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Sample records for isolated brain capillaries

  1. Developmental changes in metabolism and transport properties of capillaries isolated from rat brain.

    PubMed Central

    Betz, A L; Goldstein, G W

    1981-01-01

    1. Capillaries were isolated from the brains of 1- to 45-day-old rats in order to study the development of metabolic and transport aspects of the blood-brain barrier. 2. The hydroxyproline content of capillary hydrolysates increased nearly threefold between 5 and 45 days of age. This finding is consistent with histological studies showing thickening of capillary basement membrane during development. 3. The activities of L-DOPA decarboxylase and monoamine oxidase were greatest in capillaries from 10-day-old rat brain. Thus, the metabolic blood-brain barrier for amine precursors is present during early development. 4. Capillaries from all ages were able to metabolize glucose, beta-hydroxybutyrate and palmitate. The rate of glucose oxidation more than doubled between 21 and 30 days of age but subsequently decreased. In contrast, beta-hydroxybutyrate and palmitate oxidation increased throughout development. These data suggest a sparing effect by alternate fuels on glucose metabolism. 5. Capillary glucose uptake was similar at 10 and 30 days of age and activity of the ouabain-sensitive K+ pump (measured using 86Rb+) was relatively constant at all ages. In contrast, Na+-dependent neutral amino acid transport was not present until after 21 days of age. Since this transport system may be responsible for the active efflux of neutral amino acids from brain to blood, it is likely that this process does not occur at the immature blood-brain barrier. 6. We conclude that various aspects of brain capillary functions show distinct developmental patterns which may be related to changes in blood-brain barrier permeability during development. PMID:7264999

  2. Assessment of Ex Vivo Transport Function in Isolated Rodent Brain Capillaries.

    PubMed

    Chan, Gary N Y; Cannon, Ronald E

    2017-03-17

    The blood-brain barrier plays an important role in neuroprotection; however, it can be a major obstacle for drug delivery to the brain. This barrier primarily resides in the brain capillaries and functions as an interface between the brain and peripheral blood circulation. Several anatomical and biochemical elements of the blood-brain barrier are essential to regulate the permeability of nutrients, ions, hormones, toxic metabolites, and xenobiotics into and out of the brain. In particular, high expression of ATP-driven efflux transporters at the blood-brain barrier is a major obstacle in the delivery of CNS pharmacotherapeutics to the brain. The complete understanding of these elements can offer insights on how to modulate barrier functions for neuroprotection against CNS drug toxicity and to enhance drug delivery to the brain. In the literature, preclinical models of the blood-brain barrier are widely utilized to predict drug pharmacokinetics and pharmacodynamics properties in the brain. In addition, these models are essential tools to investigate cellular mechanisms and novel interventions that alter barrier function and permeability. This unit presents procedures to isolate fresh and viable rodent brain capillaries for the assessment of ex vivo transport activity at the blood-brain barrier. © 2017 by John Wiley & Sons, Inc.

  3. Alkamides from Echinacea angustifolia Interact with P-glycoprotein of primary brain capillary endothelial cells isolated from porcine brain blood vessels.

    PubMed

    Mahringer, Anne; Ardjomand-Woelkart, Karin; Bauer, Rudolf; Fricker, Gert; Efferth, Thomas

    2013-03-01

    The blood-brain barrier prevents the passage of toxic compounds from blood circulation into brain tissue. Unfortunately, drugs for the treatment of neurodegenerative diseases, brain tumors, and other diseases also do not cross the blood-brain barrier. In the present investigation, we used isolated porcine brain capillary endothelial cells and a flow cytometric calcein-AM assay to analyze inhibition of P-glycoprotein, a major constituent of the blood-brain barrier. We tested 8 alkamides isolated from Echinacea angustifolia and found that four of them inhibited P-glycoprotein-mediated calcein transport in porcine brain capillary endothelial cells.

  4. Absorptive-mediated endocytosis of cationized albumin and a beta-endorphin-cationized albumin chimeric peptide by isolated brain capillaries. Model system of blood-brain barrier transport

    SciTech Connect

    Kumagai, A.K.; Eisenberg, J.B.; Pardridge, W.M.

    1987-11-05

    Cationized albumin (pI greater than 8), unlike native albumin (pI approximately 4), enters cerebrospinal fluid (CSF) rapidly from blood. This suggests that a specific uptake mechanism for cationized albumin may exist at the brain capillary wall, i.e. the blood-brain barrier. Isolated bovine brain capillaries rapidly bound cationized (/sup 3/H)albumin and approximately 70% of the bound radioactivity was resistant to mild acid wash, which is assumed to represent internalized peptide. Binding was saturable and a Scatchard plot gave a maximal binding capacity (Ro) = 5.5 +/- 0.7 micrograms/mgp (79 +/- 10 pmol/mgp), and a half-saturation constant (KD) = 55 +/- 8 micrograms/ml (0.8 +/- 0.1 microM). The binding of cationized (/sup 3/H)albumin (pI = 8.5-9) was inhibited by protamine, protamine sulfate, and polylysine (molecular weight = 70,000) with a Ki of approximately 3 micrograms/ml for all three proteins. The use of cationized albumin in directed delivery of peptides through the blood-brain barrier was examined by coupling (/sup 3/H)beta-endorphin to unlabeled cationized albumin (pI = 8.5-9) using the bifunctional reagent, N-succinimidyl 3-(2-pyridyldithio)proprionate. The (/sup 3/H)beta-endorphin-cationized albumin chimeric peptide was rapidly bound and endocytosed by isolated bovine brain capillaries, and this was inhibited by unlabeled cationized albumin but not by unconjugated beta-endorphin or native bovine albumin. Cationized albumin provides a new tool for studying absorptive-mediated endocytosis at the brain capillary and may also provide a vehicle for directed drug delivery through the blood-brain barrier.

  5. Identification of Novel Site-Specific Alterations in the Modification Level of Myelin Basic Protein Isolated from Mouse Brain at Different Ages Using Capillary Electrophoresis-Mass Spectrometry.

    PubMed

    Sarg, Bettina; Faserl, Klaus; Lindner, Herbert H

    2017-10-01

    Myelin basic protein (MBP) is a multifunctional protein involved in maintaining the stability and integrity of the myelin sheath by a variety of interactions with membranes and other proteins. MBP is subjected to extensive posttranslational modifications (PTMs) that are known to be crucial for the regulation of these interactions. Here, we report capillary electrophoresis-mass spectrometric (CE-MS) analysis for the separation and identification of MBP peptides that incorporate the same PTM at different sites, creating multiple localization variants, and the ability to analyze challenging modifications such as asparagine and glutamine deamidation, isomerization, and arginine citrullination. Moreover, we observed site-specific alterations in the modification level of MBP purified from brain of mice of different age. In total, we identified 40 modifications at 33 different sites, which include both previously reported and seven novel modifications. The identified modifications include Nα-terminal acetylation, mono- and dimethylation, phosphorylation, oxidation, deamidation, and citrullination. Notably, some new sites of arginine methylation overlap with the sites of citrullination. Our results highlight the need for sensitive and efficient techniques for a comprehensive analysis of PTMs. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Reduction of functional capillary density in human brain after stroke.

    PubMed

    Gjedde, A; Kuwabara, H; Hakim, A M

    1990-05-01

    The blood flow of brain tissue often returns to normal after an ischemic episode. As "luxury" rather than "reactive" reperfusion, this hyperemia is associated with low metabolism. It is not known to what extent the high blood flow accompanies a high, normal, or low density of capillaries. The resolution of this question may indicate whether the functional capillary density is variable and, if so, whether it is coupled to blood flow or metabolism. To answer these questions, we defined functional capillaries as capillaries that transport glucose. We then calculated the density of functional capillaries (Dcap) and the mean time of transit of blood through the capillaries (tcap) from hemodynamic variables obtained in vivo by positron tomography of five patients afflicted by cerebral ischemic stroke. Each patient was studied twice, within 36 h of the insult and 1 week later. We identified nominally "ischemic" regions in the first study as cortical gray matter regions, contiguous with the ischemic focus, in which the magnitude of blood flow did not exceed 20 ml 100 g-1 min-1. In these regions, values of metabolism and functional capillary density were proportionately low compared with normal values obtained in the contralateral hemisphere. The studies revealed a reduction of the functional density of exchange vessels in postischemic brain tissue as soon as 36 h after the insult. In "ischemic" regions, within 36 h of the insult, the net extraction of oxygen was inversely related to the capillary transit time and appeared to be limited mainly by the low functional density of the capillaries. Contrary to expectations, the reduced density persisted, even when more than adequate perfusion of the tissue returned. For these reasons, we concluded that changes of the capillary density were associated with changes of the metabolism of the tissue rather than with blood flow.

  7. Absent recruitment of capillaries in brain tissue recovering from stroke.

    PubMed

    Gjedde, A; Kuwabara, H

    1993-01-01

    The density of perfused capillaries (dCAP), defined as capillaries that transport glucose, as well as the volume fraction of these capillaries in the vascular bed (fCAP), and the mean transit time of blood through the capillaries (tCAP), were calculated from hemodynamic variables obtained in vivo by positron tomography of brains of six patients affected by stroke. Each patient was studied twice, within 38 hrs of the insult, and one week later. 38 ischemic and 38 contralateral mirror regions were compared. The metabolic rate for glucose (CMRglc) was determined on the basis of regional calculations of the lumped constant. No significant change of the lumped constant was observed in any region. In normal regions, no significant differences of any variables existed between the first and second studies. In the infarct regions of the first study, CMRglc and CMRO2 (cerebral metabolic rate for oxygen) were 30-50% of control (deactivation) and CBF (cerebral blood flow), capillary density, and the capillary diffusion capacity for fluorodeoxyglucose (K1) were similarly reduced, although the oxygen/glucose ratio was only 3.75 in the ischemic regions. While fCAP decreased, tCAP doubled. One week after the first study, blood flow returned to normal in the infarct regions despite continued depression of metabolism. Capillary density and diffusion capacity remained low, indicating absent recruitment of nutrition vessels (perfusion capillaries).

  8. Purification and characterization of metabolically active capillaries of the blood-brain barrier.

    PubMed Central

    Dallaire, L; Tremblay, L; Béliveau, R

    1991-01-01

    Microvessels were isolated from bovine and rat cerebral cortex by simple procedures involving mechanical homogenization, differential and density-gradient centrifugation, and chromatography on a column of glass beads. The preparations were composed of short capillaries with a diameter of 1-10 microns. Both purifications were monitored by assaying the activity of the marker enzyme gamma-glutamyl transpeptidase (gamma-GTase). The final bovine and rat preparations were enriched 20- and 14-fold over the homogenate respectively. gamma-GTase activity was measured in different fractions after bovine and rat membranes were solubilized with 0.5% and 0.3% Triton X-100 respectively. Measurement of 5'-nucleotidase and acetylcholinesterase activities indicated very low levels of contamination of the microvessel preparations by glial cells and neurons. The integrity of the capillary membranes was confirmed by the assay of a cytosolic marker enzyme, lactate dehydrogenase. Viability of the microvessels was demonstrated by the presence of detectable levels of adenylates and by tissue respiration induced by glucose and succinate. Comparison of the proteins of homogenized bovine and rat brain cortex with those of purified capillaries separated by SDS/PAGE revealed enrichment of at least three predominant proteins of 14, 16 and 18 kDa in the capillary preparations. It is concluded that these methods allow rapid isolation of small blood vessels of the blood-brain barrier which are suitable for metabolic and structural studies in vitro. Images Fig. 2. Fig. 4. PMID:1712199

  9. Flow distributions and spatial correlations in human brain capillary networks

    NASA Astrophysics Data System (ADS)

    Lorthois, Sylvie; Peyrounette, Myriam; Larue, Anne; Le Borgne, Tanguy

    2015-11-01

    The vascular system of the human brain cortex is composed of a space filling mesh-like capillary network connected upstream and downstream to branched quasi-fractal arterioles and venules. The distribution of blood flow rates in these networks may affect the efficiency of oxygen transfer processes. Here, we investigate the distribution and correlation properties of blood flow velocities from numerical simulations in large 3D human intra-cortical vascular network (10000 segments) obtained from an anatomical database. In each segment, flow is solved from a 1D non-linear model taking account of the complex rheological properties of blood flow in microcirculation to deduce blood pressure, blood flow and red blood cell volume fraction distributions throughout the network. The network structural complexity is found to impart broad and spatially correlated Lagrangian velocity distributions, leading to power law transit time distributions. The origins of this behavior (existence of velocity correlations in capillary networks, influence of the coupling with the feeding arterioles and draining veins, topological disorder, complex blood rheology) are studied by comparison with results obtained in various model capillary networks of controlled disorder. ERC BrainMicroFlow GA615102, ERC ReactiveFronts GA648377.

  10. Diet-induced ketosis increases capillary density without altered blood flow in rat brain.

    PubMed

    Puchowicz, Michelle A; Xu, Kui; Sun, Xiaoyan; Ivy, Andre; Emancipator, Doug; LaManna, Joseph C

    2007-06-01

    It is recognized that ketone bodies, such as R-beta-hydroxybutyrate (beta-HB) and acetoacetate, are energy sources for the brain. As with glucose metabolism, monocarboxylate uptake by the brain is dependent on the function and regulation of its own transporter system. We concurrently investigated ketone body influx, blood flow, and regulation of monocarboxylate transporter (MCT-1) and glucose transporter (GLUT-1) in diet-induced ketotic (KG) rat brain. Regional blood-to-brain beta-HB influx (micromol.g(-1).min(-1)) increased 40-fold with ketosis (4.8 +/- 1.8 plasmabeta-HB; mM) in all regions compared with the nonketotic groups (standard and no-fat diets); there were no changes in regional blood flow. Immunohistochemical staining revealed that GLUT-1 density (number/mm2) in the cortex was significantly elevated (40%) in the ketotic group compared with the standard and no-fat diet groups. MCT-1 was also markedly (3-fold) upregulated in the ketotic group compared with the standard diet group. In the standard diet group, 40% of the brain capillaries stained positive for MCT-1; this amount doubled with the ketotic diet. Western blot analysis of isolated microvessels from ketotic rat brain showed an eightfold increase in GLUT-1 and a threefold increase in MCT-1 compared with the standard diet group. These data suggest that diet-induced ketosis results in increased vascular density at the blood-brain barrier without changes in blood flow. The increase in extraction fraction and capillary density with increased plasma ketone bodies indicates a significant flux of substrates available for brain energy metabolism.

  11. Brain and atrial natriuretic peptides bind to common receptors in brain capillary endothelial cells.

    PubMed

    Gelfand, R A; Frank, H J; Levin, E; Pedram, A

    1991-08-01

    The recent discovery of brain natriuretic peptides (BNP) that stimulates natriuresis, diuresis, and vascular smooth muscle relaxation in a manner similar to that of atrial natriuretic peptide (ANP) suggests the possibility that these endocrine hormones function via some common mechanism. Indirect evidence from several laboratories suggests that BNP and ANP may bind to the same receptors. We examined whether ANP and BNP bind to a common set of receptors in cultured bovine brain capillary endothelial cells and in bovine aortic endothelial cells. Scatchard plot analysis of binding data shows a similar dissociation constant (KD) of approximately 0.3 nM and a maximal binding capacity (Bmax) of 50 fmol/mg protein for both natriuretic peptides in brain capillary cells and 0.6 nM and 80 fmol/mg protein, respectively, in the aortic endothelial cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the affinity cross-linked receptor-ligand complex shows a strongly labeled 65-kDa receptor and a 125-kDa band that is likely to be a receptor of the guanylate cyclase type. ANP and BNP cross compete equally for binding to the two receptors identified on the gels. ANP and BNP also stimulate guanosine 3', 5'-cyclic monophosphate production in these cells, consistent with the presence of a functional guanylate cyclase-linked B receptor. We conclude that ANP and BNP share common receptors in brain capillary and aortic endothelial cells.

  12. Monolithic molecularly imprinted polymeric capillary columns for isolation of aflatoxins.

    PubMed

    Szumski, Michał; Grzywiński, Damian; Prus, Wojciech; Buszewski, Bogusław

    2014-10-17

    Monolithic molecularly imprinted polymers extraction columns have been prepared in fused-silica capillaries by UV or thermal polymerization in a two-step process. First, a poly-(trimethylolpropane trimethacrylate) (polyTRIM) core monolith was synthesized either by UV or thermal polymerization. Then it was grafted with the mixture of methacrylic acid (MAA) as a functional monomer, ethylene dimethacrylate (EDMA) as a cross-linking agent, 5,7-dimethoxycoumarin (DMC) as an aflatoxin-mimicking template, toluene as a porogen solvent and 2,2-azobis-(2-methylpropionitrile) (AIBN) as an initiator of the polymerization reaction. Different thermal condition of the photografting and different concentrations of the grafting mixture were tested during polymerization. The extraction capillary columns were evaluated in the terms of their hydrodynamic and chromatographic properties. Retention coefficients for aflatoxin B1 and DMC were used for assessment of the selectivity and imprinting factor. The obtained results indicate that the temperature of photografting and concentration of the grafting mixture are key parameters that determine the quality of the prepared MIPs. From the MIP columns characterized by the highest permeability the column of the highest imprinting factor was applied for isolation of aflatoxins B1, B2, G1 and G2 from the model aqueous sample followed by on-line chromatographic separation. The process was performed using a micro-MISPE-microLC-LIF system of a novel design, which allowed for detection of the eluates from the sample preparation part as well as from the chromatographic separation. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Oscillation of an isolated liquid plug inside a dry capillary

    NASA Astrophysics Data System (ADS)

    Srinivasan, Vyas; Kumar, Siddhartha; Asfer, Mohammed; Khandekar, Sameer

    2017-06-01

    The present work reports an experimental study on the dynamics of partially wetting isolated liquid plug (DI water), which is made to oscillate inside a square, glass capillary tube (1 mm × 1 mm; 60 mm length). The liquid plug is made to oscillate pneumatically at two different frequencies (0.25 and 0.35 Hz), using a cam-follower mechanism. Bright field imaging is used to visualize the three-phase contact line behavior, while, micro-Particle Imaging Velocimetry (PIV) apparatus is used to discern the nature of flow inside the oscillating liquid plug. During a cycle, due to the partial wetting nature of DI water, the three-phase contact line at the menisci gets pinned at the extreme end of each stroke, where the dynamic apparent contact angle gets drastically altered before the initiation of the next stroke. The difference between the apparent contact angle of the front and rear meniscus are seen to be a function of the oscillating frequency; the difference increasing with increasing frequency. The flow inside the liquid plug reveals unique non-Poiseuille flow features near the meniscus, due to free-slip boundary condition, which leads to formation of distinct vortex pairs behind it. The vortices too change their direction during each stroke of the oscillation, eventually leading to an alternating recirculation pattern inside the plug. The results clearly indicate that improved mathematical models are required for predicting transport parameters in such flows, which are important in engineering systems such as pulsating heat pipes, lab-on-chip devices and PEM fuel cells.

  14. Capillary transit time heterogeneity and flow-metabolism coupling after traumatic brain injury

    PubMed Central

    Østergaard, Leif; Engedal, Thorbjørn S; Aamand, Rasmus; Mikkelsen, Ronni; Iversen, Nina K; Anzabi, Maryam; Næss-Schmidt, Erhard T; Drasbek, Kim R; Bay, Vibeke; Blicher, Jakob U; Tietze, Anna; Mikkelsen, Irene K; Hansen, Brian; Jespersen, Sune N; Juul, Niels; Sørensen, Jens CH; Rasmussen, Mads

    2014-01-01

    Most patients who die after traumatic brain injury (TBI) show evidence of ischemic brain damage. Nevertheless, it has proven difficult to demonstrate cerebral ischemia in TBI patients. After TBI, both global and localized changes in cerebral blood flow (CBF) are observed, depending on the extent of diffuse brain swelling and the size and location of contusions and hematoma. These changes vary considerably over time, with most TBI patients showing reduced CBF during the first 12 hours after injury, then hyperperfusion, and in some patients vasospasms before CBF eventually normalizes. This apparent neurovascular uncoupling has been ascribed to mitochondrial dysfunction, hindered oxygen diffusion into tissue, or microthrombosis. Capillary compression by astrocytic endfeet swelling is observed in biopsies acquired from TBI patients. In animal models, elevated intracranial pressure compresses capillaries, causing redistribution of capillary flows into patterns argued to cause functional shunting of oxygenated blood through the capillary bed. We used a biophysical model of oxygen transport in tissue to examine how capillary flow disturbances may contribute to the profound changes in CBF after TBI. The analysis suggests that elevated capillary transit time heterogeneity can cause critical reductions in oxygen availability in the absence of ‘classic' ischemia. We discuss diagnostic and therapeutic consequences of these predictions. PMID:25052556

  15. Internalization of targeted quantum dots by brain capillary endothelial cells in vivo.

    PubMed

    Paris-Robidas, Sarah; Brouard, Danny; Emond, Vincent; Parent, Martin; Calon, Frédéric

    2016-04-01

    Receptors located on brain capillary endothelial cells forming the blood-brain barrier are the target of most brain drug delivery approaches. Yet, direct subcellular evidence of vectorized transport of nanoformulations into the brain is lacking. To resolve this question, quantum dots were conjugated to monoclonal antibodies (Ri7) targeting the murine transferrin receptor. Specific transferrin receptor-mediated endocytosis of Ri7-quantum dots was first confirmed in N2A and bEnd5 cells. After intravenous injection in mice, Ri7-quantum dots exhibited a fourfold higher volume of distribution in brain tissues, compared to controls. Immunofluorescence analysis showed that Ri7-quantum dots were sequestered throughout the cerebral vasculature 30 min, 1 h, and 4 h post injection, with a decline of signal intensity after 24 h. Transmission electron microscopic studies confirmed that Ri7-quantum dots were massively internalized by brain capillary endothelial cells, averaging 37 ± 4 Ri7-quantum dots/cell 1 h after injection. Most quantum dots within brain capillary endothelial cells were observed in small vesicles (58%), with a smaller proportion detected in tubular structures or in multivesicular bodies. Parenchymal penetration of Ri7-quantum dots was extremely low and comparable to control IgG. Our results show that systemically administered Ri7-quantum dots complexes undergo extensive endocytosis by brain capillary endothelial cells and open the door for novel therapeutic approaches based on brain endothelial cell drug delivery. © The Author(s) 2015.

  16. Aβ42 production in brain capillary endothelial cells after oxygen and glucose deprivation.

    PubMed

    Bulbarelli, Alessandra; Lonati, Elena; Brambilla, Anna; Orlando, Antonina; Cazzaniga, Emanuela; Piazza, Fabrizio; Ferrarese, Carlo; Masserini, Massimo; Sancini, Giulio

    2012-04-01

    Although the diverse triggers of AD are still under debate, the hypothesis of the contribution of cerebrovascular deficiencies has emerged in recent years. Cerebrovascular dysfunction may precede cognitive decline and onset of neurodegeneration. Indeed, the toxic Aβ(42) aggregates constituting senile plaques, one of AD hallmarks, is often detected as amorphous material or fine fibrils in the brain capillary of AD patients. Aβ(42) causing cerebral microangiopathy might originate either from the circulating blood, the vessel wall itself or the brain parenchyma. In the present investigation we show, for the first time, that in rat brain capillary endothelial cells (RBE4), in vitro oxygen glucose deprivation treatment elicits 250% of Aβ(42) peptide production increase through a mechanism that involves the hypoxia inducible factor-1-mediated β-secretase (BACE1) up-regulation. Furthermore, we observed a time dependent increase of amyloid protein precursor (AβPP) gene and protein expression, confirming previous reports which established the existence of AβPP in the cerebrovascular domain. Our experimental evidences point out that ischemic events may directly contribute in brain capillary endothelial cells to the enhancement of the amyloidogenic metabolism, leading to intracellular deposition of Aβ(42). This events may contribute to the impairment of Aβ brain clearance and AD related blood brain barrier dysfunctions. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Effect of carbon dioxide on the utilization of brain capillary reserve and flow

    SciTech Connect

    Frankel, H.; Dribben, J.; Kissen, I.; Gerlock, T.; Weiss, H.R. )

    1989-12-01

    This study investigated effects of increased arterial carbon dioxide on the brain capillary perfusion pattern. Conscious rats were exposed to a 0%, 8% or 12% CO{sub 2} in air gas mixture. Arterial blood pressure, heart rate, arterial blood gases and pH were recorded, and either regional cerebral blood flow or the percent of capillary volume/mm3 or number/mm2 perfused were determined in cortical, hypothalamic, pontine or medullary regions of the brain. Arterial PCO{sub 2} increased from 37 +/- 1 in control to 74 +/- 1 torr in the high CO{sub 2} group. A position linear relationship was found between cerebral blood flow and arterial PCO{sub 2} in all examined regions. Approximately half of the capillaries in the examined regions were perfused under normocapnic conditions. Increasing arterial PCO{sub 2} had no effect on the percent of the capillary bed perfused in the cortex or hypothalamus. However, there was a significant linear relationship between carbon dioxide tension and the percent of the microvasculature perfused in the hindbrain. The percent of capillaries/mm2 perfused increased significantly in the medulla (to 60 +/- 5%) and pons (70 +/- 4%) with 12% CO{sub 2} in air. These data suggest that carbon dioxide may have differential effects on diffusion distances affecting the hindbrain to a greater extent than the forebrain.

  18. Distribution of anionic sites on the capillary endothelium in an experimental brain tumor model.

    PubMed

    Vincent, S; DePace, D; Finkelstein, S

    1988-02-01

    The distribution of anionic domains on the capillary endothelium of experimental brain tumors was determined using cationic ferritin (CF) in order to ascertain whether the pattern of these domains is different from that on normal cerebral capillaries. Tumors were induced by stereotaxic injection of cultured neoplastic glial cells, A15A5, into the caudate nucleus of Sprague-Dawley rats. Following a 14-21 day growth period tumors appeared as vascularized, sharply circumscribed masses which caused compression of the surrounding brain tissue. Anionic domains were distributed in a patchy and irregular pattern on the luminal plasma membrane of the endothelia of blood vessels in the tumors. Some variability in this pattern was observed infrequently in limited regions of the tumor where there was either a continuous layer of CF or an absence of CF binding. Plasmalemmal vesicles, coated vesicles, coated pits, multivesicular bodies, and some junctional complexes showed varying degrees of labeling with the probe. Capillaries in the tumor periphery and normal cerebral vessels showed a uniform distribution of anionic groups. These results indicate that there is an altered surface charge on the endothelial luminal plasma membrane of blood vessels in brain tumors. A correlation may exist between the altered surface charge and the degree to which the blood-brain barrier is impaired in these vessels.

  19. Coincident expression and distribution of melanotransferrin and transferrin receptor in human brain capillary endothelium.

    PubMed

    Rothenberger, S; Food, M R; Gabathuler, R; Kennard, M L; Yamada, T; Yasuhara, O; McGeer, P L; Jefferies, W A

    1996-03-11

    One method of iron transport across the blood brain barrier (BBB) involves the transferrin receptor (TR), which is localized to the specialized brain capillary endothelium. The melanotransferrin (MTf) molecule, also called p97, has been widely described as a melanoma specific molecule, however, its expression in brain tissues has not been addressed. MTf has a high level of sequence homology to transferrin (Tf) and lactoferrin, but is unusual because it predominantly occurs as a membrane bound, glycosylphosphatidylinositol (GPI) anchored molecule, but can also occur as a soluble form. We have recently demonstrated that GPI-anchored MTf provides a novel route for cellular iron uptake which is independent of Tf and its receptor. Here we consider whether MTf may have a role in the transport of iron across the BBB. The distributions of MTf, Tf and the TR were studied immunohistochemically in human brain tissues. The distributions of MTf and TR were remarkably similar, and quite different from that of Tf. In all brain tissues examined, MTf and the TR were highly localized to capillary endothelium, while Tf itself was mainly localized to glial cells. These data suggest that MTf may play a role in iron transport within the human brain.

  20. Uptake Mechanism of ApoE-Modified Nanoparticles on Brain Capillary Endothelial Cells as a Blood-Brain Barrier Model

    PubMed Central

    Wagner, Sylvia; Zensi, Anja; Wien, Sascha L.; Tschickardt, Sabrina E.; Maier, Wladislaw; Vogel, Tikva; Worek, Franz; Pietrzik, Claus U.; Kreuter, Jörg; von Briesen, Hagen

    2012-01-01

    Background The blood-brain barrier (BBB) represents an insurmountable obstacle for most drugs thus obstructing an effective treatment of many brain diseases. One solution for overcoming this barrier is a transport by binding of these drugs to surface-modified nanoparticles. Especially apolipoprotein E (ApoE) appears to play a major role in the nanoparticle-mediated drug transport across the BBB. However, at present the underlying mechanism is incompletely understood. Methodology/Principal Findings In this study, the uptake of the ApoE-modified nanoparticles into the brain capillary endothelial cells was investigated to differentiate between active and passive uptake mechanism by flow cytometry and confocal laser scanning microscopy. Furthermore, different in vitro co-incubation experiments were performed with competing ligands of the respective receptor. Conclusions/Significance This study confirms an active endocytotic uptake mechanism and shows the involvement of low density lipoprotein receptor family members, notably the low density lipoprotein receptor related protein, on the uptake of the ApoE-modified nanoparticles into the brain capillary endothelial cells. This knowledge of the uptake mechanism of ApoE-modified nanoparticles enables future developments to rationally create very specific and effective carriers to overcome the blood-brain barrier. PMID:22396775

  1. Polymersomes conjugated to 83-14 monoclonal antibodies: in vitro targeting of brain capillary endothelial cells.

    PubMed

    Dieu, Le-Ha; Wu, Dalin; Palivan, Cornelia G; Balasubramanian, Vimalkumar; Huwyler, Jörg

    2014-10-01

    The blood-brain barrier (BBB) remains an obstacle for many drugs to reach the brain. A strategy to cross the BBB is to modify nanocarrier systems with ligands that bind to endogenous receptors expressed at the BBB to induce receptor-mediated transport. The aim of the present study was to investigate the potential of polymersomes composed of the amphiphilic diblock copolymer poly(dimethylsiloxane)-block-poly(2-methyl-2-oxazoline), PDMS-b-PMOXA, for active targeting of brain capillary endothelial cells. We conjugated PDMS-b-PMOXA polymersomes to the anti-human insulin receptor antibody 83-14 and studied their uptake by brain capillary endothelial cells. Transmission electron micrography and light scattering measurements revealed the self-assembly of the block copolymers into 200 nm vesicles after extrusion. Fluorescence correlation spectroscopy was employed to calculate the number of antibodies coupled to one polymersome. Binding and uptake of the polymersomes conjugated to 83-14 mAb were studied in the human BBB in vitro model hCMEC/D3 expressing the human insulin receptor. Competitive inhibition with an excess of free 83-14 mAb demonstrated the specificity of cellular binding and uptake. Our results suggest that PDMS-b-PMOXA polymersomes conjugated to 83-14 mAb may be suitable nanocarriers for drug delivery to the brain.

  2. Mapping human brain capillary water lifetime: high-resolution metabolic neuroimaging.

    PubMed

    Rooney, William D; Li, Xin; Sammi, Manoj K; Bourdette, Dennis N; Neuwelt, Edward A; Springer, Charles S

    2015-06-01

    Shutter-speed analysis of dynamic-contrast-agent (CA)-enhanced normal, multiple sclerosis (MS), and glioblastoma (GBM) human brain data gives the mean capillary water molecule lifetime (τ(b)) and blood volume fraction (v(b); capillary density-volume product (ρ(†)V)) in a high-resolution (1)H2O MRI voxel (40 μL) or ROI. The equilibrium water extravasation rate constant, k(po) (τ(b)(-1)), averages 3.2 and 2.9 s(-1) in resting-state normal white matter (NWM) and gray matter (NGM), respectively (n = 6). The results (italicized) lead to three major conclusions. (A) k(po) differences are dominated by capillary water permeability (P(W)(†)), not size, differences. NWM and NGM voxel k(po) and v(b) values are independent. Quantitative analyses of concomitant population-averaged k(po), v(b) variations in normal and normal-appearing MS brain ROIs confirm P(W)(†) dominance. (B) P(W)(†) is dominated (>95%) by a trans(endothelial)cellular pathway, not the P(CA)(†) paracellular route. In MS lesions and GBM tumors, P(CA)(†) increases but P(W)(†) decreases. (C) k(po) tracks steady-state ATP production/consumption flux per capillary. In normal, MS, and GBM brain, regional k(po) correlates with literature MRSI ATP (positively) and Na(+) (negatively) tissue concentrations. This suggests that the P(W)(†) pathway is metabolically active. Excellent agreement of the relative NGM/NWM k(po)v(b) product ratio with the literature (31)PMRSI-MT CMR(oxphos) ratio confirms the flux property. We have previously shown that the cellular water molecule efflux rate constant (k(io)) is proportional to plasma membrane P-type ATPase turnover, likely due to active trans-membrane water cycling. With synaptic proximities and synergistic metabolic cooperativities, polar brain endothelial, neuroglial, and neuronal cells form "gliovascular units." We hypothesize that a chain of water cycling processes transmits brain metabolic activity to k(po), letting it report neurogliovascular unit Na

  3. New aspects in fenestrated capillary and tissue dynamics in the sensory circumventricular organs of adult brains.

    PubMed

    Miyata, Seiji

    2015-01-01

    The blood-brain barrier (BBB) generally consists of endothelial tight junction barriers that prevent the free entry of blood-derived substances, thereby maintaining the extracellular environment of the brain. However, the circumventricular organs (CVOs), which are located along the midlines of the brain ventricles, lack these endothelial barriers and have fenestrated capillaries; therefore, they have a number of essential functions, including the transduction of information between the blood circulation and brain. Previous studies have demonstrated the extensive contribution of the CVOs to body fluid and thermal homeostasis, energy balance, the chemoreception of blood-derived substances, and neuroinflammation. In this review, recent advances have been discussed in fenestrated capillary characterization and dynamic tissue reconstruction accompanied by angiogenesis and neurogliogenesis in the sensory CVOs of adult brains. The sensory CVOs, including the organum vasculosum of the lamina terminalis (OVLT), subfornical organ (SFO), and area postrema (AP), have size-selective and heterogeneous vascular permeabilities. Astrocyte-/tanycyte-like neural stem cells (NSCs) sense blood- and cerebrospinal fluid-derived information through the transient receptor potential vanilloid 1, a mechanical/osmotic receptor, Toll-like receptor 4, a lipopolysaccharide receptor, and Nax, a Na-sensing Na channel. They also express tight junction proteins and densely and tightly surround mature neurons to protect them from blood-derived neurotoxic substances, indicating that the NSCs of the CVOs perform BBB functions while maintaining the capacity to differentiate into new neurons and glial cells. In addition to neurogliogenesis, the density of fenestrated capillaries is regulated by angiogenesis, which is accompanied by the active proliferation and sprouting of endothelial cells. Vascular endothelial growth factor (VEGF) signaling may be involved in angiogenesis and neurogliogenesis, both of

  4. Development of Ascorbate Transporters in Brain Cortical Capillary Endothelial Cells in Culture

    PubMed Central

    Qiao, Huan; May, James M.

    2008-01-01

    Ascorbic acid in its reduced form is not transported across the capillary endothelial cell blood-brain barrier. This is thought to be due to absence of the SVCT2, a specific transporter for ascorbate. To assess this directly we prepared primary cultures of mouse cortical microvascular endothelial cells. When still in the capillaries, these cells did not express the SVCT2 protein as assessed by immunocytochemistry and by immunoblotting. However, during several days in culture, they developed SVCT2 expression and showed ascorbate transport rates comparable to those in immortalized endothelial cell lines. SVCT2 expression was inversely proportional to cell density, was enhanced by culture at low physiologic plasma ascorbate concentrations, was inhibited by ascorbate concentrations expected in the brain interstitium, and was stimulated by cobalt ions. Expression of the SVCT2 was associated with ascorbate-dependent maturation and release of type IV collagen by the cells in culture. Although the SVCT2 is induced by culture of cortical capillary endothelial cells, its absence in vivo remains perplexing, given the need for intracellular ascorbate to facilitate type IV collagen maturation and release by endothelial cells. PMID:18394593

  5. Hindered transport of macromolecules in isolated glomeruli. I. Diffusion across intact and cell-free capillaries.

    PubMed Central

    Edwards, A; Deen, W M; Daniels, B S

    1997-01-01

    The filtrate formed by renal glomerular capillaries must pass through a layer of endothelial cells, the glomerular basement membrane (GBM), and a layer of epithelial cells, arranged in series. To elucidate the relative resistances of the GBM and cell layers to movement of uncharged macromolecules, we measured the diffusional permeabilities of intact and cell-free capillaries to narrow fractions of Ficoll with Stokes-Einstein radii ranging from 3.0 to 6.2 nm. Glomeruli were isolated from rat kidneys, and diffusion of fluorescein-labeled Ficoll across the walls of single capillary loops was monitored with a confocal microscopy technique. In half of the experiments the glomeruli were treated first to remove the cells, leaving skeletons that retained the general shape of the glomerulus and consisted almost entirely of GBM. The diffusional permeability of cell-free capillaries to Ficoll was approximately 10 to 20 times that of intact capillaries, depending on molecular size. Taking into account the blockage of much of the GBM surface by cells, the contribution of the GBM to the diffusional resistance of the intact barrier was calculated to be 13% to 26% of the total, increasing with molecular size. Thus, the GBM contribution, although smaller than that of the cells, was not negligible. The structure that is most likely to be responsible for the cellular part of the diffusional resistance is the slit diaphragm, which spans the filtration slit between epithelial foot processes. A novel hydrodynamic model was developed to relate the diffusional resistance of the slit diaphragm to its structure, which was idealized as a single layer of cylindrical fibers in a ladder-like arrangement. Images FIGURE 1 FIGURE 2 FIGURE 3 FIGURE 5 PMID:8994605

  6. Ultrafast Capillary Electrophoresis Isolation of DNA Aptamer for the PCR Amplification-Based Small Analyte Sensing

    NASA Astrophysics Data System (ADS)

    Fiore, Emmanuelle; Dausse, Eric; Dubouchaud, Hervé; Peyrin, Eric; Ravelet, Corinne

    2015-08-01

    Here, we report a new homogeneous DNA amplification-based aptamer assay for small analyte sensing. The aptamer of adenosine chosen as the model analyte was split into two fragments able to assemble in the presence of target. Primers were introduced at extremities of one fragment in order to generate the amplifiable DNA component. The amount of amplifiable fragment was quantifiable by Real-Time Polymerase Chain Reaction (RT-PCR) amplification and directly reliable on adenosine concentration. This approach combines the very high separation efficiency and the homogeneous format (without immobilization) of capillary electrophoresis and the sensitivity of real time PCR amplification. An ultrafast isolation of target-bound split aptamer (60 s) was developed by designing a capillary electrophoresis input/ouput scheme. Such method was successfully applied to the determination of adenosine with a LOD of 1 µM.

  7. Molecular and functional characterization of riboflavin specific transport system in rat brain capillary endothelial cells

    PubMed Central

    Patel, Mitesh; Vadlapatla, Ramya Krishna; Pal, Dhananjay; Mitra, Ashim K.

    2012-01-01

    Riboflavin is an important water soluble vitamin (B2) required for metabolic reactions, normal cellular growth, differentiation and function. Mammalian brain cells cannot synthesize riboflavin and must import from systemic circulation. However, the uptake mechanism, cellular translocation and intracellular trafficking of riboflavin in brain capillary endothelial cells are poorly understood. The primary objective of this study is to investigate the existence of riboflavin-specific transport system and delineate the uptake and intracellular regulation of riboflavin in immortalized rat brain capillary endothelial cells (RBE4). The uptake of [3H]-Riboflavin is sodium, temperature and energy dependent but pH independent. [3H]-Riboflavin uptake is saturable with Km and Vmax values of 19 ± 3 µM and 0.235 ± 0.012 picomoles/min/mg protein, respectively. The uptake process is inhibited by unlabelled structural analogs (lumiflavin, lumichrome) but not by structurally unrelated vitamins. Ca++/calmodulin and protein kinase A (PKA) pathways are found to play an important role in the intracellular regulation of [3H]-Riboflavin. Apical and baso-lateral uptake of [3H]-Riboflavin clearly indicate that riboflavin specific transport system is predominantly localized on the apical side of RBE4 cells. A 628 bp band corresponding to riboflavin transporter is revealed in RT-PCR analysis. These findings, for the first time report the existence of a specialized and high affinity transport system for riboflavin in RBE4 cells. Blood-brain barrier (BBB) is a major obstacle limiting drug transport inside the brain as it regulates drug permeation from systemic circulation. This transporter can be utilized for targeted delivery in enhancing brain permeation of highly potent drugs on systemic administration. PMID:22683359

  8. Distribution of Capillary Transit Times in Isolated Lungs of Oxygen-Tolerant Rats

    PubMed Central

    Ramakrishna, Madhavi; Gan, Zhuohui; Clough, Anne V.; Molthen, Robert C.; Roerig, David L.; Audi, Said H.

    2014-01-01

    Rats pre-exposed to 85% O2 for 5–7 days tolerate the otherwise lethal effects of 100% O2. The objective was to evaluate the effect of rat exposure to 85% O2 for 7 days on lung capillary mean transit time (t̄c) and distribution of capillary transit times (hc(t)). This information is important for subsequent evaluation of the effect of this hyperoxia model on the redox metabolic functions of the pulmonary capillary endothelium. The venous concentration vs. time outflow curves of fluorescein isothiocyanate labeled dextran (FITC-dex), an intravascular indicator, and coenzyme Q1 hydroquinone (CoQ1H2), a compound which rapidly equilibrates between blood and tissue on passage through the pulmonary circulation, were measured following their bolus injection into the pulmonary artery of isolated perfused lungs from rats exposed to room air (normoxic) or 85% O2 for 7 days (hyperoxic). The moments (mean transit time and variance) of the measured FITC-dex and CoQ1H2 outflow curves were determined for each lung, and were then used in a mathematical model [Audi et al. J. Appl. Physiol. 77: 332–351, 1994] to estimate t̄c and the relative dispersion (RDc) of hc(t). Data analysis reveals that exposure to hyperoxia decreases lung t̄c by 42% and increases RDc, a measure hc(t) heterogeneity, by 40%. PMID:20552277

  9. Distribution of capillary transit times in isolated lungs of oxygen-tolerant rats.

    PubMed

    Ramakrishna, Madhavi; Gan, Zhuohui; Clough, Anne V; Molthen, Robert C; Roerig, David L; Audi, Said H

    2010-11-01

    Rats pre-exposed to 85% O₂ for 5-7 days tolerate the otherwise lethal effects of 100% O₂. The objective was to evaluate the effect of rat exposure to 85% O₂ for 7 days on lung capillary mean transit time t(c) and distribution of capillary transit times (h(c)(t)). This information is important for subsequent evaluation of the effect of this hyperoxia model on the redox metabolic functions of the pulmonary capillary endothelium. The venous concentration vs. time outflow curves of fluorescein isothiocyanate labeled dextran (FITC-dex), an intravascular indicator, and coenzyme Q₁ hydroquinone (CoQ₁H₂), a compound which rapidly equilibrates between blood and tissue on passage through the pulmonary circulation, were measured following their bolus injection into the pulmonary artery of isolated perfused lungs from rats exposed to room air (normoxic) or 85% O₂ for 7 days (hyperoxic). The moments (mean transit time and variance) of the measured FITC-dex and CoQ₁H₂ outflow curves were determined for each lung, and were then used in a mathematical model [Audi et al. J. Appl. Physiol. 77: 332-351, 1994] to estimate t(c) and the relative dispersion (RD(c)) of h (c)(t). Data analysis reveals that exposure to hyperoxia decreases lung t(c) by 42% and increases RD(c), a measure h(c)(t) heterogeneity, by 40%.

  10. Affinity filtration coupled with capillary-based affinity purification for the isolation of protein complexes.

    PubMed

    Qureshi, M S; Sheikh, Q I; Hill, R; Brown, P E; Dickman, M J; Tzokov, S B; Rice, D W; Gjerde, D T; Hornby, D P

    2013-08-01

    The isolation of complex macromolecular assemblies at the concentrations required for structural analysis represents a major experimental challenge. Here we present a method that combines the genetic power of site-specific recombination in order to selectively "tag" one or more components of a protein complex with affinity-based rapid filtration and a final step of capillary-based enrichment. This modified form of tandem affinity purification produces highly purified protein complexes at high concentrations in a highly efficient manner. The application of the method is demonstrated for the yeast Arp2/3 heptameric protein complex involved in mediating reorganization of the actin cytoskeleton.

  11. Uptake and transport of superparamagnetic iron oxide nanoparticles through human brain capillary endothelial cells.

    PubMed

    Thomsen, L B; Linemann, T; Pondman, K M; Lichota, J; Kim, K S; Pieters, R J; Visser, G M; Moos, T

    2013-10-16

    The blood-brain barrier (BBB) formed by brain capillary endothelial cells (BCECs) constitutes a firm physical, chemical, and immunological barrier, making the brain accessible to only a few percent of potential drugs intended for treatment inside the central nervous system. With the purpose of overcoming the restraints of the BBB by allowing the transport of drugs, siRNA, or DNA into the brain, a novel approach is to use superparamagnetic iron oxide nanoparticles (SPIONs) as drug carriers. The aim of this study was to investigate the ability of fluorescent SPIONs to pass through human brain microvascular endothelial cells facilitated by an external magnet. The ability of SPIONs to penetrate the barrier was shown to be significantly stronger in the presence of an external magnetic force in an in vitro BBB model. Hence, particles added to the luminal side of the in vitro BBB model were found in astrocytes cocultured at a remote distance on the abluminal side, indicating that particles were transported through the barrier and taken up by astrocytes. Addition of the SPIONs to the culture medium did not negatively affect the viability of the endothelial cells. The magnetic force-mediated dragging of SPIONs through BCECs may denote a novel mechanism for the delivery of drugs to the brain.

  12. Ultrastructural alterations of human cortical capillary basement membrane in human brain oedema.

    PubMed

    Castejón, Orlando José

    2014-01-01

    The capillary basement membranes are examined in severe traumatic brain injuries, vascular malformation, congenital hydrocephalus and brain tumours. They exhibit homogeneous and nodular thickening, vacuolization, rarefaction, reduplication, and deposition of collagen fibers. Their average thickness varied according to the aetiology and severity of brain oedema. In moderate brain oedema the thickness ranged from 71.97 to 191.90 nm in width, and in patients with severe brain oedema it varied from 206.66 to 404.22 nm. The basement membrane complex appears apparently intact in moderate oedema, and shows glio-basal dissociation in severe oedema. In areas of highly increased cerebro-vascular permeability, the basement membrane shows matrix disorganization, reduplication, and bifurcations protruding toward the endothelial cells, and acting as abluminal transcapillary channels. In regions of total brain necrosis, its structural stability is lost showing loosening, dissolution and rupture. Basement membrane swelling is due to overhydration of its protein-complex glycoprotein matrix. The thickening, rarefaction and vacuolization are induced by the increased vacuolar and vesicular transendothelial transport. The degenerated basement membrane areas exhibit a finely granular precipitate interpreted as protein, proteoglycan, glycoprotein, and agrin degraded matrix.

  13. Mathematical model of the effect of ischemia-reperfusion on brain capillary collapse and tissue swelling.

    PubMed

    Mohamed Mokhtarudin, M J; Payne, S J

    2015-05-01

    Restoration of an adequate cerebral blood supply after an ischemic attack is a primary clinical goal. However, the blood-brain barrier may break down after a prolonged ischemia causing the fluid in the blood plasma to filtrate and accumulate into the cerebral tissue interstitial space. Accumulation of this filtration fluid causes the cerebral tissue to swell, a condition known as vasogenic oedema. Tissue swelling causes the cerebral microvessels to be compressed, which may further obstruct the blood flow into the tissue, thus leading to the no-reflow phenomenon or a secondary ischemic stroke. The actual mechanism of this however is still not fully understood. A new model is developed here to study the effect of reperfusion on the formation of vasogenic oedema and cerebral microvessel collapse. The formation of vasogenic oedema is modelled using the capillary filtration equation while vessel collapse is modelled using the tube law of microvessel. Tissue swelling is quantified in terms of displacement, which is modelled using poroelastic theory. The results show that there is an increase in tissue displacement and interstitial pressure after reperfusion. In addition, the results also show that vessel collapse can occur at high value of reperfusion pressure, low blood osmotic pressure, high cerebral capillary permeability and low cerebral capillary stiffness. This model provides insight on the formation of ischemia-reperfusion injury by tissue swelling and vessel collapse. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. Mechanisms and regulation of iron trafficking across the capillary endothelial cells of the blood-brain barrier

    PubMed Central

    McCarthy, Ryan C.; Kosman, Daniel J.

    2015-01-01

    The transcellular trafficking of iron from the blood into the brain interstitium depends on iron uptake proteins in the apical membrane of brain microvascular capillary endothelial cells and efflux proteins at the basolateral, abluminal membrane. In this review, we discuss the three mechanisms by which these cells take-up iron from the blood and the sole mechanism by which they efflux this iron into the abluminal space. We then focus on the regulation of this efflux pathway by exocrine factors that are released from neighboring astrocytes. Also discussed are the cytokines secreted by capillary cells that regulate the expression of these glial cell signals. Among the interstitial factors that regulate iron efflux into the brain is the Amyloid precursor protein (APP). The role of this amyliodogenic species in brain iron metabolism is discussed. Last, we speculate on the potential relationship between iron transport at the blood-brain barrier and neurological disorders associated with iron mismanagement. PMID:26236187

  15. Transferrin Receptor 1 Facilitates Poliovirus Permeation of Mouse Brain Capillary Endothelial Cells*

    PubMed Central

    Mizutani, Taketoshi; Ishizaka, Aya; Nihei, Coh-ichi

    2016-01-01

    As a possible route for invasion of the CNS, circulating poliovirus (PV) in the blood is believed to traverse the blood-brain barrier (BBB), resulting in paralytic poliomyelitis. However, the underlying mechanism is poorly understood. In this study, we demonstrated that mouse transferrin receptor 1 (mTfR1) is responsible for PV attachment to the cell surface, allowing invasion into the CNS via the BBB. PV interacts with the apical domain of mTfR1 on mouse brain capillary endothelial cells (MBEC4) in a dose-dependent manner via its capsid protein (VP1). We found that F-G, G-H, and H-I loops in VP1 are important for this binding. However, C-D, D-E, and E-F loops in VP1-fused Venus proteins efficiently penetrate MBEC4 cells. These results imply that the VP1 functional domain responsible for cell attachment is different from that involved in viral permeation of the brain capillary endothelium. We observed that co-treatment of MBEC4 cells with excess PV particles but not dextran resulted in blockage of transferrin transport into cells. Using the Transwell in vitro BBB model, transferrin co-treatment inhibited permeation of PV into MBEC4 cells and delayed further viral permeation via mTfR1 knockdown. With mTfR1 as a positive mediator of PV-host cell attachment and PV permeation of MBEC4 cells, our results indicate a novel role of TfR1 as a cellular receptor for human PV receptor/CD155-independent PV invasion of the CNS. PMID:26637351

  16. Ultrafast capillary electrophoresis isolation of DNA aptamer for the PCR amplification-based small analyte sensing.

    PubMed

    Fiore, Emmanuelle; Dausse, Eric; Dubouchaud, Hervé; Peyrin, Eric; Ravelet, Corinne

    2015-01-01

    Here, we report a new homogeneous DNA amplification-based aptamer assay for small analyte sensing. The aptamer of adenosine chosen as the model analyte was split into two fragments able to assemble in the presence of target. Primers were introduced at extremities of one fragment in order to generate the amplifiable DNA component. The amount of amplifiable fragment was quantifiable by Real-Time Polymerase Chain Reaction (RT-PCR) amplification and directly reliable on adenosine concentration. This approach combines the very high separation efficiency and the homogeneous format (without immobilization) of capillary electrophoresis (CE) and the sensitivity of real time PCR amplification. An ultrafast isolation of target-bound split aptamer (60 s) was developed by designing a CE input/ouput scheme. Such method was successfully applied to the determination of adenosine with a LOD of 1 μM.

