoelectric point of about 6.4 for neurothelin. Polysaccharide analysis by glycosidase digestion and lectin binding indicates that neurothelin is highly glycosylated. The relative molecular mass of glycosylated neurothelin is 41 × 10(3), whereas the peptide backbone is only 25 × 10(3). The very strict spatiotemporal regulation of neurothelin expression in the central nervous system suggests that neurothelin fulfils possibly a crucial function such as transport of low relative molecular mass components that are essential for neuronal metabolism. The proposed biological activity of neurothelin might be specifically affected by some of its distinct biochemical features.

scholarly journals Multiple pathways mediate the sex-peptide-regulated switch in female Drosophila reproductive behaviours

2013 ◽  
Vol 280 (1771) ◽  
pp. 20131938 ◽  
Author(s):  
Irmgard U. Haussmann ◽  
Yash Hemani ◽  
Thilini Wijesekera ◽  
Brigitte Dauwalder ◽  
Matthias Soller
Keyword(s):  
G Protein ◽  
Blood Brain Barrier ◽  
Egg Laying ◽  
Brain Barrier ◽  
G Protein Coupled Receptor ◽  
Sex Peptide ◽  
Barrier Membrane ◽  
Blood Brain ◽  
G Protein Coupled

Male-derived sex-peptide (SP) induces profound changes in the behaviour of Drosophila females, resulting in decreased receptivity to further mating and increased egg laying. SP can mediate the switch in female reproductive behaviours via a G protein-coupled receptor, SPR, in neurons expressing fruitless , doublesex and pickpocket . Whether SPR is the sole receptor and whether SP induces the postmating switch in a single pathway has not, to our knowledge been tested. Here we report that the SP response can be induced in the absence of SPR when SP is ectopically expressed in neurons or when SP, transferred by mating, can access neurons through a leaky blood brain barrier. Membrane-tethered SP can induce oviposition via doublesex , but not fruitless and pickpocket neurons in SPR mutant females. Although pickpocket and doublesex neurons rely on G(o) signalling to reduce receptivity and induce oviposition, G(o) signalling in fruitless neurons is required only to induce oviposition, but not to reduce receptivity. Our results show that SP's action in reducing receptivity and inducing oviposition can be separated in fruitless and doublesex neurons. Hence, the SP-induced postmating switch incorporates shared, but also distinct circuitry of fruitless , doublesex and pickpocket neurons and additional receptors.

scholarly journals Cloned Blood–Brain Barrier Adenosine Transporter is Identical to the Rat Concentrative Na+ Nucleoside Cotransporter CNT2

2001 ◽  
Vol 21 (8) ◽  
pp. 929-936 ◽  
Author(s):  
Jian Yi Li ◽  
Ruben J. Boado ◽  
William M. Pardridge
Keyword(s):  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Hill Coefficient ◽  
Expression Cloning ◽  
Nucleoside Transporter ◽  
Blood Brain ◽  
Adenosine Uptake ◽  
Sodium Dependent ◽  
Adenosine Transport

Adenosine transport into brain is regulated by the activity of the adenosine transporter located at the brain capillary endothelial wall, which forms the blood–brain barrier (BBB) in vivo. To facilitate cloning of BBB adenosine transporters, poly A+ RNA was purified from isolated rat brain capillaries for production of a rat BBB cDNA library in the pSPORT vector. The cloned RNA (cRNA) generated from in vitro transcription of this library was injected into frog oocytes followed by measurement of [3H]-adenosine uptake. After dilutional cloning, a full-length, 2905-nucleotide adenosine transporter cDNA, designated clone A-11, was isolated. The A-11 clone yielded [3H]-adenosine flux ratios of 400 to 500 after injection of cRNA in oocytes. The adenosine uptake was sodium-dependent and insensitive to inhibition by S-(4-nitrobenzyl)-6-thioinosine (NBTI). The Km and Vmax of adenosine transport in the cRNA-injected oocytes were 23.1 ± 3.7 μmol/L and 10.8 ± 0.9 pmol/oocyte · min. The K0.5 for sodium was 2.4 ± 0.1 mmol/L, with a Hill coefficient (n) of 1.06 ± 0.07. DNA sequence analysis indicated the rat BBB A-11 adenosine cDNA was identical to rat concentrative nucleoside transporter type 2 (CNT2). The adenosine transporter activity of the rat BBB A-11 CNT2 clone is 50-fold more active than previously reported rat CNT2 clones. In summary, these studies describe the expression cloning of CNT2 from a rat BBB library and show that the pattern of sodium dependency and NBTI insensitivity of the cloned CNT2 are identical to patterns of adenosine transport across the BBB in vivo. These results suggest that BBB adenosine transport in vivo is mediated by CNT2, which would make CNT2 one of the few known sodium-dependent cotransporters that mediate substrate transport across the BBB in the blood to brain direction.

scholarly journals Subtractive Expression Cloning Reveals High Expression of CD46 at the Blood-Brain Barrier

2002 ◽  
Vol 61 (7) ◽  
pp. 597-604 ◽  
Author(s):  
Eric V. Shusta ◽  
Chunni Zhu ◽  
Ruben J. Boado ◽  
William M. Pardridge
Keyword(s):  
Blood Brain Barrier ◽  
High Expression ◽  
Brain Barrier ◽  
Expression Cloning ◽  
Blood Brain
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