  17. Brain capillary transit time heterogeneity in healthy volunteers measured by dynamic contrast-enhanced T1 -weighted perfusion MRI.

    PubMed

    Larsson, Henrik B W; Vestergaard, Mark B; Lindberg, Ulrich; Iversen, Helle K; Cramer, Stig P

    2017-06-01

    Capillary transit time heterogeneity, measured as CTH, may set the upper limit for extraction of substances in brain tissue, e.g., oxygen. The purpose of this study was to investigate the feasibility of dynamic contrast-enhanced T1 weighted MRI (DCE-MRI) at 3 Tesla (T), in estimating CTH based on a gamma-variate model of the capillary transit time distribution. In addition, we wanted to investigate if a subtle increase of the blood-brain barrier permeability can be incorporated into the model, still allowing estimation of CTH. Twenty-three healthy subjects were scanned at 3.0T MRI system applying DCE-MRI and using a gamma-variate model to estimate CTH as well as cerebral blood flow (CBF), cerebral blood volume (CBV), and permeability of the blood-brain barrier, measured as the influx constant Ki . For proof of principle we also investigated three patients with recent thromboembolic events and a patient with a high grade brain tumor. In the healthy subjects, we found a narrow symmetric delta-like capillary transit time distribution in basal ganglia gray matter with median CTH of 0.93 s and interquartile range of 1.33 s. The corresponding residue impulse response function was compatible with the adiabatic tissue homogeneity model. In two patients with complete occlusion of the internal carotid artery and in the patient with a brain tumor CTH was increased with values up to 6 s in the affected brain tissue, with an exponential like residue impulse response function. Our results open the possibility of characterizing brain perfusion by the capillary transit time distribution using DCE-MRI, theoretically a determinant of efficient blood to brain transport of important substances. 2 J. MAGN. RESON. IMAGING 2017;45:1809-1820. © 2016 International Society for Magnetic Resonance in Medicine.

  18. Involvement of Proton-Coupled Organic Cation Antiporter in Varenicline Transport at Blood-Brain Barrier of Rats and in Human Brain Capillary Endothelial Cells.

    PubMed

    Kurosawa, Toshiki; Higuchi, Kei; Okura, Takashi; Kobayashi, Kazumasa; Kusuhara, Hiroyuki; Deguchi, Yoshiharu

    2017-09-01

    Varenicline is a selective partial α4β2 nicotinic acetylcholine receptor agonist, which is used to help achieve smoking cessation. Here, we investigated varenicline transport at the blood-brain barrier by means of in vivo microdialysis, in situ brain perfusion, and brain efflux index measurements in rats, and in vitro uptake studies in human brain capillary endothelial cells. Microdialysis demonstrated that varenicline is actively transported from blood to brain in rats. Blood-to-brain uptake transport of varenicline, as measured by the in situ brain perfusion technique, was strongly inhibited by diphenhydramine, a potent inhibitor of proton-coupled organic cation (H(+)/OC) antiporter. However, brain efflux index study showed that brain-to-blood efflux transport of varenicline was not inhibited by diphenhydramine. In human brain capillary endothelial cells, varenicline was taken up time- and concentration-dependently. The uptake was dependent on an oppositely directed proton gradient, but was independent of extracellular sodium and membrane potential. The uptake was inhibited by a metabolic inhibitor, and by substrates of H(+)/OC antiporter, but not by substrates or inhibitors of OCTs, OCTNs, PMAT, and MATE1, which are known organic cation transporters. The present results suggest that the H(+)/OC antiporter contributes predominantly to varenicline uptake at the blood-brain barrier. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  19. Modifications in astrocyte morphology and calcium signaling induced by a brain capillary endothelial cell line.

    PubMed

    Yoder, Elizabeth J

    2002-04-15

    Astrocytes extend specialized endfoot processes to perisynaptic and perivascular regions, and thus are positioned to mediate the bidirectional flow of metabolic, ionic, and other transmissive substances between neurons and the blood stream. While mutual structural and functional interactions between neurons and astrocytes have been documented, less is known about the interactions between astrocytes and cerebrovascular cells. For example, although the ability of astrocytes to induce structural and functional changes in endothelial cells is established, the reciprocity of brain endothelial cells to induce changes in astrocytes is undetermined. This issue is addressed in the present study. Changes in primary cultures of neonatal mouse cortical astrocytes were investigated following their coculture with mouse brain capillary endothelial (bEnd3) cells. The presence of bEnd3 cells altered the morphology of astrocytes by transforming them from confluent monolayers into networks of elongated multicellular columns. These columns did not occur when either bEnd3 cells or astrocytes were cocultured with other cell types, suggesting that astrocytes undergo specific morphological consequences when placed in close proximity to brain endothelial cells. In addition to these structural changes, the pharmacological profile of astrocytes was modified by coculture with bEnd3 cells. Astrocytes in the cocultures showed an increased Ca2+ responsiveness to bradykinin and glutamate, but no change in responsiveness to ATP, as compared to controls. Coculturing the astrocytes with a neuronal cell line resulted in increased responsiveness of the glial responses to glutamate but not to bradykinin. These studies indicate that brain endothelial cells induce changes in astrocyte morphology and pharmacology.

  20. Constitutively active Notch4 receptor elicits brain arteriovenous malformations through enlargement of capillary-like vessels.

    PubMed

    Murphy, Patrick A; Kim, Tyson N; Huang, Lawrence; Nielsen, Corinne M; Lawton, Michael T; Adams, Ralf H; Schaffer, Chris B; Wang, Rong A

    2014-12-16

    Arteriovenous (AV) malformation (AVM) is a devastating condition characterized by focal lesions of enlarged, tangled vessels that shunt blood from arteries directly to veins. AVMs can form anywhere in the body and can cause debilitating ischemia and life-threatening hemorrhagic stroke. The mechanisms that underlie AVM formation remain poorly understood. Here, we examined the cellular and hemodynamic changes at the earliest stages of brain AVM formation by time-lapse two-photon imaging through cranial windows of mice expressing constitutively active Notch4 (Notch4*). AVMs arose from enlargement of preexisting microvessels with capillary diameter and blood flow and no smooth muscle cell coverage. AV shunting began promptly after Notch4* expression in endothelial cells (ECs), accompanied by increased individual EC areas, rather than increased EC number or proliferation. Alterations in Notch signaling in ECs of all vessels, but not arteries alone, affected AVM formation, suggesting that Notch functions in the microvasculature and/or veins to induce AVM. Increased Notch signaling interfered with the normal biological control of hemodynamics, permitting a positive feedback loop of increasing blood flow and vessel diameter and driving focal AVM growth from AV connections with higher blood velocity at the expense of adjacent AV connections with lower velocity. Endothelial expression of constitutively active Notch1 also led to brain AVMs in mice. Our data shed light on cellular and hemodynamic mechanisms underlying AVM pathogenesis elicited by increased Notch signaling in the endothelium.

  1. Constitutively active Notch4 receptor elicits brain arteriovenous malformations through enlargement of capillary-like vessels

    PubMed Central

    Murphy, Patrick A.; Kim, Tyson N.; Huang, Lawrence; Nielsen, Corinne M.; Lawton, Michael T.; Adams, Ralf H.; Schaffer, Chris B.; Wang, Rong A.

    2014-01-01

    Arteriovenous (AV) malformation (AVM) is a devastating condition characterized by focal lesions of enlarged, tangled vessels that shunt blood from arteries directly to veins. AVMs can form anywhere in the body and can cause debilitating ischemia and life-threatening hemorrhagic stroke. The mechanisms that underlie AVM formation remain poorly understood. Here, we examined the cellular and hemodynamic changes at the earliest stages of brain AVM formation by time-lapse two-photon imaging through cranial windows of mice expressing constitutively active Notch4 (Notch4*). AVMs arose from enlargement of preexisting microvessels with capillary diameter and blood flow and no smooth muscle cell coverage. AV shunting began promptly after Notch4* expression in endothelial cells (ECs), accompanied by increased individual EC areas, rather than increased EC number or proliferation. Alterations in Notch signaling in ECs of all vessels, but not arteries alone, affected AVM formation, suggesting that Notch functions in the microvasculature and/or veins to induce AVM. Increased Notch signaling interfered with the normal biological control of hemodynamics, permitting a positive feedback loop of increasing blood flow and vessel diameter and driving focal AVM growth from AV connections with higher blood velocity at the expense of adjacent AV connections with lower velocity. Endothelial expression of constitutively active Notch1 also led to brain AVMs in mice. Our data shed light on cellular and hemodynamic mechanisms underlying AVM pathogenesis elicited by increased Notch signaling in the endothelium. PMID:25468970

  2. Characterization of molds isolated from smoked paprika by PCR-RFLP and micellar electrokinetic capillary electrophoresis.

    PubMed

    Ruiz-Moyano, Santiago; Benito, María J; Martín, Alberto; Aranda, Emilio; Hernández, Alejandro; Córdoba, María G

    2009-12-01

    Molds are common contaminants of paprika meat products. The drying and storage stages of paprika processing are critical because they can provide molds with the conditions particularly appropriate for their growth and proliferation. Thus, an efficient and accurate characterization of the toxigenic molds of paprika is necessary. An RFLP analysis of the rRNA genes was performed by using a TaqI restriction enzyme. In addition, a micellar electrokinetic capillary chromatography (MECC) method was tested to analyze secondary metabolites produced by mold strains commonly found in paprika. This study was confirmed with a 5.8S-ITS region sequence analysis. A total of 31 isolates were identified by RFLP and MECC analysis. These showed stable RFLP profiles that were clearly different for the different genera and species, and were grouped into clusters together with the profiles of the 16 reference strains. MECC analysis provided additional characteristic peak patterns for the characterization of the mold species present. The characterized isolates were species of the genera Fusarium spp., Aspergillus spp., Penicillium spp., Cladosporium spp., Mucor spp. and Phlebia spp. The identifications were confirmed by the 5.8S-ITS region sequence analysis and by a BLAST search of the GenBank database. RFLP patterns with TaqI restriction enzyme and MECC profiles, either singly or combined, could be of great interest to distinguish molds in paprika.

  3. Capillary Isoelectric Focusing of Akt Isoforms Identifies Highly Dynamic Phosphorylation in Neuronal Cells and Brain Tissue.

    PubMed

    Schrötter, Sandra; Leondaritis, George; Eickholt, Britta J

    2016-05-06

    The PI3K/PTEN/Akt pathway has been established as a core signaling pathway that is crucial for the integration of neurons into neuronal circuits and the maintenance of the architecture and function of neurons in the adult brain. Akt1-3 kinases are specifically activated by two phosphorylation events on residues Thr(308) and Ser(473) upon growth factor signaling, which subsequently phosphorylate a vast cohort of downstream targets. However, we still lack a clear understanding of the complexity and regulation of isoform specificity within the PI3K/PTEN/Akt pathway. We utilized a capillary-based isoelectric focusing method to study dynamics of Akt phosphorylation in neuronal cells and the developing brain and identify previously undescribed features of Akt phosphorylation and activation. First, we show that the accumulation of multiple phosphorylation events on Akt forms occur concurrently with Ser(473) and Thr(308) phosphorylation upon acute PI3K activation and provide evidence for uncoupling of Ser(473) and Thr(308) phosphorylation, as well as differential sensitivities of Akt1 forms upon PI3K inhibition. Second, we detect a transient shift in Akt isoform phosphorylation and activation pattern during early postnatal brain development, at stages corresponding to synapse development and maturation. Third, we show differential sensitivities of Ser(473)-Akt species to PTEN deletion in mature neurons, which suggests inherent differences in the Akt pools that are accessible to growth factors as compared with the pools that are controlled by PTEN. Our study demonstrates the presence of complex phosphorylation events of Akt in a time- and signal-dependent manner in neurons.

  4. Capillary Isoelectric Focusing of Akt Isoforms Identifies Highly Dynamic Phosphorylation in Neuronal Cells and Brain Tissue*

    PubMed Central

    Schrötter, Sandra; Leondaritis, George; Eickholt, Britta J.

    2016-01-01

    The PI3K/PTEN/Akt pathway has been established as a core signaling pathway that is crucial for the integration of neurons into neuronal circuits and the maintenance of the architecture and function of neurons in the adult brain. Akt1–3 kinases are specifically activated by two phosphorylation events on residues Thr308 and Ser473 upon growth factor signaling, which subsequently phosphorylate a vast cohort of downstream targets. However, we still lack a clear understanding of the complexity and regulation of isoform specificity within the PI3K/PTEN/Akt pathway. We utilized a capillary-based isoelectric focusing method to study dynamics of Akt phosphorylation in neuronal cells and the developing brain and identify previously undescribed features of Akt phosphorylation and activation. First, we show that the accumulation of multiple phosphorylation events on Akt forms occur concurrently with Ser473 and Thr308 phosphorylation upon acute PI3K activation and provide evidence for uncoupling of Ser473 and Thr308 phosphorylation, as well as differential sensitivities of Akt1 forms upon PI3K inhibition. Second, we detect a transient shift in Akt isoform phosphorylation and activation pattern during early postnatal brain development, at stages corresponding to synapse development and maturation. Third, we show differential sensitivities of Ser473-Akt species to PTEN deletion in mature neurons, which suggests inherent differences in the Akt pools that are accessible to growth factors as compared with the pools that are controlled by PTEN. Our study demonstrates the presence of complex phosphorylation events of Akt in a time- and signal-dependent manner in neurons. PMID:26945062

  5. Motion of an isolated liquid plug inside a capillary tube: effect of contact angle hysteresis

    NASA Astrophysics Data System (ADS)

    Srinivasan, Vyas; Khandekar, Sameer; Bouamrane, Nathan; Lefevre, Frederic; Bonjour, Jocelyn

    2015-01-01

    Dynamics of a single, small and isolated partially wetting liquid plug (of known length L and wettability), placed at rest inside a long, dry, circular capillary tube ( D = 1.5 mm), and subsequently quasi-statically pushed from one end by applying air pressure, the other end being kept exposed to atmosphere, are reported. The air pressure first overcomes the `static' friction manifested by the three-phase contact line at the advancing and receding menisci, and then, the plug motion gets initiated, eventually leading to a terminal velocity (Ca ~ 2.8 × 10-5), when pressure force balances net frictional resistance due to viscous and surface forces. It is seen that, under steady motion, the curvature profiles of the advancing and receding menisci of liquid plug, respectively, remain the same, independent of the plug length. Steady-state pressure drop is dominated by the contribution due to contact angle hysteresis, which is also independent of the plug length. Increasing the system wettability drastically decreased the contact angle hysteresis and the associated net pressure drop.

  6. Impaired regulation of pH homeostasis by oxidative stress in rat brain capillary endothelial cells.

    PubMed

    Sipos, Hdikó; Törocsik, Beáta; Tretter, Laszlo; Adam-Vizi, Vera

    2005-02-01

    (1) Endothelial cells are permanently challenged by altering pH in the blood, and oxidative damage could also influence the intracellular pH (pH(i)) of the endothelium. Cerebral microvascular endothelial cells form the blood-brain barrier (BBB) and pH(i) regulation of brain capillary endothelial cells is important for the maintenance of BBB integrity. The aim of this study was to address the pH regulatory mechanisms and the effect of an acute exposure to hydrogen peroxide (H2O2) on the pH regulation in primary rat brain capillary endothelial (RBCE) cells The RBCE monolayers were loaded with the fluorescent pH indicator BCECF and pH(i) was monitored by detecting the fluorescent changes. (2) The steady-state pH(i) of RBCE cells in HEPES-buffer (6.83 +/- 0.1) did not differ significantly from that found in bicarbonate-buffered medium (6.90 +/- 0.08). Cells were exposed to NH4CI to induce intracellular acidification and then the recovery to resting pH was studied. Half-recovery time after NH4Cl prepulse-induced acid load was significantly less in the bicarbonate-buffered medium than in the HEPES-medium, suggesting that in addition to the Na+ / H+ exchanger, HCO3- / Cl- exchange mechanism is also involved in the restoration of pH(i) after an intracellular acid load in primary RBCE cells. We used RT-PCR-reactions to detect the isoforms of Na+ / H+ exchanger gene family (NHE). NHE-1 -2, -3 and -4 were equally present, and there was no significant difference in the relative abundance of the four transcripts in these cells. (3) No pH(i) recovery was detected when the washout after an intracellular acid load occurred in nominally Na+ -free HEPES-buffered medium or in the presence of 10 microM 5-(N-ethyl-N-isopropyl)amiloride (EIPA), a specific inhibitor of Na+ / H+ exchanger. The new steady-state pH(i) were 6.37 +/- 0.02 and 6.60 +/- 0.02, respectively. (4) No detectable change was observed in the steady-state pH(i) in the presence of 100 microM H2O2; however, recovery from

  7. Physiological pathway for low-density lipoproteins across the blood-brain barrier: transcytosis through brain capillary endothelial cells in vitro.

    PubMed

    Candela, Pietra; Gosselet, Fabien; Miller, Florence; Buee-Scherrer, Valerie; Torpier, Gérard; Cecchelli, Roméo; Fenart, Laurence

    2008-01-01

    Although an immense knowledge has accumulated concerning regulation of cholesterol homeostasis in the body, this does not include the brain, where details are just emerging. Using an in vitro blood-brain barrier model, the authors have demonstrated that low-density lipoprotein (LDL) underwent transcytosis through the endothelial cells (ECs) by a receptor-mediated process, bypassing the lysosomal compartment. Moreover, caveolae might be involved in these blood-borne molecule transports from the blood to the brain. Although several ligands are known to be internalized through cell surface caveolae, the subsequent intracellular pathways have remained elusive. By cell fractionation experiment and Western blot, the authors have demonstrated that the LDL receptor is located in the caveolae membrane fraction. Then, LDLs internalized were detected by electron microscopy in multivesicular bodies. The authors identified in brain capillary ECs a novel endosomal compartment, mildly acidic, positive for marker Lamp-1 but devoid of any degradative capability. From the point of view of pH, cellular location, and caveolae-derived formation, the multivesicular organelles described here can be related to the caveosome structure. These results could provide clues to physiological functions of caveolae-caveosome transcellular pathway in brain capillary ECs and may help in the rational design of more effective therapeutic drugs to the brain.

  8. The roles of cerebral blood flow, capillary transit time heterogeneity, and oxygen tension in brain oxygenation and metabolism

    PubMed Central

    Jespersen, Sune N; Østergaard, Leif

    2012-01-01

    Normal brain function depends critically on moment-to-moment regulation of oxygen supply by the bloodstream to meet changing metabolic needs. Neurovascular coupling, a range of mechanisms that converge on arterioles to adjust local cerebral blood flow (CBF), represents our current framework for understanding this regulation. We modeled the combined effects of CBF and capillary transit time heterogeneity (CTTH) on the maximum oxygen extraction fraction (OEFmax) and metabolic rate of oxygen that can biophysically be supported, for a given tissue oxygen tension. Red blood cell velocity recordings in rat brain support close hemodynamic–metabolic coupling by means of CBF and CTTH across a range of physiological conditions. The CTTH reduction improves tissue oxygenation by counteracting inherent reductions in OEFmax as CBF increases, and seemingly secures sufficient oxygenation during episodes of hyperemia resulting from cortical activation or hypoxemia. In hypoperfusion and states of blocked CBF, both lower oxygen tension and CTTH may secure tissue oxygenation. Our model predicts that disturbed capillary flows may cause a condition of malignant CTTH, in which states of higher CBF display lower oxygen availability. We propose that conditions with altered capillary morphology, such as amyloid, diabetic or hypertensive microangiopathy, and ischemia–reperfusion, may disturb CTTH and thereby flow-metabolism coupling and cerebral oxygen metabolism. PMID:22044867

  9. TRPV4 partially participates in proliferation of human brain capillary endothelial cells.

    PubMed

    Hatano, Noriyuki; Suzuki, Hiroka; Itoh, Yuka; Muraki, Katsuhiko

    2013-03-12

    The vanilloid type 4 transient receptor potential channel (TRPV4) is a potential environmental sensor to multiple stimuli in many types of cells. In this study, we show that TRPV4 activated by 4α-phorbol 12,13-didecanoate (4αPDD) and hypo-osmotic stimulation (HOS) is a regulator of intracellular calcium ([Ca(2+)](i)) in human brain capillary endothelial cells (HBCEs), and its activation can partially regulate cell proliferation of HBCEs. The expression of TRPV4 in HBCEs was analyzed at the mRNA and protein levels. The function of TRPV4 in HBCEs was evaluated using a TRPV4 agonist, 4αPDD, and HOS while measuring [Ca(2+)](i) and membrane currents. Analysis of the mRNA transcripts of the TRPV subfamily revealed that TRPV2 and TRPV4 were expressed in HBCEs. Immunoreactivity to the TRPV4 protein was also detected in HBCEs, which were positively stained by von Willebrand factor and CD31. When 4αPDD was applied, [Ca(2+)](i) in HBCEs was elevated in a concentration-dependent manner. In addition, exposure of HBCEs to HOS at 228mOsm induced an elevation of [Ca(2+)](i). Application of 4αPDD also activated non-selective cation currents (NSCCs). Pretreatment of HBCEs with short interference RNA targeting TRPV4 (siRNA) significantly reduced the 4αPDD-induced elevation of [Ca(2+)](i). When HBCEs were treated for 24h with concentrations of 4αPDD between 0.3 and 3 μM, the cell proliferation was potentiated in a concentration-dependent manner. The potentiation was partially inhibited in HBCEs treated with siRNA. These data suggest that endogenous TRPV4, which functions as a regulator of [Ca(2+)](i) in HBCEs, partially controls cell proliferation. Copyright © 2013 Elsevier Inc. All rights reserved.

  10. Multiple brain abscesses from isolated cerebral mucormycosis.

    PubMed Central

    Escobar, A; Del Brutto, O H

    1990-01-01

    A report is presented of a patient with cerebral mucormycosis without rhinosinusal or systemic evidence of the disease. The predisposing condition was drug-induced immunosuppression. Computed tomography (CT) showed focal areas of abnormal enhancement which correlated with necropsy findings of localised parenchymal brain damage; this represented encapsulated brain abscesses, a rare form of presentation of cerebral mucormycosis. Images PMID:2351973

  11. Transient oxygen-glucose deprivation sensitizes brain capillary endothelial cells to rtPA at 4h of reoxygenation.

    PubMed

    Kuntz, Mélanie; Mysiorek, Caroline; Pétrault, Olivier; Boucau, Marie-Christine; Aijjou, Rachid; Uzbekov, Rustem; Bérézowski, Vincent

    2014-01-01

    Thrombolysis treatment of acute ischemic stroke is limited by the pro-edematous and hemorrhagic effects exerted by reperfusion, which disrupts the blood-brain barrier (BBB) capillary endothelium in the infarct core. Most studies of the ischemic BBB overlook the complexity of the penumbral area, where the affected brain cells are still viable following deprivation. Our present objective was to examine in vitro the kinetic impact of reoxygenation on the integrity of ischemic BBB cells after oxygen-glucose deprivation. Through the use of a co-culture of brain capillary endothelial cells and glial cells, we first showed that the transendothelial permeability increase induced by deprivation can occur with both preserved cell viability and interendothelial tight junction network. The subtle and heterogeneous alteration of the tight junctions was observable only through electron microscopy. A complete permeability recovery was then found after reoxygenation, when Vimentin and Actin networks were reordered. However, still sparse ultrastructural alterations of tight junctions suggested an acquired vulnerability. Endothelial cells were then exposed to recombinant tissue-type plasminogen activator (rtPA) to define a temporal profile for the toxic effect of this thrombolytic on transendothelial permeability. Interestingly, the reoxygenated BBB broke down with aggravated tight junction disruption when exposed to rtPA only at 4h after reoxygenation. Moreover, this breakdown was enhanced by 50% when ischemic glial cells were present during the first hours of reoxygenation. Our results suggest that post-stroke reoxygenation enables retrieval of the barrier function of brain capillary endothelium when in a non-necrotic environment, but may sensitize it to rtPA at the 4-hour time point, when both endothelial breakdown mechanisms and glial secretions could be identified and targeted in a therapeutical perspective.

  12. Isolation, synthesis and anti-hepatitis B virus evaluation of p-hydroxyacetophenone derivatives from Artemisia capillaris.

    PubMed

    Zhao, Yong; Geng, Chang-An; Chen, Hao; Ma, Yun-Bao; Huang, Xiao-Yan; Cao, Tuan-Wu; He, Kang; Wang, Hao; Zhang, Xue-Mei; Chen, Ji-Jun

    2015-04-01

    p-Hydroxyacetophenone (p-HAP), as a main hepatoprotective and choleretic constituent of Artemisia capillaris, was revealed with anti-hepatitis B virus (HBV) effects in recent investigation. In addition to p-HAP, four derivatives of p-HAP were also isolated from A. capillaris by various chromatographic methods. Subsequent structural modification on p-HAP and its glycoside led to the synthesis of 28 additional derivatives, of which 13 compounds showed activity inhibiting hepatitis B surface antigen (HBsAg) secretion; and 18 compounds possessed inhibition on HBV DNA replication. The primary structure-activity relationships (SARs) suggested that the conjugated derivatives of p-HAP glycoside and substituted cinnamic acids (2a-2i) obviously enhanced the activity against HBV DNA replication with IC50 values ranged from 5.8 to 74.4 μM.

  13. Major involvement of Na(+) -dependent multivitamin transporter (SLC5A6/SMVT) in uptake of biotin and pantothenic acid by human brain capillary endothelial cells.

    PubMed

    Uchida, Yasuo; Ito, Katsuaki; Ohtsuki, Sumio; Kubo, Yoshiyuki; Suzuki, Takashi; Terasaki, Tetsuya

    2015-07-01

    The purpose of this study was to clarify the expression of Na(+) -dependent multivitamin transporter (SLC5A6/SMVT) and its contribution to the supply of biotin and pantothenic acid to the human brain via the blood-brain barrier. DNA microarray and immunohistochemical analyses confirmed that SLC5A6 is expressed in microvessels of human brain. The absolute expression levels of SLC5A6 protein in isolated human and monkey brain microvessels were 1.19 and 0.597 fmol/μg protein, respectively, as determined by a quantitative targeted absolute proteomics technique. Using an antibody-free method established by Kubo et al. (2015), we found that SLC5A6 was preferentially localized at the luminal membrane of brain capillary endothelium. Knock-down analysis using SLC5A6 siRNA showed that SLC5A6 accounts for 88.7% and 98.6% of total [(3) H]biotin and [(3) H]pantothenic acid uptakes, respectively, by human cerebral microvascular endothelial cell line hCMEC/D3. SLC5A6-mediated transport in hCMEC/D3 was markedly inhibited not only by biotin and pantothenic acid, but also by prostaglandin E2, lipoic acid, docosahexaenoic acid, indomethacin, ketoprofen, diclofenac, ibuprofen, phenylbutazone, and flurbiprofen. This study is the first to confirm expression of SLC5A6 in human brain microvessels and to provide evidence that SLC5A6 is a major contributor to luminal uptake of biotin and pantothenic acid at the human blood-brain barrier. In humans, it was unclear (not concluded) about what transport system at the blood-brain barrier (BBB) is responsible for the brain uptakes of two vitamins, biotin and pantothenic acid, which are necessary for brain proper function. This study clarified for the first time that the solute carrier 5A6/Na(+) -dependent multivitamin transporter SLC5A6/SMVT is responsible for the supplies of biotin and pantothenic acid into brain across the BBB in humans. DHA, docosahexaenoic acid; NSAID, non-steroidal anti-inflammatory drug; PGE2, prostaglandin E2. © 2015

  14. 3D BrainCV: simultaneous visualization and analysis of cells and capillaries in a whole mouse brain with one-micron voxel resolution.

    PubMed

    Wu, Jingpeng; He, Yong; Yang, Zhongqin; Guo, Congdi; Luo, Qingming; Zhou, Wei; Chen, Shangbin; Li, Anan; Xiong, Benyi; Jiang, Tao; Gong, Hui

    2014-02-15

    Systematic cellular and vascular configurations are essential for understanding fundamental brain anatomy and metabolism. We demonstrated a 3D brainwide cellular and vascular (called 3D BrainCV) visualization and quantitative protocol for a whole mouse brain. We developed a modified Nissl staining method that quickly labeled the cells and blood vessels simultaneously in an entire mouse brain. Terabytes 3D datasets of the whole mouse brains, with unprecedented details of both individual cells and blood vessels, including capillaries, were simultaneously imaged at 1-μm voxel resolution using micro-optical sectioning tomography (MOST). For quantitative analysis, we proposed an automatic image-processing pipeline to perform brainwide vectorization and analysis of cells and blood vessels. Six representative brain regions from the cortex to the deep, including FrA, M1, PMBSF, V1, striatum, and amygdala, and six parameters, including cell number density, vascular length density, fractional vascular volume, distance from the cells to the nearest microvessel, microvascular length density, and fractional microvascular volume, had been quantitatively analyzed. The results showed that the proximity of cells to blood vessels was linearly correlated with vascular length density, rather than the cell number density. The 3D BrainCV made overall snapshots of the detailed picture of the whole brain architecture, which could be beneficial for the state comparison of the developing and diseased brain.

  15. Freeze-Drying as Sample Preparation for Micellar Electrokinetic Capillary Chromatography – Electrochemical Separations of Neurochemicals in Drosophila Brains

    PubMed Central

    Berglund, E. Carina; Kuklinski, Nicholas J.; Karagündüz, Ekin; Ucar, Kubra; Hanrieder, Jörg; Ewing, Andrew G.

    2013-01-01

    Micellar electrokinetic capillary chromatography with electrochemical detection has been used to quantify biogenic amines in freeze-dried Drosophila melanogaster brains. Freeze drying samples offers a way to preserve the biological sample while making dissection of these tiny samples easier and faster. Fly samples were extracted in cold acetone and dried in a rotary evaporator. Extraction and drying times were optimized in order to avoid contamination by red-pigment from the fly eyes and still have intact brain structures. Single freeze-dried fly-brain samples were found to produce representative electropherograms as a single hand-dissected brain sample. Utilizing the faster dissection time that freeze drying affords, the number of brains in a fixed homogenate volume can be increased to concentrate the sample. Thus, concentrated brain samples containing five or fifteen preserved brains were analyzed for their neurotransmitter content, and five analytes; dopamine N-acetyloctopamine, Nacetylserotonin, N-acetyltyramine, N-acetyldopamine were found to correspond well with previously reported values. PMID:23387977

  16. Ferritin: isolation of aluminum-ferritin complex from brain.

    PubMed Central

    Fleming, J; Joshi, J G

    1987-01-01

    Ferritin was isolated from the livers and brains of two groups of rats, one of which was fed aluminum chloride (100 microM) for 1 year in the drinking water. Brain tissue contained about one-third of the amount of ferritin found in the liver. While brain ferritin from normal rats contained 42.1 +/- 14.3 mol of aluminum, that from the aluminum-fed group contained 115.4 +/- 48.3 mol of aluminum per mol of ferritin. Liver ferritin from both groups contained similar amounts of both aluminum and iron, and the amounts were less than that found associated with brain ferritin. Ferritin isolated from the brains of patients who died of Alzheimer disease contained more aluminum and more iron than that from age-matched controls. Human brain ferritin is composed of two types of subunits--about 70% heavy chain (Mr, 22,000) and 30% light chain (Mr, 19,500). The isoelectric focusing pattern of human brain ferritin was considerably different from that of human liver. Only 5 of the 20 brain ferritin bands migrated similarly to the acidic isoferritins from the liver, and the major component of brain ferritin, representing 30% of the total ferritin, had a pI of 8.0. Images PMID:3479769

  17. Brain tumor cell line authentication, an efficient alternative to capillary electrophoresis by using a microfluidics-based system

    PubMed Central

    An, Qian; Fillmore, Helen L.; Vouri, Mikaella; Pilkington, Geoffrey J.

    2014-01-01

    Background The current method for cell line authentication is genotyping based on short tandem repeat (STR)–PCR involving coamplification of a panel of STR loci by multiplex PCR and downstream fragment length analysis (FLA), usually performed by capillary electrophoresis. FLA by capillary electrophoresis is time-consuming and can be expensive, as the facilities are generally not accessible for many research laboratories. Methods In the present study, a microfluidic electrophoresis system, the Agilent 2100 Bioanalyzer, was used to analyze the STR-PCR fragments from 10 human genomic loci of a number of human cell lines, including 6 gliomas, 1 astrocyte, 1 primary lung cancer, 1 lung brain metastatic cancer, and 1 rhabdomyosarcoma; and this was compared with the standard method, that is, capillary electrophoresis, using the Applied Biosystems 3130xl Genetic Analyzer. Results The microfluidic electrophoresis method produced highly reproducible results with good sensitivity in sizing of multiple PCR fragments, and each cell line demonstrated a unique DNA profile. Furthermore, DNA fingerprinting of samples from 5 different passage numbers of the same cell line showed excellent reproducibility when FLA was performed with the Bioanalyzer, indicating that no cross-contamination had occurred during the culture period. Conclusion This novel application provides a straightforward and cost-effective alternative to STR-based cell line authentication. In addition, this application would be of great value for cell bank repositories to maintain and distribute precious cell lines. PMID:24335698

  18. Isolation and Culture of Adult Zebrafish Brain-derived Neurospheres

    PubMed Central

    Lopez-Ramirez, Miguel A.; Calvo, Charles-Félix; Ristori, Emma; Thomas, Jean-Léon; Nicoli, Stefania

    2016-01-01

    The zebrafish is a highly relevant model organism for understanding the cellular and molecular mechanisms involved in neurogenesis and brain regeneration in vertebrates. However, an in-depth analysis of the molecular mechanisms underlying zebrafish adult neurogenesis has been limited due to the lack of a reliable protocol for isolating and culturing neural adult stem/progenitor cells. Here we provide a reproducible method to examine adult neurogenesis using a neurosphere assay derived from zebrafish whole brain or from the telencephalon, tectum and cerebellum regions of the adult zebrafish brain. The protocol involves, first the microdissection of zebrafish adult brain, then single cell dissociation and isolation of self-renewing multipotent neural stem/progenitor cells. The entire procedure takes eight days. Additionally, we describe how to manipulate gene expression in zebrafish neurospheres, which will be particularly useful to test the role of specific signaling pathways during adult neural stem/progenitor cell proliferation and differentiation in zebrafish. PMID:26967835

  19. Analysis of multiple quaternary ammonium compounds in the brain using tandem capillary column separation and high resolution mass spectrometric detection.

    PubMed

    Falasca, Sara; Petruzziello, Filomena; Kretz, Robert; Rainer, Gregor; Zhang, Xiaozhe

    2012-06-08

    Endogenous quaternary ammonium compounds are involved in various physiological processes in the central nervous system. In the present study, eleven quaternary ammonium compounds, including acetylcholine, choline, carnitine, acetylcarnitine and seven other acylcarnitines of low polarity, were analyzed from brain extracts using a two dimension capillary liquid chromatography-Fourier transform mass spectrometry method. To deal with their large difference in hydrophobicities, tandem coupling between reversed phase and hydrophilic interaction chromatography columns was used to separate all the targeted quaternary ammonium compounds. Using high accuracy mass spectrometry in selected ion monitoring mode, all the compounds could be detected from each brain sample with high selectivity. The developed method was applied for the relative quantification of these quaternary ammonium compounds in three different brain regions of tree shrews: prefrontal cortex, striatum, and hippocampus. The comparative analysis showed that quaternary ammonium compounds were differentially distributed across the three brain areas. The analytical method proved to be highly sensitive and reliable for simultaneous determination of all the targeted analytes from brain samples.

  20. The effects of capillary transit time heterogeneity (CTH) on brain oxygenation

    PubMed Central

    Angleys, Hugo; Østergaard, Leif; Jespersen, Sune N

    2015-01-01

    We recently extended the classic flow–diffusion equation, which relates blood flow to tissue oxygenation, to take capillary transit time heterogeneity (CTH) into account. Realizing that cerebral oxygen availability depends on both cerebral blood flow (CBF) and capillary flow patterns, we have speculated that CTH may be actively regulated and that changes in the capillary morphology and function, as well as in blood rheology, may be involved in the pathogenesis of conditions such as dementia and ischemia-reperfusion injury. The first extended flow–diffusion equation involved simplifying assumptions which may not hold in tissue. Here, we explicitly incorporate the effects of oxygen metabolism on tissue oxygen tension and extraction efficacy, and assess the extent to which the type of capillary transit time distribution affects the overall effects of CTH on flow–metabolism coupling reported earlier. After incorporating tissue oxygen metabolism, our model predicts changes in oxygen consumption and tissue oxygen tension during functional activation in accordance with literature reports. We find that, for large CTH values, a blood flow increase fails to cause significant improvements in oxygen delivery, and can even decrease it; a condition of malignant CTH. These results are found to be largely insensitive to the choice of the transit time distribution. PMID:25669911

  1. Isolation and characterization of intact mitochondria from neonatal rat brain.

    PubMed

    Rajapakse, N; Shimizu, K; Payne, M; Busija, D

    2001-12-01

    Poor outcome after neonatal brain injury may be associated with alterations in mitochondrial function. Thus, isolated mitochondria have been a useful tool in understanding the underlying mechanisms of mitochondrial dysfunction. However, isolation and characterization of mitochondria from neonatal rat brain are not fully described. Thus, the aim of this study was to develop a rapid method for the isolation and characterization of functional mitochondria from neonatal rat brain. Mitochondria were isolated from 7-day-old rat brain weighing approximately 500 mg using a discontinuous Percoll density gradient. Brains were homogenized in 12% Percoll/sucrose buffer and layered onto a 26% Percoll/40% Percoll gradient followed by centrifugation. Four methods were used for assessing mitochondrial integrity and function: (1) electron microscopy to assess the morphology of the mitochondria and to determine the relative purity of the preparation; (2) fluorescence of chloromethyl-X-rosamine (Mito Tracker Red) in mitochondria as an indicator of mitochondrial membrane potential (Delta psi(m)); (3) state 3 and 4 respiration; and (4) protein import into mitochondria using an in vitro-synthesized mitochondrial malate dehydrogenase (mMDH). These studies demonstrated that the morphology of mitochondria is maintained with intact outer membranes and well-developed cristae, and Delta psi(m) is preserved. Respiration measurements revealed tightly coupled mitochondria with a respiration control ratio (RCR) of 4.1+/-0.18 (n=6). Import of precursor mMDH into mitochondria increased in a time-dependent manner maximizing at 15 min. The results indicate that neonatal brain mitochondria isolated using this method are well coupled, morphologically intact and are capable of protein import across the outer and inner mitochondrial membranes.

  2. Culture and isolation of brain tumor initiating cells.

    PubMed

    Lenkiewicz, Monika; Li, Na; Singh, Sheila K

    2009-10-01

    This unit describes protocols for the culture and isolation of brain tumor initiating cells (BTIC). The cancer stem cell (CSC) hypothesis suggests that tumors are maintained exclusively by a rare fraction of cells that have stem cell properties. We applied culture conditions and assays originally used for normal neural stem cells (NSCs) in vitro to a variety of brain tumors. The BTIC were isolated by fluorescence activated cell sorting for the neural precursor cell surface marker CD133. Only the CD133(+) brain tumor fraction contains cells capable of sphere formation and sustained self-renewal in vitro, and tumor initiation in NOD-SCID mouse brains. Therefore, CD133(+) BTICs satisfy the definition of cancer stem cells in that they are able to generate a replica of the patient's tumor and they exhibit self-renewal ability through serial retransplantation. This established that only a rare subset of brain tumor cells with stem cell properties are tumor-initiating, and, in this unit, we describe their culture and isolation.

  3. Oxysterols decrease apical-to-basolateral transport of Aß peptides via an ABCB1-mediated process in an in vitro Blood-brain barrier model constituted of bovine brain capillary endothelial cells.

    PubMed

    Saint-Pol, Julien; Candela, Pietra; Boucau, Marie-Christine; Fenart, Laurence; Gosselet, Fabien

    2013-06-23

    It is known that activation of the liver X receptors (LXRs) by natural or synthetic agonists decreases the amyloid burden and enhances cognitive function in transgenic murine models of Alzheimer's disease (AD). Recent evidence suggests that LXR activation may affect the transport of amyloid ß (Aß) peptides across the blood-brain barrier (the BBB, which isolates the brain from the peripheral circulation). By using a well-characterized in vitro BBB model, we demonstrated that LXR agonists (24S-hydroxycholesterol, 27-hydroxycholesterol and T0901317) modulated the expression of target genes involved in cholesterol homeostasis (such as ATP-binding cassette sub-family A member 1 (ABCA1)) and promoted cellular cholesterol efflux to apolipoprotein A-I and high density lipoproteins. Interestingly, we also observed a decrease in Aß peptide influx across brain capillary endothelial cells, although ABCA1 did not appear to be directly involved in this process. By focusing on others receptors and transporters that are thought to have major roles in Aß peptide entry into the brain, we then demonstrated that LXR stimulation provoked an increase in expression of the ABCB1 transporter (also named P-glycoprotein (P-gp)). Further investigations confirmed ABCB1's involvement in the restriction of Aß peptide influx. Taken as a whole, our results not only reinforce the BBB's key role in cerebral cholesterol homeostasis but also demonstrate the importance of the LXR/ABCB1 axis in Aß peptide influx-highlighting an attractive new therapeutic approach whereby the brain could be protected from peripheral Aß peptide entry.

  4. Capillary Hemangioma

    MedlinePlus

    ... Why do capillary hemangiomas on the eyelids cause vision problems? Capillary Hemangiomas of the eyelid can cause ... a capillary hemangioma in the eye socket cause vision problems? A capillary hemangioma in the eye socket ( ...

  5. In vitro discrimination of the role of LRP1 at the BBB cellular level: focus on brain capillary endothelial cells and brain pericytes.

    PubMed

    Candela, Pietra; Saint-Pol, Julien; Kuntz, Mélanie; Boucau, Marie-Christine; Lamartiniere, Yordenca; Gosselet, Fabien; Fenart, Laurence

    2015-01-12

    Several studies have demonstrated that the blood-brain barrier (BBB) (dynamic cellular complex composed by brain capillary endothelial cells (BCECs) and surrounded by astrocytic end feet and pericytes) regulates the exchanges of amyloid β (Aβ) peptide between the blood and the brain. Deregulation of these exchanges seems to be a key trigger for the brain accumulation of Aβ peptide observed in Alzheimer's disease (AD). Whereas the involvement of receptor for advanced glycation end-products in Aβ peptide transcytosis has been demonstrated in our laboratory, low-density lipoprotein receptor's role at the cellular level needs to be clarified. For this, we used an in vitro BBB model that consists of a co-culture of bovine BCECs and rat glial cells. This model has already been used to characterize low-density lipoprotein receptor-related peptide (LRP)'s involvement in the transcytosis of molecules such as tPA and angiopep-2. Our results suggest that Aβ peptide efflux across the BCEC monolayer involves a transcellular transport. However, the experiments with RAP discard an involvement of LRP family members at BCECs level. In contrast, our results show a strong transcriptional expression of LRP1 in pericytes and suggest its implication in Aβ endocytosis. Moreover, the observations of pericytes contraction and local downregulation of LRP1 in response to Aβ treatment opens up perspectives for studying this cell type with respect to Aβ peptide metabolism and AD. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Hypertensive encephalopathy presenting with isolated brain stem and cerebellar edema.

    PubMed

    Bhagavati, Satyakam; Chum, Florence; Choi, Jai

    2008-10-01

    Hypertensive encephalopathy typically presents with headache and confusion and bilateral parietooccipital vasogenic edema. Brain stem and cerebellar edema in hypertensive encephalopathy usually occurs in association with these typical supratentorial changes and is usually asymptomatic. We report here an uncommon hypertensive patient with isolated, severe, and symptomatic brain stem and cerebellar edema with fourth ventricular obstruction and mild hydrocephalus. Rapid treatment of hypertension resulted in clinical and radiological improvement. Prompt recognition of the cause and aggressive treatment of hypertension in such patients are crucial to relieve edema and prevent life-threatening progression.

  7. One-step multiple component isolation from the oil of Crinitaria tatarica (Less) Sojak by preparative capillary gas chromatography with characterization by spectroscopic and spectrometric techniques

    USDA-ARS?s Scientific Manuscript database

    In the present work multiple component isolation from the oil of Crinitaria tatarica (Less.) Sojak. by Preparative Capillary Gas Chromatography (PCGC) with characterization by mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy have been carried out. Gas chromatography (GC-FID) ...

  8. Application of capillary isotachophoresis-based multidimensional separations coupled with electrospray ionization-tandem mass spectrometry for characterization of mouse brain mitochondrial proteome

    PubMed Central

    Fang, Xueping; Wang, Weijie; Yang, Li; Chandrasekaran, Krish; Kristian, Tibor; Balgley, Brian M.; Lee, Cheng S.

    2017-01-01

    By employing a capillary ITP (CITP)/CZE-based proteomic technology, a total of 1795 distinct mouse Swiss-Prot protein entries (or 1705 nonredundant proteins) are identified from synaptic mitochondria isolated from mouse brain. The ultrahigh resolving power of CITP/CZE is evidenced by the large number of distinct peptide identifications measured from each CITP fraction together with the low peptide fraction overlapping among identified peptides. The degree of peptide overlapping among CITP fractions is even lower than that achieved using combined CIEF/nano-RP LC separations for the analysis of the same mitochondrial sample. When evaluating the protein sequence coverage by the number of distinct peptides mapping to each mitochondrial protein identification, CITP/CZE similarly achieves superior performance with 1041 proteins (58%) having 3 or more distinct peptides, 233 (13%) having 2 distinct peptides, and 521 (29%) having a single distinct peptide. The reproducibility of protein identifications is found to be around 86% by comparing proteins identified from repeated runs of the same mitochondrial sample. The analysis of the mouse mitochondrial proteome by two CITP/CZE runs results in the detection of 2095 distinct mouse Swiss-Prot protein entries (or 1992 nonredundant proteins), corresponding to 59% coverage of the updated Maestro mitochondrial reference set. The collective analysis from combined CITP/CZE and CIEF-based proteomic studies yields the identification of 2191 distinct mitochondrial protein entries (or 2082 nonredundant proteins), corresponding to 76% coverage of the MitoP2-database reference set. PMID:18425750

  9. Culture and Isolation of Brain Tumor Initiating Cells.

    PubMed

    Vora, Parvez; Venugopal, Chitra; McFarlane, Nicole; Singh, Sheila K

    2015-08-03

    Brain tumors are typically composed of heterogeneous cells that exhibit distinct phenotypic characteristics and proliferative potentials. Only a relatively small fraction of cells in the tumor with stem cell properties, termed brain tumor initiating cells (BTICs), possess an ability to differentiate along multiple lineages, self-renew, and initiate tumors in vivo. This unit describes protocols for the culture and isolation BTICs. We applied culture conditions and assays originally used for normal neural stem cells (NSCs) in vitro to a variety of brain tumors. Using fluorescence-activated cell sorting for the neural precursor cell surface marker CD133/CD15, BTICs can be isolated and studied prospectively. Isolation of BTICs from GBM bulk tumor will enable examination of dissimilar morphologies, self-renewal capacities, tumorigenicity, and therapeutic sensitivities. As cancer is also considered a disease of unregulated self-renewal and differentiation, an understanding of BTICs is fundamental to understanding tumor growth. Ultimately, it will lead to novel drug discovery approaches that strategically target the functionally relevant BTIC population. Copyright © 2015 John Wiley & Sons, Inc.

  10. Neuron enriched nuclear proteome isolated from human brain.

    PubMed

    Dammer, Eric B; Duong, Duc M; Diner, Ian; Gearing, Marla; Feng, Yue; Lah, James J; Levey, Allan I; Seyfried, Nicholas T

    2013-07-05

    The brain consists of diverse cell types including neurons, astrocytes, oligodendrocytes, and microglia. The isolation of nuclei from these distinct cell populations provides an opportunity to identify cell-type-specific nuclear proteins, histone modifications, and regulation networks that are altered with normal brain aging or neurodegenerative disease. In this study, we used a method by which intact neuronal and non-neuronal nuclei were purified from human post-mortem brain employing a modification of fluorescence activated cell sorting (FACS) termed fluorescence activated nuclei sorting (FANS). An antibody against NeuN, a neuron specific splicing factor, was used to isolate neuronal nuclei. Utilizing mass spectrometry (MS) based label-free quantitative proteomics, we identified 1755 proteins from sorted NeuN-positive and negative nuclear extracts. Approximately 20% of these proteins were significantly enriched or depleted in neuronal versus non-neuronal populations. Immunoblots of primary cultured rat neuron, astrocyte, and oligodendrocyte extracts confirmed that distinct members of the major nucleocytoplasmic structural linkage complex (LINC), nesprin-1 and nesprin-3, were differentially enriched in neurons and astrocytes, respectively. These comparative proteomic data sets also reveal a number of transcription and splicing factors that are selectively enriched in a cell-type-specific manner in human brain.

  11. Transfection of primary brain capillary endothelial cells for protein synthesis and secretion of recombinant erythropoietin: a strategy to enable protein delivery to the brain.

    PubMed

    Burkhart, Annette; Andresen, Thomas Lars; Aigner, Achim; Thomsen, Louiza Bohn; Moos, Torben

    2017-03-14

    Treatment of chronic disorders affecting the central nervous system (CNS) is complicated by the inability of drugs to cross the blood-brain barrier (BBB). Non-viral gene therapy applied to brain capillary endothelial cells (BCECs) denotes a novel approach to overcome the restraints in this passage, as turning BCECs into recombinant protein factories by transfection could result in protein secretion further into the brain. The present study aims to investigate the possibility of transfecting primary rat brain endothelial cells (RBECs) for recombinant protein synthesis and secretion of the neuroprotective protein erythropoietin (EPO). We previously showed that 4% of RBECs with BBB properties can be transfected without disrupting the BBB integrity in vitro, but it can be questioned whether this is sufficient to enable protein secretion at therapeutic levels. The present study examined various transfection vectors, with regard to increasing the transfection efficiency without disrupting the BBB integrity. Lipofectamine 3000™ was the most potent vector compared to polyethylenimine (PEI) and Turbofect. When co-cultured with astrocytes, the genetically modified RBECs secreted recombinant EPO into the cell culture medium both luminally and abluminally, and despite lower levels of EPO reaching the abluminal chamber, the amount of recombinant EPO was sufficient to evolve a biological effect on astrocytes cultured at the abluminal side in terms of upregulated gene expression of brain-derived neurotropic factor (BDNF). In conclusion, non-viral gene therapy to RBECs leads to protein secretion and signifies a method for therapeutic proteins to target cells inside the CNS otherwise omitted due to the BBB.

  12. Cooling treatment transiently increases the permeability of brain capillary endothelial cells through translocation of claudin-5.

    PubMed

    Inamura, Akinori; Adachi, Yasuhiro; Inoue, Takao; He, Yeting; Tokuda, Nobuko; Nawata, Takashi; Shirao, Satoshi; Nomura, Sadahiro; Fujii, Masami; Ikeda, Eiji; Owada, Yuji; Suzuki, Michiyasu

    2013-08-01

    The blood-brain-barrier (BBB) is formed by different cell types, of which brain microvascular endothelial cells are major structural constituents. The goal of this study was to examine the effects of cooling on the permeability of the BBB with reference to tight junction formation of brain microendothelial cells. The sensorimotor cortex above the dura mater in adult male Wistar rats was focally cooled to a temperature of 5 °C for 1 h, then immunostaining for immunoglobulin G (IgG) was performed to evaluate the permeability of the BBB. Permeability produced by cooling was also evaluated in cultured murine brain endothelial cells (bEnd3) based on measurement of trans-epithelial electric resistance (TEER). Immunocytochemistry and Western blotting of proteins associated with tight junctions in bEnd3 were performed to determine protein distribution before and after cooling. After focal cooling of the rat brain cortex, diffuse immunostaining for IgG was observed primarily around the small vasculature and in the extracellular spaces of parenchyma of the cortex. In cultured bEnd3, TEER significantly decreased during cooling (15 °C) and recovered to normal levels after rewarming to 37 °C. Immunocytochemistry and Western blotting showed that claudin-5, a critical regulatory protein for tight junctions, was translocated from the membrane to the cytoplasm after cooling in cultured bEnd3 cells. These results suggest that focal brain cooling may open the BBB transiently through an effect on tight junctions of brain microendothelial cells, and that therapeutically this approach may allow control of BBB function and drug delivery through the BBB.

  13. Arterial-ventricular and interventricular interaction in isolated post-capillary and combined pulmonary hypertension in severe mitral stenosis.

    PubMed

    Venkateshvaran, Ashwin; Sola, Srikanth; Govind, Satish Chandra; Dash, Pravat Kumar; Vyavahare, Sagar; Lund, Lars H; la Merkely, Bé; Nagy, Anikó Ilona; Manouras, Aristomenis

    2016-08-01

    Isolated post-capillary pulmonary hypertension (Ipc-PH) is characterized by elevated left atrial pressures that are passively transmitted upstream, whereas combined pre- and post-capillary PH (Cpc-PH) demonstrates additional reactive changes in pulmonary vasculature. The increased load imposed on the right ventricle (RV) influences left ventricular (LV) mechanics by means of interventricular interaction. However, there is lack of evidence to substantiate the effect of possible additional alterations in the arterio-ventricular (AV) coupling and their effect on LV function. Considering the discrepant RV load in Cpc-PH and Ipc-PH, we sought to investigate whether these two conditions are also characterized by differential alterations in AV coupling. Invasive hemodynamic and echocardiographic data of 120 patients with PH due to severe rheumatic mitral stenosis before and immediately after percutaneous valvulotomy, along with 40 age-matched healthy controls, were analyzed. Effective arterial (E a) and ventricular elastance (E es) were measured. PH patients demonstrated elevated LV afterload (E a) along with AV uncoupling, and these derangements were more evident in the Cpc-PH group [E a: 3.3 (2.3-5.4) vs 2.6 (2.1-3.5) mmHg/mL, E a/E es: 0.73 (0.6-0.9) vs 0.88 (0.7-1.2), p < 0.05]. In addition, PH was associated with reduced LV deformation, which was mainly determined by elevated E a, while the effect of interventricular interaction was limited to the septal wall. Our results suggest that in addition to the interventricular interaction, an abnormal AV coupling contributes to the altered LV mechanics that has been associated with adverse prognosis in Cpc-PH.

  14. Ultrastructural pathology of cortical capillary pericytes in human traumatic brain oedema.

    PubMed

    Castejón, Orlando J

    2011-01-01

    In human traumatic brain oedema pericytes exhibit remarkable oedematous changes, increased vacuolar and vesicular transport, transient transpericytal channels, and tubular structures demonstrating pericyte brain barrier dysfunction. They show nuclear invaginations, actin and myosin-like filaments, and coupled interaction with endothelial cells through the macula occludens. Some pericytes display hypertrophic and necrotic changes, and phagocytic capacity. Hypertrophic pericytes induce basement membrane splitting. Degenerated pericytes exhibit lacunar enlargement of endoplasmic reticulum, dense osmiophilic bodies, glycogen granules, vacuolization, oedematous Golgi apparatus, and pleomorphic mitochondria. Certain micropinocytotic vesicles are orientated to the Golgi complex and multivesicular bodies, suggesting that pericytes play some role in oedema resolution.

  15. Synthesis and deposition of basement membrane proteins by primary brain capillary endothelial cells in a murine model of the blood-brain barrier.

    PubMed

    Thomsen, Maj Schneider; Birkelund, Svend; Burkhart, Annette; Stensballe, Allan; Moos, Torben

    2017-03-01

    The brain vascular basement membrane is important for both blood-brain barrier (BBB) development, stability, and barrier integrity and the contribution hereto from brain capillary endothelial cells (BCECs), pericytes, and astrocytes of the BBB is probably significant. The aim of this study was to analyse four different in vitro models of the murine BBB for expression and possible secretion of major basement membrane proteins from murine BCECs (mBCECs). mBCECs, pericytes and glial cells (mainly astrocytes and microglia) were prepared from brains of C57BL/6 mice. The mBCECs were grown as monoculture, in co-culture with pericytes or mixed glial cells, or as a triple-culture with both pericytes and mixed glial cells. The integrity of the BBB models was validated by measures of transendothelial electrical resistance (TEER) and passive permeability to mannitol. The expression of basement membrane proteins was analysed using RT-qPCR, mass spectrometry and immunocytochemistry. Co-culturing mBCECs with pericytes, mixed glial cells, or both significantly increased the TEER compared to the monoculture, and a low passive permeability was correlated with high TEER. The mBCECs expressed all major basement membrane proteins such as laminin-411, laminin-511, collagen [α1(IV)]2 α2(IV), agrin, perlecan, and nidogen 1 and 2 in vitro. Increased expression of the laminin α5 subunit correlated with the addition of BBB-inducing factors (hydrocortisone, Ro 20-1724, and pCPT-cAMP), whereas increased expression of collagen IV α1 primarily correlated with increased levels of cAMP. In conclusion, BCECs cultured in vitro coherently form a BBB and express basement membrane proteins as a feature of maturation. Cover Image for this issue: doi: 10.1111/jnc.13789. © 2016 International Society for Neurochemistry.

  16. Hypoxic stress up-regulates Kir2.1 expression and facilitates cell proliferation in brain capillary endothelial cells

    SciTech Connect

    Yamamura, Hideto; Suzuki, Yoshiaki; Yamamura, Hisao; Asai, Kiyofumi; Imaizumi, Yuji

    2016-08-05

    The blood-brain barrier (BBB) is mainly composed of brain capillary endothelial cells (BCECs), astrocytes and pericytes. Brain ischemia causes hypoxic encephalopathy and damages BBB. However, it remains still unclear how hypoxia affects BCECs. In the present study, t-BBEC117 cells, an immortalized bovine brain endothelial cell line, were cultured under hypoxic conditions at 4–5% oxygen for 72 h. This hypoxic stress caused hyperpolarization of resting membrane potential. Patch-clamp recordings revealed a marked increase in Ba{sup 2+}-sensitive inward rectifier K{sup +} current in t-BBEC117 cells after hypoxic culture. Western blot and real-time PCR analyses showed that Kir2.1 expression was significantly up-regulated at protein level but not at mRNA level after the hypoxic culture. Ca{sup 2+} imaging study revealed that the hypoxic stress enhanced store-operated Ca{sup 2+} (SOC) entry, which was significantly reduced in the presence of 100 μM Ba{sup 2+}. On the other hand, the expression of SOC channels such as Orai1, Orai2, and transient receptor potential channels was not affected by hypoxic stress. MTT assay showed that the hypoxic stress significantly enhanced t-BBEC117 cell proliferation, which was inhibited by approximately 60% in the presence of 100 μM Ba{sup 2+}. We first show here that moderate cellular stress by cultivation under hypoxic conditions hyperpolarizes membrane potential via the up-regulation of functional Kir2.1 expression and presumably enhances Ca{sup 2+} entry, resulting in the facilitation of BCEC proliferation. These findings suggest potential roles of Kir2.1 expression in functional changes of BCECs in BBB following ischemia. -- Highlights: •Hypoxic culture of brain endothelial cells (BEC) caused membrane hyperpolarization. •This hyperpolarization was due to the increased expression of Kir2.1 channels. •Hypoxia enhanced store-operated Ca{sup 2+} (SOC) entry via Kir2.1 up-regulation. •Expression levels of putative SOC

  17. Capillary sample

    MedlinePlus

    ... using capillary blood sampling. Disadvantages to capillary blood sampling include: Only a limited amount of blood can be drawn using this method. The procedure has some risks (see below). Capillary ...

  18. Multiple capillary biochemical analyzer

    DOEpatents

    Dovichi, N.J.; Zhang, J.Z.

    1995-08-08

    A multiple capillary analyzer allows detection of light from multiple capillaries with a reduced number of interfaces through which light must pass in detecting light emitted from a sample being analyzed, using a modified sheath flow cuvette. A linear or rectangular array of capillaries is introduced into a rectangular flow chamber. Sheath fluid draws individual sample streams through the cuvette. The capillaries are closely and evenly spaced and held by a transparent retainer in a fixed position in relation to an optical detection system. Collimated sample excitation radiation is applied simultaneously across the ends of the capillaries in the retainer. Light emitted from the excited sample is detected by the optical detection system. The retainer is provided by a transparent chamber having inward slanting end walls. The capillaries are wedged into the chamber. One sideways dimension of the chamber is equal to the diameter of the capillaries and one end to end dimension varies from, at the top of the chamber, slightly greater than the sum of the diameters of the capillaries to, at the bottom of the chamber, slightly smaller than the sum of the diameters of the capillaries. The optical system utilizes optic fibers to deliver light to individual photodetectors, one for each capillary tube. A filter or wavelength division demultiplexer may be used for isolating fluorescence at particular bands. 21 figs.

  19. Multiple capillary biochemical analyzer

    DOEpatents

    Dovichi, Norman J.; Zhang, Jian Z.

    1995-01-01

    A multiple capillary analyzer allows detection of light from multiple capillaries with a reduced number of interfaces through which light must pass in detecting light emitted from a sample being analyzed, using a modified sheath flow cuvette. A linear or rectangular array of capillaries is introduced into a rectangular flow chamber. Sheath fluid draws individual sample streams through the cuvette. The capillaries are closely and evenly spaced and held by a transparent retainer in a fixed position in relation to an optical detection system. Collimated sample excitation radiation is applied simultaneously across the ends of the capillaries in the retainer. Light emitted from the excited sample is detected by the optical detection system. The retainer is provided by a transparent chamber having inward slanting end walls. The capillaries are wedged into the chamber. One sideways dimension of the chamber is equal to the diameter of the capillaries and one end to end dimension varies from, at the top of the chamber, slightly greater than the sum of the diameters of the capillaries to, at the bottom of the chamber, slightly smaller than the sum of the diameters of the capillaries. The optical system utilizes optic fibres to deliver light to individual photodetectors, one for each capillary tube. A filter or wavelength division demultiplexer may be used for isolating fluorescence at particular bands.

  20. AMP kinase regulation of sugar transport in brain capillary endothelial cells during acute metabolic stress

    PubMed Central

    Cura, Anthony J.

    2012-01-01

    AMP-dependent kinase (AMPK) and GLUT1-mediated sugar transport in blood-brain barrier endothelial cells are activated during acute cellular metabolic stress. Using murine brain microvasculature endothelium bEnd.3 cells, we show that AMPK phosphorylation and stimulation of 3-O-methylglucose transport by the AMPK agonist AICAR are inhibited in a dose-dependent manner by the AMPK antagonist Compound C. AMPK α1- or AMPK α2-knockdown by RNA interference or AMPK inhibition by Compound C reduces AMPK phosphorylation and 3-O-methylglucose transport stimulation induced by cellular glucose-depletion, by potassium cyanide (KCN), or by carbonyl cyanide-p-trifluoromethoxy-phenylhydrazone (FCCP). Cell surface biotinylation studies reveal that plasma membrane GLUT1 levels are increased two- to threefold by cellular glucose depletion, AICAR or KCN treatment, and that these increases are prevented by Compound C and by AMPK α1- or α2-knockdown. These results support the hypothesis that AMPK activation in blood-brain barrier-derived endothelial cells directs the trafficking of GLUT1 intracellular pools to the plasma membrane, thereby increasing endothelial sugar transport capacity. PMID:22763120

  1. AMP kinase regulation of sugar transport in brain capillary endothelial cells during acute metabolic stress.

    PubMed

    Cura, Anthony J; Carruthers, Anthony

    2012-10-15

    AMP-dependent kinase (AMPK) and GLUT1-mediated sugar transport in blood-brain barrier endothelial cells are activated during acute cellular metabolic stress. Using murine brain microvasculature endothelium bEnd.3 cells, we show that AMPK phosphorylation and stimulation of 3-O-methylglucose transport by the AMPK agonist AICAR are inhibited in a dose-dependent manner by the AMPK antagonist Compound C. AMPK α1- or AMPK α2-knockdown by RNA interference or AMPK inhibition by Compound C reduces AMPK phosphorylation and 3-O-methylglucose transport stimulation induced by cellular glucose-depletion, by potassium cyanide (KCN), or by carbonyl cyanide-p-trifluoromethoxy-phenylhydrazone (FCCP). Cell surface biotinylation studies reveal that plasma membrane GLUT1 levels are increased two- to threefold by cellular glucose depletion, AICAR or KCN treatment, and that these increases are prevented by Compound C and by AMPK α1- or α2-knockdown. These results support the hypothesis that AMPK activation in blood-brain barrier-derived endothelial cells directs the trafficking of GLUT1 intracellular pools to the plasma membrane, thereby increasing endothelial sugar transport capacity.

  2. Isolated Brain Trauma in Cats Triggers Rapid Onset of Hypovolemia.

    PubMed

    Bentzer, Peter; Grände, Per-Olof

    2017-06-01

    Hemodynamic instability responsive to fluid resuscitation is common after a traumatic brain injury (TBI), also in the absence of systemic hemorrhage. The present study tests if an isolated severe TBI induces a decrease in plasma volume (PV). The study was performed in three groups of anesthetized and tracheostomized male cats (n = 21). In one group (n = 8), the cats were prepared with a cranial borehole (10 mm i.d) used to expose the brain to a fluid percussion brain injury (FPI) (1.90-2.20 bar), and two smaller cranial boreholes (4 mm i.d) for insertion of an intracranial pressure (ICP) and a microdialysis catheter. To differentiate the effect of FPI from that of the surgical preparation, a sham group was exposed to the same surgical preparation but no FPI trauma (n = 8). A control group had no brain trauma and no surgical preparation (n = 5). PV was determined by a (125)I-albumin dilution technique. PV, electrolytes, pH, BE (base excess), hematocrit (Hct), PaO2, and PaCO2 were measured at baseline and after 3 h. Mean arterial pressure (MAP) was measured continuously. ICP was measured in the FPI and the sham group. In the FPI group, PV decreased by 11.2 mL/kg from 31.7 mL/kg (p < 0.01) with a simultaneous increase in Hct and decrease in pH. In the sham group, PV decreased by 5.7 mL/kg from 32.7 mL/kg (p < 0.01). The control group showed no PV reduction. The results support that an isolated severe head trauma triggers a significant and rapid reduction in PV, most likely due to vascular leak.

  3. Isolation of speech area from focal brain ischemia.

    PubMed

    Bogousslavsky, J; Regli, F; Assal, G

    1985-01-01

    A patient with atrial fibrillation and internal carotid artery occlusion developed mixed transcortical aphasia. The CT scan showed two recent distinct infarcts in the dominant hemisphere, one in the precentral artery area (pial artery infarct) and one in the borderzone area between the posterior and middle cerebral arteries territories (watershed infarct). The perisylvian speech areas were spared, but probably disconnected from other areas by the infarcts. The syndrome of isolation of speech area may be caused by vascular conditions which are able to produce simultaneous pial artery and watershed infarcts, and is not necessarily related to more extensive processes of the brain.

  4. Isolated angiitis in the hypothalamus mimicking brain tumor.

    PubMed

    Tsutsumi, Satoshi; Ito, Masanori; Yasumoto, Yukimasa; Kaneda, Kazuhiko

    2008-01-01

    A 64-year-old female presented with exaggerating somnolence without contributory medical and lifestyle histories. She was not aware of any preceding infection or headache. Cerebral magnetic resonance imaging demonstrated an isolated enhanced mass in the hypothalamus without meningeal enhancement. Blood and cerebrospinal fluid examinations showed no significant findings except for hypernatremia and hyperprolactinemia. She underwent an open biopsy via the interhemispheric route. Histological examination revealed marked perivascular lymphocytic aggregation with polyclonal immunostaining both for B and T lymphocytes. No findings suggestive of underlying malignancy were recognized. Extensive work-up aiming at systemic vasculitis and lymphoma revealed no signs of extracranial lesion, so the most probable diagnosis was isolated angiitis in the hypothalamus. Angiitis may originate from the hypothalamus and should be considered in the differential diagnosis of hypothalamic lesion mimicking brain tumor on neuroimaging.

  5. On-line two-dimensional capillary electrophoresis with mass spectrometric detection using a fully electric isolated mechanical valve.

    PubMed

    Kohl, Felix J; Montealegre, Cristina; Neusüß, Christian

    2016-04-01

    CE is becoming more and more important in many fields of bioanalytical chemistry. Besides optical detection, hyphenation to ESI-MS detection is increasingly applied for sensitive identification purposes. Unfortunately, many CE techniques and methods established in research and industry are not compatible to ESI-MS since essential components of the background electrolyte interfere in ES ionization. In order to identify unknown peaks in established CE methods, here, a heart-cut 2D-CE separation system is introduced using a fully isolated mechanical valve with an internal loop of only 20 nL. In this system, the sample is separated using potentially any non-ESI compatible method in the first separation dimension. Subsequently, the portion of interest is cut by the internal sample loop of the valve and reintroduced to the second dimension where the interfering compounds are removed, followed by ESI-MS detection. When comparing the separation efficiency of the system with the valve to a system using a continuous capillary only a slight increase in peak width is observed. Ultraviolet/visible detection is integrated in the first dimension for switching time determination, enabling reproducible cutting of peaks of interest. The feasibility of the system is successfully demonstrated by a 2D analysis of a BSA tryptic digest sample using a nonvolatile (phosphate based) background electrolyte in the first dimension.

  6. On ultrasound-induced microbubble oscillation in a capillary blood vessel and its implications for the blood-brain barrier

    NASA Astrophysics Data System (ADS)

    Wiedemair, W.; Tuković, Ž.; Jasak, H.; Poulikakos, D.; Kurtcuoglu, V.

    2012-02-01

    The complex interaction between an ultrasound-driven microbubble and an enclosing capillary microvessel is investigated by means of a coupled, multi-domain numerical model using the finite volume formulation. This system is of interest in the study of transient blood-brain barrier disruption (BBBD) for drug delivery applications. The compliant vessel structure is incorporated explicitly as a distinct domain described by a dedicated physical model. Red blood cells (RBCs) are taken into account as elastic solids in the blood plasma. We report the temporal and spatial development of transmural pressure (Ptm) and wall shear stress (WSS) at the luminal endothelial interface, both of which are candidates for the yet unknown mediator of BBBD. The explicit introduction of RBCs shapes the Ptm and WSS distributions and their derivatives markedly. While the peak values of these mechanical wall parameters are not affected considerably by the presence of RBCs, a pronounced increase in their spatial gradients is observed compared to a configuration with blood plasma alone. The novelty of our work lies in the explicit treatment of the vessel wall, and in the modelling of blood as a composite fluid, which we show to be relevant for the mechanical processes at the endothelium.

  7. Isolation of a large cholecystokinin precursor from canine brain.

    PubMed Central

    Eysselein, V E; Reeve, J R; Shively, J E; Miller, C; Walsh, J H

    1984-01-01

    Cholecystokinin (CCK)-like immunoreactivity (CCK-LI) in a pool of 12 dog brains was extracted sequentially into boiling water and cold 2% trifluoroacetic acid. Gel filtration on Sephadex G-50 revealed three main molecular forms detected by a carboxyl-terminal antibody; one was eluted in the position of CCK-58 (58 amino acid residues long); a second, in the position of CCK-8; and a third, near the radioactive iodide marker. When the CCK-LI was purified by affinity chromatography using carboxyl-terminal CCK antibody followed by three steps of reversed-phase high-pressure liquid chromatography, three components were isolated and characterized by sequence microanalysis. The smallest component was the pentapeptide common to gastrin and CCK. The second peak was eluted in the same region as synthetic CCK octapeptide, and sequence analysis showed that the chemical structure of this biologically active region of canine CCK is identical to that found in sheep and pig brains. The 22-residue amino-terminal sequence of brain CCK-58 was: Ala-Val-Gln-Lys-Val-Asp-Gly-Glu-Pro-Arg-Ala-His-Leu-Gly -Ala-Leu-leu-Ala-Arg-Tyr-Ile-Gln-, the same as the sequence found for canine intestinal CCK-58 from this pool of dogs. This is the same sequence others have reported for porcine brain CCK-58 lacking nine amino acid residues (CCK-58 desnonapeptide) except that the porcine peptide had a serine in position 9. The canine CCK amino-terminal sequence differed from the sequence Ala-Gln-Lys-Val-Asn-Ser previously reported for intestinal CCK-58 purified from another pool of dog tissue, but the rest of the residues identified were identical in the two peptides. CCK-58 may be a molecular precursor of the smaller forms of CCK in brain as well as in gut. PMID:6093106

  8. Isolation of primary murine brain microvascular endothelial cells.

    PubMed

    Ruck, Tobias; Bittner, Stefan; Epping, Lisa; Herrmann, Alexander M; Meuth, Sven G

    2014-11-14

    The blood-brain-barrier is ultrastructurally assembled by a monolayer of brain microvascular endothelial cells (BMEC) interconnected by a junctional complex of tight and adherens junctions. Together with other cell-types such as astrocytes or pericytes, they form the neurovascular unit (NVU), which specifically regulates the interchange of fluids, molecules and cells between the peripheral blood and the CNS. Through this complex and dynamic system BMECs are involved in various processes maintaining the homeostasis of the CNS. A dysfunction of the BBB is observed as an essential step in the pathogenesis of many severe CNS diseases. However, specific and targeted therapies are very limited, as the underlying mechanisms are still far from being understood. Animal and in vitro models have been extensively used to gain in-depth understanding of complex physiological and pathophysiological processes. By reduction and simplification it is possible to focus the investigation on the subject of interest and to exclude a variety of confounding factors. However, comparability and transferability are also reduced in model systems, which have to be taken into account for evaluation. The most common animal models are based on mice, among other reasons, mainly due to the constantly increasing possibilities of methodology. In vitro studies of isolated murine BMECs might enable an in-depth analysis of their properties and of the blood-brain-barrier under physiological and pathophysiological conditions. Further insights into the complex mechanisms at the BBB potentially provide the basis for new therapeutic strategies. This protocol describes a method to isolate primary murine microvascular endothelial cells by a sequence of physical and chemical purification steps. Special considerations for purity and cultivation of MBMECs as well as quality control, potential applications and limitations are discussed.

  9. Regional Blood Flow in the Normal and Ischemic Brain Is Controlled by Arteriolar Smooth Muscle Cell Contractility and Not by Capillary Pericytes.

    PubMed

    Hill, Robert A; Tong, Lei; Yuan, Peng; Murikinati, Sasidhar; Gupta, Shobhana; Grutzendler, Jaime

    2015-07-01

    The precise regulation of cerebral blood flow is critical for normal brain function, and its disruption underlies many neuropathologies. The extent to which smooth muscle-covered arterioles or pericyte-covered capillaries control vasomotion during neurovascular coupling remains controversial. We found that capillary pericytes in mice and humans do not express smooth muscle actin and are morphologically and functionally distinct from adjacent precapillary smooth muscle cells (SMCs). Using optical imaging we investigated blood flow regulation at various sites on the vascular tree in living mice. Optogenetic, whisker stimulation, or cortical spreading depolarization caused microvascular diameter or flow changes in SMC but not pericyte-covered microvessels. During early stages of brain ischemia, transient SMC but not pericyte constrictions were a major cause of hypoperfusion leading to thrombosis and distal microvascular occlusions. Thus, capillary pericytes are not contractile, and regulation of cerebral blood flow in physiological and pathological conditions is mediated by arteriolar SMCs. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. Antagonistic regulation of native Ca2+- and ATP-sensitive cation channels in brain capillaries by nucleotides and decavanadate.

    PubMed

    Csanády, László; Adam-Vizi, Vera

    2004-06-01

    Regulation by cytosolic nucleotides of Ca2+- and ATP-sensitive nonselective cation channels (CA-NSCs) in rat brain capillary endothelial cells was studied in excised inside-out patches. Open probability (Po) was suppressed by cytosolic nucleotides with apparent KI values of 17, 9, and 2 microM for ATP, ADP, and AMP, as a consequence of high-affinity inhibition of channel opening rate and low-affinity stimulation of closing rate. Cytosolic [Ca2+] and voltage affected inhibition of Po, but not of opening rate, by ATP, suggesting that the conformation of the nucleotide binding site is influenced only by the state of the channel gate, not by that of the Ca2+ and voltage sensors. ATP inhibition was unaltered by channel rundown. Nucleotide structure affected inhibitory potency that was little sensitive to base substitutions, but was greatly diminished by 3'-5' cyclization, removal of all phosphates, or complete omission of the base. In contrast, decavanadate potently (K1/2 = 90 nM) and robustly stimulated Po, and functionally competed with inhibitory nucleotides. From kinetic analyses we conclude that (a) ATP, ADP, and AMP bind to a common site; (b) inhibition by nucleotides occurs through simple reversible binding, as a consequence of tighter binding to the closed-channel relative to the open-channel conformation; (c) the conformation of the nucleotide binding site is not directly modulated by Ca2+ and voltage; (d) the differences in inhibitory potency of ATP, ADP, and AMP reflect their different affinities for the closed channel; and (e) though decavanadate is the only example found to date of a compound that stimulates Po with high affinity even in the presence of millimolar nucleotides, apparently by competing for the nucleotide binding site, a comparable mechanism might allow CA-NSC channels to open in living cells despite physiological levels of nucleotides. Decavanadate now provides a valuable tool for studying native CA-NSC channels and for screening cloned

  11. Activity of adenosine diphosphates and triphosphates on a P2YT-type receptor in brain capillary endothelial cells

    PubMed Central

    Simon, J; Vigne, P; Eklund, K M; Michel, A D; Carruthers, A M; Humphrey, P P A; Frelin, C; Barnard, E A

    2001-01-01

    A P2Y (nucleotide) receptor activity in a clonal population (B10) of rat brain capillary endothelial cells is coupled to inhibition of adenylyl cyclase and has functional similarities to the P2YT (previously designated ‘P2T') receptor for ADP of blood platelets. However, the only P2Y receptor which was detectable in a previous study of B10 cells by mRNA analysis was the P2Y1 receptor, which elsewhere shows no transduction via cyclic nucleotides. We have sought here to clarify these issues. The inhibition of forskolin-stimulated adenylyl cyclase induced by purified nucleotides was measured on B10 cells. The EC50 value for 2-methylthioADP (2-MeSADP) was 2.2 nM and, surprisingly, 2-MeSATP was an almost equally strong agonist (EC50=3.5 nM). ATP and 2-ClATP were weak partial agonists (EC50=26 μM and 10 μM respectively) and under appropriate conditions could antagonise the activity on 2-MeSADP. A known selective antagonist of the platelet P2YT receptor, 2-propylthioadenosine-5′-(β,γ)-difluoromethylene) triphosphonate (AR-C 66096), was a competitive antagonist of this B10 cell receptor, with pKB=7.6. That ligand is inactive at the P2Y1 receptor in the same cells. Conversely, the competitive P2Y1 receptor antagonists, the 3′, 5′- and 2′, 5′-adenosine bis-monophosphates, are, instead, weak agonists at the adenylyl cyclase-inhibitory receptor. The inhibition of adenylyl cyclase by 2-MeSADP was completely abolished by pertussis toxin. In summary, these brain endothelial cells possess a P2YT-type receptor in addition to the P2Y1 receptor. The two have similarities in agonist profiles but are clearly distinguishable by antagonists and by their second messenger activations. The possible relationships between the B10 and platelet P2YT receptors are discussed. PMID:11156575

  12. ISOLATION AND STRUCTURAL STUDIES ON SYNAPTIC COMPLEXES FROM RAT BRAIN

    PubMed Central

    Cotman, Carl W.; Taylor, Dwan

    1972-01-01

    A fraction enriched in synaptic complexes has been isolated from rat brain. The major structural elements of synaptic complexes after isolation are a sector of pre- and postsynaptic plasma membranes joined together by a synaptic cleft and a postsynaptic density (PSD) located on the inner surface of the postsynaptic membrane. On its outer surface, the postsynaptic membrane has a series of projections which extend about halfway into the cleft and which occur along the entire length of the PSD. Proteolytic enzymes at high concentrations remove the PSD and open the synaptic cleft; at low concentrations the PSD is selectively destroyed. By contrast, the structural integrity of the PSD is resistant to treatment with NaCl, EGTA, and low concentrations of urea. Pre- and postsynaptic membranes also remain joined by the synaptic cleft after NaCl, EGTA, or mild urea treatment. High concentrations of urea cause the partial dissociation of the PSD. We conclude that polypeptides are probably one of the major components of the PSD and that the structural integrity of the PSD depends on polypeptides because disruption of the covalent or hydrophobic bonding of these polypeptides leads to a progressive loss of PSD structure. PMID:4656707

  13. Quantification of Transporter and Receptor Proteins in Dog Brain Capillaries and Choroid Plexus: Relevance for the Distribution in Brain and CSF of Selected BCRP and P-gp Substrates.

    PubMed

    Braun, Clemens; Sakamoto, Atsushi; Fuchs, Holger; Ishiguro, Naoki; Suzuki, Shinobu; Cui, Yunhai; Klinder, Klaus; Watanabe, Michitoshi; Terasaki, Tetsuya; Sauer, Achim

    2017-10-02

    Transporters at the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB) play a pivotal role as gatekeepers for efflux or uptake of endogenous and exogenous molecules. The protein expression of a number of them has already been determined in the brains of rodents, nonhuman primates, and humans using quantitative targeted absolute proteomics (QTAP). The dog is an important animal model for drug discovery and development, especially for safety evaluations. The purpose of the present study was to clarify the relevance of the transporter protein expression for drug distribution in the dog brain and CSF. We used QTAP to examine the protein expression of 17 selected transporters and receptors at the dog BBB and BCSFB. For the first time, we directly linked the expression of two efflux transporters, P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), to regional brain and CSF distribution using specific substrates. Two cocktails, each containing one P-gp substrate (quinidine or apafant) and one BCRP substrate (dantrolene or daidzein) were infused intravenously prior to collection of the brain. Transporter expression varied only slightly between the capillaries of different brain regions and did not result in region-specific distribution of the investigated substrates. There were, however, distinct differences between brain capillaries and choroid plexus. Largest differences were observed for BCRP and P-gp: both were highly expressed in brain capillaries, but no BCRP and only low amounts of P-gp were detected in the choroid plexus. Kp,uu,brain and Kp,uu,CSF of both P-gp substrates were indicative of drug efflux. Also, Kp,uu,brain for the BCRP substrates was low. In contrast, Kp,uu,CSF for both BCRP substrates was close to unity, resulting in Kp,uu,CSF/Kp,uu,brain ratios of 7 and 8, respectively. We conclude that the drug transporter expression profiles differ between the BBB and BCSFB in dogs, that there are species differences in the

  14. Transport of monocarboxylic acids at the blood-brain barrier: Studies with monolayers of primary cultured bovine brain capillary endothelial cells

    SciTech Connect

    Terasaki, T.; Takakuwa, S.; Moritani, S.; Tsuji, A. )

    1991-09-01

    The kinetics and mechanism of the transport of monocarboxylic acids (MCAs) were studied by using primary cultured bovine brain capillary endothelial cells. Concentration-dependent uptake of acetic acid was observed, and the kinetic parameters were estimated as follows: the Michaelis constant, Kt, was 3.41 {plus minus} 1.87 mM, the maximum uptake rate, Jmax, was 144.7 {plus minus} 55.7 nmol/mg of protein/min and the nonsaturable first-order rate constant, Kd, was 6.66 {plus minus} 1.98 microliters/mg of protein/min. At medium pH below 7.0, the uptake rate of (3H)acetic acid increased markedly with decreasing medium pH, whereas pH-independent uptake was observed in the presence of 10 mM acetic acid. An energy requirement for (3H)acetic acid uptake was also demonstrated, because metabolic inhibitors (2,4-dinitrophenol and rotenone) reduced significantly the uptake rate (P less than .05). Carbonylcyanide-p-trifluoro-methoxyphenylhydrazone, a protonophore, inhibited significantly the uptake of (3H)acetic acid at medium pH of 5.0 and 6.0, whereas 4,4{prime}-diisothiocyanostilben-2,2{prime}-disulfonic acid did not. Several MCAs inhibited significantly the uptake rate of (3H)acetic acid, whereas di- and tricarboxylic acids did not. The uptake of (3H)acetic acid was competitively inhibited by salicylic acid, with an inhibition constant, Ki, of 3.60 mM, suggesting a common transport system between acetic acid and salicylic acid. Moreover, at the medium pH of 7.4, salicylic acid and valproic acid inhibited significantly the uptake of (3H)acetic acid, demonstrating that the transport of MCA drugs could also be ascribed to the MCA transport system at the physiologic pH.

  15. Regulation of store-operated Ca{sup 2+} entry activity by cell cycle dependent up-regulation of Orai2 in brain capillary endothelial cells

    SciTech Connect

    Kito, Hiroaki; Yamamura, Hisao; Suzuki, Yoshiaki; Yamamura, Hideto; Ohya, Susumu; Asai, Kiyofumi; Imaizumi, Yuji

    2015-04-10

    Store-operated Ca{sup 2+} entry (SOCE) via Orai1 and STIM1 complex is supposed to have obligatory roles in the regulation of cellular functions of vascular endothelial cells, while little is known about the contribution of Orai2. Quantitative PCR and Western blot analyses indicated the expression of Orai2 and STIM2, in addition to Orai1 and STIM1 in bovine brain capillary endothelial cell line, t-BBEC117. During the exponential growth of t-BBEC117, the knockdown of Orai1 and STIM1 significantly reduced the SOCE activity, whereas Orai2 and STIM2 siRNAs had no effect. To examine whether endogenous SOCE activity contributes to the regulation of cell cycle progression, t-BBEC117 were synchronized using double thymidine blockage. At the G2/M phase, Ca{sup 2+} influx via SOCE was decreased and Orai2 expression was increased compared to the G0/G1 phase. When Orai2 was knocked down at the G2/M phase, the decrease in SOCE was removed, and cell proliferation was partly attenuated. Taken together, Orai1 significantly contributes to cell proliferation via the functional expression, which is presumably independent of the cell cycle phases. In construct, Orai2 is specifically up-regulated during the G2/M phase, negatively modulates the SOCE activity, and may contribute to the regulation of cell cycle progression in brain capillary endothelial cells. - Highlights: • Orai1 is essential for SOCE activity in brain capillary endothelial cells (BCECs). • Cell cycle independent expression of Orai1 regulated SOCE and cell proliferation. • Orai2 was up-regulated only at G2/M phase and this consequently reduced SOCE. • Orai2 as well as Orai1 is a key player controlling SOCE and proliferation in BCECs.

  16. Characteristics of L-citrulline transport through blood-brain barrier in the brain capillary endothelial cell line (TR-BBB cells).

    PubMed

    Lee, Kyeong-Eun; Kang, Young-Sook

    2017-05-10

    L-Citrulline is a neutral amino acid and a major precursor of L-arginine in the nitric oxide (NO) cycle. Recently it has been reported that L-citrulline prevents neuronal cell death and protects cerebrovascular injury, therefore, L-citrulline may have a neuroprotective effect to improve cerebrovascular dysfunction. Therefore, we aimed to clarify the brain transport mechanism of L-citrulline through blood-brain barrier (BBB) using the conditionally immortalized rat brain capillary endothelial cell line (TR-BBB cells), as an in vitro model of the BBB. The uptake study of [(14)C] L-citrulline, quantitative real-time polymerase chain reaction (PCR) analysis, and rLAT1, system b(0,+), and CAT1 small interfering RNA study were performed in TR-BBB cells. The uptake of [(14)C] L-citrulline was a time-dependent, but ion-independent manner in TR-BBB cells. The transport process involved two saturable components with a Michaelis-Menten constant of 30.9 ± 1.0 μM (Km1) and 1.69 ± 0.43 mM (Km2). The uptake of [(14)C] L-citrulline in TR-BBB cells was significantly inhibited by neutral and cationic amino acids, but not by anionic amino acids. In addition, [(14)C]L-citrulline uptake in the cells was markedly inhibited by 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH), which is the inhibitor of the large neutral amino acid transporter 1 (LAT1), B(0), B(0,+) and harmaline, the inhibitor of system b(0,+). Gabapentin and L-dopa as the substrates of LAT1 competitively inhibited the uptake of [(14)C] L-citrulline. IC50 values for L-dopa, gabapentin, L-phenylalanine and L-arginine were 501 μM, 223 μM, 68.9 μM and 33.4 mM, respectively. The expression of mRNA for LAT1 was predominantly increased 187-fold in comparison with that of system b(0,+) in TR-BBB cells. In the studies of LAT1, system b(0,+) and CAT1 knockdown via siRNA transfection into TR-BBB cells, the transcript level of LAT1 and [(14)C] L-citrulline uptake by LAT1 siRNA were significantly reduced

  17. Involvement of insulin-degrading enzyme in insulin- and atrial natriuretic peptide-sensitive internalization of amyloid-β peptide in mouse brain capillary endothelial cells.

    PubMed

    Ito, Shingo; Ohtsuki, Sumio; Murata, Sho; Katsukura, Yuki; Suzuki, Hiroya; Funaki, Miho; Tachikawa, Masanori; Terasaki, Tetsuya

    2014-01-01

    Cerebral clearance of amyloid-β peptide (Aβ), which is implicated in Alzheimer's disease, involves elimination across the blood-brain barrier (BBB), and we previously showed that an insulin-sensitive process is involved in the case of Aβ1-40. The purpose of this study was to clarify the molecular mechanism of the insulin-sensitive Aβ1-40 elimination across mouse BBB. An in vivo cerebral microinjection study demonstrated that [125I]hAβ1-40 elimination from mouse brain was inhibited by human natriuretic peptide (hANP), and [125I]hANP elimination was inhibited by hAβ1-40, suggesting that hAβ1-40 and hANP share a common elimination process. Internalization of [125I]hAβ1-40 into cultured mouse brain capillary endothelial cells (TM-BBB4) was significantly inhibited by either insulin, hANP, other natriuretic peptides or insulin-degrading enzyme (IDE) inhibitors, but was not inhibited by phosphoramidon or thiorphan. Although we have reported the involvement of natriuretic peptide receptor C (Npr-C) in hANP internalization, cells stably expressing Npr-C internalized [125I]hANP but not [125I]hAβ1-40, suggesting that there is no direct interaction between Npr-C and hAβ1-40. IDE was detected in plasma membrane of TM-BBB4 cells, and internalization of [125I]hAβ1-40 by TM-BBB4 cells was reduced by IDE-targeted siRNAs. We conclude that elimination of hAβ1-40 from mouse brain across the BBB involves an insulin- and ANP-sensitive process, mediated by IDE expressed in brain capillary endothelial cells.

  18. Extraction of water labeled with oxygen 15 during single-capillary transit. Influence of blood pressure, osmolarity, and blood-brain barrier damage

    SciTech Connect

    Go, K.G.; Lammertsma, A.A.; Paans, A.M.; Vaalburg, W.; Woldring, M.G.

    1981-09-01

    By external detection, the influence of arterial blood pressure (BP), osmolarity, and cold-induced blood-brain barrier damage was assessed on the extraction of water labeled with oxygen 15 during single-capillary transit in the rat. There was an inverse relation between arterial BP and extraction that was attributable to the influence of arterial BP on cerebral blood flow (CBF) and the relation between CBF and extraction. Neither arterial BP nor osmolarity of the injected bolus had any direct effect on extraction of water 15O, signifying that the diffusional exchange component (determined by blood flow) of extraction greatly surpasses the convection flow contribution by hydrostatic or osmotic forces. Damage to the blood-brain barrier did not change its permeability to water.

  19. Regulation of store-operated Ca2+ entry activity by cell cycle dependent up-regulation of Orai2 in brain capillary endothelial cells.

    PubMed

    Kito, Hiroaki; Yamamura, Hisao; Suzuki, Yoshiaki; Yamamura, Hideto; Ohya, Susumu; Asai, Kiyofumi; Imaizumi, Yuji

    2015-04-10

    Store-operated Ca(2+) entry (SOCE) via Orai1 and STIM1 complex is supposed to have obligatory roles in the regulation of cellular functions of vascular endothelial cells, while little is known about the contribution of Orai2. Quantitative PCR and Western blot analyses indicated the expression of Orai2 and STIM2, in addition to Orai1 and STIM1 in bovine brain capillary endothelial cell line, t-BBEC117. During the exponential growth of t-BBEC117, the knockdown of Orai1 and STIM1 significantly reduced the SOCE activity, whereas Orai2 and STIM2 siRNAs had no effect. To examine whether endogenous SOCE activity contributes to the regulation of cell cycle progression, t-BBEC117 were synchronized using double thymidine blockage. At the G2/M phase, Ca(2+) influx via SOCE was decreased and Orai2 expression was increased compared to the G0/G1 phase. When Orai2 was knocked down at the G2/M phase, the decrease in SOCE was removed, and cell proliferation was partly attenuated. Taken together, Orai1 significantly contributes to cell proliferation via the functional expression, which is presumably independent of the cell cycle phases. In construct, Orai2 is specifically up-regulated during the G2/M phase, negatively modulates the SOCE activity, and may contribute to the regulation of cell cycle progression in brain capillary endothelial cells. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Up-regulation of K{sub ir}2.1 by ER stress facilitates cell death of brain capillary endothelial cells

    SciTech Connect

    Kito, Hiroaki; Yamazaki, Daiju; Ohya, Susumu; Yamamura, Hisao; Asai, Kiyofumi; Imaizumi, Yuji

    2011-07-29

    Highlights: {yields} We found that application of endoplasmic reticulum (ER) stress with tunicamycin to brain capillary endothelial cells (BCECs) induced cell death. {yields} The ER stress facilitated the expression of inward rectifier K{sup +} channel (K{sub ir}2.1) and induced sustained membrane hyperpolarization. {yields} The membrane hyperpolarization induced sustained Ca{sup 2+} entry through voltage-independent nonspecific cation channels and consequently facilitated cell death. {yields} The K{sub ir}2.1 up-regulation by ER stress is, at least in part, responsible for cell death of BCECs under pathological conditions. -- Abstract: Brain capillary endothelial cells (BCECs) form blood brain barrier (BBB) to maintain brain homeostasis. Cell turnover of BCECs by the balance of cell proliferation and cell death is critical for maintaining the integrity of BBB. Here we found that stimuli with tunicamycin, endoplasmic reticulum (ER) stress inducer, up-regulated inward rectifier K{sup +} channel (K{sub ir}2.1) and facilitated cell death in t-BBEC117, a cell line derived from bovine BCECs. The activation of K{sub ir} channels contributed to the establishment of deeply negative resting membrane potential in t-BBEC117. The deep resting membrane potential increased the resting intracellular Ca{sup 2+} concentration due to Ca{sup 2+} influx through non-selective cation channels and thereby partly but significantly regulated cell death in t-BBEC117. The present results suggest that the up-regulation of K{sub ir}2.1 is, at least in part, responsible for cell death/cell turnover of BCECs induced by a variety of cellular stresses, particularly ER stress, under pathological conditions.

  1. Structural and functional effects of social isolation on the hippocampus of rats with traumatic brain injury.

    PubMed

    Khodaie, Babak; Lotfinia, Ahmad Ali; Ahmadi, Milad; Lotfinia, Mahmoud; Jafarian, Maryam; Karimzadeh, Fariba; Coulon, Philippe; Gorji, Ali

    2015-02-01

    Social isolation has significant long-term psychological and physiological consequences. Both social isolation and traumatic brain injury (TBI) alter normal brain function and structure. However, the influence of social isolation on recovery from TBI is unclear. This study aims to evaluate if social isolation exacerbates the anatomical and functional deficits after TBI in young rats. Juvenile male rats were divided into four groups; sham operated control with social contacts, sham control with social isolation, TBI with social contacts, and TBI with social isolation. During four weeks after brain injury in juvenile rats, we evaluated the animal behaviors by T-maze and open-field tests, recorded brain activity with electrocorticograms and assessed structural changes by histological procedures in the hippocampal dentate gyrus, CA1, and CA3 areas. Our findings revealed significant memory impairments and hyperactivity conditions in rats with TBI and social isolation compared to the other groups. Histological assessments showed an increase of the mean number of dark neurons, apoptotic cells, and caspase-3 positive cells in all tested areas of the hippocampus in TBI rats with and without social isolation compared to sham rats. Furthermore, social isolation significantly increased the number of dark cells, apoptotic neurons, and caspase-3 positive cells in the hippocampal CA3 region in rats with TBI. This study indicates the harmful effect of social isolation on anatomical and functional deficits induced by TBI in juvenile rats. Prevention of social isolation may improve the outcome of TBI.

  2. UFLC/MS-IT-TOF guided isolation of anti-HBV active chlorogenic acid analogues from Artemisia capillaris as a traditional Chinese herb for the treatment of hepatitis.

    PubMed

    Zhao, Yong; Geng, Chang-An; Ma, Yun-Bao; Huang, Xiao-Yan; Chen, Hao; Cao, Tuan-Wu; He, Kang; Wang, Hao; Zhang, Xue-Mei; Chen, Ji-Jun

    2014-10-28

    Hepatitis B induced by HBV is a serious health problem. Artemisia capillaris (Yin-Chen) has long been used to treat hepatitis in traditional Chinese medicine. Coumarins, flavonoids and organic acids were revealed as its hepatoprotective and choleretic components, but its anti-HBV active components remain unknown. This current study focused on its anti-HBV active constituents by various chromatographic methods. LC/MS and bioassay-guided fractionation on the active extract of Artemisia capillaris led to the isolation of nine chlorogenic acid analogues. Structures of the isolates were elucidated by MS/MS and NMR techniques. Anti-HBV assay was performed on HepG 2.2.15 cell line in vitro: reduction of HBsAg and HBeAg secretions was measured by an ELISA method; inhibition of HBV DNA replication was monitored by real-time quantitative PCR and cellular toxicity was assessed by a MTT method. The 90% ethanol extract of Artemisia capillaris (Fr. AC) showed significantly inhibitory activity on HBV DNA replication with an IC₅₀ value of 76.1 ± 3.9 μg/mL and low cytotoxic effects (SI>20.1). To clarify its active constituents, the extract was further separated into 3 sub-fractions (AC-1, AC-2 and AC-3), of which Fr. AC-2 was the most active fraction against HBeAg secretion and HBV DNA replication with IC50 values of 44.2 ± 2.8 and 23.2 ± 1.9 μg/mL. Nine chlorogenic acid analogues were detected from the active part (Fr. AC-2) by a LC/MS technique and further separated by a HPLC method. The isolates were determined as chlorogenic acid (1), cryptochlorogenic acid (2), neochlorogenic acid (3), 3,5-dicaffeoylquinic acid (4), 4,5-dicaffeoylquinic acid (5), 3,4-dicaffeoylquinic acid (6), chlorogenic acid methyl ester (7), cryptochlorogenic acid methyl ester (8), neochlorogenic acid methyl ester (9). Compounds 1-6 possessed potent activity against HBV DNA replication with IC50 values in the range of 5.5 ± 0.9-13.7 ± 1.3 μM. Di-caffeoyl analogues (4-6) also exhibited activity

  3. Targeting anti-transferrin receptor antibody (OX26) and OX26-conjugated liposomes to brain capillary endothelial cells using in situ perfusion.

    PubMed

    Gosk, Sara; Vermehren, Charlotte; Storm, Gert; Moos, Torben

    2004-11-01

    Brain capillary endothelial cells (BCECs) express transferrin receptors. The uptake of a potential drug vector (OX26, or anti-transferrin receptor antibody IgG2a) conjugated to polyethyleneglycol-coated liposomes by BCECs was studied using in situ perfusion in 18-day-old rats in which the uptake of OX26 is almost twice as high as in the adult rat. Using radio-labeling, the uptake of OX26 by BCECs after 15-minute perfusion was approximately 16 times higher than that of nonimmune IgG2a (Ni-IgG2a). OX26 and OX26-conjugated liposomes selectively distributed to BCECs, leaving choroid plexus epithelium, neurons, and glia unlabeled. Ni-IgG2a and unconjugated liposomes did not reveal any labeling of BCECs. The labeling of BCECs by OX26 was profoundly higher than that of transferrin. Perfusion with albumin for 15 minutes did not reveal any labeling of neurons or glia, thus confirming the integrity of the blood-brain barrier. The failure to label neurons and glia shows that OX26 and OX26-conjugated liposomes did not pass through BCECs. The expression of transferrin receptors by endothelial cells selective to the brain qualifies OX26 as a candidate for blood-to-endothelium transport. A specifically designed formulation of liposomes may allow for their degradation within BCECs, leading to subsequent transport of liposomal cargo further into the brain.

  4. Strawberries on the brain--intracranial capillary hemangioma: two case reports and systematic literature review in children and adults.

    PubMed

    Mirza, Bilal; Shi, William Y; Phadke, Rahul; Holton, Janice L; Turner, Chris; Plant, Gordon T; Brew, Stefan; Kitchen, Neil; Zrinzo, Ludvic

    2013-12-01

    Capillary hemangioma in the cranial cavity is rare. This report describes 2 additional cases presenting shortly after pregnancy and provides a systematic review summarizing clinical experience to date. Case reports were compiled retrospectively. Patient 1 was a 28-year-old woman who presented with a simple partial seizure associated with left-sided visual distortions that progressed to a secondary generalized tonic-clonic seizure. Imaging revealed a contrast-enhancing lesion in the right temporal region adjacent to the transverse sinus, with vasogenic edema and scalloping of the inner table. The angiographic appearance suggested the correct diagnosis. Patient 2 was a 41-year-old woman who presented with progressive visual disturbance 6 months after giving birth to her second child. Subtle symptoms of headache and visual disturbance had commenced during pregnancy. Noncontrasted imaging displayed a homogenous tumor with surrounding vasogenic edema in the occipital region. PubMed and Science Citation Index were reviewed systematically for prior publications. Complete surgical excision was performed in both patients. Neuropathological examination confirmed benign capillary hemangioma consisting of a meshwork of capillary-sized and larger feeder vessels in both instances. Systematic review was based on 20 publications and a total of 24 patients; their clinical, radiological, and surgical features as well as management are summarized. At follow-up, symptoms had resolved and magnetic resonance imaging confirmed complete removal in both patients. Intracranial capillary hemangioma is rare, and treatment is empirical. Review of limited published reports suggests that surgery is the most commonly used treatment and that complete excision seems most likely to prevent further recurrence. Copyright © 2013 Elsevier Inc. All rights reserved.

  5. Preservation of mitochondrial functional integrity in mitochondria isolated from small cryopreserved mouse brain areas.

    PubMed

    Valenti, Daniela; de Bari, Lidia; De Filippis, Bianca; Ricceri, Laura; Vacca, Rosa Anna

    2014-01-01

    Studies of mitochondrial bioenergetics in brain pathophysiology are often precluded by the need to isolate mitochondria immediately after tissue dissection from a large number of brain biopsies for comparative studies. Here we present a procedure of cryopreservation of small brain areas from which mitochondrial enriched fractions (crude mitochondria) with high oxidative phosphorylation efficiency can be isolated. Small mouse brain areas were frozen and stored in a solution containing glycerol as cryoprotectant. Crude mitochondria were isolated by differential centrifugation from both cryopreserved and freshly explanted brain samples and were compared with respect to their ability to generate membrane potential and produce ATP. Intactness of outer and inner mitochondrial membranes was verified by polarographic ascorbate and cytochrome c tests and spectrophotometric assay of citrate synthase activity. Preservation of structural integrity and oxidative phosphorylation efficiency was successfully obtained in crude mitochondria isolated from different areas of cryopreserved mouse brain samples. Long-term cryopreservation of small brain areas from which intact and phosphorylating mitochondria can be isolated for the study of mitochondrial bioenergetics will significantly expand the study of mitochondrial defects in neurological pathologies, allowing large comparative studies and favoring interlaboratory and interdisciplinary analyses.

  6. Isolation and Functional Assessment of Mitochondria from Small Amounts of Mouse Brain Tissue

    PubMed Central

    Chinopoulos, Christos; Zhang, Steven F.; Thomas, Bobby; Ten, Vadim; Starkov, Anatoly A.

    2013-01-01

    Recent discoveries have brought mitochondria functions in focus of the neuroscience research community and greatly stimulated the demand for approaches to study mitochondria dysfunction in neurodegenerative diseases. Many mouse disease models have been generated, but studying mitochondria isolated from individual mouse brain regions is a challenge because of small amount of the available brain tissue. Conventional techniques for isolation and purification of mitochondria from mouse brain subregions, such as ventral midbrain, hippocampus, or striatum, require pooling brain tissue from six to nine animals for a single mitochondrial preparation. Working with pooled tissue significantly decreases the quality of data because of the time required to dissect several brains. It also greatly increases the labor intensity and the cost of experiments as several animals are required per single data point. We describe a method for isolation of brain mitochondria from mouse striata or other 7–12 mg brain samples. The method utilizes a refrigerated table-top microtube centrifuge, and produces research grade quality mitochondria in amounts sufficient for performing multiple enzymatic and functional assays, thereby eliminating the necessity for pooling mouse brain tissue. We also include a method of measuring ADP-ATP exchange rate as a function of mitochondrial membrane potential (ΔΨm) in small amounts of isolated mitochondria, adapted to a plate reader format. PMID:21913109

  7. Isolation and functional assessment of mitochondria from small amounts of mouse brain tissue.

    PubMed

    Chinopoulos, Christos; Zhang, Steven F; Thomas, Bobby; Ten, Vadim; Starkov, Anatoly A

    2011-01-01

    Recent discoveries have brought mitochondria functions in focus of the neuroscience research community and greatly stimulated the demand for approaches to study mitochondria dysfunction in neurodegenerative diseases. Many mouse disease models have been generated, but studying mitochondria isolated from individual mouse brain regions is a challenge because of small amount of the available brain tissue. Conventional techniques for isolation and purification of mitochondria from mouse brain subregions, such as ventral midbrain, hippocampus, or striatum, require pooling brain tissue from six to nine animals for a single mitochondrial preparation. Working with pooled tissue significantly decreases the quality of data because of the time required to dissect several brains. It also greatly increases the labor intensity and the cost of experiments as several animals are required per single data point. We describe a method for isolation of brain mitochondria from mouse striata or other 7-12 mg brain samples. The method utilizes a refrigerated table-top microtube centrifuge, and produces research grade quality mitochondria in amounts sufficient for performing multiple enzymatic and functional assays, thereby eliminating the necessity for pooling mouse brain tissue. We also include a method of measuring ADP-ATP exchange rate as a function of mitochondrial membrane potential (ΔΨm) in small amounts of isolated mitochondria, adapted to a plate reader format.

  8. The in vitro isolated whole guinea pig brain as a model to study epileptiform activity patterns.

    PubMed

    de Curtis, Marco; Librizzi, Laura; Uva, Laura

    2016-02-15

    Research on ictogenesis is based on the study of activity between seizures and during seizures in animal models of epilepsy (chronic condition) or in in vitro slices obtained from naïve non-epileptic brains after treatment with pro-convulsive drugs, manipulations of the extracellular medium and specific stimulation protocols. The in vitro isolated guinea pig brain retains the functional connectivity between brain structures and maintains interactions between neuronal, glial and vascular compartments. It is a close-to-in vivo preparation that offers experimental advantages not achieved with the use of other experimental models. Neurophysiological and imaging techniques can be utilized in this preparation to study brain activity during and between seizures induced by pharmacological or functional manipulations. Cellular and network determinants of interictal and ictal discharges that reproduce abnormal patterns observed in human focal epilepsies and the associated changes in extracellular ion and blood-brain permeability can be identified and analyzed in the isolated guinea pig brain. Ictal and interictal patterns recorded in in vitro slices may show substantial differences from seizure activity recorded in vivo due to slicing procedure itself. The isolated guinea pig brain maintained in vitro by arterial perfusion combines the typical facilitated access of in vitro preparations, that are difficult to approach during in vivo experiments, with the preservation of larger neuronal networks. The in vitro whole isolated guinea pig brain preparation offers an unique experimental model to study systemic and neurovascular changes during ictogenesis. Published by Elsevier B.V.

  9. A sensitive capillary LC-UV method for the simultaneous analysis of olanzapine, chlorpromazine and their FMO-mediated N-oxidation products in brain microdialysates.

    PubMed

    Hendrickx, Stijn; Uğur, Duygu Yeniceli; Yilmaz, Işil Tan; Şener, Erol; Van Schepdael, Ann; Adams, Erwin; Broeckhoven, Ken; Cabooter, Deirdre

    2017-01-01

    A specific and sensitive capillary liquid chromatography-ultraviolet detection (cap-LC-UV) method in combination with a micro-extraction by packed sorbent (MEPS) sample clean-up procedure has been developed and validated for the simultaneous analysis of chlorpromazine, olanzapine and their flavin-containing monooxygenase (FMO) mediated N-oxides in rat brain microdialysates. Chromatographic separation was obtained on an Acclaim Pepmap RP C18 column with an ID of 300µm. An injection volume of 20µL was used to inject the largely aqueous samples and was shown to have no influence on the obtained peak shape of the compounds of interest. Optimal conditions for MEPS extraction were obtained on a mixed-mode M1 (80% C8, 20% SCX) cartridge after diluting microdialysate samples with phosphate buffer pH 2.5 (1:3 v/v). The method was validated and lower limits of quantification (LLOQ) were determined at 0.5nM for all compounds. Linearity was demonstrated between the LLOQ and 1µM for all compounds (R(2)>0.995). MEPS recoveries were between 92% and 98%, with intra- and interday variabilities below 15%. The applicability of the developed method was successfully demonstrated by analysing rat brain microdialysates. The capillary LC-UV method in combination with MEPS sample treatment provides a simple, sensitive method to quantify all compounds of interest in 45min and can be applied for routine therapeutic monitoring and pharmacokinetic studies of olanzapine, chlorpromazine and their respective N-oxides. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Isolation of individual fatty acids in sediments using preparative capillary gas chromatography (PCGC) for radiocarbon analysis at NIES-TERRA

    NASA Astrophysics Data System (ADS)

    Uchida, Masao; Shibata, Yasuyuki; Kawamura, Kimitaka; Yoneda, Minoru; Mukai, Hitoshi; Tanaka, Atsushi; Uehiro, Takashi; Morita, Masatoshi

    2000-10-01

    Compound-specific radiocarbon analysis (CSRA) of individual fatty acids (140-1190 μg C) in an estuarine sediment sample collected from Tokyo Bay was carried out using a recently developed preparative capillary gas chromatography (PCGC) system and accelerator mass spectrometry (AMS). The results showed that the estimated 14C ages of four components greatly varied from modern age (combined iso and anteiso C 15:0, C 16:0) to 17 000 years BP (C 22:0), while a bulk-phase 14C age of organic matter is 5000 years BP. The 14C ages of the fatty acids derived from phytoplankton and bacteria are much younger than that of the bulk phase. On the other hand, the fatty acid originated from terrestrial higher plants (C 22:0) shows an older 14C age of 17 000 years BP.

  11. A large-scale electrophoresis- and chromatography-based determination of gene expression profiles in bovine brain capillary endothelial cells after the re-induction of blood-brain barrier properties

    PubMed Central

    2010-01-01

    Background Brain capillary endothelial cells (BCECs) form the physiological basis of the blood-brain barrier (BBB). The barrier function is (at least in part) due to well-known proteins such as transporters, tight junctions and metabolic barrier proteins (e.g. monoamine oxidase, gamma glutamyltranspeptidase and P-glycoprotein). Our previous 2-dimensional gel proteome analysis had identified a large number of proteins and revealed the major role of dynamic cytoskeletal remodelling in the differentiation of bovine BCECs. The aim of the present study was to elaborate a reference proteome of Triton X-100-soluble species from bovine BCECs cultured in the well-established in vitro BBB model developed in our laboratory. Results A total of 215 protein spots (corresponding to 130 distinct proteins) were identified by 2-dimensional gel electrophoresis, whereas over 350 proteins were identified by a shotgun approach. We classified around 430 distinct proteins expressed by bovine BCECs. Our large-scale gene expression analysis enabled the correction of mistakes referenced into protein databases (e.g. bovine vinculin) and constitutes valuable evidence for predictions based on genome annotation. Conclusions Elaboration of a reference proteome constitutes the first step in creating a gene expression database dedicated to capillary endothelial cells displaying BBB characteristics. It improves of our knowledge of the BBB and the key proteins in cell structures, cytoskeleton organization, metabolism, detoxification and drug resistance. Moreover, our results emphasize the need for both appropriate experimental design and correct interpretation of proteome datasets. PMID:21078152

  12. Functional expression of a proton-coupled organic cation (H+/OC) antiporter in human brain capillary endothelial cell line hCMEC/D3, a human blood–brain barrier model

    PubMed Central

    2013-01-01

    Background Knowledge of the molecular basis and transport function of the human blood–brain barrier (BBB) is important for not only understanding human cerebral physiology, but also development of new central nervous system (CNS)-acting drugs. However, few studies have been done using human brain capillary endothelial cells, because human brain materials are difficult to obtain. The purpose of this study is to clarify the functional expression of a proton-coupled organic cation (H+/OC) antiporter in human brain capillary endothelial cell line hCMEC/D3, which has been recently developed as an in vitro human BBB model. Methods Diphenhydramine, [3H]pyrilamine and oxycodone were used as cationic drugs that proved to be H+/OC antiporter substrates. The in vitro uptake experiments by hCMEC/D3 cells were carried out under several conditions. Results Diphenhydramine and [3H]pyrilamine were both transported into hCMEC/D3 cells in a time- and concentration-dependent manner with Km values of 59 μM and 19 μM, respectively. Each inhibited uptake of the other in a competitive manner, suggesting that a common mechanism is involved in their transport. The diphenhydramine uptake was significantly inhibited by amantadine and quinidine, but not tetraethylammonium and 1-methyl-4-phenylpyridinium (substrates for well-known organic cation transporters). The uptake was inhibited by metabolic inhibitors, but was insensitive to extracellular sodium and membrane potential. Further, the uptake was increased by extracellular alkalization and intracellular acidification. These transport properties are completely consistent with those of previously characterized H+/OC antiporter in rat BBB. Conclusions The present results suggest that H+/OC antiporter is functionally expressed in hCMEC/D3 cells. PMID:23351963

  13. One-step multiple component isolation from the oil of Crinitaria tatarica (Less.) Sojak. by preparative capillary gas with characterization by spectroscopic and spectrometric techniques and evaluation of biological activity

    USDA-ARS?s Scientific Manuscript database

    In the present work multiple component isolation from the oil of Crinitaria tatarica (Less.) Sojak. by Preparative Capillary Gas Chromatography (PCGC) with characterization by mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy have been carried out. Gas chromatography (GC-FID) ...

  14. Isolation and the Symptoms of Chronic Brain Syndrome.

    ERIC Educational Resources Information Center

    Ernst, Philip; And Others

    1978-01-01

    A literature review of internal and external changes on the mental and emotional status of elderly persons reveals physical, social, and emotional isolation to be the key intervening variables which disclose functional symptoms of mental disorders. (Author)

  15. Reduced cerebral glucose metabolism and increased brain capillary permeability following high-dose methotrexate chemotherapy: a positron emission tomographic study

    SciTech Connect

    Phillips, P.C.; Dhawan, V.; Strother, S.C.; Sidtis, J.J.; Evans, A.C.; Allen, J.C.; Rottenberg, D.A.

    1987-01-01

    Regional glucose metabolic rate constants and blood-to-brain transport of rubidium were estimated using positron emission tomography in an adolescent patient with a brain tumor, before and after chemotherapy with intravenous high-dose methotrexate. Widespread depression of cerebral glucose metabolism was apparent 24 hours after drug administration, which may reflect reduced glucose phosphorylation, and the influx rate constant for /sup 82/Rb was increased, indicating a drug-induced alteration in blood-brain barrier function. Associated changes in neuropsychological performance, electroencephalogram, and plasma amino acid concentration were identified in the absence of evidence of systemic methotrexate toxicity, suggesting primary methotrexate neurotoxicity.

  16. Simultaneous determination of biogenic monoamines in rat brain dialysates using capillary high-performance liquid chromatography with photoluminescence following electron transfer.

    PubMed

    Jung, Moon Chul; Shi, Guoyue; Borland, Laura; Michael, Adrian C; Weber, Stephen G

    2006-03-15

    Simultaneous determination of biogenic monoamines such as dopamine, serotonin, and 3-methoxytyramine in brain is important in understanding neurotransmitter activity. This study presents a sensitive determination of biogenic monoamines in rat brain striatum microdialysates using capillary high-performance liquid chromatography with the photoluminescence following electron-transfer detection technique. Separation conditions were optimized by changing the concentration of an ion-interaction agent and the percentage of an organic modifier. The high concentration of ion-interaction agent enabled the amines as a class to be separated from interfering acids, but also made the separation very long. To shorten the separation time, 10% (v/v) acetonitrile was used as the organic modifier. Eight chromatographic runs during a 3-h period were analyzed in terms of retention times, peak heights, and peak widths. Chromatograms are very reproducible, with less than 1% changes in peak height over 3 h. Typical concentration detection limits at the optimum separation conditions were less than 100 pM for metabolic acids and approximately 200 pM for monoamines. The injection volume of the sample was 500 nL. Thus, the mass detection limits were less than 50 amol for metabolic acids and approximately 100 amol for monoamines. Typical separation time was less than 10 min. To validate the technique, the separation method was applied to the observation of drug-induced changes of monoamine concentrations in rat brain microdialysis samples. Local perfusion of tetrodotoxin, a sodium channel blocker, into the striatum of an anesthetized rat decreased dopamine, 3-methoxytyramine, and serotonin concentrations in dialysates. Successive monitoring of striatal dialysates at a temporal resolution of 7.7 min showed that the injection of nomifensine transiently increased dopamine and 3-methoxytyramine concentrations in rat brain dialysate.

  17. Targeting Cells With MR Imaging Probes: Cellular Interaction And Intracellular Magnetic Iron Oxide Nanoparticles Uptake In Brain Capillary Endothelial and Choroidal Plexus Epithelial Cells

    NASA Astrophysics Data System (ADS)

    Cambianica, I.; Bossi, M.; Gasco, P.; Gonzalez, W.; Idee, J. M.; Miserocchi, G.; Rigolio, R.; Chanana, M.; Morjan, I.; Wang, D.; Sancini, G.

    2010-10-01

    Magnetic iron oxide nanoparticles (NPs) are considered for various diagnostic and therapeutic applications in brain including their use as contrast agent for magnetic resonance imaging. In delivery application, the critical step is the transport across cell layers and the internalization of NPs into specific cells, a process often limited by poor targeting specificity and low internalization efficiency. The development of the models of brain endothelial cells and choroidal plexus epithelial cells in culture has allowed us to investigate into these mechanisms. Our strategy is aimed at exploring different routes to the entrapment of iron oxide NPs in these brain related cells. Here we demonstrated that not only cells endowed with a good phagocytic activity like activated macrophages but also endothelial brain capillary and choroidal plexus epithelial cells do internalize iron oxide NPs. Our study of the intracellular trafficking of NPs by TEM, and confocal microscopy revealed that NPs are mainly internalized by the endocytic pathway. Iron oxide NPs were dispersed in water and coated with 3,4-dihydroxyl-L-phenylalanine (L-DOPA) using standard procedures. Magnetic lipid NPs were prepared by NANOVECTOR: water in oil in water (W/O/W) microemulsion process has been applied to directly coat different iron based NPs by lipid layer or to encapsulate them into Solid Lipid Nanoparticles (SLNs). By these coating/loading the colloidal stability was improved without strong alteration of the particle size distribution. Magnetic lipid NPs could be reconstituted after freeze drying without appreciable changes in stability. L-DOPA coated NPs are stable in PBS and in MEM (Modified Eagle Medium) medium. The magnetic properties of these NPs were not altered by the coating processes. We investigated the cellular uptake, cytotoxicity, and interaction of these NPs with rat brain capillary endothelial (REB4) and choroidal plexus epithelial (Z310) cells. By means of widefield, confocal

  18. Drug-induced trafficking of p-glycoprotein in human brain capillary endothelial cells as demonstrated by exposure to mitomycin C.

    PubMed

    Noack, Andreas; Noack, Sandra; Hoffmann, Andrea; Maalouf, Katia; Buettner, Manuela; Couraud, Pierre-Olivier; Romero, Ignacio A; Weksler, Babette; Alms, Dana; Römermann, Kerstin; Naim, Hassan Y; Löscher, Wolfgang

    2014-01-01

    P-glycoprotein (Pgp; ABCB1/MDR1) is a major efflux transporter at the blood-brain barrier (BBB), restricting the penetration of various compounds. In other tissues, trafficking of Pgp from subcellular stores to the cell surface has been demonstrated and may constitute a rapid way of the cell to respond to toxic compounds by functional membrane insertion of the transporter. It is not known whether drug-induced Pgp trafficking also occurs in brain capillary endothelial cells that form the BBB. In this study, trafficking of Pgp was investigated in human brain capillary endothelial cells (hCMEC/D3) that were stably transfected with a doxycycline-inducible MDR1-EGFP fusion plasmid. In the presence of doxycycline, these cells exhibited a 15-fold increase in Pgp-EGFP fusion protein expression, which was associated with an increased efflux of the Pgp substrate rhodamine 123 (Rho123). The chemotherapeutic agent mitomycin C (MMC) was used to study drug-induced trafficking of Pgp. Confocal fluorescence microscopy of single hCMEC/D3-MDR1-EGFP cells revealed that Pgp redistribution from intracellular pools to the cell surface occurred within 2 h of MMC exposure. Pgp-EGFP exhibited a punctuate pattern at the cell surface compatible with concentrated regions of the fusion protein in membrane microdomains, i.e., lipid rafts, which was confirmed by Western blot analysis of biotinylated cell surface proteins in Lubrol-resistant membranes. MMC exposure also increased the functionality of Pgp as assessed in three functional assays with Pgp substrates (Rho123, eFluxx-ID Gold, calcein-AM). However, this increase occurred with some delay after the increased Pgp expression and coincided with the release of Pgp from the Lubrol-resistant membrane complexes. Disrupting rafts by depleting the membrane of cholesterol increased the functionality of Pgp. Our data present the first direct evidence of drug-induced Pgp trafficking at the human BBB and indicate that Pgp has to be released from lipid

  19. Drug-Induced Trafficking of P-Glycoprotein in Human Brain Capillary Endothelial Cells as Demonstrated by Exposure to Mitomycin C

    PubMed Central

    Noack, Andreas; Noack, Sandra; Hoffmann, Andrea; Maalouf, Katia; Buettner, Manuela; Couraud, Pierre-Olivier; Romero, Ignacio A.; Weksler, Babette; Alms, Dana; Römermann, Kerstin; Naim, Hassan Y.; Löscher, Wolfgang

    2014-01-01

    P-glycoprotein (Pgp; ABCB1/MDR1) is a major efflux transporter at the blood-brain barrier (BBB), restricting the penetration of various compounds. In other tissues, trafficking of Pgp from subcellular stores to the cell surface has been demonstrated and may constitute a rapid way of the cell to respond to toxic compounds by functional membrane insertion of the transporter. It is not known whether drug-induced Pgp trafficking also occurs in brain capillary endothelial cells that form the BBB. In this study, trafficking of Pgp was investigated in human brain capillary endothelial cells (hCMEC/D3) that were stably transfected with a doxycycline-inducible MDR1-EGFP fusion plasmid. In the presence of doxycycline, these cells exhibited a 15-fold increase in Pgp-EGFP fusion protein expression, which was associated with an increased efflux of the Pgp substrate rhodamine 123 (Rho123). The chemotherapeutic agent mitomycin C (MMC) was used to study drug-induced trafficking of Pgp. Confocal fluorescence microscopy of single hCMEC/D3-MDR1-EGFP cells revealed that Pgp redistribution from intracellular pools to the cell surface occurred within 2 h of MMC exposure. Pgp-EGFP exhibited a punctuate pattern at the cell surface compatible with concentrated regions of the fusion protein in membrane microdomains, i.e., lipid rafts, which was confirmed by Western blot analysis of biotinylated cell surface proteins in Lubrol-resistant membranes. MMC exposure also increased the functionality of Pgp as assessed in three functional assays with Pgp substrates (Rho123, eFluxx-ID Gold, calcein-AM). However, this increase occurred with some delay after the increased Pgp expression and coincided with the release of Pgp from the Lubrol-resistant membrane complexes. Disrupting rafts by depleting the membrane of cholesterol increased the functionality of Pgp. Our data present the first direct evidence of drug-induced Pgp trafficking at the human BBB and indicate that Pgp has to be released from lipid

  20. Isolation and Perivascular Localization of Mesenchymal Stem Cells From Mouse Brain

    PubMed Central

    Kang, Seok-Gu; Shinojima, Naoki; Hossain, Anwar; Gumin, Joy; Yong, Raymund L.; Colman, Howard; Marini, Frank; Andreeff, Michael; Lang, Frederick F.

    2012-01-01

    BACKGROUND Although originally isolated from the bone marrow, mesenchymal stem cells (MSCs) have recently been detected in other tissues. However, little is known about MSCs in the brain. OBJECTIVE To determine the extent to which cells with the features of MSCs exist in normal brain tissue and to determine the location of these cells in the brain. METHODS Single-cell suspensions from mouse brains were cultured according to the same methods used for culturing bone marrow–derived MSCs (BM-MSCs). These brain-derived cells were analyzed by fluorescence-activated cell sorting for surface markers associated with BM-MSCs (stem cell antigen 1 [Sca-1+], CD9+, CD45−, CD11b−, and CD31−). Brain-derived cells were exposed to mesenchymal differentiation conditions. To determine the locations of these cells within the brain, sections of normal brains were analyzed by immunostaining for Sca-1, CD31, and nerve/glial antigen 2. RESULTS Cells morphologically similar to mouse BM-MSCs were identified and called brain-derived MSCs (Br-MSCs). Fluorescence-activated cell sorting indicated that the isolated cells had a surface marker profile similar to BM-MSCs, ie, Sca-1+, CD9+, CD45−, and CD11b−. Like BM-MSCs, Br-MSCs were capable of differentiation into adipocytes, osteocytes, and chondrocytes. Immunostaining indicated that Sca-1+ Br-MSCs are located around blood vessels and may represent progenitor cells that serve as a source of mesenchymal elements (eg, pericytes) within the brain. CONCLUSION Our results indicate that cells similar to BM-MSCs exist in the brain. These Br-MSCs appear to be located within the vascular niche and may provide the mesenchymal elements of this niche. Because MSCs may be part of the cellular response to tissue injury, Br-MSCs may represent targets in the therapy of pathological processes such as stroke, trauma, and tumorigenesis. PMID:20651630

  1. Capillary electrophoresis.

    PubMed

    Compton, S W; Brownlee, R G

    1988-05-01

    While capillary electrophoresis, or historically related techniques, have been used for over a century, and recognition of the value of this separation methodology has certainly grown rapidly in the past few years, the technique has generally been used by analytical chemists, particularly in Europe and Japan, and small groups of researchers in the United States. Many of the basic instrumentation problems have been solved only relatively recently, and researchers using capillary electrophoresis are now turning their attention to studying specific applications which demonstrate the potential versatility of this electrophoretic technique. The appearance of standardized commercial instrumentation is imminent. With the availability of such technology, capillary electrophoresis will no longer be an academic curiosity, but rather a tool with the potential for routine separations of diverse samples of interest to analyst, researcher, and clinician.

  2. Isolation of viable Neospora caninum from brains of wild gray wolves (Canis lupus).

    PubMed

    Dubey, J P; Jenkins, M C; Ferreira, L R; Choudhary, S; Verma, S K; Kwok, O C H; Fetterer, R; Butler, E; Carstensen, M

    2014-03-17

    Neospora caninum is a common cause of abortion in cattle worldwide. Canids, including the dog and the dingo (Canis familiaris), the coyote (Canis latrans), and the gray wolf (Canis lupus) are its definitive hosts that can excrete environmentally resistant oocysts in the environment, but also can act as intermediate hosts, harboring tissue stages of the parasite. In an attempt to isolate viable N. caninum from tissues of naturally infected wolves, brain and heart tissue from 109 wolves from Minnesota were bioassayed in mice. Viable N. caninum (NcWolfMn1, NcWolfMn2) was isolated from the brains of two wolves by bioassays in interferon gamma gene knockout mice. DNA obtained from culture-derived N. caninum tachyzoites of the two isolates were analyzed by N. caninum-specific Nc5 polymerase chain reaction and confirmed diagnosis. This is the first report of isolation of N. caninum from tissues of any wild canid host. Published by Elsevier B.V.

  3. Social isolation and brain development in the ant Camponotus floridanus

    NASA Astrophysics Data System (ADS)

    Seid, Marc A.; Junge, Erich

    2016-06-01

    Social interactions play a key role in the healthy development of social animals and are most pronounced in species with complex social networks. When developing offspring do not receive proper social interaction, they show developmental impairments. This effect is well documented in mammalian species but controversial in social insects. It has been hypothesized that the enlargement of the mushroom bodies, responsible for learning and memory, observed in social insects is needed for maintaining the large social networks and/or task allocation. This study examines the impact of social isolation on the development of mushroom bodies of the ant Camponotus floridanus. Ants raised in isolation were shown to exhibit impairment in the growth of the mushroom bodies as well as behavioral differences when compared to ants raised in social groups. These results indicate that social interaction is necessary for the proper development of C. floridanus mushroom bodies.

  4. Characterization of isolates of Citrus tristeza virus by sequential analyses of enzyme immunoassays and capillary electrophoresis-single-strand conformation polymorphisms.

    PubMed

    Licciardello, G; Raspagliesi, D; Bar-Joseph, M; Catara, A

    2012-05-01

    Citrus tristeza virus (CTV) is the causal agent of tristeza disease, which is one of the most devastating diseases of citrus crops worldwide. This paper describes a method for the rapid detection and genotyping of naturally spreading CTV isolates. This method uses ELISA or dot-blot immunological tests to detect trees infected with CTV. The reaction wells or membrane spots for which there is a positive reaction are sequentially treated by (i) washing and elution of viral RNA from the trapped samples, (ii) one-step synthesis of cDNA and PCR and (iii) automated fluorescence-based capillary electrophoresis single-strand conformation polymorphism (CE-SSCP) analysis of amplification products. Comparative CE-SSCP results are presented for CTV RNA extracted directly from infected leaves and ELISA plates or from membranes. In the analyses of all of these RNA samples, the p18, p27 and p23 CTV genes were targeted for amplification. Specific profiles of forward and reverse strands were obtained from a group of eight CTV isolates collected in Sicily, each with distinct biological characteristics, which were analyzed using the conventional two-step procedure (immunological detection followed by CE-SSCP molecular characterization after RNA isolation) or in a continuous process of ELISA/CE-SSCP or dot-blot/CE-SSCP starting from infected plant material. The combined method is simple, highly sensitive and reproducible, thus allowing the processing of numerous field samples for a variety of epidemiological needs. The sequential processing of an ELISA or dot-blot/ELISA followed by CE-SSCP is expected to allow the rapid detection of recent CTV infections along with the simultaneous characterization of the genetic diversity and structure of the population of newly invading CTV.

  5. An Isolation Method for Assessment of Brain Mitochondria Function in Neonatal Mice with Hypoxic-Ischemic Brain Injury

    PubMed Central

    Caspersen, Casper S.; Sosunov, Alexander; Utkina-Sosunova, Irina; Ratner, Veniamin I.; Starkov, Anatoly A.; Ten, Vadim S.

    2010-01-01

    This work was undertaken to develop a method for the isolation of mitochondria from a single cerebral hemisphere in neonatal mice. Mitochondria from the normal mouse brain hemisphere isolated by the proposed method exhibited a good respiratory control ratio of 6.39 ± 0.53 during glutamate-malate-induced phosphorylating respiration. Electron microscopy showed intact mitochondria. The applicability of this method was tested on mitochondria isolated from naïve mice and their littermates subjected to hypoxic-ischemic insult. Hypoxic-ischemic insult prior to reperfusion resulted in a significant (p < 0.01) inhibition of phosphorylating respiration compared to naïve littermates. This was associated with a profound depletion of the ATP content in the ischemic hemisphere. The expression for Mn superoxide dismutase and cytochrome C (markers for the integrity of the mitochondrial matrix and outer membrane) was determined by Western blot to control for mitochondrial integrity and quantity in the compared samples. Thus, we have developed a method for the isolation of the cerebral mitochondria from a single hemisphere adapted to neonatal mice. This method may serve as a valuable tool to study mitochondrial function in a mouse model of immature brain injury. In addition, the suggested method enables us to examine the mitochondrial functional phenotype in immature mice with a targeted genetic alteration. PMID:18349523

  6. Surgical treatment of non-small cell lung cancer with isolated synchronous brain metastases.

    PubMed

    I, Hoseok; Lee, Jung Il; Nam, Do Hyun; Ahn, Yong Chan; Shim, Young Mog; Kim, Kwhanmien; Choi, Yong Soo; Kim, Jhingook

    2006-04-01

    This study is a retrospective examination of our experiences with patients who underwent treatment of isolated synchronous brain metastases coupled with primary non-small cell lung cancer. From January 1995 to June 2004, 12 patients presented with isolated synchronous brain metastases coupled with primary non-small cell lung cancer. The patient was comprised of 8 men and 4 women. The median age was 52 yr, in a range of 32 to 75 yr. Median follow-up duration was 10.6 months, in a range of 2 to 55.8 months. Recurrence developed in 7 patients, and the median interval from 1st treatment to recurrence was 4.5 months (2.8-6.5 months). The overall 1-yr survival rate was 61.7%. The 1-yr survival rates for pathologic N0 and N1 cases were 75% and 66.7%, respectively. The median survival duration for pathologic N2 was 6.2 months (95% CI, 4.8-7.5 months). The 1-yr survival rate for cases of single brain metastasis was 75%. Based on our current observations, we could speculate that aggressive management of primary non-small cell lung cancer and isolated synchronous brain metastases was beneficial in a selected group of patients, as long as the brain lesions and pulmonary lesions were limited or resectable.

  7. Surgical Treatment of Non-Small Cell Lung Cancer with Isolated Synchronous Brain Metastases

    PubMed Central

    I, Hoseok; Lee, Jung Il; Nam, Do Hyun; Ahn, Yong Chan; Shim, Young Mog; Kim, Kwhanmien; Choi, Yong Soo

    2006-01-01

    This study is a retrospective examination of our experiences with patients who underwent treatment of isolated synchronous brain metastases coupled with primary non-small cell lung cancer. From January 1995 to June 2004, 12 patients presented with isolated synchronous brain metastases coupled with primary non-small cell lung cancer. The patient was comprised of 8 men and 4 women. The median age was 52 yr, in a range of 32 to 75 yr. Median follow-up duration was 10.6 months, in a range of 2 to 55.8 months. Recurrence developed in 7 patients, and the median interval from 1st treatment to recurrence was 4.5 months (2.8-6.5 months). The overall 1-yr survival rate was 61.7%. The 1-yr survival rates for pathologic N0 and N1 cases were 75% and 66.7%, respectively. The median survival duration for pathologic N2 was 6.2 months (95% CI, 4.8-7.5 months). The 1-yr survival rate for cases of single brain metastasis was 75%. Based on our current observations, we could speculate that aggressive management of primary non-small cell lung cancer and isolated synchronous brain metastases was beneficial in a selected group of patients, as long as the brain lesions and pulmonary lesions were limited or resectable. PMID:16614507

  8. ALTERATIONS IN BRAIN CREATINE CONCENTRATIONS UNDER LONG-TERM SOCIAL ISOLATION (EXPERIMENTAL STUDY).

    PubMed

    Koshoridze, N; Kuchukashvili, Z; Menabde, K; Lekiashvili, Sh; Koshoridze, M

    2016-02-01

    Stress represents one of the main problems of modern humanity. This study was done for understanding more clearly alterations in creatine content of the brain under psycho-emotional stress induced by long-term social isolation. It was shown that under 30 days social isolation creatine amount in the brain was arisen, while decreasing concentrations of synthesizing enzymes (AGAT, GAMT) and creatine transporter protein (CrT). Another important point was that such changes were accompanied by down-regulation of creatine kinase (CK), therefore the enzyme's concentration was lowered. In addition, it was observed that content of phosphocreatine (PCr) and ATP were also reduced, thus indicating down-regulation of energy metabolism of brain that is really a crucial point for its normal functioning. To sum up the results it can be underlined that long-term social isolation has negative influence on energy metabolism of brain; and as a result reduce ATP content, while increase of free creatine concentration, supposedly maintaining maximal balance for ATP amount, but here must be also noted that up-regulated oxidative pathways might have impact on blood brain barrier, resulting on its permeability.

  9. Distinct Patterns of Brain Function in Children with Isolated Spelling Impairment: New Insights

    ERIC Educational Resources Information Center

    Gebauer, Daniela; Enzinger, Christian; Kronbichler, Martin; Schurz, Matthias; Reishofer, Gernot; Koschutnig, Karl; Kargl, Reinhard; Purgstaller, Christian; Fazekas, Franz; Fink, Andreas

    2012-01-01

    Studies investigating reading and spelling difficulties heavily focused on the neural correlates of reading impairments, whereas spelling impairments have been largely neglected so far. Hence, the aim of the present study was to investigate brain structure and function of children with isolated spelling difficulties. Therefore, 31 children, aged…

  10. Distinct Patterns of Brain Function in Children with Isolated Spelling Impairment: New Insights

    ERIC Educational Resources Information Center

    Gebauer, Daniela; Enzinger, Christian; Kronbichler, Martin; Schurz, Matthias; Reishofer, Gernot; Koschutnig, Karl; Kargl, Reinhard; Purgstaller, Christian; Fazekas, Franz; Fink, Andreas

    2012-01-01

    Studies investigating reading and spelling difficulties heavily focused on the neural correlates of reading impairments, whereas spelling impairments have been largely neglected so far. Hence, the aim of the present study was to investigate brain structure and function of children with isolated spelling difficulties. Therefore, 31 children, aged…

  11. Method for isolation and molecular characterization of extracellular microvesicles released from brain endothelial cells.

    PubMed

    Haqqani, Arsalan S; Delaney, Christie E; Tremblay, Tammy-Lynn; Sodja, Caroline; Sandhu, Jagdeep K; Stanimirovic, Danica B

    2013-01-10

    In addition to possessing intracellular vesicles, eukaryotic cells also produce extracellular microvesicles, ranging from 50 to 1000 nm in diameter that are released or shed into the microenvironment under physiological and pathological conditions. These membranous extracellular organelles include both exosomes (originating from internal vesicles of endosomes) and ectosomes (originating from direct budding/shedding of plasma membranes). Extracellular microvesicles contain cell-specific collections of proteins, glycoproteins, lipids, nucleic acids and other molecules. These vesicles play important roles in intercellular communication by acting as carrier for essential cell-specific information to target cells. Endothelial cells in the brain form the blood-brain barrier, a specialized interface between the blood and the brain that tightly controls traffic of nutrients and macromolecules between two compartments and interacts closely with other cells forming the neurovascular unit. Therefore, brain endothelial cell extracellular microvesicles could potentially play important roles in 'externalizing' brain-specific biomarkers into the blood stream during pathological conditions, in transcytosis of blood-borne molecules into the brain, and in cell-cell communication within the neurovascular unit. To study cell-specific molecular make-up and functions of brain endothelial cell exosomes, methods for isolation of extracellular microvesicles using mass spectrometry-compatible protocols and the characterization of their signature profiles using mass spectrometry -based proteomics were developed. A total of 1179 proteins were identified in the isolated extracellular microvesicles from brain endothelial cells. The microvesicles were validated by identification of almost 60 known markers, including Alix, TSG101 and the tetraspanin proteins CD81 and CD9. The surface proteins on isolated microvesicles could potentially interact with both primary astrocytes and cortical neurons

  12. Isolation and Flow Cytometric Analysis of Immune Cells from the Ischemic Mouse Brain

    PubMed Central

    Boltze, Johannes; Wagner, Daniel-Christoph; Weise, Gesa

    2016-01-01

    Ischemic stroke initiates a robust inflammatory response that starts in the intravascular compartment and involves rapid activation of brain resident cells. A key mechanism of this inflammatory response is the migration of circulating immune cells to the ischemic brain facilitated by chemokine release and increased endothelial adhesion molecule expression. Brain-invading leukocytes are well-known contributing to early-stage secondary ischemic injury, but their significance for the termination of inflammation and later brain repair has only recently been noticed. Here, a simple protocol for the efficient isolation of immune cells from the ischemic mouse brain is provided. After transcardial perfusion, brain hemispheres are dissected and mechanically dissociated. Enzymatic digestion with Liberase is followed by density gradient (such as Percoll) centrifugation to remove myelin and cell debris. One major advantage of this protocol is the single-layer density gradient procedure which does not require time-consuming preparation of gradients and can be reliably performed. The approach yields highly reproducible cell counts per brain hemisphere and allows for measuring several flow cytometry panels in one biological replicate. Phenotypic characterization and quantification of brain-invading leukocytes after experimental stroke may contribute to a better understanding of their multifaceted roles in ischemic injury and repair. PMID:26967380

  13. Electro membrane isolation of nerve agent degradation products across a supported liquid membrane followed by capillary electrophoresis with contactless conductivity detection.

    PubMed

    Xu, Li; Hauser, Peter C; Lee, Hian Kee

    2008-12-19

    In the present study, electro membrane isolation (EMI) of four nerve agent degradation products has been successfully explored. In the procedure, a polypropylene sheet membrane folded into an envelope with an open end with its wall pores impregnated with 1-octanol was employed as the artificial supported liquid membrane (SLM). The envelope containing the extractant or aqueous acceptor phase (at pH 6.8) was immersed in the sample or donor phase (also aqueous at a pH of 6.8) for extraction. This ensured that the target analytes were fully ionized. A voltage was then applied, with the negative electrode placed in the donor phase with agitation, and the positive electrode in the acceptor phase. The ionized analytes were thus driven to migrate from the donor phase across the SLM to the acceptor phase. The factors influential to extraction: type of organic solvent, voltage, agitation speed, extraction time, pH of the donor and acceptor phase and concentration of humic acids were investigated in detail. After extraction, the acceptor phase was collected and directly injected for capillary electrophoretic (CE) analysis. Combined with capacitively coupled contactless conductivity detection (C(4)D), the direct detection of these compounds could be achieved. Moreover, large-volume sample injection was employed to further enhance the sensitivity of this method. Limits of detection (LODs) as low as ng/mL were reached for the studied analytes, with overall LOD enhancements of four orders of magnitude.

  14. Acute modulation of sugar transport in brain capillary endothelial cell cultures during activation of the metabolic stress pathway.

    PubMed

    Cura, Anthony J; Carruthers, Anthony

    2010-05-14

    GLUT1-catalyzed equilibrative sugar transport across the mammalian blood-brain barrier is stimulated during acute and chronic metabolic stress; however, the mechanism of acute transport regulation is unknown. We have examined acute sugar transport regulation in the murine brain microvasculature endothelial cell line bEnd.3. Acute cellular metabolic stress was induced by glucose depletion, by potassium cyanide, or by carbonyl cyanide p-trifluoromethoxyphenylhydrazone, which reduce or deplete intracellular ATP within 15 min. This results in a 1.7-7-fold increase in V(max) for zero-trans 3-O-methylglucose uptake (sugar uptake into sugar-free cells) and a 3-10-fold increase in V(max) for equilibrium exchange transport (intracellular [sugar] = extracellular [sugar]). GLUT1, GLUT8, and GLUT9 mRNAs are detected in bEnd.3 cells where GLUT1 mRNA levels are 33-fold greater than levels of GLUT8 or GLUT9 mRNA. Neither GLUT1 mRNA nor total protein levels are affected by acute metabolic stress. Cell surface biotinylation reveals that plasma membrane GLUT1 levels are increased 2-3-fold by metabolic depletion, although cell surface Na(+),K(+)-ATPase levels remain unaffected by ATP depletion. Treatment with the AMP-activated kinase agonist, AICAR, increases V(max) for net 3-O-methylglucose uptake by 2-fold. Glucose depletion and treatment with potassium cyanide, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, and AICAR also increase AMP-dependent kinase phosphorylation in bEnd.3 cells. These results suggest that metabolic stress rapidly stimulates blood-brain barrier endothelial cell sugar transport by acute up-regulation of plasma membrane GLUT1 levels, possibly involving AMP-activated kinase activity.

  15. Acute Modulation of Sugar Transport in Brain Capillary Endothelial Cell Cultures during Activation of the Metabolic Stress Pathway*

    PubMed Central

    Cura, Anthony J.; Carruthers, Anthony

    2010-01-01

    GLUT1-catalyzed equilibrative sugar transport across the mammalian blood-brain barrier is stimulated during acute and chronic metabolic stress; however, the mechanism of acute transport regulation is unknown. We have examined acute sugar transport regulation in the murine brain microvasculature endothelial cell line bEnd.3. Acute cellular metabolic stress was induced by glucose depletion, by potassium cyanide, or by carbonyl cyanide p-trifluoromethoxyphenylhydrazone, which reduce or deplete intracellular ATP within 15 min. This results in a 1.7–7-fold increase in Vmax for zero-trans 3-O-methylglucose uptake (sugar uptake into sugar-free cells) and a 3–10-fold increase in Vmax for equilibrium exchange transport (intracellular [sugar] = extracellular [sugar]). GLUT1, GLUT8, and GLUT9 mRNAs are detected in bEnd.3 cells where GLUT1 mRNA levels are 33-fold greater than levels of GLUT8 or GLUT9 mRNA. Neither GLUT1 mRNA nor total protein levels are affected by acute metabolic stress. Cell surface biotinylation reveals that plasma membrane GLUT1 levels are increased 2–3-fold by metabolic depletion, although cell surface Na+,K+-ATPase levels remain unaffected by ATP depletion. Treatment with the AMP-activated kinase agonist, AICAR, increases Vmax for net 3-O-methylglucose uptake by 2-fold. Glucose depletion and treatment with potassium cyanide, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, and AICAR also increase AMP-dependent kinase phosphorylation in bEnd.3 cells. These results suggest that metabolic stress rapidly stimulates blood-brain barrier endothelial cell sugar transport by acute up-regulation of plasma membrane GLUT1 levels, possibly involving AMP-activated kinase activity. PMID:20231288

  16. Quantitation of dopamine, serotonin and adenosine content in a tissue punch from a brain slice using capillary electrophoresis with fast-scan cyclic voltammetry detection.

    PubMed

    Fang, Huaifang; Pajski, Megan L; Ross, Ashley E; Venton, B Jill

    2013-01-01

    Methods to determine neurochemical concentrations in small samples of tissue are needed to map interactions among neurotransmitters. In particular, correlating physiological measurements of neurotransmitter release and the tissue content in a small region would be valuable. HPLC is the standard method for tissue content analysis but it requires microliter samples and the detector often varies by the class of compound being quantified; thus detecting molecules from different classes can be difficult. In this paper, we develop capillary electrophoresis with fast-scan cyclic voltammetry detection (CE-FSCV) for analysis of dopamine, serotonin, and adenosine content in tissue punches from rat brain slices. Using field-amplified sample stacking, the limit of detection was 5 nM for dopamine, 10 nM for serotonin, and 50 nM for adenosine. Neurotransmitters could be measured from a tissue punch as small as 7 µg (7 nL) of tissue, three orders of magnitude smaller than a typical HPLC sample. Tissue content analysis of punches in successive slices through the striatum revealed higher dopamine but lower adenosine content in the anterior striatum. Stimulated dopamine release was measured in a brain slice, then a tissue punch collected from the recording region. Dopamine content and release had a correlation coefficient of 0.71, which indicates much of the variance in stimulated release is due to variance in tissue content. CE-FSCV should facilitate measurements of tissue content in nanoliter samples, leading to a better understanding of how diseases or drugs affect dopamine, serotonin, and adenosine content.

  17. Isolation, purification, and characterization of bovine brain cathepsin D

    SciTech Connect

    Karunatilake, C.

    1989-01-01

    Bovine brain cathepsin D was purified 1120-fold from enriched white matter with an overall yield of 46%. The enzyme is a single polypeptide chain with an apparent molecular weight of 42,000 as estimated by SDS-polyacrylamide gel electrophoresis and sephadex G-100 gel filtration. This is in sharp contrast to porcine spleen cathepsin D which consist of two polypeptide chains held together by noncovalent associations. Secretion of cathepsin D by cultured rat peritoneal macrophages was studied. These cells secreted the lysosomal marker enzyme {beta}-glucuronidase and lactic acid when challenged with derivatized, noningestible central nervous system tissue in a dose- and time-dependent manner. Secreted cathepsin D levels, however, were insignificant when measured with {sup 14}C-hemoglobin substrate. It was observed, however, that certain monosaccharides such as mannose and N-acetyl glucosamine were able to release cell surface bound cathepsin D. The observed specificity and the calculated binding constant indicated that cathepsin D bound to the mannose/N-acetyl glucosamine receptor of macrophages.

  18. [Rapid isolation of 4-hydroxyphenylethanol from the bovine brain using Water Sep-Pak Plus cartridges with lipophilic sorbent (Florisil)].

    PubMed

    Panova, N G; Veselovskaia, N V; Medvedev, A E

    1996-01-01

    4-Hydroxypherlylethanol, MAO-A Inhibitory component of endogenous MAO Inhibitory activity, tribulin, was isolated from a small quantity of bovine brain using Maters Sep-Pak Florisil cartridges. Procedure included isolation of crude tribulin fraction, which was passed through the cartridge. Preliminary data revealed that this substance is not absorbed on Florisil. The employment of this type of cartridges essentially accelerates isolation of 4-hydroxyphenyethanol from brain samples for quantitative determination. Pilot experiments also revealed that Sep-Pak Folrisil cartridges could be used for the rapid isolation of isatin, another endogenous MAO (B) inhibitor, from brain tissue.

  19. [Electrophysiological features (EEG) of ethanol withdrawal syndromes on isolated perfused rat brain].

    PubMed

    Tezikov, E B; Litvicki, P F

    2015-01-01

    On isolated rat brains we studied native EEC and its derivates (mean EEC amplitude and power spectrums - Fourier transformation) during perfusion with ethanol (65 Mm/ L) and after its withdrawal. Previously rats were undergone ethanol burden for 6 days according to Majchrowicz procedures to get alcohol withdrawal syndrome. Duration perfusion without ethanol was 5, 10 and 20 min depending on the experimental schedule. Ethanol infusion between periods of withdrawal comprised 20 min. 55% of isolated brains shown epileptiform activity after 1-2 min of ethanol withdrawal but others manifested only increased mean amplitude and the power spectrums of EEC as well as an appearance of single or batch spikes. Differences between in vivo and in vitro conditions can be explained by the accelerated rate of ethanol elimination. The high positive correlation was obtained between EEC findings at the 5-th min of the first ethanol withdrawal and the same findings at the 5-th min of ethanol withdrawal in the second and the third episodes of ethanol withdrawal. Prolongation of withdrawal period more than 5th min caused brain death showing epileptiform activity. Isolated rat brain is the convenient subject to study pathogenesis of excitability of neurons and examination of drugs to treat alcohol withdrawal syndrome.

  20. Metabolism of deoxypyrimidines and deoxypyrimidine antiviral analogs in isolated brain mitochondria

    PubMed Central

    McCann, Kathleen A.; Williams, David W.; McKee, Edward E.

    2012-01-01

    The goal of this project was to characterize deoxypyrimidine salvage pathways used to maintain deoxynucleoside triphosphate pools in isolated brain mitochondria and to determine the extent that antiviral pyrimidine analogs utilize or affect these pathways. Mitochondria from rat brains were incubated in media with labeled and unlabeled deoxynucleosides and deoxynucleoside analogs. Products were analyzed by HPLC coupled to an inline UV monitor and liquid scintillation counter. Isolated mitochondria transported thymidine and deoxycytidine into the matrix, and readily phosphorylated both of these to mono-, di, and tri-phosphate nucleotides. Rates of phosphorylation were much higher than rates observed in mitochondria from heart and liver. Deoxyuridine was phosphorylated much more slowly than thymidine and only to dUMP. AZT, an antiviral thymidine analog, was phosphorylated to AZT-MP as readily as thymidine was phosphorylated to TMP, but little if any AZT-DP or AZT-TP was observed. AZT at 5.5 ± 1.7 μM was shown to inhibit thymidine phosphorylation by 50%, but was not observed to inhibit deoxycytidine phosphorylation except at levels > 100 μM. Stavudine and lamivudine were inert when incubated with isolated brain mitochondria. The kinetics of phosphorylation of thymidine, dC and AZT were significantly different in brain mitochondria compared to mitochondria from liver and heart. PMID:22530558

  1. Exercise reduces activation of microglia isolated from hippocampus and brain of aged mice

    PubMed Central

    2013-01-01

    Background Aging is associated with low-grade neuroinflammation that includes basal increases in proinflammatory cytokines and expression of inflammatory markers on microglia. Exercise can reduce neuroinflammation following infection in aged animals, but whether exercise modulates basal changes in microglia activation is unknown. Therefore, we evaluated changes in basal microglia activation in cells isolated from the hippocampus and remaining brain following running-wheel access. Methods Adult (4 months) and aged (22 months) male and female BALB/c mice were housed with or without running wheels for 10 weeks. Microglia were isolated from the hippocampus or remaining brain. Flow cytometry was used to determine microglia (CD11b+ and CD45low) that co-labeled with CD86, CD206, and MHC II. Results Aged mice showed a greater proportion of CD86 and MHC II positive microglia. In aged females, access to a running wheel decreased proportion of CD86+ and MHC II+ microglia in the hippocampus whereas aged males in the running group showed a decrease in the proportion of CD86+ microglia in the brain and an increase in the proportion of MHC II+ microglia in hippocampus and brain. Conclusion Overall, these data indicate that running-wheel access modulates microglia activation, but these effects vary by age, sex, and brain region. PMID:24044641

  2. Isolation of murine postnatal brain microglia for phenotypic characterization using magnetic cell separation technology.

    PubMed

    Harms, Ashley S; Tansey, Malú G

    2013-01-01

    To shorten the time between brain harvesting and microglia isolation, and characterization, we utilized the MACS(®) neural dissociation kit followed by OctoMACS(®) CD11b magnetic bead isolation technique to positively select for brain microglia expressing the pan-microglial marker CD11b, a key subunit of the membrane attack complex (MAC). This protocol yields a viable and highly pure (>95%) microglial population of approximately 500,000 cells per pup that is amenable for in vitro characterization within hours or days after being harvested from brain tissue. Primary microglia from C57Bl/6 mice were plated for next-day analyses of morphology and cellular markers by immunocytochemistry or for analysis of gene expression under resting or LPS-stimulated conditions. The ease of isolation enables investigators to perform molecular and cellular analyses without having to wait 1-2 weeks to isolate microglia by conventional methods involving mechanical agitation to dislodge these from astrocyte beds.

  3. The Brain Tourniquet: Physiological Isolation of Brain Regions Damaged by Traumatic Head Injury

    DTIC Science & Technology

    2008-06-19

    Richardson, B., Necrotic cell injury in the preterm and near-term ovine fetal brain after intermittent umbilical cord occlusion, American Journal of...physiological saline. We had originally proposed to use whole-cell, patch- clamp techniques to record from neocortical neurons, but found that these techniques...in layer V of neocortex (Fig I A and 1 E). Membrane potentials were measured using a high impedance amplifier operating in current- clamp mode

  4. Isolated spinal cord contusion in rats induces chronic brain neuroinflammation, neurodegeneration, and cognitive impairment

    PubMed Central

    Wu, Junfang; Stoica, Bogdan A; Luo, Tao; Sabirzhanov, Boris; Zhao, Zaorui; Guanciale, Kelsey; Nayar, Suresh K; Foss, Catherine A; Pomper, Martin G; Faden, Alan I

    2014-01-01

    Cognitive dysfunction has been reported in patients with spinal cord injury (SCI), but it has been questioned whether such changes may reflect concurrent head injury, and the issue has not been addressed mechanistically or in a well-controlled experimental model. Our recent rodent studies examining SCI-induced hyperesthesia revealed neuroinflammatory changes not only in supratentorial pain-regulatory sites, but also in other brain regions, suggesting that additional brain functions may be impacted following SCI. Here we examined effects of isolated thoracic SCI in rats on cognition, brain inflammation, and neurodegeneration. We show for the first time that SCI causes widespread microglial activation in the brain, with increased expression of markers for activated microglia/macrophages, including translocator protein and chemokine ligand 21 (C–C motif). Stereological analysis demonstrated significant neuronal loss in the cortex, thalamus, and hippocampus. SCI caused chronic impairment in spatial, retention, contextual, and fear-related emotional memory—evidenced by poor performance in the Morris water maze, novel objective recognition, and passive avoidance tests. Based on our prior work implicating cell cycle activation (CCA) in chronic neuroinflammation after SCI or traumatic brain injury, we evaluated whether CCA contributed to the observed changes. Increased expression of cell cycle-related genes and proteins was found in hippocampus and cortex after SCI. Posttraumatic brain inflammation, neuronal loss, and cognitive changes were attenuated by systemic post-injury administration of a selective cyclin-dependent kinase inhibitor. These studies demonstrate that chronic brain neurodegeneration occurs after isolated SCI, likely related to sustained microglial activation mediated by cell cycle activation. PMID:25483194

  5. Isolation of virus from brain after immunosuppression of mice with latent herpes simplex

    NASA Astrophysics Data System (ADS)

    Kastrukoff, Lorne; Long, Carol; Doherty, Peter C.; Wroblewska, Zofia; Koprowski, Hilary

    1981-06-01

    Herpes simplex virus (HSV) is usually present in a latent form in the trigeminal ganglion of man1-3. Various stress factors may induce virus reactivation, which is manifest by a lip lesion (innervated from the trigeminal ganglion) and the production of infectious virus. The considerable experimental efforts to define the conditions that lead to the reactivation of latent HSV have concentrated on isolating virus either from the original extraneural site of virus inoculation, or from cell-free homogenates of sensory ganglia from latently infected animals4-15. Recent DNA hybridization experiments resulted in the demonstration of the presence of HSV genomes in the brain tissue of both latently infected mice, and of humans who showed no clinical symptoms of HSV (ref. 16 and N. Fraser, personal communication). This led us to consider the possibility that HSV may be present in brain tissue as the result of either reactivation of the virus in brain cells or the passage of reactivated virus from trigeminal ganglia through the brain stem to the brain. The presence of infectious HSV in brain tissue has not previously been demonstrated; yet this could be a factor in chronic, relapsing neurological diseases such as multiple sclerosis. We have now shown experimentally that mice carrying latent HSV in their trigeminal ganglia may, following massive immunosuppression, express infectious virus in the central nervous system (CNS).

  6. Effect of re-expansion after short-period lung collapse on pulmonary capillary permeability and pro-inflammatory cytokine gene expression in isolated rabbit lungs.

    PubMed

    Funakoshi, T; Ishibe, Y; Okazaki, N; Miura, K; Liu, R; Nagai, S; Minami, Y

    2004-04-01

    Re-expansion pulmonary oedema is a rare complication caused by rapid re-expansion of a chronically collapsed lung. Several cases of pulmonary oedema associated with one-lung ventilation (OLV) have been reported recently. Elevated levels of pro-inflammatory cytokines in pulmonary oedema fluid are suggested to play important roles in its development. Activation of cytokines after re-expansion of collapsed lung during OLV has not been thoroughly investigated. Here we investigated the effects of re-expansion of the collapsed lung on pulmonary oedema formation and pro-inflammatory cytokine expression. Lungs isolated from female white Japanese rabbits were perfused and divided into a basal (BAS) group (n=7, baseline measurement alone), a control (CONT) group (n=9, ventilated without lung collapse for 120 min) and an atelectasis (ATEL) group (n=9, lung collapsed for 55 min followed by re-expansion and ventilation for 65 min). Pulmonary vascular resistance (PVR) and the coefficient of filtration (Kfc) were measured at baseline and 60 and 120 min. At the end of perfusion, bronchoalveolar lavage fluid/plasma protein ratio (B/P), wet/dry lung weight ratio (W/D) and mRNA expressions of tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta and myeloperoxidase (MPO) were determined. TNF-alpha and IL-1beta mRNA were significantly up-regulated in lungs of the ATEL group compared with BAS and CONT, though no significant differences were noted in PVR, Kfc, B/P and W/D within and between groups. MPO increased at 120 min in CONT and ATEL groups. Pro-inflammatory cytokines were up-regulated upon re-expansion and ventilation after short-period lung collapse, though no changes were noted in pulmonary capillary permeability.

  7. Recent Zika Virus Isolates Induce Premature Differentiation of Neural Progenitors in Human Brain Organoids.

    PubMed

    Gabriel, Elke; Ramani, Anand; Karow, Ulrike; Gottardo, Marco; Natarajan, Karthick; Gooi, Li Ming; Goranci-Buzhala, Gladiola; Krut, Oleg; Peters, Franziska; Nikolic, Milos; Kuivanen, Suvi; Korhonen, Essi; Smura, Teemu; Vapalahti, Olli; Papantonis, Argyris; Schmidt-Chanasit, Jonas; Riparbelli, Maria; Callaini, Giuliano; Krönke, Martin; Utermöhlen, Olaf; Gopalakrishnan, Jay

    2017-03-02

    The recent Zika virus (ZIKV) epidemic is associated with microcephaly in newborns. Although the connection between ZIKV and neurodevelopmental defects is widely recognized, the underlying mechanisms are poorly understood. Here we show that two recently isolated strains of ZIKV, an American strain from an infected fetal brain (FB-GWUH-2016) and a closely-related Asian strain (H/PF/2013), productively infect human iPSC-derived brain organoids. Both of these strains readily target to and replicate in proliferating ventricular zone (VZ) apical progenitors. The main phenotypic effect was premature differentiation of neural progenitors associated with centrosome perturbation, even during early stages of infection, leading to progenitor depletion, disruption of the VZ, impaired neurogenesis, and cortical thinning. The infection pattern and cellular outcome differ from those seen with the extensively passaged ZIKV strain MR766. The structural changes we see after infection with these more recently isolated viral strains closely resemble those seen in ZIKV-associated microcephaly.

  8. Diadenosine polyphosphates as antagonists of the endogenous P2Y1 receptor in rat brain capillary endothelial cells of the B7 and B10 clones

    PubMed Central

    Vigne, Paul; Breittmayer, Jean Philippe; Frelin, Christian

    2000-01-01

    Diadenosine polyphosphates (ApnAs, n=2–7) are considered as stress mediators in the cardiovascular system. They act both via identified P2 purinoceptors and via yet to be characterized receptors. This study analyses the actions of ApnAs in clones of rat brain capillary endothelial cells that express P2Y1 receptors (B10 cells) or both P2Y1 and P2Y2 receptors (B7 cells).B10 cells responded to Ap3A with rises in intracellular Ca2+ concentration ([Ca2+]i). This response was prevented by adenosine-3′-phosphate-5′-phosphate, an antagonist of P2Y1 receptors. It was largely suppressed by a treatment with apyrase VII or with creatine phosphokinase/creatine phosphate to degrade contaminating ADP.ApnAs inhibited ADP induced increases in [Ca2+]i mediated by P2Y1 receptors by shifting ADP concentration-response curves to larger concentrations. Apparent Ki values were estimated to be 6 μM for Ap4A, 10 μM for Ap5A and 47 μM for Ap6A. Ap2A and Ap3A were much less active.ApnAs were neither agonists nor antagonists of the endogenous P2Y2 receptor in B7 cells.ApnAs are neither agonists nor antagonists of the Gi-coupled, ADP receptor in B10 cells.The results suggest that most actions of ApnAs in B7 and B10 cells can be accounted for by endogenous P2Y1 receptors. Ap4A, Ap5A and Ap6A are specific antagonists of endogenous Ca2+-coupled P2Y1 receptors. PMID:10742308

  9. Viral determinants that control the neuropathogenicity of PVC-211 murine leukemia virus in vivo determine brain capillary endothelial cell tropism of the virus in vitro.

    PubMed Central

    Masuda, M; Hoffman, P M; Ruscetti, S K

    1993-01-01

    PVC-211 murine leukemia virus (MuLV) is a neuropathogenic, weakly leukemogenic variant of the nonneuropathogenic, highly leukemogenic Friend MuLV (F-MuLV). Chimeric viruses constructed from PVC-211 MuLV clone 3d and F-MuLV clone 57 indicate that the env gene of PVC-211 MuLV contains the determinant(s) responsible for pathological changes in the central nervous system. However, sequences within the 5' one-third (AatII-EcoRI region) of the PVC-211 MuLV genome, which include the 5' leader sequence, the gag gene, and the 5' quarter of the pol gene, are also needed in conjunction with the env gene determinant(s) to cause clinically evident neurological disease in the majority of virus-infected animals after a short latency. In the presence of the AatII-EcoRI region of the PVC-211 MuLV genome, the PVC-211 MuLV env gene sequences encoding the amino-terminal half of the SU protein, which contains the receptor-binding region of the protein, were sufficient to cause rapidly progressive neurological disease. When PVC-211 MuLV, F-MuLV, and various chimeric viruses were tested for their ability to replicate in cultured brain capillary endothelial cells (BCEC), the primary site of PVC-211 MuLV replication within the central nervous system, there was a direct correlation between the replication efficiency of a virus in BCEC in vitro and its ability to cause neurological disease in vivo. This observation indicates that the sequences in PVC-211 MuLV that render it neuropathogenic affect its replication in BCEC and suggests that rapid and efficient replication of the virus in BCEC is crucial for the pathological changes in the central nervous system that result in development of neurological disease. Images PMID:8392599

  10. INTESTINAL CAPILLARIES

    PubMed Central

    Clementi, Francesco; Palade, George E.

    1969-01-01

    Perfusion of the fenestrated capillaries of the intestinal mucosa of the rat with 0.05–0.1 M EDTA removes the diaphragms of the endothelial cells and detaches these cells from one another and from the basement membrane. The latter, even when completely denuded, retains effectively particles of 340 A (average) diameter. Perfusion with histamine (1 µg/ml) results in partial removal of fenestral diaphragms, occasional detachment of the endothelium from the basement membrane, and focal separation of endothelial intercellular junctions. PMID:4979362

  11. Melatonin modulates permeability transition pore and 5-hydroxydecanoate induced KATP channel inhibition in isolated brain mitochondria.

    PubMed

    Waseem, Mohammad; Tabassum, Heena; Parvez, Suhel

    2016-11-01

    There is increasing recognition of the magnitude of mitochondria in neurodegenerative disorders. Mitochondria play a key role in apoptotic and necrotic cell death. Melatonin (Mel), an indoleamine produced in several organs including the pineal gland has been known for its neuroprotective actions. In our study, we have investigated whether the mitochondrial ATP sensitive potassium (mtKATP) channel blocker 5-hydroxydecanoate (5-HD) and calcium (Ca(2+)) affects permeability transition pore (PTP) alterations in isolated brain mitochondria treated with melatonin (Mel) and cyclosporin A (CsA). Mitochondrial swelling, mitochondrial membrane potential (Δψm), ROS measurement and mitochondrial respiration were evaluated in isolated brain mitochondria. In our results, mitochondrial swelling stimulated by exposing Ca(2+) ions and 5-HD associated by mPTP opening as depicted by modulation of CsA and Mel. In addition, Ca(2+) and 5-HD decreased Δψm, depleted intracellular ROS, and inhibition of mitochondrial respiration (state 3 and state 4) in isolated brain mitochondria. Addition of Mel and CsA has shown significant restoration in mitochondrial swelling, Δψm, intracellular ROS measurement and mitochondrial respiration in isolated brain mitochondria. Therefore, we speculate the modulatory effect of Mel and CsA in mitochondria treated with 5-HD and Ca(2+) hinders the mPTP-mediated mitochondrial dysfunction and cellular oxidative stress. We conclude that inhibition of mPT is one likely mechanism of CsA's and its neuroprotective actions. Development of neuroprotective agents including Mel targeting the mPTP therefore bears hope for future treatment of severe neurodegenerative diseases. Copyright © 2016 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

  12. Diagnosis of a Cerebral Arteriovenous Malformation Using Isolated Brain Computed Tomography Angiography: Case Report.

    PubMed

    Qian, Hui; Shao, Yu; Li, Zhengdong; Huang, Ping; Zou, Donghua; Liu, Ningguo; Chen, Yijiu; Wan, Lei

    2016-09-01

    This report presents a case of a 40-year-old woman who was found dead in her house. The examination of the body revealed no external injuries. The whole body was scanned by multi-detector-row computed tomography (CT) before autopsy, revealing massive hemorrhage in the right frontal extending into the ventricular system. At autopsy, the brain parenchyma was removed. Then CT angiography was carried on the isolated brain. Computed tomography angiography suggested a mass of irregular, tortuous vessels in areas of hemorrhage in the right frontal lobe of the brain. Finally, histological examination confirmed the result of CT angiography due to an arteriovenous malformation. Hence, postmortem CT angiography played an important role in diagnosis of the cerebral arteriovenous malformation that was responsible for a massive hemorrhage in the skull.

  13. Isolation and purification of monosialotetrahexosylgangliosides from pig brain by extraction and liquid chromatography.

    PubMed

    Bian, Liujiao; Yang, Jianting; Sun, Yu

    2015-10-01

    Monosialotetrahexosylganglioside (GM1), one of glycosphingolipids containing sialic acid, plays particularly important role in fighting against paralysis, dementia and other diseases caused by brain and nerve damage. In this work, a simple and highly efficient method with high yield was developed for isolation and purification of GM1 from pig brain. The method consisted of an extraction by chloroform-methanol-water and a two-step chromatographic separation by DEAE-Sepharose Fast Flow anion-exchange medium and Sephacryl S-100 HR size-exclusion medium. The purified GM1 was proved to be homogeneous and had a purity of >98.0% by high-performance anion-exchange and size-exclusion chromatography. The molecular weight was 30.0 kDa by high-performance size-exclusion chromatography and 1546.9 Da by electrospray ionization mass spectrometry. The chromogenic reaction by resorcinol-hydrochloric acid solution indicated that the purified GM1 showed a specific chromogenic reaction of sialic acid. Through this isolation and purification program, ~1.0 mg of pure GM1 could be captured from 500 g wet pig brain tissue and the yield of GM1 was around 0.022%, which was higher than the yields by other methods. The method may provide an alternative for isolation and purification of GM1 in other biological tissues.

  14. Superior isolation of antigen-specific brain infiltrating T cells using manual homogenization technique.

    PubMed

    Cumba Garcia, Luz M; Huseby Kelcher, April M; Malo, Courtney S; Johnson, Aaron J

    2016-12-01

    Effective recovery of activated brain infiltrating lymphocytes is critical for investigations involving murine neurological disease models. To optimize lymphocyte recovery, we compared two isolation methods using brains harvested from seven-day Theiler's murine encephalomyelitis virus (TMEV) and TMEV-OVA infected mice. Brains were processed using either a manual dounce based approach or enzymatic digestion using type IV collagenase. The resulting cell suspensions from these two techniques were transferred to a percoll gradient, centrifuged, and lymphocytes were recovered. Flow cytometric analysis of CD45(hi) cells showed greater percentage of CD44(hi)CD62(lo) activated lymphocytes and CD19+ B cells using the dounce method. In addition, we achieved a 3-fold greater recovery of activated virus-specific CD8 T cells specific for the immunodominant D(b):VP2121-130 and engineered K(b):OVA257-264 epitopes through manual dounce homogenization approach as compared to collagenase digest. A greater percentage of viable cells was also achieved through dounce homogenization. Therefore, we conclude that manual homogenization is a superior approach to isolate activated T cells from the mouse brain. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. A simple method for isolating and culturing the rat brain microvascular endothelial cells.

    PubMed

    Liu, Yang; Xue, Qiang; Tang, Qing; Hou, Min; Qi, Hongyi; Chen, Gang; Chen, Weihai; Zhang, Jifen; Chen, Yi; Xu, Xiaoyu

    2013-11-01

    Brain microvascular endothelial cells (BMECs), a main component of the blood-brain barrier, play a critical role in the pathogenesis of many brain diseases. The primary culture of BMECs has been used in various models for studying cerebrovascular diseases in vitro. However, there are still several problems existing in the isolation and cultivation of primary rat BMECs, such as low yield, contamination with other cell types, and requirement of a large number of animals and expensive growth factor. In this study, we describe a simple, economical (without any growth factor) and repeatable method to obtain endothelial cells with high purity (>99%) and yield (about 2.2×10(7) per rat) from cerebral cortexes of neonatal rat, mainly from gray matter. In vitro examinations determined that the isolated cells expressed typical phenotypic markers of differentiated brain endothelium such as multiple drug resistant protein, von Willebrand factor, platelet endothelial cell adhesion molecule 1 (PECAM-1/CD31), and intercellular adhesion molecule (ICAM). These cells also possessed morphological and ultra-structural characteristics that were observed by phase contrast microscope and electric microscope. Then GFAP and α-SMA were used, respectively, to identify astrocyte and pericyte which were potential to contaminate primary culturing of BMECs. And specific reaction of endothelial cells to external stimulation was tested by culture with TNF-α for 24h. All these results of our experiments supply that our protocol provides an effective and reliable method to obtain high purity and yield of rat BMECs and offers a useful tool for studying cellular physiology, cerebrovascular diseases, brain tumors, blood-brain barrier and neurovascular units, etc.

  16. Normal-appearing brain tissue analysis in radiologically isolated syndrome using 3 T MRI

    PubMed Central

    Labiano-Fontcuberta, Andrés; Mato-Abad, Virginia; Álvarez-Linera, Juan; Hernández-Tamames, Juan Antonio; Martínez-Ginés, María Luisa; Aladro, Yolanda; Ayuso, Lucía; Domingo-Santos, Ángela; Benito-León, Julián

    2016-01-01

    Abstract To date, it remains largely unknown whether there is in radiologically isolated syndrome (RIS) brain damage beyond visible T2 white matter lesions. We used single- voxel proton magnetic resonance spectroscopy and diffusion tensor imaging (3 T MRI) to analyze normal-appearing brain tissue regions in 18 RIS patients and 18 matched healthy controls. T2-hyperintense lesion volumes and structural brain volumes were also measured. The absolute metabolite concentrations and ratios of total N-acetylaspartate+N-acetylaspartyl glutamate (NAA), choline-containing compounds, myoinositol, and glutamine-glutamate complex to creatine were calculated. Spectral analysis was performed by LCModel. Voxelwise morphometry analysis was performed to localize regions of brain tissue showing significant changes of fractional anisotropy or mean diffusivity. Compared with healthy controls, RIS patients did not show any significant differences in either the absolute concentration of NAA or NAA/Cr ratio in mid-parietal gray matter. A trend toward lower NAA concentrations (–3.35%) was observed among RIS patients with high risk for conversion to multiple sclerosis. No differences in the other metabolites or their ratios were observed. RIS patients showed lower fractional anisotropy only in clusters overlapping lesional areas, namely in the cingulate gyrus bilaterally and the frontal lobe subgyral bilaterally (P < 0.001). Normalized brain and cortical volumes were significantly lower in RIS patients than in controls (P = 0.01 and P = 0.03, respectively). Our results suggest that in RIS, global brain and cortical atrophy are not primarily driven by significant occult microstructural normal appearing brain damage. Longitudinal MRI studies are needed to better understand the pathological processes underlying this novel entity. PMID:27399108

  17. Mechanistic approach for the toxic effects of perfluorooctanoic acid on isolated rat liver and brain mitochondria.

    PubMed

    Mashayekhi, V; Tehrani, K Haj Mohammad Ebrahim; Hashemzaei, M; Tabrizian, K; Shahraki, J; Hosseini, M-J

    2015-10-01

    Perfluorooctanoic acid (PFOA) is one of the most widely used perfluoroalkanes as surfactants, lubricants and processing aids in the production of polymers, which has also been detected in the environment, wildlife and human body. Animal studies indicated that PFOA caused a wide array of toxic effects including liver and brain dysfunction, carcinogenicity and reproductive and developmental toxicity. Based on the established role of mitochondria-mediated pathways in the observed toxic effects of many drugs and chemicals, in this study, the potential toxic effects of PFOA on mitochondria isolated from rat liver and brain have been investigated. Mitochondria were isolated by differential centrifugation method and incubated with different concentrations of PFOA (0.5-1.5 mM). The effects of PFOA were assessed on a series of mitochondrial parameters including reactive oxygen species (ROS) formation, activities of mitochondrial complexes I/II/III, reduced glutathione (GSH) content, adenosine triphosphate (ATP) level, membrane potential, lipid peroxidation (LPO), mitochondrial swelling and cytochrome c release. The data on liver mitochondria indicated that PFOA-induced ROS elevation in both mitochondrial complexes I and III, mitochondrial membrane potential collapse, swelling, cytochrome c release and decreased ATP level which induces apoptosis or necrosis. On brain mitochondria, PFOA showed fairly similar effects on the above-mentioned parameters. However, different results were obtained when the effect of PFOA was assessed on LPO and complex II activity. Due to the fact that PFOA had toxic effects on the mitochondria isolated, it could be suggested that mitochondrial toxicity could be a plausible mechanism for the toxic effects of this fluorochemical on liver and brain function. © The Author(s) 2015.

  18. Isolating dividing neural and brain tumour cells for gene expression profiling.

    PubMed

    Endaya, Berwini; Cavanagh, Brenton; Alowaidi, Faisal; Walker, Tom; de Pennington, Nicholas; Ng, Jin-Ming A; Lam, Paula Y P; Mackay-Sim, Alan; Neuzil, Jiri; Meedeniya, Adrian C B

    2016-01-15

    The characterisation of dividing brain cells is fundamental for studies ranging from developmental and stem cell biology, to brain cancers. Whilst there is extensive anatomical data on these dividing cells, limited gene transcription data is available due to technical constraints. We focally isolated dividing cells whilst conserving RNA, from culture, primary neural tissue and xenografted glioma tumours, using a thymidine analogue that enables gene transcription analysis. 5-ethynyl-2-deoxyuridine labels the replicating DNA of dividing cells. Once labelled, cultured cells and tissues were dissociated, fluorescently tagged with a revised click chemistry technique and the dividing cells isolated using fluorescence-assisted cell sorting. RNA was extracted and analysed using real time PCR. Proliferation and maturation related gene expression in neurogenic tissues was demonstrated in acutely and 3 day old labelled cells, respectively. An elevated expression of marker and pathway genes was demonstrated in the dividing cells of xenografted brain tumours, with the non-dividing cells showing relatively low levels of expression. BrdU "immune-labelling", the most frequently used protocol for detecting cell proliferation, causes complete denaturation of RNA, precluding gene transcription analysis. This EdU labelling technique, maintained cell integrity during dissociation, minimized copper exposure during labelling and used a cell isolation protocol that avoided cell lysis, thus conserving RNA. The technique conserves RNA, enabling the definition of cell proliferation-related changes in gene transcription of neural and pathological brain cells in cells harvested immediately after division, or following a period of maturation. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Capillary muscle

    PubMed Central

    Cohen, Caroline; Mouterde, Timothée; Quéré, David; Clanet, Christophe

    2015-01-01

    The contraction of a muscle generates a force that decreases when increasing the contraction velocity. This “hyperbolic” force–velocity relationship has been known since the seminal work of A. V. Hill in 1938 [Hill AV (1938) Proc R Soc Lond B Biol Sci 126(843):136–195]. Hill’s heuristic equation is still used, and the sliding-filament theory for the sarcomere [Huxley H, Hanson J (1954) Nature 173(4412):973–976; Huxley AF, Niedergerke R (1954) Nature 173(4412):971–973] suggested how its different parameters can be related to the molecular origin of the force generator [Huxley AF (1957) Prog Biophys Biophys Chem 7:255–318; Deshcherevskiĭ VI (1968) Biofizika 13(5):928–935]. Here, we develop a capillary analog of the sarcomere obeying Hill’s equation and discuss its analogy with muscles. PMID:25944938

  20. Distribution of PCR ribotypes among recent Clostridium difficile isolates collected in two districts of Hungary using capillary gel electrophoresis and review of changes in the circulating ribotypes over time.

    PubMed

    Tóth, Judith; Urbán, Edith; Osztie, Hilda; Benczik, Márta; Indra, Alexander; Nagy, Elisabeth; Allerberger, Franz

    2016-10-01

    Following the first description of a Clostridium difficile case caused by ribotype 027 in Hungary in 2007, the rapid spread of C. difficile infection in different hospitals within the country was observed. The aim of this pilot study was to investigate the distribution of different PCR ribotypes among inpatient and outpatient isolates obtained in two geographically different parts of Hungary. One hundred and ninety-two toxigenic C. difficile isolates collected between 1 October and 1 December 2014 were PCR ribotyped using capillary gel electrophoresis and the database of WEBRIBO (http://webribo.ages.at), which allows the automatic analysis and comparison of capillary-sequencer-based PCR ribotyping data. Altogether, 31 different known ribotypes were found, and 16 isolates showed a novel banding pattern, not included in the current library. Besides the dominance of 027 (33.3 %) among all isolates, there were differences in its presence among isolates obtained from the two regions (45.8 % in the central region and 20.8 % in the south-east region, respectively), whereas the second most prevalent ribotype 036 (19.8 %) was more frequently found among isolates obtained in the south-east region compared with the central region of Hungary (29.1 versus 10.4 %). Similar differences in the spread of different ribotypes, in particular 027, which were found during earlier studies in Hungary may be due to the existing order for admissions of patients to hospitals. We also summarized the changing pattern of PCR ribotypes of Hungarian C. difficile isolates over time, based on earlier published data.

  1. Isolation rearing significantly perturbs brain metabolism in the thalamus and hippocampus.

    PubMed

    Bonab, A A; Fricchione, J G; Gorantla, S; Vitalo, A G; Auster, M E; Levine, S J; Scichilone, J M; Hegde, M; Foote, W; Fricchione, G L; Denninger, J W; Yarmush, D M; Fischman, A J; Yarmush, M L; Levine, J B

    2012-10-25

    Psychosocial neglect during childhood severely impairs both behavioral and physical health. The isolation rearing model in rodents has been employed by our group and others to study this clinical problem at a basic level. We previously showed that immediate early gene (IEG) expression in the hippocampus and medial prefrontal cortex (mPFC) is decreased in isolation-reared (IR) compared to group-reared (GR) rats. In the current study, we sought to evaluate: (1) whether these changes in IEG expression would be detected by the measurement of brain glucose metabolism using positron emission tomography (PET) with fluorodeoxyglucose (FDG) and (2) whether PET FDG could illuminate other brain regions with different glucose metabolism in IR compared to GR rats. We found that there were significant differences in FDG uptake in the hippocampus that were consistent with our findings for IEG expression (decreased mean FDG uptake in IR rats). In contrast, in the mPFC, the FDG uptake between IR and GR rats did not differ. Finally, we found decreased mean FDG uptake in the thalamus of the IR rats, a region we had not previously examined. The results suggest that PET FDG has the potential to be utilized as a biomarker of molecular changes in the hippocampus. Further, the differences found in thalamic brain FDG uptake suggest that further investigation of this region at the molecular and cellular levels may provide an important insight into the neurobiological basis of the adverse clinical outcomes found in children exposed to psychosocial deprivation.

  2. Capillary interactions in Pickering emulsions

    NASA Astrophysics Data System (ADS)

    Guzowski, J.; Tasinkevych, M.; Dietrich, S.

    2011-09-01

    The effective capillary interaction potentials for small colloidal particles trapped at the surface of liquid droplets are calculated analytically. Pair potentials between capillary monopoles and dipoles, corresponding to particles floating on a droplet with a fixed center of mass and subjected to external forces and torques, respectively, exhibit a repulsion at large angular separations and an attraction at smaller separations, with the latter resembling the typical behavior for flat interfaces. This change of character is not observed for quadrupoles, corresponding to free particles on a mechanically isolated droplet. The analytical results are compared with the numerical minimization of the surface free energy of the droplet in the presence of spherical or ellipsoidal particles.

  3. A Method for Isolation of Extracellular Vesicles and Characterization of Exosomes from Brain Extracellular Space.

    PubMed

    Pérez-González, Rocío; Gauthier, Sebastien A; Kumar, Asok; Saito, Mitsuo; Saito, Mariko; Levy, Efrat

    2017-01-01

    Extracellular vesicles (EV), including exosomes, secreted vesicles of endocytic origin, and microvesicles derived from the plasma membrane, have been widely isolated and characterized from conditioned culture media and bodily fluids. The difficulty in isolating EV from tissues, however, has hindered their study in vivo. Here, we describe a novel method designed to isolate EV and characterize exosomes from the extracellular space of brain tissues. The purification of EV is achieved by gentle dissociation of the tissue to free the brain extracellular space, followed by sequential low-speed centrifugations, filtration, and ultracentrifugations. To further purify EV from other extracellular components, they are separated on a sucrose step gradient. Characterization of the sucrose step gradient fractions by electron microscopy demonstrates that this method yields pure EV preparations free of large vesicles, subcellular organelles, or debris. The level of EV secretion and content are determined by assays for acetylcholinesterase activity and total protein estimation, and exosomal identification and protein content are analyzed by Western blot and immuno-electron microscopy. Additionally, we present here a method to delipidate EV in order to improve the resolution of downstream electrophoretic analysis of EV proteins.

  4. Mitochondrial oxidative stress and dysfunction induced by isoniazid: study on isolated rat liver and brain mitochondria.

    PubMed

    Ahadpour, Morteza; Eskandari, Mohammad Reza; Mashayekhi, Vida; Haj Mohammad Ebrahim Tehrani, Kamaleddin; Jafarian, Iman; Naserzadeh, Parvaneh; Hosseini, Mir-Jamal

    2016-01-01

    Isoniazid (INH or isonicotinic hydrazide) is used for the treatment and prophylaxis of tuberculosis. Liver and brain are two important target organs in INH toxicity. However, the exact mechanisms behind the INH hepatotoxicity or neurotoxicity have not yet been completely understood. Considering the mitochondria as one of the possible molecular targets for INH toxicity, the aim of this study was to evaluate the mechanisms of INH mitochondrial toxicity on isolated mitochondria. Mitochondria were isolated by differential ultracentrifugation from male Sprague-Dawley rats and incubated with different concentrations of INH (25-2000 μM) for the investigation of mitochondrial parameters. The results indicated that INH could interact with mitochondrial respiratory chain and inhibit its activity. Our results showed an elevation in mitochondrial reactive oxygen species (ROS) formation, lipid peroxidation and mitochondrial membrane potential collapse after exposure of isolated liver mitochondria in INH. However, different results were obtained in brain mitochondria. Noteworthy, significant glutathione oxidation, adenosine triphosphate (ATP) depletion and lipid peroxidation were observed in higher concentration of INH, as compared to liver mitochondria. In conclusion, our results suggest that INH may initiate its toxicity in liver mitochondria through interaction with electron transfer chain, lipid peroxidation, mitochondrial membrane potential decline and cytochrome c expulsion which ultimately lead to cell death signaling.

  5. Membrane lipids and morphology of brain cortex synaptosomes isolated from hibernating Yakutian ground squirrel.

    PubMed

    Kolomiytseva, Iskra K; Perepelkina, Natalia I; Zharikova, Alevtina D; Popov, Victor I

    2008-12-01

    Synaptosomes were isolated from Yakutian ground squirrel brain cortex of summer and winter hibernating animals in active and torpor states. Synaptosomal membrane cholesterol and phospholipids were determined. The seasonal changes of synaptosomal lipid composition were found. Synaptosomes isolated from hibernating Yakutian ground squirrel brain cortex maintained the cholesterol sphingomyelin, phosphatidylethanolamine, lysophosphatidylcholine, cardiolipin, phosphatidylinositol and phosphatidylserine contents 2.5, 1.8, 2.6, 1.8, 1.6, and 1.3 times less, respectively, and the content of phosphatidylcholine twice as much as the one in summer season. The synaptosomal membrane lipid composition of summer animals was shown to be markedly different from that as hibernating ground squirrels and non-hibernating rodents. It is believed that phenotypic changes of synaptosomal membrane lipid composition in summer Yakutian ground squirrel are the important preparation step for hibernation. The phosphatidylethanolamine content was increased in torpor state compared with winter-active state and the molar ratio of cholesterol/phospholipids in synaptosomal membrane of winter torpor ground squirrels was lower than that in active winter and summer animals. These events were supposed to lead to increase of the synaptosomal membrane fluidity during torpor. Synaptosomes isolated from torpor animals have larger sizes and contain a greater number of synaptic vesicles on the synaptosomal profile area. The synaptosomal membrane lipid composition and synaptosome morphology were involved in phenotypic adaptation of Yakutian ground squirrel to hibernation.

  6. One-step multiple component isolation from the oil of Crinitaria tatarica (Less.) Sojak by preparative capillary gas chromatography with characterization by spectroscopic and spectrometric techniques and evaluation of biological activity.

    PubMed

    Özek, Gulmira; Ishmuratova, Margarita; Tabanca, Nurhayat; Radwan, Mohammed M; Göger, Fatih; Özek, Temel; Wedge, David E; Becnel, James J; Cutler, Stephen J; Can Başer, Kemal H

    2012-03-01

    Gas chromatographic analysis revealed that the oil of Crinitaria tatarica was rich in sabinene (32.1%), β-pinene (8.8%), and two unknown (M+200) compounds (I) and (II) (21.4% and 3.4%). One-step multiple fractionation of the oil and separation of two unknown constituents were performed using preparative capillary gas chromatography connected to preparative fraction collector system. This combination allowed separation and recover of sufficient quantities of two unknown compounds with high purity from complex oil matrix. Separation conditions (column temperature, cooling temperature, flow rate, injection volume, cut time) were optimized to achieve the best isolation and successful collection. The target compounds were separated from the oil using a HP Innowax (Walt & Jennings Scientific, Wilmington, DE, USA) preparative capillary column in rapid one-step manner with 95.0% purity. Trapping of the isolated compounds in collector system was facilitated by cooling with liquid nitrogen. Structure determination was accomplished by spectral analysis including ultraviolet, nuclear magnetic rezonance, and high-resolution electrospray ionization mass spectrometry. Z- (I) and E-artemidin (II) were isolated for the first time from this species. Crinitaria tatarica oil and Z- (I) and E-artemidin (II) were evaluated for biological activity.

  7. Isolation of Toxoplasma gondii from the brain of a dog in Australia and its biological and molecular characterisation

    USDA-ARS?s Scientific Manuscript database

    Toxoplasma gondii was isolated from the brain of a young dog for the first time in Australia. The identity of the parasite was confirmed by PCR, Western blotting, electron microscopy and cat bioassay. Genotyping of the isolate (TgDgAu1) was determined by PCR-RFLP markers that showed it to be a Type...

  8. Isolation and separation of alpha and beta-tubulin from chick-embryo brain, muscle and skin.

    PubMed Central

    el-Hamalawi, A R

    1978-01-01

    Two kinds of tubulin, alpha and beta, have been described in microtubules from many different systems. In the present study a new method is described for isolating and separating these two kinds of tubulin from chick-embryo brain, muscle and skin. The isolated tubulins were characterized by polyacrylamide-gel disc electrophoresis in the presence of urea and sodium dodecyl sulphate. PMID:646795

  9. Distinct characteristics of Ca(2+)-induced depolarization of isolated brain and liver mitochondria.

    PubMed

    Vergun, Olga; Reynolds, Ian J

    2005-09-05

    Ca(2+)-induced mitochondrial depolarization was studied in single isolated rat brain and liver mitochondria. Digital imaging techniques and rhodamine 123 were used for mitochondrial membrane potential measurements. Low Ca(2+) concentrations (about 30--100 nM) initiated oscillations of the membrane potential followed by complete depolarization in brain mitochondria. In contrast, liver mitochondria were less sensitive to Ca(2+); 20 microm Ca(2+) was required to depolarize liver mitochondria. Ca(2+) did not initiate oscillatory depolarizations in liver mitochondria, where each individual mitochondrion depolarized abruptly and irreversibly. Adenine nucleotides dramatically reduced the oscillatory depolarization in brain mitochondria and delayed the onset of the depolarization in liver mitochondria. In both type of mitochondria, the stabilizing effect of adenine nucleotides completely abolished by an inhibition of adenine nucleotide translocator function with carboxyatractyloside, but was not sensitive to bongkrekic acid. Inhibitors of mitochondrial permeability transition cyclosporine A and bongkrekic acid also delayed Ca(2+)-depolarization. We hypothesize that the oscillatory depolarization in brain mitochondria is associated with the transient conformational change of the adenine nucleotide translocator from a specific transporter to a non-specific pore, whereas the non-oscillatory depolarization in liver mitochondria is caused by the irreversible opening of the pore.

  10. Isolating human brain functional connectivity associated with a specific cognitive process

    NASA Astrophysics Data System (ADS)

    Silver, Michael A.; Landau, Ayelet N.; Lauritzen, Thomas Z.; Prinzmetal, William; Robertson, Lynn C.

    2010-02-01

    The use of functional magnetic resonance imaging (fMRI) to measure functional connectivity among brain areas has the potential to identify neural networks associated with particular cognitive processes. However, fMRI signals are not a direct measure of neural activity but rather represent blood oxygenation level-dependent (BOLD) signals. Correlated BOLD signals between two brain regions are therefore a combination of neural, neurovascular, and vascular coupling. Here, we describe a procedure for isolating brain functional connectivity associated with a specific cognitive process. Coherency magnitude (measuring the strength of coupling between two time series) and phase (measuring the temporal latency differences between two time series) are computed during performance of a particular cognitive task and also for a control condition. Subtraction of the coherency magnitude and phase differences for the two conditions removes sources of correlated BOLD signals that do not modulate as a function of cognitive task, resulting in a more direct measure of functional connectivity associated with changes in neuronal activity. We present two applications of this task subtraction procedure, one to measure changes in strength of coupling associated with sustained visual spatial attention, and one to measure changes in temporal latencies between brain areas associated with voluntary visual spatial attention.

  11. Comparison of proteins expressed by Pseudomonas aeruginosa strains representing initial and chronic isolates from a cystic fibrosis patient: an analysis by 2-D gel electrophoresis and capillary column liquid chromatography-tandem mass spectrometry.

    PubMed

    Hanna, S L; Sherman, N E; Kinter, M T; Goldberg, J B

    2000-10-01

    Isolates of Pseudomonas aeruginosa from chronic lung infections in cystic fibrosis (CF) patients have phenotypes distinct from those initially infecting CF patients, as well as from other clinical or environmental isolates. To gain a better understanding of the differences in these isolates, protein expression was followed using two-dimensional (2-D) gel electrophoresis and protein identification by peptide sequencing using micro-capillary column liquid chromatography-tandem mass spectrometry (microLC/MS/MS). The isolates selected for this analysis were from the sputum of a CF patient: strain 383 had a nonmucoid phenotype typical of isolates from the environment, and strain 2192, obtained from the same patient, had a mucoid phenotype typical of isolates from chronic CF lung infections. Strains 383 and 2192 were confirmed to be genetically identical by restriction endonuclease analysis, random amplified polymorphic DNA-PCR, and pulsed-field gel electrophoresis. Conditions of protein extraction were optimized for consistent high-resolution separation of several hundred proteins from these clinical isolates as detected by Coomassie staining of 2-D gels. Fourteen proteins were selected for analysis; this group included those whose expression was common between both strains as well as unique for each strain. The proteins were identified by microLC/MS/MS of the peptides produced by an in-gel tryptic digestion and compared to translated data from the Pseudomonas Genome Project; optimization of this technique has allowed for the comparison of proteins expressed by strains 383 and 2192.

  12. Optical coherence tomography angiography-based capillary velocimetry

    NASA Astrophysics Data System (ADS)

    Wang, Ruikang K.; Zhang, Qinqin; Li, Yuandong; Song, Shaozhen

    2017-06-01

    Challenge persists in the field of optical coherence tomography (OCT) when it is required to quantify capillary blood flow within tissue beds in vivo. We propose a useful approach to statistically estimate the mean capillary flow velocity using a model-based statistical method of eigendecomposition (ED) analysis of the complex OCT signals obtained with the OCT angiography (OCTA) scanning protocol. ED-based analysis is achieved by the covariance matrix of the ensemble complex OCT signals, upon which the eigenvalues and eigenvectors that represent the subsets of the signal makeup are calculated. From this analysis, the signals due to moving particles can be isolated by employing an adaptive regression filter to remove the eigencomponents that represent static tissue signals. The mean frequency (MF) of moving particles can be estimated by the first lag-one autocorrelation of the corresponding eigenvectors. Three important parameters are introduced, including the blood flow signal power representing the presence of blood flow (i.e., OCTA signals), the MF indicating the mean velocity of blood flow, and the frequency bandwidth describing the temporal flow heterogeneity within a scanned tissue volume. The proposed approach is tested using scattering phantoms, in which microfluidic channels are used to simulate the functional capillary vessels that are perfused with the scattering intralipid solution. The results indicate a linear relationship between the MF and mean flow velocity. In vivo animal experiments are also conducted by imaging mouse brain with distal middle cerebral artery ligation to test the capability of the method to image the changes in capillary flows in response to an ischemic insult, demonstrating the practical usefulness of the proposed method for providing important quantifiable information about capillary tissue beds in the investigations of neurological conditions in vivo.

  13. [Conversion to clinically definite multiple sclerosis after isolated spinal cord syndrome: value of brain and spinal MRI].

    PubMed

    Pericot, I; Tintoré, M; Grivé, E; Briev, L; Rovira, A; Montalbán, X

    2001-02-17

    Isolated spinal cord syndrome might be due to a first episode of multiple sclerosis. The aim of the study was to determine the clinical usefulness and paraclinical characteristics and of spinal and brain MR imaging predicting conversion to clinically definite multiple sclerosis (CDMS) in patients with an isolated spinal cord syndrome. We have evaluate thirty-eight patients with isolated spinal cord syndrome. A clinical protocol, lumbar puncture, evoked potential and brain-spinal cord MRI were performed. Twenty two percent of the patients fulfilling brain MRI Paty criteria (p < 0.01), 54.5% Fazekas (p = 0.007) and 80% of patients fulfilling Barkhof criteria (p = 0.009) presented CDMS. The spinal MR imaging from CDMS patients was always abnormal, showing cervical and marginal location with a diameter < 2 cm. Brain MRI is strongly predictive of the risk of developing CDMS and spinal cord MRI may increase the sensitivity to detect conversion to CDMS.

  14. A new protein of the brain myelin: isolation and chemical characterization.

    PubMed

    Riccio, P; Tsugita, A; Bobba, A; Vilbois, F; Quagliariello, E

    1985-03-15

    An unknown protein has been isolated from bovine brain myelin. This protein, purified in the nonionic detergent n-octylpolydisperse oligooxyethylene, reveals on SDS gel electrophoresis a large number of bands in the higher MW region. However, chemical analysis and gel chromatography indicate the presence of a single, small protein containing large amounts of bound phosphatidylserine. N-terminal and C-terminal sequences, aminoacid composition, and the anomalous electrophoretic behaviour led us to exclude the protein as a fragment of other already known myelin proteins.

  15. Brain metastases as site of first and isolated recurrence of breast cancer: the role of systemic therapy after local treatment.

    PubMed

    Niwińska, Anna

    2016-10-01

    The role of systemic treatment was assessed after local therapy for breast cancer patients who developed central nervous system (CNS) metastases as a first and isolated recurrence. Subjects were 128 breast cancer patients with brain metastases as the first and isolated site of recurrence that were selected from 673 consecutive breast cancer patients with brain metastases treated at the same institution. Median survival from brain metastases in patients with and without systemic treatment after local therapy was respectively 15 and 4 months (p < 0.001). In patients with a Karnofsky Performance Status ≥70 and those <70, survival was respectively 16 and 5.5 months (p < 0.001). The median survival from brain metastasis in patients with solitary brain metastasis, with and without systemic treatment after local therapy, was respectively 22 and 7 months (p = 0.003). Cox multivariate analysis demonstrated that good performance status, solitary brain metastasis and systemic therapy undertaken after local treatment were factors which prolonged survival. However patient survival was adversely affected by those having leptomeningeal metastasis associated with brain parenchymal lesions. Systemic therapy, undertaken after local treatment improved survival in those patients with breast cancer and brain metastases as the site of first and isolated recurrence. Further study is required in order to fully establish the role of systemic treatment for this patient group.

  16. Intrahippocampal Infusion of Crotamine Isolated from Crotalus durissus terrificus Alters Plasma and Brain Biochemical Parameters †

    PubMed Central

    Gonçalves, Rithiele; Vargas, Liane S.; Lara, Marcus V. S.; Güllich, Angélica; Mandredini, Vanusa; Ponce-Soto, Luis; Marangoni, Sergio; Dal Belo, Cháriston A.; Mello-Carpes, Pâmela B.

    2014-01-01

    Crotamine is one of the main constituents of the venom of the South American rattlesnake Crotalus durissus terrificus. Here we sought to investigate the inflammatory and toxicological effects induced by the intrahippocampal administration of crotamine isolated from Crotalus whole venom. Adult rats received an intrahippocampal infusion of crotamine or vehicle and were euthanized 24 h or 21 days after infusion. Plasma and brain tissue were collected for biochemical analysis. Complete blood count, creatinine, urea, glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), creatine-kinase (CK), creatine kinase-muscle B (CK-MB) and oxidative parameters (assessed by DNA damage and micronucleus frequency in leukocytes, lipid peroxidation and protein carbonyls in plasma and brain) were quantified. Unpaired and paired t-tests were used for comparisons between saline and crotamine groups, and within groups (24 h vs. 21 days), respectively. After 24 h crotamine infusion promoted an increase of urea, GOT, GPT, CK, and platelets values (p ≤ 0.01), while red blood cells, hematocrit and leukocytes values decreased (p ≤ 0.01). Additionally, 21 days after infusion crotamine group showed increased creatinine, leukocytes, TBARS (plasma and brain), carbonyl (plasma and brain) and micronucleus compared to the saline-group (p ≤ 0.01). Our findings show that crotamine infusion alter hematological parameters and cardiac markers, as well as oxidative parameters, not only in the brain, but also in the blood, indicating a systemic pro-inflammatory and toxicological activity. A further scientific attempt in terms of preserving the beneficial activity over toxicity is required. PMID:25380458

  17. Actin filament-associated protein 1 (AFAP-1) is a key mediator in inflammatory signaling-induced rapid attenuation of intrinsic P-gp function in human brain capillary endothelial cells.

    PubMed

    Hoshi, Yutaro; Uchida, Yasuo; Tachikawa, Masanori; Ohtsuki, Sumio; Terasaki, Tetsuya

    2017-04-01

    The purpose of this study was to identify regulatory molecule(s) involved in the inflammatory signaling-induced decrease in P-glycoprotein (P-gp) efflux function at the blood-brain barrier (BBB) that may occur in brain diseases. We confirmed that in vivo P-gp efflux activity at the BBB was decreased without any change in P-gp protein expression level in a mouse model of acute inflammation induced by 3 mg/kg lipopolysaccharide. In a human BBB model cell line (human brain capillary endothelial cells; hCMEC/D3), 1-h treatment with 10 ng/mL tumor necrosis factor-α (TNF-α; an inflammatory mediator) rapidly reduced P-gp efflux activity, but had no effect on P-gp protein expression level. To clarify the non-transcriptional mechanism that causes the decrease in intrinsic efflux activity of P-gp in acute inflammation, we applied comprehensive quantitative phosphoproteomics to compare hCMEC/D3 cells treated with TNF-α and vehicle (control). Actin filament-associated protein-1 (AFAP-1), MAPK1, and transcription factor AP-1 (AP-1) were significantly phosphorylated in TNF-α-treated cells, and were selected as candidate proteins. In validation experiments, knockdown of AFAP-1 expression blocked the reduction in P-gp efflux activity by TNF-α treatment, whereas inhibition of MAPK function or knockdown of AP-1 expression did not. Quantitative targeted absolute proteomics revealed that the reduction in P-gp activity by TNF-α did not require any change in P-gp protein expression levels in the plasma membrane. Our results demonstrate that AFAP-1 is a key mediator in the inflammatory signaling-induced, translocation-independent rapid attenuation of P-gp efflux activity in human brain capillary endothelial cells. © 2017 International Society for Neurochemistry.

  18. Long-term Survival after Lapatinib Rechallenge in Isolated Brain Metastasis of HER2-positive Breast Cancer

    PubMed Central

    Karagöz, Bülent; Özgün, Alpaslan; Emirzeoğlu, Levent; Tunçel, Tolga; Çelik, Serkan; Bilgi, Oğuz; Kara, Kemal

    2015-01-01

    Lapatinib is an effective drug in HER2-positive breast cancer. We present a case with successful treatment of lapatinib in brain metastasis of HER2+ breast cancer. Forty-eight years old woman was admitted our clinic with early breast cancer. In third years after adjuvant chemotherapy and trastuzumab, isolated and multiple brain metastasis were detected. After whole brain RT, lapatinib (with capecitabine for 10 months and with letrozole for 3 months) has been used. Volumetric reduction of lesions was achieved and symptoms disappeared. When lapatinib discontinued, brain metastasis relapses. Lapatinib plus capecitabine reinduction has been started. Totally, longer survival than 45 months was achieved after first brain metastasis detection. Because both combinations of lapatinib with capecitabine and letrozole were effective and reinduction treatment was successful, presented case has strongly supported activity of lapatinib treatment in brain metastasis of HER2+ breast cancer.

  19. Prevalence of and Risk Factors for Poor Functioning after Isolated Mild Traumatic Brain Injury in Children

    PubMed Central

    Durbin, Dennis R.; Koepsell, Thomas D.; Wang, Jin; Temkin, Nancy R.; Dorsch, Andrea M.; Vavilala, Monica S.; Jaffe, Kenneth M.; Rivara, Frederick P.

    2014-01-01

    Abstract This study aimed to determine the prevalence and predictors of poor 3 and 12 month quality of life outcomes in a cohort of pediatric patients with isolated mild TBI. We conducted a prospective cohort study of children and adolescents <18 years of age treated for an isolated mild TBI, defined as “no radiographically apparent intracranial injury” or “an isolated skull fracture, and no other clinically significant non-brain injuries.” The main outcome measure was the change in quality of life from baseline at 3 and 12 months following injury, as measured by the Pediatric Quality of Life index (PedsQL). Poor functioning was defined as a decrease in total PedsQL score of >15 points between baseline and follow-up scores (at 3 and 12 months). Of the 329 patients who met inclusion criteria, 11.3% (95% CI 8.3–15.3%) at 3 months and 12.9% (95% CI 9.6–17.2%) at 12 months following injury had relatively poor functioning. Significant predictors of poor functioning included less parental education, Hispanic ethnicity (at 3 months following injury, but not at 12 months); low household income (at 3 and 12 months), and Medicaid insurance (at 12 months only). Children and adolescents sustaining a mild TBI who are socioeconomically disadvantaged may require additional intervention to mitigate the effects of mild TBI on their functioning. PMID:24294826

  20. "Relating through sameness": a qualitative study of friendship and social isolation in chronic traumatic brain injury.

    PubMed

    Salas, Christian E; Casassus, Martin; Rowlands, Leanne; Pimm, Steve; Flanagan, Desmond A J

    2016-11-01

    Social isolation has been described as a common problem among traumatic brain injury (TBI) survivors during the chronic phase. Due to physical, cognitive and behavioural changes, survivors become less socially active and experience a marked decrease in the number of friends. The goal of this investigation is to explore TBI survivors' subjective account of the challenges encountered in sustaining friendships, as well as gaining insight into their particular understanding of such difficulties. Using a thematic analysis approach, 11 survivors of TBI were interviewed in relation to their experience of social isolation and friendship during the chronic stage. Four main themes emerged from the interviews: (1) The impact of long-term cognitive and behavioural problems on relationships; (2) Loss of old friends; (3) Difficulties making new friends, and (4) Relating to other survivors in order to fight social isolation (sameness). Clinical implications of these findings, as well as their relevance in the design of long-term rehabilitation programmes, are discussed. Particular emphasis is placed on the need to acknowledge the value of relating to other survivors, as a way of resisting cultural discourses about disability, and as a source of self-cohesion in the process of identity re-construction.

  1. A comparison of mitochondrial DNA isolation methods in frozen post-mortem human brain tissue--applications for studies of mitochondrial genetics in brain disorders.

    PubMed

    Devall, Matthew; Burrage, Joe; Caswell, Richard; Johnson, Matthew; Troakes, Claire; Al-Sarraj, Safa; Jeffries, Aaron R; Mill, Jonathan; Lunnon, Katie

    2015-10-01

    Given that many brain disorders are characterized by mitochondrial dysfunction, there is a growing interest in investigating genetic and epigenetic variation in mitochondrial DNA (mtDNA). One major caveat for such studies is the presence of nuclear-mitochondrial pseudogenes (NUMTs), which are regions of the mitochondrial genome that have been inserted into the nuclear genome over evolution and, if not accounted for, can confound genetic studies of mtDNA. Here we provide the first systematic comparison of methods for isolating mtDNA from frozen post-mortem human brain tissue. Our data show that a commercial method from Miltenyi Biotec, which magnetically isolates mitochondria using antibodies raised against the mitochondrial import receptor subunit TOM22, gives significant mtDNA enrichment and should be considered the method of choice for mtDNA studies in frozen brain tissue.

  2. The electrophysiology of the olfactory-hippocampal circuit in the isolated and perfused adult mammalian brain in vitro.

    PubMed

    de Curtis, M; Paré, D; Llinás, R R

    1991-10-01

    The viability and general electrophysiological properties of the limbic system in the adult mammalian brain isolated and maintained in vitro by arterial perfusion are described. The isolated brain preparation combines the advantages of intact synaptic connectivity and accessibility of different areas of the encephalic mass with those of the in vitro approach, i.e., stability and control of the ionic environment. Extracellular field potential as well as intracellular recordings were performed at different levels in the limbic system of isolated adult guinea pig brains. The results demonstrate that in the piriform, entorhinal, and hippocampal cortices, the intrinsic electrical properties of individual cells as well as the spontaneous and evoked electrical activity in the neuronal ensembles they comprise, were virtually identical to those observed in vivo. The properties of the limbic system loop were determined.

  3. A glass capillary microelectrode based on capillarity and its application to the detection of L-glutamate release from mouse brain slices.

    PubMed

    Nakajima, Kumiko; Yamagiwa, Takashi; Hirano, Ayumi; Sugawara, Masao

    2003-01-01

    A new glass capillary microelectrode for L-glutamate is described using pulled glass capillaries (tip size, approximately 12.5 microm) with a very small volume (approximately 2 microl) of inner solution containing glutamate oxidase (GluOx) and ascorbate oxidase. The operation of the electrode is based on capillary action that samples L-glutamate into the inner solution. The enzyme reaction by GluOx generates hydrogen peroxide that is detected at an Os-gel-HRP polymer modified Pt electrode in a three-electrode configuration. The amperometric response behavior of the electrode was characterized in terms of the capillarity, response time, sensitivity and selectivity for measurements of L-glutamate. The currents at 0 V vs. Ag/AgCl increased linearly with the L-glutamate concentration from 10 to 150 microM for in vitro and in situ calibrations. The response was highly selective to L-glutamate over ascorbate, dopamine, serotonin and other amino acids. The detection of L-glutamate in the extracellular fluids of different regions of mouse hippocampal slices under stimulation of KCl was demonstrated.

  4. Simultaneous investigation of the neuronal and vascular compartments in the guinea pig brain isolated in vitro.

    PubMed

    de Curtis, M; Biella, G; Buccellati, C; Folco, G

    1998-11-01

    We describe a new method for studying the interactions between vascular tone changes and neuronal activity in the arterially perfused isolated brain of the adult guinea pig maintained in vitro. Electrophysiological recordings were performed in the piriform and entorhinal cortices with the entire arterial bed preserved or after vascular restriction to the territories of median and posterior cerebral arteries of one hemisphere. The changes in vascular tone were measured by means of a pressure transducer. The arterial pressure was 53.77+/-12.74 mmHg in control conditions at 30 degreesC. Intraluminal application of vasoactive drugs, such as the tromboxane A2 receptor agonist U46619 (0.1 microM) and 5-HT (3 microM), induced an increase in the resistance to perfusion pressure that was prevented by the selective antagonists. The preservation of the endothelial function was verified by inducing the release of endogenous endothelial relaxant factor after intraluminal application of 1 microM acetylcholine. The study of the reciprocal interactions between neuronal activity and vascular tone modifications demonstrated that evoked responses in the piriform and entorhinal cortices were not modulated by rapid changes of the vascular tone. A sustained and elevated plateau of vasoconstriction maintained for several minutes determined a cortical spreading depression. Epileptiform discharges induced in limbic cortices by GABAa receptor blockade were consistently associated with a vasodilation (8.26+/-2.8 mmHg). The results demonstrate that the in vitro isolated guinea pig brain preparation can be exploited for studying simultaneously neuronal activity and cerebrovascular motility. Copyright 1998 Elsevier Science B.V.

  5. Isolation of Peroxisomes from Mouse Brain Using a Continuous Nycodenz Gradient: A Comparison to the Isolation of Liver and Kidney Peroxisomes.

    PubMed

    Schönenberger, Miriam J; Kovacs, Werner J

    2017-01-01

    In the central nervous system (CNS) peroxisomes are present in all cell types, namely neurons, oligodendrocytes, astrocytes, microglia, and endothelial cells. Brain peroxisomes are smaller in size compared to peroxisomes from other tissues and are therefore referred to as microperoxisomes. We have established a purification procedure to isolate highly purified peroxisomes from the central nervous system that are well separated from the endoplasmic reticulum and mitochondria and are free of myelin contamination. The major difficulty in purification of brain peroxisomes compared to peroxisomes from liver or kidney is the presence of large amounts of myelin in the CNS, which results in contamination of the subcellular fractions. Hence, the crucial step of the isolation procedure is the elimination of myelin by the use of a sucrose gradient, since without the elimination of myelin no significant enrichment of purified peroxisomes can be achieved. Another difficulty is that in brain tissue the abundance of peroxisomes decreases significantly during postnatal development. We provide a detailed protocol for the isolation of peroxisomes from mouse central nervous system as well as a protocol for the isolation of peroxisomes from the liver and kidney using a continuous Nycodenz gradient.

  6. Transcriptome analysis of amoeboid and ramified microglia isolated from the corpus callosum of rat brain

    PubMed Central

    2012-01-01

    Background Microglia, the resident immune cells of the central nervous system (CNS), have two distinct phenotypes in the developing brain: amoeboid form, known to be amoeboid microglial cells (AMC) and ramified form, known to be ramified microglial cells (RMC). The AMC are characterized by being proliferative, phagocytic and migratory whereas the RMC are quiescent and exhibit a slow turnover rate. The AMC transform into RMC with advancing age, and this transformation is indicative of the gradual shift in the microglial functions. Both AMC and RMC respond to CNS inflammation, and they become hypertrophic when activated by trauma, infection or neurodegenerative stimuli. The molecular mechanisms and functional significance of morphological transformation of microglia during normal development and in disease conditions is not clear. It is hypothesized that AMC and RMC are functionally regulated by a specific set of genes encoding various signaling molecules and transcription factors. Results To address this, we carried out cDNA microarray analysis using lectin-labeled AMC and RMC isolated from frozen tissue sections of the corpus callosum of 5-day and 4-week old rat brain respectively, by laser capture microdissection. The global gene expression profiles of both microglial phenotypes were compared and the differentially expressed genes in AMC and RMC were clustered based on their functional annotations. This genome wide comparative analysis identified genes that are specific to AMC and RMC. Conclusions The novel and specific molecules identified from the trancriptome explains the quiescent state functioning of microglia in its two distinct morphological states. PMID:22697290

  7. Isolation and Characterization of δ-Subspecies of Protein Kinase C from Rat Brain

    NASA Astrophysics Data System (ADS)

    Ogita, Kouji; Miyamoto, Shin-Ichi; Yamaguchi, Keizo; Koide, Hiroshi; Fujisawa, Naoko; Kikkawa, Ushio; Sahara, Setsuko; Fukami, Yasuo; Nishizuka, Yasutomi

    1992-03-01

    The δ-subspecies of protein kinase C (δ PKC) was purified to near homogeneity from the Triton X-100 extract of the rat brain particulate fraction by successive chromatographies on S-Sepharose fast flow, phenyl 5PW, heparin 5PW, hydroxyapatite, and Mono Q columns. The purified enzyme was a doublet with molecular masses of 78 and 76 kDa on SDS/PAGE. The doublet proteins were separated partially by Mono Q column chromatography; both were recognized by the antibodies raised against synthetic oligopeptides, parts of the deduced amino acid sequence of the rat δ PKC. Protein phosphatase 2A treatment suggested that the 78-kDa protein was a phosphorylated form of the 76-kDa protein. To confirm the structural and genetic identity of the doublet proteins, δ PKC was expressed in COS 7 cells by transfecting its cDNA-constructed plasmid and was purified for comparison. This recombinant enzyme was also a doublet. The enzymes isolated from the brain and COS 7 cells showed identical reactivities with δ PKC-specific antibodies, chromatographic behaviors, and V8 protease peptide mappings. In addition, these two enzyme preparations were indistinguishable from each other in their responses to phosphatidylserine, diacylglycerol, phorbol esters, free fatty acids, Ca2+, and enzyme inhibitors. Comparison was also made between the enzymologic properties of δ PKC and α PKC, which were distinctly different from each other.

  8. Isolating response inhibition in the brain: Parietal versus frontal contribution.

    PubMed

    Kolodny, Tamar; Mevorach, Carmel; Shalev, Lilach

    2017-03-01

    Response inhibition is a main function of cognitive control and its neural substrates have been studied extensively. However, it is still a question whether previous brain imaging investigations were successful in isolating specific response inhibition activation. In the current study we attempted to pinpoint response inhibition in the brain using a Go/No-go task and fMRI, by contrasting rare-No-go trials with prevalent-No-go trials. Although inhibition is required in all No-go trials, task variants with rare-No-go cases (25%) create a prepotent response which elicits a strong demand for inhibition, while task variants with prevalent-No-go cases (75%) require very little inhibition effort. Since the neural activation in this design is extracted solely from No-go trials, differing only in the extent of inhibitory demand, the analysis avoids contamination of the data with motor effects or visual factors. Using this experimental design we highlight the contribution of the parietal cortex (bilaterally) to inhibitory processes, while casting doubts about the specificity of frontal activation in such processes. Future studies are required to verify that bilateral intraparietal sulcus and left temporo-parietal junction activations could be markers of inhibitory control.

  9. Sympathoadrenal Activation is Associated with Acute Traumatic Coagulopathy and Endotheliopathy in Isolated Brain Injury

    PubMed Central

    Di Battista, Alex P.; Rizoli, Sandro B.; Lejnieks, Brandon; Min, Arimie; Shiu, Maria Y.; Peng, Henry T.; Baker, Andrew J.; Hutchison, Michael G.; Churchill, Nathan; Inaba, Kenji; Nascimento, Bartolomeu B.; de Oliveira Manoel, Airton Leonardo; Beckett, Andrew; Rhind, Shawn G.

    2016-01-01

    ABSTRACT Background: Acute coagulopathy after traumatic brain injury (TBI) involves a complex multifactorial hemostatic response that is poorly characterized. Objectives: To examine early posttraumatic alterations in coagulofibrinolytic, endothelial, and inflammatory blood biomarkers in relation to sympathetic nervous system (SNS) activation and 6-month patient outcomes, using multivariate partial least-squares (PLS) analysis. Patients and Methods: A multicenter observational study of 159 adult isolated TBI patients admitted to the emergency department at an urban level I trauma center, was performed. Plasma concentrations of 6 coagulofibrinolytic, 10 vascular endothelial, 19 inflammatory, and 2 catecholamine biomarkers were measured by immunoassay on admission and 24 h postinjury. Neurological outcome at 6 months was assessed using the Extended Glasgow Outcome Scale. PLS-discriminant analysis was used to identify salient biomarker contributions to unfavorable outcome, whereas PLS regression analysis was used to evaluate the covariance between SNS correlates (catecholamines) and biomarkers of coagulopathy, endotheliopathy, and inflammation. Results: Biomarker profiles in patients with an unfavorable outcome displayed procoagulation, hyperfibrinolysis, glycocalyx and endothelial damage, vasculature activation, and inflammation. A strong covariant relationship was evident between catecholamines and biomarkers of coagulopathy, endotheliopathy, and inflammation at both admission and 24 h postinjury. Conclusions: Biomarkers of coagulopathy and endotheliopathy are associated with poor outcome after TBI. Catecholamine levels were highly correlated with endotheliopathy and coagulopathy markers within the first 24 h after injury. Further research is warranted to characterize the pathogenic role of SNS-mediated hemostatic alterations in isolated TBI. PMID:27206278

  10. Distinct patterns of brain function in children with isolated spelling impairment: new insights.

    PubMed

    Gebauer, Daniela; Enzinger, Christian; Kronbichler, Martin; Schurz, Matthias; Reishofer, Gernot; Koschutnig, Karl; Kargl, Reinhard; Purgstaller, Christian; Fazekas, Franz; Fink, Andreas

    2012-06-01

    Studies investigating reading and spelling difficulties heavily focused on the neural correlates of reading impairments, whereas spelling impairments have been largely neglected so far. Hence, the aim of the present study was to investigate brain structure and function of children with isolated spelling difficulties. Therefore, 31 children, aged ten to 15 years, were investigated by means of functional MRI and DTI. This study revealed that children with isolated spelling impairment exhibit a stronger right hemispheric activation compared to children with reading and spelling difficulties and controls, when engaged in an orthographic decision task, presumably reflecting a highly efficient serial grapheme-phoneme decoding compensation strategy. In addition, children with spelling impairment activated bilateral inferior and middle frontal gyri during processing correctly spelled words and misspelled words, whereas the other two groups showed bilateral activation only in the misspelled condition, suggesting that additional right frontal engagement could be related to generally higher task demand and effort. DTI analyses revealed stronger frontal white matter integrity (fractional anisotropy) in controls (compared to spelling and reading impaired children), whereas no structural differences between controls and spelling impaired children were observed.

  11. Genetic similarity between cysticerci of Taenia solium isolated from human brain and from pigs.

    PubMed

    Hinojosa-Juarez, Araceli Consuelo; Sandoval-Balanzario, Miguel; McManus, Donald Peter; Monroy-Ostria, Amalia

    2008-09-01

    Mitochondrial (mt) cox1 and ribosomal ITS1 DNA sequences from Taenia solium cysticercus isolates from pigs and cysticerci (racemose and cellulose types) from patients with neurocysticercosis were amplified by the polymerase chain reaction (PCR). The amplicons were sequenced in order to determine the genetic relationship between these types of cysticerci. Phylogenetic trees were constructed and evolutionary distances were calculated. ITS1 and mt cox1 cysticerci sequence data were compared with previously published Taenia spp. sequences. The variation in the ITS1 and cox1 sequences of samples collected from Mexico was minimal, regardless of geographical origin, size or colour of cysticerci from either pigs or human brain. These results suggest that the racemose and cellulose types represent genetically identical metacestodes of T. solium. Alignment of the mt cox1 sequences of the Mexican samples with sequences of other Taenia taxa showed that most were very similar to T. solium from Mexico and T. solium from Colombia; one T. solium Mexican isolate and Taenia hydatigena were placed in the same group close to Taenia crassiceps. The ITS1 sequences for the Mexican T. solium samples indicated the majority were in the same group as the Latin American T. solium. Two Mexican T. solium samples and T. solium from Philippines were placed together in a different group.

  12. Word meaning acquisition is reflected in brain potentials of isolated words

    PubMed Central

    Kuipers, Jan Rouke; Uminski, Anastasia; Green, Zoe; Hughes, David; Aglietti, Tommaso

    2017-01-01

    Learning a new concept and corresponding word typically involves repeated exposure to the word in the same or a similar context until the link crystallizes in long term memory. Although electrophysiological indices of the result of learning are well documented, there is currently no measure of the process of conceptually-mediated learning itself. Here, we recorded event-related brain potentials from participants who read unfamiliar words presented in isolation followed by a definition that either explained the meaning of the word or was a true, but uninformative statement. Self-reported word knowledge ratings increased for those words that were followed by meaningful definitions and were correlated with a decrease in ERP amplitude of a late frontal negativity (LFN) elicited by the isolated word. Importantly, the rate of LFN amplitude change predicted post-hoc learning outcome measures. Therefore, the LFN is real-time measure that is not under conscious control and which reflects conceptually-mediated learning. We propose that the LFN provides for the first time the opportunity to assess learning during study. PMID:28256517

  13. Catecholamines as outcome markers in isolated traumatic brain injury: the COMA-TBI study.

    PubMed

    Rizoli, Sandro B; Jaja, Blessing N R; Di Battista, Alex P; Rhind, Shawn G; Neto, Antonio Capone; da Costa, Leodante; Inaba, Kenji; da Luz, Luis Teodoro; Nascimento, Bartolomeu; Perez, Adic; Baker, Andrew J; de Oliveira Manoel, Airton Leonardo

    2017-02-23

    Elevated catecholamine levels might be associated with unfavorable outcome after traumatic brain injury (TBI). We investigated the association between catecholamine levels in the first 24 h post-trauma and functional outcome in patients with isolated moderate-to-severe TBI. A cohort of 174 patients who sustained isolated blunt TBI was prospectively enrolled from three Level-1 Trauma Centers. Epinephrine (Epi) and norepinephrine (NE) concentrations were measured at admission (baseline), 6, 12 and 24 h post-injury. Outcome was assessed at 6 months by the extended Glasgow Outcome Scale (GOSE) score. Fractional polynomial plots and logistic regression models (fixed and random effects) were used to study the association between catecholamine levels and outcome. Effect size was reported as the odds ratio (OR) associated with one logarithmic change in catecholamine level. At 6 months, 109 patients (62.6%) had an unfavorable outcome (GOSE 5-8 vs. 1-4), including 51 deaths (29.3%). Higher admission levels of Epi were associated with a higher risk of unfavorable outcome (OR, 2.04, 95% CI: 1.31-3.18, p = 0.002) and mortality (OR, 2.86, 95% CI: 1.62-5.01, p = 0.001). Higher admission levels of NE were associated with higher risk of unfavorable outcome (OR, 1.59, 95% CI: 1.07-2.35, p = 0.022) but not mortality (OR, 1.45, 95% CI: 0.98-2.17, p = 0.07). There was no relationship between the changes in Epi levels over time and mortality or unfavorable outcome. Changes in NE levels with time were statistically associated with a higher risk of mortality, but the changes had no relation to unfavorable outcome. Elevated circulating catecholamines, especially Epi levels on hospital admission, are independently associated with functional outcome and mortality after isolated moderate-to-severe TBI.

  14. Imaging MALDI mass spectrometry using an oscillating capillary nebulizer matrix coating system and its application to analysis of lipids in brain from a mouse model of Tay-Sachs/Sandhoff disease.

    PubMed

    Chen, Yanfeng; Allegood, Jeremy; Liu, Ying; Wang, Elaine; Cachón-Gonzalez, Begoña; Cox, Timothy M; Merrill, Alfred H; Sullards, M Cameron

    2008-04-15

    The quality of tissue imaging by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) depends on the effectiveness of the matrix deposition, especially for lipids that may dissolve in the solvent used for the matrix application. This article describes the use of an oscillating capillary nebulizer (OCN) to spray small droplets of matrix aerosol onto the sample surface for improved matrix homogeneity, reduced crystal size, and controlled solvent effects. This system was then applied to the analysis of histological slices of brains from mice with homozygous disruption of the hexb gene (hexb-/-), a model of Tay-Sachs and Sandhoff disease, versus the functionally normal heterozygote (hexb+/-) by imaging MALDI-MS. This allowed profiling and localization of many different lipid species, and of particular interest, ganglioside GM2, asialo-GM2 (GA2), and sulfatides (ST). The presence of these compounds was confirmed by analysis of brain extracts using electrospray ionization in conjunction with tandem mass spectrometry (MS/MS). The major fatty acid of the ceramide backbone of both GM2 and GA2 was identified as stearic acid (18:0) versus nervonic acid (24:1) for ST by both tissue-imaging MS and ESI-MS/MS. GM2 and GA2 were highly elevated in hexb-/- and were both localized in the granular cell region of the cerebellum. ST, however, was localized mainly in myelinated fiber (white matter) region of the cerebellum as well as in the brain stem with a relatively uniform distribution and had similar relative signal intensity for both hexb+/- and hexb-/- brain. It was also observed that there were distinct localizations for numerous other lipid subclasses; hence, imaging MALDI-MS could be used for "lipidomic" studies. These results illustrate the usefulness of tissue-imaging MALDI-MS with matrix deposition by OCN for histologic comparison of lipids in tissues such as brains from this mouse model of Tay-Sachs and Sandhoff disease.

  15. Brain dopamine response in isolated 10-day-old rats: assessment using D2 binding and dopamine turnover.

    PubMed

    Kehoe, P; Clash, K; Skipsey, K; Shoemaker, W J

    1996-01-01

    A single 5-min isolation from the nest, dam, and siblings in 10-day-old rat pups was investigated for its effect on brain dopamine systems. The release of dopamine in innervated brain regions was measured in separate studies using in vivo ligand binding of 3H-raclopride, ex vivo binding using 3H-raclopride, and neurochemical measurement of the dopamine turnover using levels of DOPAC and dopamine. In addition, in vitro homogenate binding was performed to determine baseline Bmax and Kd values for 3H-raclopride binding sites across treatments. Isolation for 5 min in a "novel" environment resulted in decreased 3H-raclopride binding in striatum and septum as determined by both in vivo and ex vivo binding, as well as increased dopamine turnover. There was no difference in Bmax and Kd values for 3H-raclopride in these brain regions after the 5-min isolation, indicating that the binding decreases were due to an increase of available dopamine, presumably from terminal release. The convergence of results from three different techniques supports the interpretation that dopamine is released during the 5-min isolation in both brain regions.

  16. Acute and delayed mild coagulopathy are related to outcome in patients with isolated traumatic brain injury

    PubMed Central

    2011-01-01

    Introduction The relationship between isolated traumatic brain injury (TBI) associated coagulopathy and patient prognosis frequently lacks information regarding the time course of coagulation disorders throughout the post-traumatic period. This study was conducted to assess the prevalence and time course of post-traumatic coagulopathy in patients with isolated TBI and the relationship of these hemostatic disorders with outcome. Methods The local Human Subjects Committee approved the study. We retrospectively studied the medical records of computed tomography (CT)-confirmed isolated TBI patients with an extracranial abbreviated injury scale (AIS) <3 who were primarily referred to a Level 1 trauma centre in Amsterdam (n = 107). Hemostatic parameters including activated partial thromboplastin time (aPTT), prothrombin time (PT), platelet count, hemoglobin, hematocrit, glucose, pH and lactate levels were recorded throughout a 72-hour period as part of a routine standardized follow-up of TBI. Coagulopathy was defined as a aPPT >40 seconds and/or a PTT in International Normalized Ratio (INR) >1.2 and/or a platelet count <120*109/l. Results Patients were mostly male, aged 48 ± 20 years with a median injury severity score of 25 (range 20 to 25). Early coagulopathy as diagnosed in the emergency department (ED) occurred in 24% of all patients. The occurrence of TBI-related coagulopathy increased to 54% in the first 24 hours post-trauma. In addition to an increased age and disturbed pupillary reflex, both coagulopathy upon ED arrival and during the first 24 hours post-trauma provided an independent prognostic factor for unfavorable outcome (odds ratio (OR) 3.75 (95% CI 1.07 to 12.51; P = 0.04) and OR 11.61 (2.79 to 48.34); P = 0.003). Conclusions Our study confirms a high prevalence of early and delayed coagulopathy in patients with isolated TBI, which is strongly associated with an unfavorable outcome. These data support close monitoring of hemostasis after TBI and indicate

  17. Age-associated changes in gene expression in human brain and isolated neurons.

    PubMed

    Kumar, Azad; Gibbs, J Raphael; Beilina, Alexandra; Dillman, Allissa; Kumaran, Ravindran; Trabzuni, Daniah; Ryten, Mina; Walker, Robert; Smith, Colin; Traynor, Bryan J; Hardy, John; Singleton, Andrew B; Cookson, Mark R

    2013-04-01

    Previous studies have suggested that there are genes whose expression levels are associated with chronological age. However, which genes show consistent age association across studies, and which are specific to a given organism or tissue remains unresolved. Here, we reassessed this question using 2 independently ascertained series of human brain samples from 2 anatomic regions, the frontal lobe of the cerebral cortex and cerebellum. Using microarrays to estimate gene expression, we found 60 associations between expression and chronological age that were statistically significant and were replicated in both series in at least 1 tissue. There were a greater number of significant associations in the frontal cortex compared with the cerebellum. We then repeated the analysis in a subset of samples using laser capture microdissection to isolate Purkinje neurons from the cerebellum. We were able to replicate 5 gene associations from either frontal cortex or cerebellum in the Purkinje cell dataset, suggesting that there is a subset of genes which have robust changes with aging. Of these, the most consistent and strongest association was with expression of RHBDL3, a rhomboid protease family member. We confirmed several hits using an independent technique (quantitative reverse transcriptase polymerase chain reaction) and in an independent published sample series that used a different array platform. We also interrogated larger patterns of age-related gene expression using weighted gene correlation network analysis. We found several modules that showed significant associations with chronological age and, of these, several that showed negative associations were enriched for genes encoding components of mitochondria. Overall, our results show that there is a distinct and reproducible gene signature for aging in the human brain. Published by Elsevier Inc.

  18. Social isolation stress-induced oxidative damage in mouse brain and its modulation by majonoside-R2, a Vietnamese ginseng saponin.

    PubMed

    Huong, Nguyen Thi Thu; Murakami, Yukihisa; Tohda, Michihisa; Watanabe, Hiroshi; Matsumoto, Kinzo

    2005-08-01

    Stressors with a physical factor such as immobilization, electric foot shock, cold swim, etc., have been shown to produce oxidative damage to membrane lipids in the brain. In this study, we investigated the effect of protracted social isolation stress on lipid peroxidation activity in the mouse brain and elucidated the protective effect of majonoside-R2, a major saponin component of Vietnamese ginseng, in mice exposed to social isolation stress. Thiobarbituric acid reactive substance levels, one of the end products of lipid peroxidation reaction, were increased in the brains of mice subjected to 6-8 weeks of social isolation stress. Measurements of nitric oxide (NO) metabolites (NO(x)(-)) also revealed a significant increase of NO production in the brains of socially isolated mice. Moreover, the depletion of brain glutathione content, an endogenous antioxidant, in socially isolated animals occurred in association with the rise in lipid peroxidation. The intraperitoneal administration of majonoside-R2 (10-50 mg/kg) had no effect on thiobarbituric acid reactive substances (TBARS), NO, or glutathione levels in the brains of group-housed control mice but it significantly suppressed the increase in TBARS and NO levels and the decrease in glutathione levels caused by social isolation stress. These results suggest that mice subjected to 6-8 weeks of social isolation stress produces oxidative damage in the brain partly via enhancement of NO production, and that majonoside-R2 exerts a protective effect by modulating NO and glutathione systems in the brain.

  19. Comparing brain activation associated with isolated upper and lower limb movement across corresponding joints.

    PubMed

    Luft, Andreas R; Smith, Gerald V; Forrester, Larry; Whitall, Jill; Macko, Richard F; Hauser, Till-Karsten; Goldberg, Andrew P; Hanley, Daniel F

    2002-10-01

    It was shown recently that functional activation across brain motor areas during locomotion and foot movements are similar but differ substantially from activation related to upper extremity movement (Miyai [2001]: Neuroimage 14:1186-1192). The activation pattern may be a function of the behavioral context of the movement rather than of its mechanical properties. We compare motor system activation patterns associated with isolated single-joint movement of corresponding joints in arm and leg carried out in equal frequency and range. Eleven healthy volunteers underwent BOLD-weighted fMRI while performing repetitive elbow or knee extension/flexion. To relate elbow and knee activation to the well-described patterns of finger movement, serial finger-to-thumb opposition was assessed in addition. After identifying task-related voxels using statistical parametric mapping, activation was measured in five regions of interest (ROI; primary motor [M1] and somatosensory cortex [S1], premotor cortex, supplementary motor area [SMA] divided into preSMA and SMA-proper, and cerebellum). Differences in the degree of activation across ROIs were found between elbow and knee movement. SMA-proper activation was prominent for knee, but almost absent for elbow movement (P < 0.05); finger movement produced small but constant SMA-proper activation. Ipsilateral M1 activation was detected during knee and finger movement, but was absent for the elbow task (P < 0.05). Knee movement showed less lateralization in M1 and S1 than other tasks (P < 0.05). The data demonstrate that central motor structures contribute differently to isolated elbow and knee movement. Activation during knee movement shows similarities to gait-related activation patterns.

  20. Measurement of hydraulic conductivity in isolated arterioles of rat brain cortex.

    PubMed

    Kimura, M; Dietrich, H H; Huxley, V H; Reichner, D R; Dacey, R G

    1993-06-01

    We have developed a new method for quantification of arteriolar hydraulic conductivity (Lp) from isolated rat brain vessels. The volume flux of water per unit surface area across the arteriole wall (Jv/S) was assessed from measurements of silicon oil drop movement within an occluded vessel at two to three pressures (between 20 and 70 mmHg); the Lp was derived from the slope of the relationship between Jv/S and applied pressure. Lp was measured in isolated cerebral arterioles 1) at room temperature (22 degrees C) without spontaneous vessel tone (control Lp; n = 11), 2) at room temperature with 10(-4) M adenosine (n = 5), and 3) at 37 degrees C with vessels dilated submaximally with 10(-4) M adenosine (n = 6). Lp at 22 degrees C without adenosine was 13.2 +/- 4.2 x 10(-9) (+/- SE) cm.s-1.cmH2O-1 for all vessels studied. Lp values ranged from 1.2 to 44.1 x 10(-9) cm.s-1.cmH2O-1 with a median value that was 5.9 x 10(-9) cm.s-1.cmH2O-1. Lp increased significantly (on average, 2.6-fold) with adenosine at 37 degrees C but not with adenosine at 22 degrees C. Control Lp bore no relationship to either the development of spontaneous tone or the diameter response to pH change, two recognized indicators of vessel viability.

  1. Intranasal lobular capillary haemangioma

    PubMed Central

    Nayak, Dipak Ranjan; Bhandarkar, Ajay M; Shivamurthy, Archana; Joy, Jasmi

    2014-01-01

    Lobular capillary haemangioma (LCH) is a benign proliferation of capillaries with a characteristic lobular architecture on microscopy; it has an affinity for mucous membrane and skin of the head and neck. It is extremely rare in the nasal cavity. We present the case of a 45-year-old man who presented with epistaxis without any predisposing factors, which was diagnosed as lobular capillary haemangioma. PMID:25304675

  2. [Transvascular fluid exchange disturbed by capillary injuries].

    PubMed

    Lugrin, D; Chave, S; Raucoules, M; Grimaud, D

    1996-01-01

    Fluid exchange disorders due to capillary lesions are numerous and their extent depends on the underlying disease as well as the capillary structure of the affected organ. The inflammatory cascade, triggered by sepsis or reperfusion injury, is mediated by several humoral mediators and activated blood cells. These include pro-inflammatory cytokines, arachidonic acid, proteases, oxygen free radicals, polymorphonuclears, procoagulant, complement and fibrinolytic system. The interaction between these mediators leads to a loss of endothelial integrity, a loss of basment membrane and a disruption of the interstitial matrix, with wasting of the endothelial cytoskeleton. The alteration in permeability induces transcapillary exudation of water and protein in the interstitial space, leading to organ dysfunction, mainly the lungs and splanchnic organs. Nitric oxyde, by modulating the response of the endothelium to the cellular interaction may protect against capillary injury. Capillary "stress lesions" following microvascular hypertension are the physiological basis of neurogenic or high altitude pulmonary oedema, and overinflation injury from mechanical ventilation. The anatomic specific features of the cerebral capillaries resulted in the well known concept of blood brain barrier with it's changeing morphology. Under the effect of humoral mediators and cellular interactions, the endothelial cells are able, via a calcium-mediated mechanism, to contract and to modify capillary permeability, leading to vasogenic oedema.

  3. Regulation of respiratory and vocal motor pools in the isolated brain of Xenopus laevis.

    PubMed

    Zornik, Erik; Kelley, Darcy B

    2008-01-16

    The aquatic frog Xenopus laevis uses a complex vocal repertoire during mating and male-male interactions. Calls are produced without breathing, allowing the frogs to vocalize for long periods underwater. The Xenopus vocal organ, the larynx, is innervated by neurons in cranial motor nucleus (n.) IX-X, which contains both vocal (laryngeal) and respiratory (glottal) motor neurons. The primary descending input to n.IX-X comes from the pretrigeminal nucleus of the dorsal tegmental area of the medulla (DTAM), located in the rostral hindbrain. We wanted to characterize premotor inputs to respiratory and vocal motor neurons and to determine what mechanisms might be involved in regulating two temporally distinct rhythmic behaviors: breathing and calling. Using isolated brain and larynx preparations, we recorded extracellular activity from the laryngeal nerve and muscles and intracellular activity in laryngeal and glottal motor neurons. Spontaneous nerve activities mimicking respiratory and vocal patterns were observed. DTAM projection neurons (DTAM(IX-X) neurons) provide direct input to glottal and laryngeal motor neurons. Electrical stimulation produced short-latency coordinated activity in the laryngeal nerve. DTAM(IX-X) neurons provide excitatory monosynaptic inputs to laryngeal motor neurons and mixed excitatory and inhibitory inputs to glottal motor neurons. DTAM stimulation also produced a delayed burst of glottal motor neuron activity. Together, our data suggest that neurons in DTAM produce vocal motor output by directly activating laryngeal motor neurons and that DTAM may coordinate vocal and respiratory motor activity.

  4. Isoosmotic isolation of rat brain synaptic vesicles, some of which contain tyrosine hydroxylase.

    PubMed

    Tsudzuki, Toshihiro; Tsujita, Maki

    2004-08-01

    Rat brain synaptic vesicles were isoosmotically isolated and examined for Mg(2+)-ATPase [EC 3.6.1.3.] and tyrosine hydroxylase [EC 1.14.16.2.] associated with the synaptic vesicles. Synaptosomes in 0.32 M sucrose were disrupted by freezing and thawing treatment, and the cytosol fraction was fractionated on a Sephacryl S-500 column with a mean exclusion size of 200 nm. Peak I at the void volume was a mixture of large vesicular membranes, small amounts of synaptic vesicles and coated vesicles, etc. Peak II consisted of non- and granulated synaptic vesicles of 35-40 nm diameter, and peak III of soluble proteins. The synaptic vesicles in peak II reacted with antibodies against the H(+)-ATPase A-subunit, vesicular acetylcholine transporter, and vesicular monoamine transporter. However, they showed little Mg(2+)-ATPase activity. Tyrosine hydroxylase was observed in either peak II or III on blotting with an anti-tyrosine hydroxylase antibody. These results imply that tyrosine hydroxylase exists in soluble and bound forms to synaptic vesicles in nerve terminals.

  5. A simple method for isolating rat brain mitochondria with high metabolic activity: effects of EDTA and EGTA.

    PubMed

    Iglesias-González, Javier; Sánchez-Iglesias, Sofía; Beiras-Iglesias, Andrés; Soto-Otero, Ramón; Méndez-Álvarez, Estefanía

    2013-02-15

    Isolated mitochondria are widely used in metabolic and oxidative stress studies for neurodegenerative diseases. In the present work, the influence of EGTA and EDTA has been tested on a sucrose-based differential centrifugation protocol in order to establish the optimal concentrations to be used in this process. Our results showed alterations in both active and resting respiration, which were dependent on both the addition of EDTA or EGTA to the isolation buffer and the chelator concentration used. However, the addition of chelator to the isolation medium does not modify the mitochondria structure as assessed by both distribution of biological markers and electron micrography in the final pellet. Our results endorse this protocol as the method of choice for metabolic and oxidative stress experiments with fresh isolated rat brain mitochondria. Copyright © 2012 Elsevier B.V. All rights reserved.

  6. Isolation of 4,5-O-Dicaffeoylquinic Acid as a Pigmentation Inhibitor Occurring in Artemisia capillaris Thunberg and Its Validation In Vivo

    PubMed Central

    Tabassum, Nadia; Lee, Ji-Hyung; Yim, Soon-Ho; Batkhuu, Galzad Javzan

    2016-01-01

    There is a continual need to develop novel and effective melanogenesis inhibitors for the prevention of hyperpigmentation disorders. The plant Artemisia capillaris Thunberg (Oriental Wormwood) was screened for antipigmentation activity using murine cultured cells (B16-F10 malignant melanocytes). Activity-based fractionation using HPLC and NMR analyses identified the compound 4,5-O-dicaffeoylquinic acid as an active component in this plant. 4,5-O-Dicaffeoylquinic acid significantly reduced melanin synthesis and tyrosinase activity in a dose-dependent manner in the melanocytes. In addition, 4,5-O-dicaffeoylquinic acid treatment reduced the expression of tyrosinase-related protein-1. Significantly, we could validate the antipigmentation activity of this compound in vivo, using a zebrafish model. Moreover, 4,5-O-dicaffeoylquinic acid did not show toxicity in this animal model. Our discovery of 4,5-O-dicaffeoylquinic acid as an inhibitor of pigmentation that is active in vivo shows that this compound can be developed as an active component for formulations to treat pigmentation disorders. PMID:27528883

  7. Aβ40 Reduces P-Glycoprotein at the Blood–Brain Barrier through the Ubiquitin–Proteasome Pathway

    PubMed Central

    Zhong, Yu; Wolf, Andrea; LeVine, Harry; Miller, David S.; Bauer, Björn

    2016-01-01

    Failure to clear amyloid-β (Aβ) from the brain is in part responsible for Aβ brain accumulation in Alzheimer's disease (AD). A critical protein for clearing Aβ across the blood–brain barrier is the efflux transporter P-glycoprotein (P-gp) in the luminal plasma membrane of the brain capillary endothelium. P-gp is reduced at the blood–brain barrier in AD, which has been shown to be associated with Aβ brain accumulation. However, the mechanism responsible for P-gp reduction in AD is not well understood. Here we focused on identifying critical mechanistic steps involved in reducing P-gp in AD. We exposed isolated rat brain capillaries to 100 nm Aβ40, Aβ40, aggregated Aβ40, and Aβ42. We observed that only Aβ40 triggered reduction of P-gp protein expression and transport activity levels; this occurred in a dose- and time-dependent manner. To identify the steps involved in Aβ-mediated P-gp reduction, we inhibited protein ubiquitination, protein trafficking, and the ubiquitin–proteasome system, and monitored P-gp protein expression, transport activity, and P-gp-ubiquitin levels. Thus, exposing brain capillaries to Aβ40 triggers ubiquitination, internalization, and proteasomal degradation of P-gp. These findings may provide potential therapeutic targets within the blood–brain barrier to limit P-gp degradation in AD and improve Aβ brain clearance. SIGNIFICANCE STATEMENT The mechanism reducing blood–brain barrier P-glycoprotein (P-gp) in Alzheimer's disease is poorly understood. In the present study, we focused on defining this mechanism. We demonstrate that Aβ40 drives P-gp ubiquitination, internalization, and proteasome-dependent degradation, reducing P-gp protein expression and transport activity in isolated brain capillaries. These findings may provide potential therapeutic avenues within the blood–brain barrier to limit P-gp degradation in Alzheimer's disease and improve Aβ brain clearance. PMID:26865616

  8. ACUTE INDUCTION OF EPILEPTIFORM DISCHARGES BY PILOCARPINE IN THE IN VITRO ISOLATED GUINEA-PIG BRAIN REQUIRES ENHANCEMENT OF BLOOD–BRAIN BARRIER PERMEABILITY

    PubMed Central

    UVA, L.; LIBRIZZI, L.; MARCHI, N.; NOE, F.; BONGIOVANNI, R.; VEZZANI, A.; JANIGRO, D.; DE CURTIS, M.

    2008-01-01

    Systemic application of the muscarinic agonist, pilocarpine, is commonly utilized to induce an acute status epilepticus that evolves into a chronic epileptic condition characterized by spontaneous seizures. Recent findings suggest that the status epilepticus induced by pilocarpine may be triggered by changes in the blood–brain barrier (BBB) permeability. We tested the role of the BBB in an acute pilocarpine model by using the in vitro model brain preparation and compared our finding with in vivo data. Arterial perfusion of the in vitro isolated guinea-pig brain with <1 mM pilocarpine did not cause epileptiform activity, but rather reduced synaptic transmission and induced steady fast (20–25 Hz) oscillatory activity in limbic cortices. These effects were reversibly blocked by co-perfusion of the muscarinic antagonist atropine sulfate (5 μM). Brain pilocarpine measurements in vivo and in vitro suggested modest BBB penetration. Pilocarpine induced epileptiform discharges only when perfused with compounds that enhance BBB permeability, such as bradykinin (n=2) or histamine (n=10). This pro-epileptic effect was abolished when the BBB-impermeable muscarinic antagonist atropine methyl bromide (5 μM) was co-perfused with histamine and pilocarpine. In the absence of BBB permeability enhancing drugs, pilocarpine induced epileptiform activity only after arterial perfusion at concentrations >10 mM. Ictal discharges correlated with a high intracerebral pilocarpine concentration measured by high pressure liquid chromatography. We propose that acute epileptiform discharges induced by pilocarpine treatment in the in vitro isolated brain preparation are mediated by a dose-dependent, atropine-sensitive muscarinic effect promoted by an increase in BBB permeability. Pilocarpine accumulation secondary to BBB permeability changes may contribute to in vivo ictogenesis in the pilocarpine epilepsy model. PMID:18082973

  9. Porosity, single-phase permeability, and capillary pressure data from preliminary laboratory experiments on selected samples from Marker Bed 139 at the Waste Isolation Pilot Plant. Volume 1 of 3: Main report, appendix A

    SciTech Connect

    Howarth, S.M.; Christian-Frear, T.

    1997-08-01

    Three groups of core samples from Marker Bed 139 of the Salado Formation at the Waste Isolation Pilot Plant (WIPP) were analyzed to provide data to support the development of numerical models used to predict the long-term hydrologic and structural response of the WIPP repository. These laboratory experiments, part of the FY93 Experimental Scoping Activities of the Salado Two-Phase Flow Laboratory Program, were designed to (1) generate WIPP-specific porosity and single-phase permeability data, (2) provide information needed to design and implement planned tests to measure two-phase flow properties, including threshold pressure, capillary pressure, and relative permeability, and (3) evaluate the suitability of using analog correlations for the Salado Formation to assess the long-term performance of the WIPP. This report contains a description of the boreholes core samples, the core preparation techniques used, sample sizes, testing procedures, test conditions, and results of porosity and single-phase permeability tests performed at three laboratories: TerraTek, Inc. (Salt Lake City, UT), RE/SPEC, Inc. (Rapid City, SD), and Core Laboratories-Special Core Analysis Laboratory (Carrollton, TX) for Rock Physics Associates. In addition, this report contains the only WIPP-specific two-phase-flow capillary-pressure data for twelve core samples. The WIPP-specific data generated in this laboratory study and in WIPP field-test programs and information from suitable analogs will form the basis for specification of single- and two-phase flow parameters for anhydrite markers beds for WIPP performance assessment calculations.

  10. Exploring the Use of Isolated Expressions and Film Clips to Evaluate Emotion Recognition by People with Traumatic Brain Injury.

    PubMed

    Zupan, Barbra; Neumann, Dawn

    2016-05-15

    The current study presented 60 people with traumatic brain injury (TBI) and 60 controls with isolated facial emotion expressions, isolated vocal emotion expressions, and multimodal (i.e., film clips) stimuli that included contextual cues. All stimuli were presented via computer. Participants were required to indicate how the person in each stimulus was feeling using a forced-choice format. Additionally, for the film clips, participants had to indicate how they felt in response to the stimulus, and the level of intensity with which they experienced that emotion.

  11. Vaccine-induced rabies in a red fox (Vulpes vulpes): isolation of vaccine virus in brain tissue and salivary glands.

    PubMed

    Hostnik, Peter; Picard-Meyer, Evelyne; Rihtarič, Danijela; Toplak, Ivan; Cliquet, Florence

    2014-04-01

    Oral vaccination campaigns to eliminate fox rabies were initiated in Slovenia in 1995. In May 2012, a young fox (Vulpes vulpes) with typical rabies signs was captured. Its brain and salivary gland tissues were found to contain vaccine strain SAD B19. The Basic Logical Alignment Search Tool alignment of 589 nucleotides determined from the N gene of the virus isolated from the brain and salivary glands of the affected fox was 100% identical to the GenBank reference SAD B19 strain. Sequence analysis of the N and M genes (4,351 nucleotides) showed two nucleotide modifications at position 1335 (N gene) and 3114 (M gene) in the KC522613 isolate identified in the fox compared to SAD B19.

  12. Capillary rheometry for thermosets

    NASA Technical Reports Server (NTRS)

    Malguarnera, S. C.; Carroll, D. R.

    1982-01-01

    Capillary rheometry is effectively used with thermosets. Most important is providing a uniform temperature in the barrel. This was successfully accomplished by using a circulating hot oil system. Standard capillary rheometry methods provide the dependence of thermoset apparent viscosity on shear rate, temperature and time. Such information is very useful in evaluating resin processability and in setting preliminary fabrication conditions.

  13. Low-dose recombinant factor VIIa for reversing coagulopathy in patients with isolated traumatic brain injury.

    PubMed

    Yuan, Qiang; Wu, Xing; Du, Zhuo-Ying; Sun, Yi-Rui; Yu, Jian; Li, Zhi-Qi; Wu, Xue-Hai; Mao, Ying; Zhou, Liang-Fu; Hu, Jin

    2015-02-01

    The purpose of this study was to investigate the role of low-dose recombinant factor VIIa (rFVIIa) (20 μg/kg) in reversing coagulopathy in patients with isolated traumatic brain injury (TBI). Patients with isolated TBI and coagulopathy at admission were enrolled prospectively from January 2010 to December 2011. The patients were divided into 2 groups: the rFVIIa and the no-rFVIIa groups. In the rFVIIa group, patients received a single dose of 20 μg/kg rFVIIa intravenously to reverse their coagulopathy in addition to blood products. Patients in the no-rFVIIa group received only blood products to correct the coagulopathy. The clinical outcome variables evaluated included changes in coagulation parameters after administration for reversing coagulopathy, the occurrence of progressive hemorrhagic injury (PHI), intensive care unit length of stay, the incidence of thromboembolic complications, inhospital mortality, and 90-day Glasgow Outcome Scale. Eighty-seven patients were ultimately included in this study. Of them, 49 patients were treated with blood products alone, whereas 38 patients also received rFVIIa to reverse their coagulopathy. The improvement in international normalized ratio was greater in the rFVIIa group (0.26 [0.18-0.39]) than in the no-rFVIIa group (0.06 [-0.11 to 0.30]) (P = .001). In addition, the improvement in lactate was also greater in the rFVIIa group (0.33 [-0.18 to 0.54]) than in the no-rFVIIa group (0.04 [-0.25 to 0.20]) (P = .029). During the period after we began to correct the coagulopathy, PHI occurred in 19 patients (38.8%) in the no-rFVIIa group, which was significantly higher than that in the rFVIIa group (7, 18.4%; P = .040). The rate of cerebral infarction was similar in both groups (10.2% vs 5.3%). There was a trend indicating that low-dose rFVIIa therapy was associated with a lower mortality, but the association was not statistically significant (P = .266). The use of low-dose rFVIIa (20 μg/kg) is effective for correcting

  14. Predictors of mortality in patients with isolated severe traumatic brain injury.

    PubMed

    Strnad, Matej; Borovnik Lesjak, Vesna; Vujanović, Vitka; Križmarić, Miljenko

    2017-02-01

    Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. Many prognostic models predicting mortality in patients with TBI were developed, which also include patients with mild or moderate TBI and patients who suffered major extracranial injuries. From a prospective database, we conducted a retrospective medical chart review covering the period between January 2000 and December 2012 of patients with isolated severe TBI (Abbreviated Injury Score for head, AISH ≥ 3) without extracranial injuries, who were intubated in the field using the rapid sequence intubation method and were of age 16 or more. Prehospital vital signs, Injury Severity Score (ISS) and laboratory tests were compared in two study groups: survivors (n = 25) and non-survivors (n = 27). Selected variables identified during univariate analysis (p < 0.1) were then subjected to multivariate analysis logistic regression model. Univariate analysis showed that in-hospital mortality was statistically significantly associated with male sex (p = 0.040), ISS (p = 0.005) and mydriasis (p = 0.012). For predicting mortality, area under the curve (AUC) was calculated: for ISS 0.76 (95 % confidence interval, CI; 0.63-0.90; p < 0.001) and for initial Glasgow Coma Scale (GCS) 0.64 (95 % CI, 0.49-0.80, p = 0.079). In the multivariate analysis, ISS (odds ratio, OR; 1.19, 95 % CI, 1.06-1.35; p = 0.004) and mydriasis (OR, 5.73; 95 % CI, 1.06-30.88; p = 0.042) were identified as independent risk factors for in-hospital mortality. The AUC for the regression model was 0.83 (95 % CI, 0.71-0.94; p < 0.001). In prehospital intubated patients with isolated severe TBI only ISS and mydriasis were found to be independent predictors of in-hospital mortality.

  15. Osteoporosis Self-Assessment Tool for Asians Can Predict Neurologic Prognosis in Patients with Isolated Moderate Traumatic Brain Injury.

    PubMed

    Chao, Chia-Hung; Su, Yu-Feng; Chan, Hon-Man; Huang, Shiuh-Lin; Lin, Chih-Lung; Kwan, Aij-Lie; Lou, Yun-Ting; Chen, Chao-Wen

    2015-01-01

    Osteoporosis Self-Assessment Tool for Asians (OSTA) has been proved to be a simple and effective tool for recognizing osteoporosis risk. Our previous study has demonstrated that the preoperative OSTA index was a good prognostic predictor for stage II and III colon cancer patients after surgery. We aim to evaluate the value of OSTA index in prognostication of isolated traumatic brain injury with moderate severity (GCS 9-13). We retrospectively reviewed all patients visiting Kaohsiung Medical University Hospital emergency department due to isolated moderate traumatic brain injury from Jan. 2010 to Dec. 2012. Background data (including the OSTA index), clinical presentations, management and outcomes (ICU admission days, total admission days, complications, Glasgow outcome score (GOS) at discharge, mortality) of the patients were recorded for further analysis. Our major outcome was good neurologic recovery defined as GOS of 5. Pearson chi-square test and the Mann-Whitney U test were used to compare demographic features. Multiple logistic regression was used to identify independent risk factors. 107 isolated moderate TBI patients were studied. 40 patients (37.4%) showed good recovery and 10 (9.3%) died at discharge. The univariate analysis revealed that younger age, higher OSTA index, lower ISS, lower AIS-H, and avoidance to neurosurgery were associated with better neurologic outcome for all moderate TBI patients. Multivariate analysis revealed that lower ISS, higher OSTA, and the avoidance of neurosurgery were independent risk factors predicting good neurologic recovery. Higher ISS, lower OSTA index and exposure to neurosurgery were the independent risk factors for poorer recovery from isolated moderate TBI. In addition to labeling the cohort harboring osteoporotic risk, OSTA index could predict neurologic prognosis in patients with isolated moderate traumatic brain injury.

  16. Osteoporosis Self-Assessment Tool for Asians Can Predict Neurologic Prognosis in Patients with Isolated Moderate Traumatic Brain Injury

    PubMed Central

    Chan, Hon-Man; Huang, Shiuh-Lin; Lin, Chih-Lung; Kwan, Aij-Lie; Lou, Yun-Ting; Chen, Chao-Wen

    2015-01-01

    Objectives Osteoporosis Self-Assessment Tool for Asians (OSTA) has been proved to be a simple and effective tool for recognizing osteoporosis risk. Our previous study has demonstrated that the preoperative OSTA index was a good prognostic predictor for stage II and III colon cancer patients after surgery. We aim to evaluate the value of OSTA index in prognostication of isolated traumatic brain injury with moderate severity (GCS 9-13). Methods We retrospectively reviewed all patients visiting Kaohsiung Medical University Hospital emergency department due to isolated moderate traumatic brain injury from Jan. 2010 to Dec. 2012. Background data (including the OSTA index), clinical presentations, management and outcomes (ICU admission days, total admission days, complications, Glasgow outcome score (GOS) at discharge, mortality) of the patients were recorded for further analysis. Our major outcome was good neurologic recovery defined as GOS of 5. Pearson chi-square test and the Mann-Whitney U test were used to compare demographic features. Multiple logistic regression was used to identify independent risk factors. Results 107 isolated moderate TBI patients were studied. 40 patients (37.4%) showed good recovery and 10 (9.3%) died at discharge. The univariate analysis revealed that younger age, higher OSTA index, lower ISS, lower AIS-H, and avoidance to neurosurgery were associated with better neurologic outcome for all moderate TBI patients. Multivariate analysis revealed that lower ISS, higher OSTA, and the avoidance of neurosurgery were independent risk factors predicting good neurologic recovery. Conclusion Higher ISS, lower OSTA index and exposure to neurosurgery were the independent risk factors for poorer recovery from isolated moderate TBI. In addition to labeling the cohort harboring osteoporotic risk, OSTA index could predict neurologic prognosis in patients with isolated moderate traumatic brain injury. PMID:26186582

  17. Normalization of coagulopathy is associated with improved outcome after isolated traumatic brain injury.

    PubMed

    Epstein, Daniel S; Mitra, Biswadev; Cameron, Peter A; Fitzgerald, Mark; Rosenfeld, Jeffrey V

    2016-07-01

    Acute traumatic coagulopathy (ATC) has been reported in the setting of isolated traumatic brain injury (iTBI) and is associated with poor outcomes. We aimed to evaluate the effectiveness of procoagulant agents administered to patients with ATC and iTBI during resuscitation, hypothesizing that timely normalization of coagulopathy may be associated with a decrease in mortality. A retrospective review of the Alfred Hospital trauma registry, Australia, was conducted and patients with iTBI (head Abbreviated Injury Score [AIS] ⩾3 and all other body AIS <3) and coagulopathy (international normalized ratio ⩾1.3) were selected for analysis. Data on procoagulant agents used (fresh frozen plasma, platelets, cryoprecipitate, prothrombin complex concentrates, tranexamic acid, vitamin K) were extracted. Among patients who had achieved normalization of INR or survived beyond 24hours and were not taking oral anticoagulants, the association of normalization of INR and death at hospital discharge was analyzed using multivariable logistic regression analysis. There were 157 patients with ATC of whom 68 (43.3%) received procoagulant products within 24hours of presentation. The median time to delivery of first products was 182.5 (interquartile range [IQR] 115-375) minutes, and following administration of coagulants, time to normalization of INR was 605 (IQR 274-1146) minutes. Normalization of INR was independently associated with significantly lower mortality (adjusted odds ratio 0.10; 95% confidence interval 0.03-0.38). Normalization of INR was associated with improved mortality in patients with ATC in the setting of iTBI. As there was a substantial time lag between delivery of products and eventual normalization of coagulation, specific management of coagulopathy should be implemented as early as possible. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Comorbid mild traumatic brain injury increases pain symptoms in patients suffering from an isolated limb fracture.

    PubMed

    Jodoin, Marianne; Rouleau, Dominique M; Gosselin, Nadia; Benoit, Benoit; Leduc, Stéphane; Laflamme, Yves; Larson-Dupuis, Camille; De Beaumont, Louis

    2017-09-01

    This study seeks to evaluate the effects of a mild traumatic brain injury (mTBI) on pain in patients with an isolated limb fracture (ILF) when compared to a matched cohort group with no mTBI (control group). All subjects included in this observational study suffered from an ILF. Groups were matched according to the type of injury, sex, age, and time since the accident. Main outcome measurements were: Standardized semi-structured interviews at follow-up of a Level I Trauma Center, and a questionnaire on fracture-related pain symptoms. Factors susceptible to influence the perception of pain, such as age, sex, severity of post-concussive symptoms, and worker compensation were also assessed. A total of 68 subjects (36 females; 45 years old) with an ILF were selected, 34 with a comorbid mTBI and 34 without (24/34 with an upper limb fracture per group, 71% of total sample). Patients with mTBI and an ILF reported significantly higher pain scores at the time of assessment (mean: 49days, SD: 34.9), compared to the control group (p<0.0001; mean difference 2.8, 95% confidence interval 1.8-4.0). Correlational analyses show no significant association between the level of pain and factors such as age, sex, severity of post-concussive symptoms, and worker compensation. Results suggest that mTBI exacerbate perception of pain in the acute phase when occurring with an ILF, and were not explained by age, sex, post-concussive symptoms, or worker compensation. Rather, it appears possible that neurological sequelae induced by mTBI may interfere with the normal recovery of pain following trauma. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Separation and identification of peptides from gel-isolated membrane proteins using a microfabricated device for combined capillary electrophoresis/nanoelectrospray mass spectrometry.

    PubMed

    Li, J; Kelly, J F; Chernushevich, I; Harrison, D J; Thibault, P

    2000-02-01

    The coupling of microfabricated devices to nanoelectrospray mass spectrometers using both a triple quadrupole and a quadrupole time-of-flight mass spectrometer (QqTOF MS) is presented for the analysis of trace-level membrane proteins. Short disposable nanoelectrospray emitters were directly coupled to the chip device via a low dead volume connection. The analytical performance of this integrated device in terms of sensitivity and reproducibility was evaluated for standard peptide mixtures. A concentration detection limit ranging from 3.2 to 43.5 nM for different peptides was achieved in selected ion monitoring, thus representing a 10-fold improvement in sensitivity compared to that of microelectrospray using the same chip/mass spectrometer. Replicate injections indicated that reproducibility of migration time was typically less than 3.1% RSD whereas RSD values of 6-13% were observed on peak areas. Although complete resolution of individual components is not typically achieved for complex digests, the present chip capillary electrophoresis (chip-CE) device enabled proper sample cleanup and partial separation of multicomponent samples prior to mass spectral identification. Analyses of protein digests were typically achieved in less than 1.5 min with peak widths of 1.8-2.5 s (half-height definition) as indicated from individual reconstructed ion electropherograms. The application of this chip-CE/QqTOF MS system is further demonstrated for the identification of membrane proteins which form a subset of the Haemophilus influenzae proteome. Bands first separated by 1D-gel electrophoresis were excised and digested, and extracted tryptic peptides were loaded on the chip without any further sample cleanup or on-line adsorption preconcentration. Accurate molecular mass determination (< 5 ppm) in peptide-mapping experiments was obtained by introducing an internal standard via a postseparation channel. The analytical potential of this integrated device for the identification of

  20. Repeated isolation stress in the neonatal rat: relation to brain dopamine systems in the 10-day-old rat.

    PubMed

    Kehoe, P; Shoemaker, W J; Arons, C; Triano, L; Suresh, G

    1998-12-01

    Isolation of the rat pup from the nest and dam for one hour per day from PN 2-9 is a useful paradigm for producing stress in the neonate. These previously isolated rats respond to an amphetamine challenge with alterations in activity at the juvenile stage or as adults. Furthermore, when dopamine release is measured in the nucleus accumbens, juveniles release 3 times more dopamine after amphetamine than do controls. This study describes changes in behavior and brain dopamine systems at PN 10. Experiment 1 determined an appropriate amphetamine dose that could be used for behavioral activation at PN 10. Experiment 2 produced significant evidence of enhanced behavioral activation after the isolation paradigm and indicated that brain regions innervated by the mesolimbic dopamine system, septum, and hypothalamus display increased dopamine turnover and that the nigrostriatal pathway is less active. Likewise, in Experiment 3, in vivo microdialysis of the nucleus accumbens indicated that previously isolated pups respond to an amphetamine challenge with a several-fold increase in dopamine release over a 4-hour session.

  1. Oxytocin-Induced Changes in Monoamine Level in Symmetric Brain Structures of Isolated Aggressive C57Bl/6 Mice.

    PubMed

    Karpova, I V; Mikheev, V V; Marysheva, V V; Bychkov, E R; Proshin, S N

    2016-03-01

    Changes in activity of monoaminergic systems of the left and right brain hemispheres after administration of saline and oxytocin were studied in male C57Bl/6 mice subjected to social isolation. The concentrations of dopamine, norepinephrine, serotonin, and their metabolites dihydroxyphenylacetic, homovanillic, and 5-hydroxyindoleacetic acids were measured in the cerebral cortex, hippocampus, olfactory tubercle, and striatum of the left and right brain hemispheres by HPLC. In isolated aggressive males treated intranasally with saline, the content of serotonin and 5-hydroxyindoleacetic acid was significantly higher in the right hippocampus. Oxytocin reduces aggression caused by long-term social isolation, but has no absolute ability to suppress this type of behavior. Oxytocin reduced dopamine content in the left cortex and serotonin content in the right hippocampus and left striatum. Furthermore, oxytocin evened the revealed asymmetry in serotonin and 5-hydroxyindoleacetic acid concentrations in the hippocampus. At the same time, asymmetry in dopamine concentration appeared in the cortex with predominance of this transmitter in the right hemisphere. The data are discussed in the context of lateralization of neurotransmitter systems responsible for intraspecific aggression caused by long-term social isolation.

  2. Isolated Adrenocorticotropic Hormone or Thyrotropin Deficiency Following Mild Traumatic Brain Injury: Three Cases with Long-Term Follow-Up

    PubMed Central

    Baek, Cho-Ok; Kim, Yu Ji; Kim, Ji Hye

    2015-01-01

    Few studies have examined the clinical features and long-term outcomes of isolated pituitary hormone deficiencies after traumatic brain injury (TBI). Such deficiencies typically present at time intervals after TBI, especially after mild injuries such as concussions, which makes their diagnosis difficult without careful history taking. It is necessary to improve diagnosis and prevent life threatening or morbid conditions such as those that may occur in deficiencies of adrenocorticotropic hormone (ACTH) or thyroid-stimulating hormone (as known as thyrotropin, TSH), the two most important pituitary hormones in hypopituitarism treatment. Here, we report two cases of isolated ACTH deficiency and one case of isolated TSH deficiency. These patients presented at different time points after concussion and underwent long-term follow-ups. PMID:27169080

  3. Isolated Adrenocorticotropic Hormone or Thyrotropin Deficiency Following Mild Traumatic Brain Injury: Three Cases with Long-Term Follow-Up.

    PubMed

    Baek, Cho-Ok; Kim, Yu Ji; Kim, Ji Hye; Park, Ji Hyun

    2015-10-01

    Few studies have examined the clinical features and long-term outcomes of isolated pituitary hormone deficiencies after traumatic brain injury (TBI). Such deficiencies typically present at time intervals after TBI, especially after mild injuries such as concussions, which makes their diagnosis difficult without careful history taking. It is necessary to improve diagnosis and prevent life threatening or morbid conditions such as those that may occur in deficiencies of adrenocorticotropic hormone (ACTH) or thyroid-stimulating hormone (as known as thyrotropin, TSH), the two most important pituitary hormones in hypopituitarism treatment. Here, we report two cases of isolated ACTH deficiency and one case of isolated TSH deficiency. These patients presented at different time points after concussion and underwent long-term follow-ups.

  4. Cognitive impairment and structural brain changes in patients with clinically isolated syndrome at high risk for multiple sclerosis.

    PubMed

    Hynčicová, Eva; Vyhnálek, Martin; Kalina, Adam; Martinkovič, Lukáš; Nikolai, Tomáš; Lisý, Jiří; Hort, Jakub; Meluzínová, Eva; Laczó, Jan

    2017-03-01

    Patients with clinically isolated syndrome (CIS), unlike those with multiple sclerosis (MS), have a selective cognitive impairment which is not consistently related to structural brain changes. Our objective was to characterize a profile of cognitive impairment and its association with structural brain changes in patients with CIS who are at high risk of developing MS. Patients with CIS at high risk for MS on interferon-beta (n = 51) and age-, gender-, and education-matched controls (n = 44) underwent comprehensive neuropsychological testing and MRI brain scan with voxel-based morphometry. The CIS group had lower cognitive performance in verbal and nonverbal memory, information processing speed/attention/working memory, and executive and visuo-spatial functions compared to controls (p ≤ 0.040). Lower cognitive performance was present in 18-37 and 14-26% of patients with CIS at high risk for MS depending on the criteria used. Brain volume was reduced predominantly in fronto-temporal regions and the thalamus in the CIS group (p ≤ 0.019). Cognitive performance was not associated with structural brain changes except for the association between worse visuo-spatial performance and lower white matter volume in the CIS group (β = 0.29; p = 0.042). Our results indicated that patients with CIS at high risk for MS may have a pattern of lower cognitive performance and regional brain atrophy similar to that found in patients with MS. Lower cognitive performance may be present in up to one-third of patients with CIS at high risk for MS, but, unlike patients with MS, variability in their cognitive performance may lead to a lack of consistent associations with structural brain changes.

  5. Multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, Edward S.; Li, Qingbo; Lu, Xiandan

    1998-04-21

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification ("base calling") is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations.

  6. Multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, Edward S.; Chang, Huan-Tsang; Fung, Eliza N.; Li, Qingbo; Lu, Xiandan

    1996-12-10

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification ("base calling") is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations.

  7. Multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, E.S.; Li, Q.; Lu, X.

    1998-04-21

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification (``base calling``) is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations. 19 figs.

  8. Multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, E.S.; Chang, H.T.; Fung, E.N.; Li, Q.; Lu, X.

    1996-12-10

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification (``base calling``) is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations. 19 figs.

  9. Isolation and characterization of extrachromosomal circular DNAs in mouse heart, brain and liver tissues at various ages

    SciTech Connect

    Flores, S.C.

    1988-01-01

    Eucaryotic cells contains extrachromosomal circular (eccDNAs) which can be separated and distinguished from chromosomal DNA. Using alkaline denaturation-renaturation, exonuclease III digestion and density gradient centrifugations, covalently closed circular DNA (cc-cDNA) molecules were isolated from 1-, 8-, 16-, and 24-month C57BL/6 mouse heart, brain and liver organs. Restriction enzyme analyses and other enzymatic treatments established the covalently closed nature of the isolated molecules. Electron microscopic analyses of heart eccDNAs showed similar size distributions at all ages, but more discrete size classes and slightly larger circles were observed in 24-month heart eccDNA preparations. Heart contained more circles per cell than either liver or brain, which contained approximately the same amount of eccDNAs per genome. Furthermore, ({sup 3}H)-pBR322 recovery studies revealed no endogenous factors that might have affected the yields of eccDNAs from young and old tissues. To determine if there were any age-related or tissue-specific differences in repetitive sequences in eccDNAs, heart, brain and liver eccDNAs were probed with B1, B2, IAP, L1 and satellite sequences of the mouse genome. The hybridization results showed that these sequence families were differentially represented at all ages in eccDNAs. B2 sequences were the highest in heart, while satellite sequences were the highest in liver and brain. In heart, very little age-related change was observed in the quantity of repetitive sequences. Nevertheless, a tendency to decrease for B1 and B2 sequences at 24 months was observed. In liver, repetitive sequences decreased from 1 to 8 months of age, with very little change beyond that time point. Brain eccDNA repetitive sequences did not change significantly with age.

  10. Capillary rise of superspreaders.

    PubMed

    Radulovic, Jovana; Sefiane, Khellil; Shanahan, Martin E R

    2011-09-15

    Trisiloxane surfactants, known as 'superspreaders', are commonly employed in numerous applications where enhanced wetting is of the utmost importance. The underlying mechanisms of superspreader wetting have been a focus of scientific interest for ca. 2 decades, and a number of mechanisms have been proposed to explain the unique trisiloxane dynamics. We have studied trisiloxane behaviour in thin capillaries to get further insight into their interfacial activity. Additionally, our knowledge of the capillary rise of superspreaders is surprisingly limited, and the effect of this extraordinary group of surfactants on capillary phenomena has been largely overlooked. Diffusion was confirmed to be the limiting factor of trisiloxane behaviour. A tentative theoretical explanation for the phenomenon studied and an appropriate mathematical model are presented. It is concluded that the enhancement of wetting due to surfactant addition is also a function of geometry: the effect is clear for a sessile drop, but more complex and less beneficial in a capillary.

  11. Displaced capillary dies

    DOEpatents

    Kalejs, Juris P.; Chalmers, Bruce; Surek, Thomas

    1982-01-01

    An asymmetrical shaped capillary die made exclusively of graphite is used to grow silicon ribbon which is capable of being made into solar cells that are more efficient than cells produced from ribbon made using a symmetrically shaped die.

  12. Displaced capillary dies

    DOEpatents

    Kalejs, Juris P.; Chalmers, Bruce; Surek, Thomas

    1984-01-01

    An asymmetrical shaped capillary die made exclusively of graphite is used to grow silicon ribbon which is capable of being made into solar cells that are more efficient than cells produced from ribbon made using a symmetrically shaped die.

  13. Capillary nail refill test

    MedlinePlus

    ... may indicate: Dehydration Shock Peripheral vascular disease (PVD) Hypothermia Alternative Names Nail blanch test; Capillary refill time ... Elsevier Saunders; 2016:chap 79. Read More Dehydration Hypothermia Peripheral artery disease - legs Shock Review Date 4/ ...

  14. Development of a procedure for the isolation and enrichment of modified nucleosides and nucleobases from urine prior to their determination by capillary electrophoresis-mass spectrometry.

    PubMed

    Rodríguez-Gonzalo, Encarnación; Hernández-Prieto, Raquel; García-Gómez, Diego; Carabias-Martínez, Rita

    2014-01-01

    A sample treatment step based on solid-phase extraction (SPE) with polymeric sorbents has been developed for the simultaneous isolation and preconcentration of nucleosides and nucleobases from urine prior to analyses by CE-ESI-MS. In most reported methods nucleosides are isolated from urine by SPE in affinity mode, using an immobilized phenylboronic acid group, which specifically binds cis-diols. However, this is not applicable to non-cis-diol compounds. Here, different types of polymeric sorbents were evaluated for the simultaneous extraction of nucleosides and nucleobases from urine. The best results were obtained with Isolute ENV+, a hydroxylated styrene-divylbenzene polymer, whose retention capacity can be attributed mainly to hydrophobic interactions, and thus it can be applied to a broad range of compounds, regardless of whether they present or not to the cis-diol group in their structure. Other parameters such as the elution solvent and sample volume were optimized. We also studied the influence of the addition of isotopically labeled internal standards (ILISs) before or after the extraction step. The detection limits achieved were in the 0.04-0.17μg/mL range for a sample size of 2.0mL and relative standard deviations were 4-22%. The whole method developed, SPE prior to CE-ESI-MS, was applied to human urine samples from healthy volunteers. We conclude that SPE with polymeric sorbents prior to the electrophoretic CE-ESI-MS methodology constitutes a fast, valid and reliable approach for the simultaneously extraction of urinary nucleosides and nucleobases. Copyright © 2013 Elsevier B.V. All rights reserved.

  15. Capillary saturation and desaturation

    NASA Astrophysics Data System (ADS)

    Hilfer, R.; Armstrong, R. T.; Berg, S.; Georgiadis, A.; Ott, H.

    2015-12-01

    Capillary desaturation experiments produce disconnected (trapped) ganglia of mesoscopic sizes intermediate between pore size and system size. Experimental evidence for interactions between these mesoscale clusters during desaturation is analyzed and discussed within the established microscopic and macroscopic laws of Newton, Young-Laplace, and Darcy. A theoretical expression for capillary number correlations is introduced that seems to have remained unnoticed. It expresses capillary desaturation curves in terms of stationary capillary pressures and relative permeabilities. The theoretical expression shows that the plateau saturation in capillary desaturation curves may in general differ from the residual nonwetting saturation defined through the saturation limit of the main hysteresis loop. Hysteresis effects as well as the difference between wetting and nonwetting fluids are introduced into the analysis of capillary desaturation experiments. The article examines experiments with different desaturation protocols and discusses the existence of a mesoscopic length scale intermediate between pore scale and sample scale. The theoretical expression is derived entirely within the existing traditional theory of two-phase flow in porous media and compared to a recent experiment.

  16. Capillary discharge source

    DOEpatents

    Bender, III, Howard Albert

    2003-11-25

    Debris generation from an EUV electric discharge plasma source device can be significantly reduced or essentially eliminated by encasing the electrodes with dielectric or electrically insulating material so that the electrodes are shielded from the plasma, and additionally by providing a path for the radiation to exit wherein the electrodes are not exposed to the area where the radiation is collected. The device includes: (a) a body, which is made of an electrically insulating material, that defines a capillary bore that has a proximal end and a distal end and that defines at least one radiation exit; (b) a first electrode that defines a first channel that has a first inlet end that is connected to a source of gas and a first outlet end that is in communication with the capillary bore, wherein the first electrode is positioned at the distal end of the capillary bore; (c) a second electrode that defines a second channel that has a second inlet end that is in communication with the capillary bore and an outlet end, wherein the second electrode is positioned at the proximal end of the capillary bore; and (d) a source of electric potential that is connected across the first and second electrodes, wherein radiation generated within the capillary bore is emitted through the at least one radiation exit and wherein the first electrode and second electrode are shielded from the emitted radiation.

  17. Capillary saturation and desaturation.

    PubMed

    Hilfer, R; Armstrong, R T; Berg, S; Georgiadis, A; Ott, H

    2015-12-01

    Capillary desaturation experiments produce disconnected (trapped) ganglia of mesoscopic sizes intermediate between pore size and system size. Experimental evidence for interactions between these mesoscale clusters during desaturation is analyzed and discussed within the established microscopic and macroscopic laws of Newton, Young-Laplace, and Darcy. A theoretical expression for capillary number correlations is introduced that seems to have remained unnoticed. It expresses capillary desaturation curves in terms of stationary capillary pressures and relative permeabilities. The theoretical expression shows that the plateau saturation in capillary desaturation curves may in general differ from the residual nonwetting saturation defined through the saturation limit of the main hysteresis loop. Hysteresis effects as well as the difference between wetting and nonwetting fluids are introduced into the analysis of capillary desaturation experiments. The article examines experiments with different desaturation protocols and discusses the existence of a mesoscopic length scale intermediate between pore scale and sample scale. The theoretical expression is derived entirely within the existing traditional theory of two-phase flow in porous media and compared to a recent experiment.

  18. Membrane hyperpolarization induced by endoplasmic reticulum stress facilitates ca(2+) influx to regulate cell cycle progression in brain capillary endothelial cells.

    PubMed

    Kito, Hiroaki; Yamamura, Hisao; Suzuki, Yoshiaki; Ohya, Susumu; Asai, Kiyofumi; Imaizumi, Yuji

    2014-01-01

    Upregulation of the Kir2.1 channel during endoplasmic reticulum (ER) stress in t-BBEC117, an immortalized bovine brain endothelial cell line, caused a sustained increase in intracellular Ca(2+) concentration ([Ca(2+)]i) and a facilitation of cell death. Expressions of Ca(2+) influx channels (TRPC, Orai1, STIM1) were unchanged by ER stress. The ER stress-induced [Ca(2+)]i increase was mainly attributed to the deeper resting membrane potential due to Kir2.1 upregulation. ER stress arrested at the G2/M phase and it was attenuated by an inhibitor of Kir2.1. These results indicate that Kir2.1 upregulation by ER stress facilitates cell death via regulation of cell cycle progression in t-BBEC117.

  19. Volumetric and surface-based 3D MRI analyses of fetal isolated mild ventriculomegaly: brain morphometry in ventriculomegaly.

    PubMed

    Scott, Julia A; Habas, Piotr A; Rajagopalan, Vidya; Kim, Kio; Barkovich, A James; Glenn, Orit A; Studholme, Colin

    2013-05-01

    Diagnosis of fetal isolated mild ventriculomegaly (IMVM) is the most common brain abnormality on prenatal ultrasound. We have set to identify potential alterations in brain development specific to IMVM in tissue volume and cortical and ventricular local surface curvature derived from in utero magnetic resonance imaging (MRI). Multislice 2D T2-weighted MRI were acquired from 32 fetuses (16 IMVM, 16 controls) between 22 and 25.5 gestational weeks. The images were motion-corrected and reconstructed into 3D volumes for volumetric and curvature analyses. The brain images were automatically segmented into cortical plate, cerebral mantle, deep gray nuclei, and ventricles. Volumes were compared between IMVM and control subjects. Surfaces were extracted from the segmentations for local mean surface curvature measurement on the inner cortical plate and the ventricles. Linear models were estimated for age-related and ventricular volume-associated changes in local curvature in both the inner cortical plate and ventricles. While ventricular volume was enlarged in IMVM, all other tissue volumes were not different from the control group. Ventricles increased in curvature with age along the atrium and anterior body. Increasing ventricular volume was associated with reduced curvature over most of the ventricular surface. The cortical plate changed in curvature with age at multiple sites of primary sulcal formation. Reduced cortical folding was detected near the parieto-occipital sulcus in IMVM subjects. While tissue volume appears to be preserved in brains with IMVM, cortical folding may be affected in regions where ventricles are dilated.

  20. Biochemical properties of Na+/K(+)-ATPase in axonal growth cone particles isolated from fetal rat brain.

    PubMed

    Mercado, R; Hernández, J

    1994-08-01

    Axonal growth cones (AGC) isolated from fetal rat brain have an important specific activity of N+/K(+)-ATPase. Kinetic assays of the enzyme in AGC showed that Km values for ATP or K+ are similar to those reported for the adult brain enzyme. For Na+ the affinity (Km) was lower. Vmax for the three substrates was several times lower in AGC as compared to the adult value. We also observed two apparent inhibition constants of Na+/K(+)-ATPase by ouabain, one of low affinity, possibly corresponding to the alpha 1 isoform and another of high affinity which is different to that described for the alpha 2 isoform of the enzyme. These results support an important role for the sodium pump in the maintainance of volume and cationic balance in neuronal differentiating structures. The functional differences observed also suggest that the enzymatic complex of Na+/K(+)-ATPase in AGC is in a transitional state towards the adult configuration.

  1. Gene Expression Analysis of Neurons and Astrocytes Isolated by Laser Capture Microdissection from Frozen Human Brain Tissues

    PubMed Central

    Tagliafierro, Lidia; Bonawitz, Kirsten; Glenn, Omolara C.; Chiba-Falek, Ornit

    2016-01-01

    Different cell types and multiple cellular connections characterize the human brain. Gene expression analysis using a specific population of cells is more accurate than conducting analysis of the whole tissue homogenate, particularly in the context of neurodegenerative diseases, where a specific subset of cells is affected by the different pathology. Due to the difficulty of obtaining homogenous cell populations, gene expression in specific cell-types (neurons, astrocytes, etc.) has been understudied. To leverage the use of archive resources of frozen human brains in studies of neurodegenerative diseases, we developed and calibrated a method to quantify cell-type specific—neuronal, astrocytes—expression profiles of genes implicated in neurodegenerative diseases, including Parkinson's and Alzheimer's diseases. Archive human frozen brain tissues were used to prepare slides for rapid immunostaining using cell-specific antibodies. The immunoreactive-cells were isolated by Laser Capture Microdissection (LCM). The enrichment for a particular cell-type of interest was validated in post-analysis stage by the expression of cell-specific markers. We optimized the technique to preserve the RNA integrity, so that the RNA was suitable for downstream expression analyses. Following RNA extraction, the expression levels were determined digitally using nCounter Single Cell Gene Expression assay (NanoString Technologies®). The results demonstrated that using our optimized technique we successfully isolated single neurons and astrocytes from human frozen brain tissues and obtained RNA of a good quality that was suitable for mRNA expression analysis. We present here new advancements compared to previous reported methods, which improve the method's feasibility and its applicability for a variety of downstream molecular analyses. Our new developed method can be implemented in genetic and functional genomic research of neurodegenerative diseases and has the potential to

  2. Cerebral capillary velocimetry based on temporal OCT speckle contrast

    PubMed Central

    Choi, Woo June; Li, Yuandong; Qin, Wan; Wang, Ruikang K.

    2016-01-01

    We propose a new optical coherence tomography (OCT) based method to measure red blood cell (RBC) velocities of single capillaries in the cortex of rodent brain. This OCT capillary velocimetry exploits quantitative laser speckle contrast analysis to estimate speckle decorrelation rate from the measured temporal OCT speckle signals, which is related to microcirculatory flow velocity. We hypothesize that OCT signal due to sub-surface capillary flow can be treated as the speckle signal in the single scattering regime and thus its time scale of speckle fluctuations can be subjected to single scattering laser speckle contrast analysis to derive characteristic decorrelation time. To validate this hypothesis, OCT measurements are conducted on a single capillary flow phantom operating at preset velocities, in which M-mode B-frames are acquired using a high-speed OCT system. Analysis is then performed on the time-varying OCT signals extracted at the capillary flow, exhibiting a typical inverse relationship between the estimated decorrelation time and absolute RBC velocity, which is then used to deduce the capillary velocities. We apply the method to in vivo measurements of mouse brain, demonstrating that the proposed approach provides additional useful information in the quantitative assessment of capillary hemodynamics, complementary to that of OCT angiography. PMID:28018711

  3. Complete genome sequences of two bovine viral diarrhea viruses isolated from brain tissues of nonambulatory (downer) cattle.

    PubMed

    Oem, Jae-Ku; Joo, Soo-Kyung; An, Dong-Jun

    2013-09-12

    Here, we report the complete genome sequences of two bovine viral diarrhea viruses (BVDVs) (strains 11F011 and 12F004) isolated from brain tissues from nonambulatory (downer) cattle. The complete genomes of strains 11F011 and 12F004 contain 12,287 nucleotides (nt) with a single large open reading frame and 12,301 nt with a single large open reading frame, respectively. Phylogenetic analysis indicated that these strains belong to the BVDV-2a and -1b genotypes, respectively.

  4. Isolation of primary microglia from the human post-mortem brain: effects of ante- and post-mortem variables.

    PubMed

    Mizee, Mark R; Miedema, Suzanne S M; van der Poel, Marlijn; Adelia; Schuurman, Karianne G; van Strien, Miriam E; Melief, Jeroen; Smolders, Joost; Hendrickx, Debbie A; Heutinck, Kirstin M; Hamann, Jörg; Huitinga, Inge

    2017-02-17

    Microglia are key players in the central nervous system in health and disease. Much pioneering research on microglia function has been carried out in vivo with the use of genetic animal models. However, to fully understand the role of microglia in neurological and psychiatric disorders, it is crucial to study primary human microglia from brain donors. We have developed a rapid procedure for the isolation of pure human microglia from autopsy tissue using density gradient centrifugation followed by CD11b-specific cell selection. The protocol can be completed in 4 h, with an average yield of 450,000 and 145,000 viable cells per gram of white and grey matter tissue respectively. This method allows for the immediate phenotyping of microglia in relation to brain donor clinical variables, and shows the microglia population to be distinguishable from autologous choroid plexus macrophages. This protocol has been applied to samples from over 100 brain donors from the Netherlands Brain Bank, providing a robust dataset to analyze the effects of age, post-mortem delay, brain acidity, and neurological diagnosis on microglia yield and phenotype. Our data show that cerebrospinal fluid pH is positively correlated to microglial cell yield, but donor age and post-mortem delay do not negatively affect viable microglia yield. Analysis of CD45 and CD11b expression showed that changes in microglia phenotype can be attributed to a neurological diagnosis, and are not influenced by variation in ante- and post-mortem parameters. Cryogenic storage of primary microglia was shown to be possible, albeit with variable levels of recovery and effects on phenotype and RNA quality. Microglial gene expression substantially changed due to culture, including the loss of the microglia-specific markers, showing the importance of immediate microglia phenotyping. We conclude that primary microglia can be isolated effectively and rapidly from human post-mortem brain tissue, allowing for the study of the

  5. Calcium-activated potassium channels in isolated presynaptic nerve terminals from rat brain.

    PubMed Central

    Bartschat, D K; Blaustein, M P

    1985-01-01

    86Rb efflux was examined in isolated presynaptic nerve terminals (synaptosomes) from rat brain in a study designed to assess K permeability (PK) changes sensitive to alterations in internal Ca activity. Rb efflux from 86Rb-loaded synaptosomes into nominally Ca-free physiological saline (PSS) containing 5 mM-K was about 0.3-0.4%/s. Raising extracellular K concentration [( K]o), to depolarize the synaptosomes, stimulated the 86Rb efflux. Addition of Ca to the 5 mM-K PSS had no effect, but Ca did further stimulate 86Rb efflux into K-rich solutions. The effect of Ca was graded, with apparent half-maximal activation, KA approximately equal to 0.5 mM-Ca. These data fit the view that, during depolarization, Ca enters the terminals through voltage-regulated Ca channels, and that the rise in intracellular Ca concentration opens certain (Ca-activated) K channels. The Ca-dependent stimulation of 86Rb efflux was greatest during the initial seconds of incubation (component CT), and then declined to a much lower rate (component CS). Much of this change in rate could be attributed to inactivation of voltage-regulated Ca channels and reduced entry of Ca. The Ca-dependent increase in 86Rb efflux was completely inhibited by 100 microM-La. In the presence of Ca, but not in its absence, the Ca ionophore A23187 stimulated 86Rb efflux both in 5 and 100 mM-K PSS. The effect in 100 mM-K was quantitatively greater, perhaps because of the increased outward driving force on Rb in depolarized synaptosomes. When synaptosomes were suspended in media containing the voltage-sensitive fluorescent dye, DiS-C3-(5) (1,1'-dipentyl-2,2'-thiocarbocyanine), the addition of Ca+ A23187 decreased the fluorescence intensity (= synaptosome hyperpolarization) when the media contained 5 mM-K but not 100 mM-K. This implies that in the presence of Ca + A23187, PK was increased, and the membrane potential moved closer to the K equilibrium potential, EK. Quinine sulphate, a blocker of Ca-activated K channels

  6. Presence of hormonally-sensitive adenylate cyclase receptors in capillary-enriched fractions from rat cerebral cortex.

    PubMed

    Baca, G M; Palmer, G C

    1978-01-01

    The 10 000 g particulate fraction from capillary-enriched fractions isolated from rat cerebral cortex was shown to possess an adenylate cyclase highly sensitive to activation by sodium fluoride, norepinephrine, epinephrine, isoproterenol and dopamine. To a lesser extent histamine and three dopamine agonists, namely M-7 (5,6-dihydroxy-2-dimethylamino tetralin), ET-495 (methane sulfonate of pyribedil), and S-584 (metabolite of pyribedil) stimulated the enzyme preparation. The action of norepinephrine was blocked by propanolol while phenotolamine and haloperidol were relatively ineffective except at highest concentrations. Phentolamine and propanolol at only highest concentrations (10(-4) M) antagonized the action of dopamine. Haloperidol was seen to be a potent inhibitor of either dopamine- or dopamine agonist-sensitive adenylate cyclase. No effects on the enzyme were observed with methoxamine, octopamine or serotonin. These preliminary data suggest the presence of a mixed population of receptors for adenylate cyclase in rat brain capillaries.

  7. Structural changes in Alzheimer's disease brain microvessels.

    PubMed

    Christov, Alexander; Ottman, J; Hamdheydari, L; Grammas, Paula

    2008-08-01

    Brain microvascular alterations are thought to contribute to the development of stroke and dementia. Structural changes in capillaries of elderly patients correlate positively with advanced age and dementia. The objective of this study is to use laser-induced fluorescence spectroscopy to compare structural (collagen content) and functional (apoptosis) parameters in brain tissues and isolated vessels of AD patients to age-matched controls. Our results show significantly higher fluorescent labeling for apoptosis in AD vessels compared to controls. Also, there is significantly higher autofluorescence (reflecting levels of collagen and other proteins that autofluoresce) in AD brain and vessels compared to controls. Western blot analysis of collagen subtypes shows elevated type I and type III and reduced type IV levels in AD vessels. These data demonstrate that changes in the amount and type of collagen occur in AD brain and suggest that cerebral vessel injury is part of AD pathology.

  8. Gas-Filled Capillary Model

    NASA Astrophysics Data System (ADS)

    Steinhauer, L. C.; Kimura, W. D.

    2006-11-01

    We have developed a 1-D, quasi-steady-state numerical model for a gas-filled capillary discharge that is designed to aid in selecting the optimum capillary radius in order to guide a laser beam with the required intensity through the capillary. The model also includes the option for an external solenoid B-field around the capillary, which increases the depth of the parabolic density channel in the capillary, thereby allowing for propagation of smaller laser beam waists. The model has been used to select the parameters for gas-filled capillaries to be utilized during the Staged Electron Laser Acceleration — Laser Wakefield (STELLA-LW) experiment.

  9. Isolation from bovine brain of substances inhibiting specific binding of imipramine and serotonin uptake

    SciTech Connect

    Mukhin, A.G.; Kladnitskii, A.V.; Kovaleva, E.S.; Kudryakova, T.B.

    1986-01-01

    The authors search for endogenous ligands of the ''imipramine receptor'' in brain tissue. Binding of tritium-imipramine with the fraction of unpruified bovine brain synaptic membranes was carried out by the method of Raisman et'al. Uptake of tritium-serotonin by synaptosomes of rat cerebral cortex was estimated. The results do not give a final anser to the question of the existence of an endogenous ligand of the ''imipramine receptor'' but they can serve as the basis for research aimed at purifying the active fractions already obtained and identifying the compounds containined in them.

  10. [Role of small-dose recombinant human coagulation factor VIIa for coagulopathy in patients with isolated traumatic brain injury].

    PubMed

    Pei, Bing-bing; Li, Chao-yue; Lu, Xin; Wu, Xing; Gao, Liang; Yu, Jian; Wu, Xue-hai; Jin, Yi; Sun, Yi-rui; Du, Zhuo-ying; Mao, Ying; Hu, Jin; Zhou, Liang-fu

    2013-06-18

    To explore the role of small-dose recombinant human coagulation factor VIIa (rFVIIa) for coagulopathy in patients with isolated traumatic brain injury. A total of 86 isolated traumatic brain patients with coagulopathy were treated at our neurosurgery intensive care unit (NICU) from January 2010 to December 2012. Their trauma registry data included mortality, pre-and post-rFVIIa coagulation parameters. Two-tailed paired t-test was used to determine significant changes in coagulation parameters and other major clinical parameters. Twenty-seven patients made up the low-dose rFVIIa (20 µg/kg) group. And the control group had 59 well-matched subjects. At admission, age, blood pressure, Glasgow coma scale score, hemoglobin, platelets and international normalize ratio were similar in both groups. After treatment, the INR of patients on rFVIIa was lower than that of the conventional treatment group (1.1 ± 0.2 vs 1.2 ± 0.2, P < 0.01) and it declined more in the rFVIIa group (0.3 ± 0.2 vs 0.1 ± 0.4, P = 0.05). No significant difference existed in mortality or length of stay between two groups.There was no occurrence of subsequent thromboembolic events. The application of small-dose rFVIIa can effectively reduce the value of INR and improve the coagulation status of patients. During the course of treatment, no major adverse events occur.

  11. COMPARATIVE STUDIES ON MITOCHONDRIA ISOLATED FROM NEURON-ENRICHED AND GLIA-ENRICHED FRACTIONS OF RABBIT AND BEEF BRAIN

    PubMed Central

    Hamberger, Anders; Blomstrand, Christian; Lehninger, Albert L.

    1970-01-01

    Fractions enriched in neuronal and glial cells were obtained from dispersions of whole beef brain and rabbit cerebral cortex by large-scale density gradient centrifugation procedures. The fractions were characterized by appropriate microscopic observation. Mitochondria were then isolated from these fractions by differential centrifugation of their homogenates. The two different types of mitochondria were characterized with respect to certain enzyme activities, respiratory rate, rate of protein synthesis, and their buoyant density in sucrose gradients. The mitochondria from the neuron-enriched fraction were distinguished by a higher rate of incorporation of amino acids into protein, higher cytochrome oxidase activity, and a higher buoyant density in sucrose density gradients. Mitochondria from the glia-enriched fraction showed relatively high monoamine oxidase and Na+- and K+-stimulated ATPase activities. The rates of oxidation of various substrates and the acceptor control ratios did not differ appreciably between the two types of mitochondria. The difference in the buoyant density of mitochondria isolated from the neuron-enriched and glia-enriched cell fractions was utilized in attempts to separate neuronal and glial mitochondria from the mixed mitochondria obtained from whole brain homogenates in shallow sucrose gradients. The appearance of two peaks of cytochrome oxidase, monoamine oxidase, and protein concentration in such gradients shows the potential feasibility of such an approach. PMID:5513605

  12. Variation in vasotocin immunoreactivity in the brain of recently isolated populations of a death valley pupfish, Cyprinodon nevadensis.

    PubMed

    Lema, Sean C; Nevitt, Gabrielle A

    2004-02-01

    Pupfishes in the Death Valley region of California and Nevada comprise a monophyletic group of populations that became isolated in remote streams and springs over the past 20,000 years. These aquatic habitats show considerable ecological diversity, and allopatric populations have evolved differences in morphology and behavior. Here we investigated whether the divergence of pupfish populations in Death Valley might be associated with changes in arginine vasotocin (AVT). We used immunocytochemistry to compare the expression of AVT in the brain of Amargosa pupfish (Cyprinodon nevadensis) from two Death Valley populations: (1) the Amargosa River-a highly variable desert stream containing the Cyprinodon nevadensis amargosae subspecies, and (2) Big Spring-a comparatively stable springhead and outflow inhabited by Cyprinodon nevadensis mionectes. These particular populations have been isolated from each other for only 400-4000 years. In both populations AVT-immunoreactive somata localized to parvocellular and magnocellular neurons in the preoptic area, with AVT-immunoreactive fibers extending ventrolaterally to innervate the pituitary. We found that both parvocellular and magnocellular AVT-immunoreactive neurons were significantly larger in males and females from the Amargosa River population than in same sex pupfish from Big Spring. Our findings suggest that the divergent ecological conditions of these two habitats have brought about changes in AVT pathways in the brain.

  13. Comparative studies on mitochondria isolated from neuron-enriched and glia-enriched fractions of rabbit and beef brain.

    PubMed

    Hamberger, A; Blomstrand, C; Lehninger, A L

    1970-05-01

    Fractions enriched in neuronal and glial cells were obtained from dispersions of whole beef brain and rabbit cerebral cortex by large-scale density gradient centrifugation procedures. The fractions were characterized by appropriate microscopic observation. Mitochondria were then isolated from these fractions by differential centrifugation of their homogenates. The two different types of mitochondria were characterized with respect to certain enzyme activities, respiratory rate, rate of protein synthesis, and their buoyant density in sucrose gradients. The mitochondria from the neuron-enriched fraction were distinguished by a higher rate of incorporation of amino acids into protein, higher cytochrome oxidase activity, and a higher buoyant density in sucrose density gradients. Mitochondria from the glia-enriched fraction showed relatively high monoamine oxidase and Na(+)- and K(+)-stimulated ATPase activities. The rates of oxidation of various substrates and the acceptor control ratios did not differ appreciably between the two types of mitochondria. The difference in the buoyant density of mitochondria isolated from the neuron-enriched and glia-enriched cell fractions was utilized in attempts to separate neuronal and glial mitochondria from the mixed mitochondria obtained from whole brain homogenates in shallow sucrose gradients. The appearance of two peaks of cytochrome oxidase, monoamine oxidase, and protein concentration in such gradients shows the potential feasibility of such an approach.

  14. Derivatization in Capillary Electrophoresis.

    PubMed

    Marina, M Luisa; Castro-Puyana, María

    2016-01-01

    Capillary electrophoresis is a well-established separation technique in analytical research laboratories worldwide. Its interesting advantages make CE an efficient and potent alternative to other chromatographic techniques. However, it is also recognized that its main drawback is the relatively poor sensitivity when using optical detection. One way to overcome this limitation is to perform a derivatization reaction which is intended to provide the analyte more suitable analytical characteristics enabling a high sensitive detection. Based on the analytical step where the CE derivatization takes place, it can be classified as precapillary (before separation), in-capillary (during separation), or postcapillary (after separation). This chapter describes the application of four different derivatization protocols (in-capillary and precapillary modes) to carry out the achiral and chiral analysis of different compounds in food and biological samples with three different detection modes (UV, LIF, and MS).

  15. Capillary condenser/evaporator

    NASA Technical Reports Server (NTRS)

    Valenzuela, Javier A. (Inventor)

    2010-01-01

    A heat transfer device is disclosed for transferring heat to or from a fluid that is undergoing a phase change. The heat transfer device includes a liquid-vapor manifold in fluid communication with a capillary structure thermally connected to a heat transfer interface, all of which are disposed in a housing to contain the vapor. The liquid-vapor manifold transports liquid in a first direction and conducts vapor in a second, opposite direction. The manifold provides a distributed supply of fluid (vapor or liquid) over the surface of the capillary structure. In one embodiment, the manifold has a fractal structure including one or more layers, each layer having one or more conduits for transporting liquid and one or more openings for conducting vapor. Adjacent layers have an increasing number of openings with decreasing area, and an increasing number of conduits with decreasing cross-sectional area, moving in a direction toward the capillary structure.

  16. Activity of factor VII in patients with isolated blunt traumatic brain injury: association with coagulopathy and progressive hemorrhagic injury.

    PubMed

    Wu, Xing; Du, Zhuoying; Yu, Jian; Sun, Yirui; Pei, Bingbing; Lu, Xin; Tang, Zhengyu; Yin, Mao; Zhou, Liangfu; Hu, Jin

    2014-01-01

    Given the importance of factor VII (FVII) in extrinsic pathway of coagulation cascade, we sought to elucidate the relationship between FVII and traumatic brain injury-induced coagulopathy and progressive hemorrhagic injury (PHI). Eighty-one patients with isolated traumatic brain injury, 16 years or older, were recruited between 2010 and 2012. Blood was collected on arrival in the emergency department and analyzed with activated partial thromboplastic time, international normalized ratio, platelet count, and activity of FVII. Coagulopathy was defined as thrombocytopenia (platelet count < 120,000/μL) or elevated international normalized ratio of greater than 1.2 or prolonged activated partial thromboplastin time greater than 40 seconds at admission. PHI was present when the follow-up computed tomographic scan reported any increase in size or number of the hemorrhagic lesions. Logistic regression examined the risks for coagulopathy and PHI. Mean (SD) FVII activity in patients with coagulopathy was 85.69% (34.88%), which was significantly lower than patients without coagulopathy (99.57% [29.37%], p = 0.04). Isolated traumatic brain injury patients with FVII activity less than 77.5% have an odds ratio for coagulopathy of 5.52 (95% confidence interval, 1.82-16.68; p = 0.03) relative to patients with FVII activity of 77.5% or greater. Mean (SD) FVII activity in patients with PHI was 70.76% (18.21%), which was significantly lower than patients without PHI (105.76% [32.27%], p < 0.001). A stepwise logistic regression analysis identified FVII less than 77.5% (odds ratio, 4.53; 95% confidence interval, 1.62-12.67; p = 0.004) as a predisposing risk factors independently associated with the presence of PHI. The overall mortality rate in the study population was 7.4% (n = 6). The plasma FVII in death patients (91.44% [47.19%]) was slightly lower than that in survival patients (92.01% [32.04%]). However, there was no statistical difference between the two groups (p = 0.95). A

  17. Capillary Structures for Exploration Life Support (Capillary Structures)

    NASA Image and Video Library

    2017-07-10

    iss052e013146 (July 10, 2017) --- Astronaut Jack Fischer is photographed during setup of hardware for the Capillary Structures for Exploration Life Support (Capillary Structures) two sorbent demonstrations. The Capillary Structures for Exploration Life Support (Capillary Structures) investigation studies a new method using structures of specific shapes to manage fluid and gas mixtures. The investigation studies water recycling and carbon dioxide removal, benefiting future efforts to design lightweight, more reliable life support systems for future space missions.

  18. Behavioral tests and oxidative stress evaluation in mitochondria isolated from the brain and liver of mice treated with riparin A.

    PubMed

    Nunes, Geandra Batista Lima; Costa, Luciana Muratori; Gutierrez, Stanley Juan Chávez; Satyal, Prabodh; de Freitas, Rivelilson Mendes

    2015-01-15

    This study aimed at evaluating the oxidative stress in mitochondria isolated from the brain and liver of mice treated with riparin A, as well as the locomotor activity and myorelaxant effect of this compound. Behavioral models of rota rod and open field tests were used for locomotor activity and myorelaxant effect evaluation. The animals were divided into five groups (n=8), which were treated with: diazepam (1mg/kg, i.p), riparin A (5, 10, and 20mg/kg, o.r.) or vehicle (0.9% saline, o.r.). The oxidative stress evaluation was carried out in mitochondria isolated from the brain and liver of mice from five experimental groups (n=8), which were treated with: ascorbic acid (250 mg/kg; positive control), vehicle (0.9% saline; negative control) and riparin A (5, 10 and 20mg/kg). In an open field and rota rod test a significant difference in the number of crossings, in time of permanence on the swivel bar and in the number of falls in riparin A treated animals (5, 10 and 20mg/kg) was not observed, when compared with negative control (vehicle) (p>0.05). In comparison to the negative control, there was a reduction of lipid peroxidation levels and nitrite content in mice treated with riparin A (p<0.05). Reduced glutathione (GSH) levels (p<0.05), superoxide dismutase (SOD) and catalase activities increased in the brain (rip A 5mg/kg; p<0.05), while in the liver SOD remained unchanged (p>0.05) and catalase activity (p<0.05) was reduced. Riparin A was presented as a bioactive molecule devoid of adverse effects of alteration of motor activity. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Prevalence of clinically important traumatic brain injuries in children with minor blunt head trauma and isolated severe injury mechanisms.

    PubMed

    Nigrovic, Lise E; Lee, Lois K; Hoyle, John; Stanley, Rachel M; Gorelick, Marc H; Miskin, Michelle; Atabaki, Shireen M; Dayan, Peter S; Holmes, James F; Kuppermann, Nathan

    2012-04-01

    To determine the prevalence of clinically important traumatic brain injuries (TBIs) with severe injury mechanisms in children with minor blunt head trauma but with no other risk factors from the Pediatric Emergency Care Applied Research Network (PECARN) TBI prediction rules (defined as isolated severe injury mechanisms). Secondary analysis of a large prospective observational cohort study. Twenty-five emergency departments participating in the PECARN. Children with minor blunt head trauma and Glasgow Coma Scale scores of at least 14. Treating clinicians completed a structured data form that included injury mechanism (severity categories defined a priori). Clinically important TBIs were defined as intracranial injuries resulting in death, neurosurgical intervention, intubation for more than 24 hours, or hospital admission for at least 2 nights. We investigated the rate of clinically important TBIs in children with either severe injury mechanisms or isolated severe injury mechanisms. Of the 42,412 patients enrolled in the overall study, 42,099 (99%) had injury mechanisms recorded, and their data were included for analysis. Of all study patients, 5869 (14%) had severe injury mechanisms, and 3302 (8%) had isolated severe injury mechanisms. Overall, 367 children had clinically important TBIs (0.9%; 95% CI, 0.8%-1.0%). Of the 1327 children younger than 2 years with isolated severe injury mechanisms, 4 (0.3%; 95% CI, 0.1%-0.8%) had clinically important TBIs, as did 12 of the 1975 children 2 years or older (0.6%; 95% CI, 0.3%-1.1%). Children with isolated severe injury mechanisms are at low risk of clinically important TBI, and many do not require emergent neuroimaging.

  20. Capillary action liquid chromatography.

    PubMed

    Zhang, Bo; Bergström, Edmund T; Goodall, David M; Myers, Peter

    2009-06-01

    Capillary action LC (caLC) is introduced as a technique using capillary action as the driving force to perform LC in capillary columns packed with HPLC type microparticulate materials. A dry packing method with centrifugal force was developed to prepare capillary columns in parallel (10 columns per 3 min) to support their disposable use in caLC. Using a digital microscope for real-time imaging and recording separations of components in a dye mixture, caLC was found to have flow characteristics similar to TLC. Based on the investigation of microparticulate HPLC silica gels of different size (1.5-10 microm) and a typical TLC grade irregular medium, Merck 60G silica, the van Deemter curves suggested molecular diffusion as the major contribution to band broadening in caLC. With Waters Xbridge 2.6 microm silica, plate heights down to 8.8 microm were obtained, comparable to those achievable in HPLC. Assisted by an image-processing method, the visual caLC separation was converted to a classical chromatogram for further data analysis and such a facility confirmed the observation of highly efficient bands.

  1. Capillary blood glucose monitoring.

    PubMed

    Wallymahmed, M

    This article, the first in a series of articles relating to clinical skills in nursing, outlines the procedure of capillary blood glucose monitoring. This is a convenient way of monitoring blood glucose patterns and can be a useful aid in guiding treatment changes in patients with type 1 and type 2 diabetes, especially during periods of illness or frequent hypoglycaemia.

  2. Depleted uranium induces disruption of energy homeostasis and oxidative stress in isolated rat brain mitochondria.

    PubMed

    Shaki, Fatemeh; Hosseini, Mir-Jamal; Ghazi-Khansari, Mahmoud; Pourahmad, Jalal

    2013-06-01

    Depleted uranium (DU) is emerging as an environmental pollutant primarily due to its military applications. Gulf War veterans with embedded DU showed cognitive disorders that suggest that the central nervous system is a target of DU. Recent evidence has suggested that DU could induce oxidative stress and mitochondrial dysfunction in brain tissue. However, the underlying mechanisms of DU toxicity in brain mitochondria are not yet well understood. Brain mitochondria were obtained using differential centrifugation and were incubated with different concentrations (50, 100 and 200 μM) of uranyl acetate (UA) as a soluble salt of U(238) for 1 h. In this research, mitochondrial ROS production, collapse of mitochondrial membrane potential and mitochondrial swelling were examined by flow cytometry following the addition of UA. Meanwhile, mitochondrial sources of ROS formation were determined using specific substrates and inhibitors. Complex II and IV activity and also the extent of lipid peroxidation and glutathione (GSH) oxidation were detected via spectroscopy. Furthermore, we investigated the concentration of ATP and ATP/ADP ratio using luciferase enzyme and cytochrome c release from mitochondria which was detected by ELISA kit. UA caused concentration-dependent elevation of succinate-linked mitochondrial ROS production, lipid peroxidation, GSH oxidation and inhibition of mitochondrial complex II. UA also induced mitochondrial permeability transition, ATP production decrease and increase in cytochrome c release. Pre-treatment with antioxidants significantly inhibited all the above mentioned toxic effects of UA. This study suggests that mitochondrial oxidative stress and impairment of oxidative phosphorylation in brain mitochondria may play a key role in DU neurotoxicity as reported in Gulf War Syndrome.

  3. Noise suppressing capillary separation system

    DOEpatents

    Yeung, Edward S.; Xue, Yongjun

    1996-07-30

    A noise-suppressing capillary separation system for detecting the real-time presence or concentration of an analyte in a sample is provided. The system contains a capillary separation means through which the analyte is moved, a coherent light source that generates a beam which is split into a reference beam and a sample beam that irradiate the capillary, and a detector for detecting the reference beam and the sample beam light that transmits through the capillary. The laser beam is of a wavelength effective to be absorbed by a chromophore in the capillary. The system includes a noise suppressing system to improve performance and accuracy without signal averaging or multiple scans.

  4. Isolation of NPY-25 (neuropeptide Y[12-36]), a potent inhibitor of calmodulin, from porcine brain.

    PubMed

    Kitamura, K; Kangawa, K; Tanaka, K; Matsuo, H

    1990-06-29

    In the present purification of low molecular weight fractions (Mr: 2000-4000) containing basic peptides, twenty nmol of novel calmodulin binding peptide, possessing a potent affinity for calmodulin, was isolated from 18 kg of porcine brain. By analysis with gas phase sequencer, the sequence was determined to be APAEDLARYYSALRHYINLITRQRY. Carboxy terminus of the peptide was determined to be Tyr-NH2. The peptide was a carboxy terminal pentacosanepeptide of neuropeptide Y and was termed NPY-25. NPY-25 competitively inhibited the activation of cAMP-phosphodiesterase through CaM binding in a Ca++ dependent fashion, but did not inhibit the basal activity of cAMP phosphodiesterase. NPY-25 elicited a more potent activity than did neuropeptide Y. IC50 values of NPY-25 and Neuropeptide Y were 0.06 microM and 0.54 microM respectively.

  5. Effect of Alkaloids Isolated from Phyllodium pulchellum on Monoamine Levels and Monoamine Oxidase Activity in Rat Brain

    PubMed Central

    Cai, Lu; Wang, Chao; Dong, Pei-pei; Zhang, Bao-jing; Zhang, Hou-Li; Huang, Shan-shan; Zhang, Bo; Yu, Sheng-ming; Zhong, Ming; Ma, Xiao-Chi

    2016-01-01

    Phyllodium pulchellum (P. pulchellum) is a folk medicine with a significant number of bioactivities. The aim of this study was to investigate the effects displayed by alkaloids fractions, isolated from the roots of P. pulchellum, on neurotransmitters monoamine levels and on monoamine oxidase (MAO) activity. Six alkaloids, which had indolealkylamine or β-carboline skeleton, were obtained by chromatographic technologies and identified by spectroscopic methods such as NMR and MS. After treatment with alkaloids of P. pulchellum, the reduction of DA levels (54.55%) and 5-HT levels (35.01%) in rat brain was observed by HPLC-FLD. The effect of alkaloids on the monoamines metabolism was mainly related to MAO inhibition, characterized by IC50 values of 37.35 ± 6.41 and 126.53 ± 5.39 μg/mL for MAO-A and MAO-B, respectively. The acute toxicity indicated that P. pulchellum extract was nontoxic. PMID:27195015

  6. Expression of Iron-Related Proteins at the Neurovascular Unit Supports Reduction and Reoxidation of Iron for Transport Through the Blood-Brain Barrier.

    PubMed

    Burkhart, Annette; Skjørringe, Tina; Johnsen, Kasper Bendix; Siupka, Piotr; Thomsen, Louiza Bohn; Nielsen, Morten Schallburg; Thomsen, Lars Lykke; Moos, Torben

    2016-12-01

    The mechanisms for iron transport through the blood-brain barrier (BBB) remain a controversy. We analyzed for expression of mRNA and proteins involved in oxidation and transport of iron in isolated brain capillaries from dietary normal, iron-deficient, and iron-reverted rats. The expression was also investigated in isolated rat brain endothelial cells (RBECs) and in immortalized rat brain endothelial (RBE4) cells grown as monoculture or in hanging culture inserts with defined BBB properties. Transferrin receptor 1, ferrireductases Steap 2 and 3, divalent metal transporter 1 (DMT1), ferroportin, soluble and glycosylphosphatidylinositol (GPI)-anchored ceruloplasmin, and hephaestin were all expressed in brain capillaries in vivo and in isolated RBECs and RBE4 cells. Gene expression of DMT1, ferroportin, and soluble and GPI-anchored ceruloplasmin were significantly higher in isolated RBECs with induced BBB properties. Primary pericytes and astrocytes both expressed ceruloplasmin and hephaestin, and RBECs, pericytes, and astrocytes all exhibited ferrous oxidase activity. The coherent protein expression of these genes was demonstrated by immunocytochemistry. The data show that brain endothelial cells provide the machinery for receptor-mediated uptake of ferric iron-containing transferrin. Ferric iron can then undergo reduction to ferrous iron by ferrireductases inside endosomes followed by DMT1-mediated pumping into the cytosol and subsequently cellular export by ferroportin. The expression of soluble ceruloplasmin by brain endothelial cells, pericytes, and astrocytes that together form the neurovascular unit (NVU) provides the ferroxidase activity necessary to reoxidize ferrous iron once released inside the brain.

  7. Regulated release of serotonin from axonal growth cones isolated from the fetal rat brain.

    PubMed

    Mercado, R; Floran, B; Hernandez, J

    1998-01-01

    In the present work we propose an hypothetical model related to a molecular recognizing system for serotonin in isolated growth cone particles. This model is supported by previous results from our laboratory plus new ones which show that growth cones release serotonin tonically and such release can be stimulated by potassium in a calcium-dependent manner. The present results, together with other author's data, suggest a physiological basis for the putative role of serotonin as a trophic factor during nervous system development.

  8. Genes that mediate breast cancer metastasis to the brain.

    PubMed

    Bos, Paula D; Zhang, Xiang H-F; Nadal, Cristina; Shu, Weiping; Gomis, Roger R; Nguyen, Don X; Minn, Andy J; van de Vijver, Marc J; Gerald, William L; Foekens, John A; Massagué, Joan

    2009-06-18

    The molecular basis for breast cancer metastasis to the brain is largely unknown. Brain relapse typically occurs years after the removal of a breast tumour, suggesting that disseminated cancer cells must acquire specialized functions to take over this organ. Here we show that breast cancer metastasis to the brain involves mediators of extravasation through non-fenestrated capillaries, complemented by specific enhancers of blood-brain barrier crossing and brain colonization. We isolated cells that preferentially infiltrate the brain from patients with advanced disease. Gene expression analysis of these cells and of clinical samples, coupled with functional analysis, identified the cyclooxygenase COX2 (also known as PTGS2), the epidermal growth factor receptor (EGFR) ligand HBEGF, and the alpha2,6-sialyltransferase ST6GALNAC5 as mediators of cancer cell passage through the blood-brain barrier. EGFR ligands and COX2 were previously linked to breast cancer infiltration of the lungs, but not the bones or liver, suggesting a sharing of these mediators in cerebral and pulmonary metastases. In contrast, ST6GALNAC5 specifically mediates brain metastasis. Normally restricted to the brain, the expression of ST6GALNAC5 in breast cancer cells enhances their adhesion to brain endothelial cells and their passage through the blood-brain barrier. This co-option of a brain sialyltransferase highlights the role of cell-surface glycosylation in organ-specific metastatic interactions.

  9. Isolation of synaptic junctional complexes of high structural integrity from rat brain

    PubMed Central

    1976-01-01

    A new method has been developed for isolating synaptic junctional complexes (SJC) of high structural integrity. The major step in the isolation involves homogenization of a synaptosomal membrane (SM) fraction in a biphasic system consisting of Freon 113 and an aqueous phase containing 0.2% Triton X-100. Well-preserved SJCs, along with membrane vesicles, were recovered in the aqueous phase after low-speed centrifugation of the homogenate. The membranes were subsequently separated from the SJCs by centrifugation on a discontinuous sucrose density gradient. The purity and identity of subcellular fractions were monitored by thin sectioning electron microscopy, using specific and nonspecific staining methods. From the electron microscope studies we conclude that SJCs and their components occupy about 65% of the area covered by structures in this fraction. The assay of enzyme activities indicates that homogenization in Triton-Freon and subsequent steps of the isolation procedure affect the activities of Na, K-ATPase, cytochrome oxidase, and acid phosphatase to different extents, but do not cause total inactivation. Electrophoresis of the SJC-enriched fraction on sodium dodecyl sulfate-polyacrylamide gels has demonstrated that a polypeptide which co-migrates with tubulin is the major component in this fraction, and that a polypeptide co-migrating with actin is also present. PMID:186464

  10. Effect of isolation conditions on brain regional enkephalin and beta-endorphin levels and vocalizations in 10-day-old rat pups.

    PubMed

    Shoemaker, W J; Kehoe, P

    1995-02-01

    Young rat pups were isolated from their dams under different conditions. The endogenous opioid peptides were measured in brain regions after isolation. Because there is no uptake mechanism for peptides released at the synapse and because released peptide is rapidly degraded enzymatically, decreases in peptide levels over this time course can be interpreted as release from terminals. No change was observed in either peptide in the hypothalamus, septum, or amygdala after isolation compared with controls. Significant decreases were seen in the midbrain after isolation. A comparison of peptide levels and ultrasonic vocalizations in the pups isolated in familiar, novel, or control conditions was also performed. Enkephalin levels in the midbrain were decreased in familiar and novel conditions, but in the brainstem opioid peptides were decreased only in the familiar condition. The greater involvement of the opioid peptides in the pups isolated in familiar conditions may contribute to the ability of naltrexone to block vocalization.

  11. Brain-derived neurotrophic factor controls functional differentiation and microcircuit formation of selectively isolated fast-spiking GABAergic interneurons.

    PubMed

    Berghuis, Paul; Dobszay, Marton B; Sousa, Kyle M; Schulte, Gunnar; Mager, Peter P; Härtig, Wolfgang; Görcs, Tamás J; Zilberter, Yuri; Ernfors, Patrik; Harkany, Tibor

    2004-09-01

    GABAergic interneurons with high-frequency firing, fast-spiking (FS) cells, form synapses on perisomatic regions of principal cells in the neocortex and hippocampus to control the excitability of cortical networks. Brain-derived neurotrophic factor (BDNF) is essential for the differentiation of multiple interneuron subtypes and the formation of their synaptic contacts. Here, we examined whether BDNF, alone or in conjunction with sustained KCl-induced depolarization, drives functional FS cell differentiation and the formation of inhibitory microcircuits. Homogeneous FS cell cultures were established by target-specific isolation using the voltage-gated potassium channel 3.1b subunit as the selection marker. Isolated FS cells expressed parvalbumin, were surrounded by perineuronal nets, formed immature inhibitory connections and generated slow action potentials at 12 days in vitro. Brain-derived neurotrophic factor (BDNF) promoted FS cell differentiation by increasing the somatic diameter, dendritic branching and the frequency of action potential firing. In addition, BDNF treatment led to a significant up-regulation of synaptophysin and vesicular GABA transporter expression, components of the synaptic machinery critical for GABA release, which was paralleled by an increase in synaptic strength. Long-term membrane depolarization alone was detrimental to dendritic branching. However, we observed that BDNF and KCl exerted additive effects, as reflected by the significantly accelerated maturation of synaptic contacts and high discharge frequencies, and was required for the formation of reciprocal connections between FS cells. Our results show that BDNF, along with membrane depolarization, is critical for FS cells to establish inhibitory circuitries during corticogenesis.

  12. Biapigenin modulates the activity of the adenine nucleotide translocator in isolated rat brain mitochondria.

    PubMed

    Silva, Bruno A; Oliveira, Paulo J; Cristóvão, Armando; Dias, Alberto C P; Malva, João O

    2010-01-01

    In this study, we investigated the effects of biapigenin, a biflavone present in the extracts of Hypericum perforatum, in rat brain mitochondrial bioenergetics and calcium homeostasis. We found that biapigenin significantly decreased adenosine diphosphate (ADP)-induced membrane depolarization and increased repolarization (by 68 and 37%, respectively). These effects were blocked by atractyloside and bongkrekic acid, but not oligomycin. In the presence of biapigenin, an ADP-stimulated state 3 respiration was still noticeable, which did not happen in the presence of adenine nucleotide translocator (ANT) inhibitors. Taking in consideration the relevance of the ANT in the modulation of the mitochondrial permeability transition pore (mPTP), mitochondrial calcium homeostasis was evaluated alone or in the presence of biapigenin. We found that biapigenin reduces mitochondrial calcium retention by increasing calcium efflux, an effect that was blocked by ADP plus oligomycin, an efficient blocker of the mPTP in brain mitochondria. Taken together, the results in this article suggest that biapigenin modulates mPTP opening, possibly by modulating ANT function, contributing for enhanced mitochondrial calcium efflux, thereby reducing calcium burden and contributing for neuroprotection against excitotoxicity.

  13. Isolation of pseudoperoxidase-positive astrocyte granules from intact rat brain and cysteamine-treated neuroglial cultures.

    PubMed

    Cissé, S; Schipper, H M

    1993-06-25

    A subpopulation of astrocytes in periventricular regions of aging brain and in cysteamine (CSH)-treated glial cultures contain autofluorescent cytoplasmic granules that exhibit an affinity for Gomori's chrome alum hematoxylin (CAH), and non-enzymatic peroxidase activity. Although shown to be histochemically distinct from lipofuscin, the lack of pure preparations of these glial inclusions has hindered the elucidation of their precise chemical constituents. Using sucrose gradient fractionation and density centrifugation on percoll, we obtained enriched preparations of astrocyte cytoplasmic granules from intact rat brain and CSH-treated astrocyte cultures. The presence and relative purity of these inclusions were confirmed by laser scanning confocal microscopy for red autofluorescent granules, diaminobenzidine histochemistry for non-enzymatic peroxidase activity and chrome alum hematoxylin (Gomori) staining. In the enriched fractions, the smaller granules (0.5-4.0 microns) were spherical and weakly autofluorescent, whereas larger inclusions (5.0-10.0 microns) tended to be intensely autofluorescent and pleomorphic. As in situ, the purified material was argyrophilic and did not stain for lipids. Isolation of these astrocytic inclusions should permit a more thorough characterization of their biochemical contents.

  14. The clinical significance of isolated traumatic subarachnoid hemorrhage in mild traumatic brain injury: A meta-analysis.

    PubMed

    Nassiri, Farshad; Badhiwala, Jetan H; Witiw, Christopher D; Mansouri, Alireza; Davidson, Benjamin; Almenawer, Saleh A; Lipsman, Nir; Da Costa, Leodante; Pirouzmand, Farhad; Nathens, Avery B

    2017-10-01

    The rates of clinical and radiographic progression and the need for neurosurgical intervention in patients with isolated traumatic subarachnoid hemorrhage (itSAH) after mild traumatic brain injury (Glasgow Coma Scale score, 13-15) has not been well established.The aim of this work was to review the evidence regarding patient outcomes after mild traumatic brain injury with itSAH. Two authors independently extracted the data according to a predefined protocol. The proportions of patients who had the outcomes of interest were pooled using random-effects model. The quality of included studies was assessed using the methodological index for nonrandomized studies scale. Thirteen studies reporting on 15,327 patients met inclusion criteria, and outcomes were pooled where available. No patient required neurosurgical intervention on presentation to hospital, and the incidence of need for eventual neurosurgical intervention was 0.0017% (95% confidence interval [CI], 0-0.39%). Moreover, the pooled analyses for all patients with available data showed an incidence of 5.76% (95% CI, 1.18-12.94%) for radiographic progression, 0.75% (95% CI, 0-2.39%) for neurologic deterioration, and 0.60% (95% CI, 0.09-1.41%) for mortality. Only one patient had died due to neurological injury. These patients experience very low rates of radiographic progression and neurologic deterioration and rarely require neurosurgical intervention or die due to neurological injury. Meta-analysis, level III.

  15. Monitoring of the velocity of high-affinity glutamate uptake by isolated brain nerve terminals using amperometric glutamate biosensor.

    PubMed

    Soldatkin, O; Nazarova, A; Krisanova, N; Borуsov, A; Kucherenko, D; Kucherenko, I; Pozdnyakova, N; Soldatkin, A; Borisova, T

    2015-04-01

    Glutamate is the major excitatory neurotransmitter in the central nervous system, which is involved in the main aspects of normal brain functioning. High-affinity Na(+)-dependent glutamate transporters is key proteins, which transport extracellular glutamate to the cytoplasm of nerve cells, thereby preventing continuous activation of glutamate receptors, and thus the development of neurotoxicity. Disturbance in glutamate uptake is involved in the pathogenesis of major neurological disorders. Amperometric biosensors are the most promising and successful among electrochemical biosensors. In this study, we developed (1) amperometric glutamate biosensor, (2) methodological approach for the analysis of glutamate uptake in liquid samples of isolated rat brain nerve terminals (synaptosomes). The basal level of glutamate, the initial velocity of glutamate uptake and time-dependent accumulation of glutamate by synaptosomes were determined using developed glutamate biosensor. Comparative analysis of the data with those obtained by radioactive analysis, spectrofluorimetry and ion exchange chromatography was performed. Therefore, the methodological approach for monitoring of the velocity of glutamate uptake, which takes into consideration the definite level of endogenous glutamate in nerve terminals, was developed using glutamate biosensor. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Ca(2+) handling in isolated brain mitochondria and cultured neurons derived from the YAC128 mouse model of Huntington's disease.

    PubMed

    Pellman, Jessica J; Hamilton, James; Brustovetsky, Tatiana; Brustovetsky, Nickolay

    2015-08-01

    We investigated Ca(2+) handling in isolated brain synaptic and non-synaptic mitochondria and in cultured striatal neurons from the YAC128 mouse model of Huntington's disease. Both synaptic and non-synaptic mitochondria from 2- and 12-month-old YAC128 mice had larger Ca(2+) uptake capacity than mitochondria from YAC18 and wild-type FVB/NJ mice. Synaptic mitochondria from 12-month-old YAC128 mice had further augmented Ca(2+) capacity compared with mitochondria from 2-month-old YAC128 mice and age-matched YAC18 and FVB/NJ mice. This increase in Ca(2+) uptake capacity correlated with an increase in the amount of mutant huntingtin protein (mHtt) associated with mitochondria from 12-month-old YAC128 mice. We speculate that this may happen because of mHtt-mediated sequestration of free fatty acids thereby increasing resistance of mitochondria to Ca(2+)-induced damage. In experiments with striatal neurons from YAC128 and FVB/NJ mice, brief exposure to 25 or 100 μM glutamate produced transient elevations in cytosolic Ca(2+) followed by recovery to near resting levels. Following recovery of cytosolic Ca(2+), mitochondrial depolarization with FCCP produced comparable elevations in cytosolic Ca(2+), suggesting similar Ca(2+) release and, consequently, Ca(2+) loads in neuronal mitochondria from YAC128 and FVB/NJ mice. Together, our data argue against a detrimental effect of mHtt on Ca(2+) handling in brain mitochondria of YAC128 mice. We demonstrate that mutant huntingtin (mHtt) binds to brain synaptic and nonsynaptic mitochondria and the amount of mitochondria-bound mHtt correlates with increased mitochondrial Ca(2+) uptake capacity. We propose that this may happen due to mHtt-mediated sequestration of free fatty acids thereby increasing resistance of mitochondria to Ca(2+)-induced damage. © 2015 International Society for Neurochemistry.

  17. Capillary Contact Angle in a Completely Wet Groove

    NASA Astrophysics Data System (ADS)

    Parry, A. O.; Malijevský, A.; Rascón, C.

    2014-10-01

    We consider the phase equilibria of a fluid confined in a deep capillary groove of width L with identical side walls and a bottom made of a different material. All walls are completely wet by the liquid. Using density functional theory and interfacial models, we show that the meniscus separating liquid and gas phases at two phase capillary coexistence meets the bottom capped end of the groove at a capillary contact angle θcap(L) which depends on the difference between the Hamaker constants. If the bottom wall has a weaker wall-fluid attraction than the side walls, then θcap>0 even though all the isolated walls are themselves completely wet. This alters the capillary condensation transition which is now first order; this would be continuous in a capped capillary made wholly of either type of material. We show that the capillary contact angle θcap(L) vanishes in two limits, corresponding to different capillary wetting transitions. These occur as the width (i) becomes macroscopically large, and (ii) is reduced to a microscopic value determined by the difference in Hamaker constants. This second wetting transition is characterized by large scale fluctuations and essential critical singularities arising from marginal interfacial interactions.

  18. Capillary contact angle in a completely wet groove.

    PubMed

    Parry, A O; Malijevský, A; Rascón, C

    2014-10-03

    We consider the phase equilibria of a fluid confined in a deep capillary groove of width L with identical side walls and a bottom made of a different material. All walls are completely wet by the liquid. Using density functional theory and interfacial models, we show that the meniscus separating liquid and gas phases at two phase capillary coexistence meets the bottom capped end of the groove at a capillary contact angle θ(cap)(L) which depends on the difference between the Hamaker constants. If the bottom wall has a weaker wall-fluid attraction than the side walls, then θ(cap) > 0 even though all the isolated walls are themselves completely wet. This alters the capillary condensation transition which is now first order; this would be continuous in a capped capillary made wholly of either type of material. We show that the capillary contact angle θ(cap)(L) vanishes in two limits, corresponding to different capillary wetting transitions. These occur as the width (i) becomes macroscopically large, and (ii) is reduced to a microscopic value determined by the difference in Hamaker constants. This second wetting transition is characterized by large scale fluctuations and essential critical singularities arising from marginal interfacial interactions.

  19. Capillary Isoelectric Focusing

    NASA Astrophysics Data System (ADS)

    Markuszewski, Michał J.; Bujak, Renata; Daghir, Emilia

    Capillary isoelectric focusing (CIEF) is a widespread technique for the analysis of peptides and proteins in biological samples. CIEF is used to separate mixtures of compounds on the basis of differences in their isoelectric point. Aspects of sample preparation, capillary selection, zone mobilization procedures as well as various detection modes used have been described and discussed. Moreover CIEF, coupled to various types of detection techniques (MALDI or LIF), has increasingly been applied to the analysis of variety different high-molecular compounds. CIEF is considered as a highly specific analytical method which may be routinely used in the separation of rare hemoglobin variants. In addition, the application of CIEF in proteomic field have been discussed on the examples of analyses of glycoproteins and immunoglobins due to the meaning in clinical diagnostic.

  20. Capillary Flow Experiment

    NASA Image and Video Library

    2014-06-19

    ISS040-E-015545 (19 June 2014) --- European Space Agency astronaut Alexander Gerst, Expedition 40 flight engineer, conducts a session with the Capillary Flow Experiment (CFE-2) in the Harmony node of the International Space Station. CFE is a suite of fluid physics experiments that investigate how fluids behave in microgravity which could benefit water and fuel delivery systems on future spacecraft. Scientists designed the CFE-2 to study properties of fluids and bubbles inside containers with a specific 3-D geometry.

  1. Capillary Flow Experiment

    NASA Image and Video Library

    2014-06-19

    ISS040-E-015539 (19 June 2014) --- NASA astronaut Reid Wiseman, Expedition 40 flight engineer, conducts a session with the Capillary Flow Experiment (CFE-2) in the Harmony node of the International Space Station. CFE is a suite of fluid physics experiments that investigate how fluids behave in microgravity which could benefit water and fuel delivery systems on future spacecraft. Scientists designed the CFE-2 to study properties of fluids and bubbles inside containers with a specific 3-D geometry.

  2. Enantioseparations by capillary electrochromatography.

    PubMed

    Fanali, S; Catarcini, P; Blaschke, G; Chankvetadze, B

    2001-09-01

    The review summarizes recent developments in enantioseparations by capillary electrochromatography (CEC). Selected fundamental aspects of CEC are discussed in order to stress those features which may allow the success of this technique in the competitive field of enantioseparations. In addition, the comparative characteristics of the different modes of chiral CEC and the stationary phases are presented. The effects of the characteristics of the stationary and liquid phases and operational conditions on the separation results are discussed. Finally, some future trends are briefly addressed.

  3. Capillary Flow Experiment

    NASA Image and Video Library

    2014-06-19

    ISS040-E-015523 (19 June 2014) --- European Space Agency astronaut Alexander Gerst, Expedition 40 flight engineer, conducts a session with the Capillary Flow Experiment (CFE-2) in the Harmony node of the International Space Station. CFE is a suite of fluid physics experiments that investigate how fluids behave in microgravity which could benefit water and fuel delivery systems on future spacecraft. Scientists designed the CFE-2 to study properties of fluids and bubbles inside containers with a specific 3-D geometry.

  4. Capillary Flow Experiment

    NASA Image and Video Library

    2014-06-19

    ISS040-E-015536 (19 June 2014) --- European Space Agency astronaut Alexander Gerst, Expedition 40 flight engineer, conducts a session with the Capillary Flow Experiment (CFE-2) in the Harmony node of the International Space Station. CFE is a suite of fluid physics experiments that investigate how fluids behave in microgravity which could benefit water and fuel delivery systems on future spacecraft. Scientists designed the CFE-2 to study properties of fluids and bubbles inside containers with a specific 3-D geometry.

  5. Capillary Flow Experiment

    NASA Image and Video Library

    2014-06-19

    ISS040-E-015532 (19 June 2014) --- European Space Agency astronaut Alexander Gerst, Expedition 40 flight engineer, conducts a session with the Capillary Flow Experiment (CFE-2) in the Harmony node of the International Space Station. CFE is a suite of fluid physics experiments that investigate how fluids behave in microgravity which could benefit water and fuel delivery systems on future spacecraft. Scientists designed the CFE-2 to study properties of fluids and bubbles inside containers with a specific 3-D geometry.

  6. Capillary Flow Experiment

    NASA Image and Video Library

    2014-06-19

    ISS040-E-015543 (19 June 2014) --- European Space Agency astronaut Alexander Gerst, Expedition 40 flight engineer, conducts a session with the Capillary Flow Experiment (CFE-2) in the Harmony node of the International Space Station. CFE is a suite of fluid physics experiments that investigate how fluids behave in microgravity which could benefit water and fuel delivery systems on future spacecraft. Scientists designed the CFE-2 to study properties of fluids and bubbles inside containers with a specific 3-D geometry.

  7. Effects of concomitant mild traumatic brain injury on resuming work after suffering from an isolated limb fracture: A cohort study.

    PubMed

    Jodoin, Marianne; Rouleau, Dominique M; Larson-Dupuis, Camille; Benoit, Benoit; Leduc, Stéphane; Laflamme, G-Yves; Gosselin, Nadia; Sabir, Meriem; De Beaumont, Louis

    2017-09-06

    The objective is to explore the effects of concomitant mild traumatic brain injury (mTBI) on return to work (RTW), among patients suffering from an isolated limb fracture. This follow-up study included a total of 170 working age subjects with an isolated limb fracture, and was conducted in a phone interview approximately 1-year post trauma. 41 had experienced an mTBI and 129 did not. Data were obtained through a phone interview conducted on average 20.7 months (SD = 9.6 months) post-accident. The main outcome measure was the number of days taken to RTW after the injury. Demographic information was also gathered during the phone interview. Workers' compensation status was obtained through the hospitals' orthopaedic clinic data. The mTBI group took on average 329.7 days (SD = 298.0) to RTW after the injury, as opposed to 150.3 days (SD = 171.3) for the control group (p < 0.001). After excluding patients who received workers' compensation, the mTBI group still missed significantly more days of work (M = 299.4 days; SD = 333.0) than the control group (M = 105.2 days; SD = 121.6) (p < 0.0001). This study shows that mTBI increases work disability by preventing working-age individuals from rapidly returning to work.

  8. Effects of 2-substituted-4-phenylquinolines on uptake of serotonin and norepinephrine by isolated brain synaptosomes

    SciTech Connect

    Alhaider, A.A.; Lein, E.J.; Ransom, R.W.; Bolger, M.B.

    1987-03-02

    In this present communication, the in vitro inhibition of the uptake of (/sup 3/H)-L-norepinephrine ((/sup 3/H) NE) and (/sup 3/H)-Serotonin ((/sup 3/H) 5-HT) by eleven synthesized 2-substituted-4-phenylquionlines were studied using rate brain synaptosomal preparations. Compounds with an open side chain were relatively weak inhibitors of the synaptosomal uptake of (/sup 3/H) NE and (/sup 3/H) 5HT. Compounds having a distance of three atoms between the terminal basic nitrogen of the side chain and the quinoline ring were better inhibitors of serotonin uptake than those compounds having a four-atom distance. The replacement of the side chain with a piperazine ring produced compounds which were more potent and selective inhibitors of the uptake of either (/sup 3/H) 5-HT or (/sup 3/H) NE. Further structure-activity relationships are also discussed. 13 references, 1 table.

  9. Automated Parallel Capillary Electrophoretic System

    DOEpatents

    Li, Qingbo; Kane, Thomas E.; Liu, Changsheng; Sonnenschein, Bernard; Sharer, Michael V.; Kernan, John R.

    2000-02-22

    An automated electrophoretic system is disclosed. The system employs a capillary cartridge having a plurality of capillary tubes. The cartridge has a first array of capillary ends projecting from one side of a plate. The first array of capillary ends are spaced apart in substantially the same manner as the wells of a microtitre tray of standard size. This allows one to simultaneously perform capillary electrophoresis on samples present in each of the wells of the tray. The system includes a stacked, dual carousel arrangement to eliminate cross-contamination resulting from reuse of the same buffer tray on consecutive executions from electrophoresis. The system also has a gel delivery module containing a gel syringe/a stepper motor or a high pressure chamber with a pump to quickly and uniformly deliver gel through the capillary tubes. The system further includes a multi-wavelength beam generator to generate a laser beam which produces a beam with a wide range of wavelengths. An off-line capillary reconditioner thoroughly cleans a capillary cartridge to enable simultaneous execution of electrophoresis with another capillary cartridge. The streamlined nature of the off-line capillary reconditioner offers the advantage of increased system throughput with a minimal increase in system cost.

  10. Comparison of post-mortem metabolic changes in sheep brain tissue in isolated heads and whole animals using 1H-MR spectroscopy--preliminary results.

    PubMed

    Musshoff, Frank; Klotzbach, Heike; Block, Wolfgang; Traeber, Frank; Schild, Hans; Madea, Burkhard

    2011-09-01

    By means of in situ proton magnetic resonance spectroscopy ((1)H-MRS) time-dependent metabolic changes in brain are measured for the estimation of post-mortem intervals (PMIs). An isolated whole head and a whole young sheep from an abattoir with known time of death were stored at approximately 21°C, and localized (1)H-MRS (TR/TE 2,500/25 ms) of the brain was performed at various PMIs with a clinical 1.5 T whole-body scanner using a quadratur head coil. In the isolated head, additional metabolites including free trimethylammonium (fTMA), propionate, and butyrate, and especially acetate, could be detected as described in previous studies. In the head of the whole animal, especially fTMAs and lactate were found in the late PMI but no further metabolites of relevance. These preliminary findings support the hypothesis that in a whole-body corpse, the distribution of bacteria from the gastrointestinal tract up to the brain in the late post-mortem interval can influence the metabolic decomposition of brain tissues. In further studies, it should be kept in mind that an isolated head does not represent authentic circumstances and is, therefore, not an ideal model for brain decomposition in intact corpses.

  11. Isolated blunt severe traumatic brain injury in Bern, Switzerland, and the United States: A matched cohort study.

    PubMed

    Haltmeier, Tobias; Schnüriger, Beat; Benjamin, Elizabeth; Brodmann Maeder, Monika; Künzler, Michael; Siboni, Stefano; Inaba, Kenji; Demetriades, Demetrios

    2016-02-01

    The ideal prehospital management of patients with severe traumatic brain injury (TBI) including the impact of endotracheal intubation (ETI) and physicians on scene is unclear. Prehospital management differs substantially in Switzerland and the United States: in Switzerland, there is usually a physician on scene who may provide ETI and other advanced life support procedures, whereas in the United States, prehospital management (including ETI) is performed by paramedics. This is a retrospective cohort-matched study of patients with isolated blunt severe TBI (head Abbreviated Injury Scale [AIS] score, 4-5) and no major extracranial injuries, using Bern University Hospital data from the Swiss PEBITA [Patient-relevant Endpoints after Brain Injury from Traumatic Accidents] (TBI-specific) database and the US National Trauma Data Bank from 2009 to 2010. A 1:4 cohort matching of Bern and US patients was performed. Matching criteria were sex, age (±10 years), exact field Glasgow Coma Scale (GCS) score, exact head AIS score, and injury type (subdural hematoma, epidural hematoma, intraparenchymal hemorrhage, intraventricular hemorrhage, brain edema/swelling, brain stem injury). The matched cohorts were compared with univariable analysis (Fisher's exact test and Mann-Whitney U-test). Matching of the Bern (n = 128) and US (n = 86,375) cohort resulted in 355 matched cases (71 Bern and 284 US patients). Bern patients had significantly longer scene times (median, 23.0 minutes vs. 9.0 minutes, p < 0.001) and more frequent prehospital ETI (31.0% vs. 18.7%, p = 0.034) and air transportation (39.4% vs. 19.4%, p < 0.001). No significant difference in procedures (craniotomy/craniectomy, intracranial pressure monitoring, tracheotomy), intensive care unit and total hospital lengths of stay, ventilator days, and in-hospital mortality (14.1% vs. 15.8%, p = 0.855) was found between the two cohorts. When taking into account the limitation that patient- and injury-related factors, but not in

  12. One-step Multiple Component Isolation from the Oil of Crinitaria tatarica (Less.) Sojak by Preparative Capillary Gas Chromatography with Characterization by Spectroscopic and Spectrometric Techniques and Evaluation of Biological Activity

    DTIC Science & Technology

    2012-01-01

    20] Adams, R. P., Identification of Essential Oil Components by Gas Chromatography /Mass Spectrometry, Allured Publishing Corporation, Carol Stream... chromatography with characterization by spectroscopic and spectrometric techniques and evaluation of biological activity Gas chromatographic analysis revealed...step multiple fractionation of the oil and separation of two unknown constituents were performed using preparative capillary gas chromatography

  13. Isolation, culture and long-term maintenance of primary mesencephalic dopaminergic neurons from embryonic rodent brains.

    PubMed

    Weinert, Maria; Selvakumar, Tharakeswari; Tierney, Travis S; Alavian, Kambiz N

    2015-02-19

    Degeneration of mesencephalic dopaminergic (mesDA) neurons is the pathological hallmark of Parkinson's diseae. Study of the biological processes involved in physiological functions and vulnerability and death of these neurons is imparative to understanding the underlying causes and unraveling the cure for this common neurodegenerative disorder. Primary cultures of mesDA neurons provide a tool for investigation of the molecular, biochemical and electrophysiological properties, in order to understand the development, long-term survival and degeneration of these neurons during the course of disease. Here we present a detailed method for the isolation, culturing and maintenance of midbrain dopaminergic neurons from E12.5 mouse (or E14.5 rat) embryos. Optimized cell culture conditions in this protocol result in presence of axonal and dendritic projections, synaptic connections and other neuronal morphological properties, which make the cultures suitable for study of the physiological, cell biological and molecular characteristics of this neuronal population.

  14. CD4 and MHC class I down-modulation activities of nef alleles from brain- and lymphoid tissue-derived primary HIV-1 isolates

    PubMed Central

    Gray, Lachlan R.; Gabuzda, Dana; Cowley, Daniel; Ellett, Anne; Chiavaroli, Lisa; Wesselingh, Steven L.; Churchill, Melissa J.; Gorry, Paul R.

    2015-01-01

    HIV-1 nef undergoes adaptive evolution in the CNS, reflecting altered requirements for HIV-1 replication in macrophages/microglia and brain-specific immune selection pressures. The role of Nef in HIV-1 neurotropism and the pathogenesis of HIV-associated dementia (HAD) is unclear. In this study, we characterized 82 nef alleles cloned from brain, CSF, spinal cord and blood/lymphoid tissue-derived HIV-1 isolates from 7 subjects with HAD. CNS isolate-derived nef alleles were genetically compartmentalized and had reduced sequence diversity compared to those from lymphoid tissue isolates. Defective nef alleles predominated in a brain-derived isolate from one of the 7 subjects (MACS2-br). The ability of Nef to down-modulate CD4 and MHC class 1 (MHC-1) was generally conserved among nef alleles from both CNS and lymphoid tissues. However, the potency of CD4 and MHC-1 down-modulation was variable, which was associated with sequence alterations known to influence these Nef functions. These results suggest that CD4 and MHC-1 down-modulation are highly conserved functions among nef alleles from CNS- and lymphoid tissue-derived HIV-1 isolates that may contribute to viral replication and escape from immune surveillance in the CNS. PMID:21165790

  15. Polyamine synthesis in mammalian tissues. Isolation and characterization of spermidine synthase from bovine brain.

    PubMed Central

    Raina, A; Hyvönen, T; Eloranta, T; Voutilainen, M; Samejima, K; Yamanoha, B

    1984-01-01

    Spermidine synthase (EC 2.5.1.16) was purified to apparent homogeneity (about 11 000-fold) from bovine brain by affinity chromatography, with S-adenosyl-(5')-3-thiopropylamine linked to Sepharose as the adsorbent. The enzyme preparation was free from S-adenosylmethionine decarboxylase (EC 4.1.1.50) and spermine synthase (EC 2.5.1.22) activities. The native enzyme had an apparent Mr of 70 000, was composed of two subunits of equal size, and had an isoelectric point at pH 5.22. The apparent Km values for putrescine and decarboxylated adenosylmethionine [S-adenosyl-(5')-3-methylthiopropylamine] were 40 microM and 0.3 microM respectively. Cadaverine and 1,6-diaminohexane could replace putrescine as the aminopropyl acceptor, although the reaction rates were only 6% and 1% respectively of that obtained with putrescine. Ethyl, propyl and carboxymethyl analogues of decarboxy-S-adenosylmethionine could act as propylamine donors. Both the reaction products, spermidine and 5'-methylthioadenosine, were mixed-type inhibitors of the enzyme. On the basis of initial-velocity and product-inhibition studies, a ping-pong reaction mechanism for the spermidine synthase reaction was ruled out. Images Fig. 1. PMID:6743257

  16. Hyperfibrinolysis in severe isolated traumatic brain injury may occur without tissue hypoperfusion: a retrospective observational multicentre study.

    PubMed

    Hayakawa, Mineji; Maekawa, Kunihiko; Kushimoto, Shigeki; Kato, Hiroshi; Sasaki, Junichi; Ogura, Hiroshi; Matsuoka, Tetsuya; Uejima, Toshifumi; Morimura, Naoto; Ishikura, Hiroyasu; Hagiwara, Akiyoshi; Takeda, Munekazu; Kaneko, Naoyuki; Saitoh, Daizoh; Kudo, Daisuke; Kanemura, Takashi; Shibusawa, Takayuki; Furugori, Shintaro; Nakamura, Yoshihiko; Shiraishi, Atsushi; Murata, Kiyoshi; Mayama, Gou; Yaguchi, Arino; Kim, Shiei; Takasu, Osamu; Nishiyama, Kazutaka

    2017-08-23

    Hyperfibrinolysis is a critical complication in severe trauma. Hyperfibrinolysis is traditionally diagnosed via elevated D-dimer or fibrin/fibrinogen degradation product levels, and recently, using thromboelastometry. Although hyperfibrinolysis is observed in patients with severe isolated traumatic brain injury (TBI) on arrival at the emergency department (ED), it is unclear which factors induce hyperfibrinolysis. The present study aimed to investigate the factors associated with hyperfibrinolysis in patients with isolated severe TBI. We conducted a multicentre retrospective review of data for adult trauma patients with an injury severity score ≥ 16, and selected patients with isolated TBI (TBI group) and extra-cranial trauma (non-TBI group). The TBI group included patients with an abbreviated injury score (AIS) for the head ≥ 4 and an extra-cranial AIS < 2. The non-TBI group included patients with an extra-cranial AIS ≥ 3 and head AIS < 2. Hyperfibrinolysis was defined as a D-dimer level ≥ 38 mg/L on arrival at the ED. We evaluated the relationships between hyperfibrinolysis and injury severity/tissue injury/tissue perfusion in TBI patients by comparing them with non-TBI patients. We enrolled 111 patients in the TBI group and 126 in the non-TBI group. In both groups, patients with hyperfibrinolysis had more severe injuries and received transfusion more frequently than patients without hyperfibrinolysis. Tissue injury, evaluated on the basis of lactate dehydrogenase and creatine kinase levels, was associated with hyperfibrinolysis in both groups. Among patients with TBI, the mortality rate was higher in those with hyperfibrinolysis than in those without hyperfibrinolysis. Tissue hypoperfusion, evaluated on the basis of lactate level, was associated with hyperfibrinolysis in only the non-TBI group. Although the increase in lactate level was correlated with the deterioration of coagulofibrinolytic variables (prolonged prothrombin time

  17. A novel dipeptidyl aminopeptidase in rat brain membranes. Its isolation, purification, and characterization.

    PubMed

    Hui, K S

    1988-05-15

    A new type of dipeptidyl aminopeptidase, which releases basic aminoacyl dipeptides from the NH2-terminal end of oligopeptides, was purified about 2100-fold with 6.8% recovery from rat brain membranes by column chromatography on Cellex D, Arg-Tyr-AH-Sepharose 4B, hydroxylapatite, and Sephadex G-75, after the membranes were solubilized with Nonidet P-40. Activity was assayed by high performance liquid chromatography (HPLC) using Arg0-Met5-enkephalin (Arg0-enk)* as substrate in the presence of bestatin, thiorphan, and captopril. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the purified enzyme is apparently homogeneous with a mass of 64,000 daltons. This thiol enzyme is optimally active at pH 7 and is selectively activated by Mn(II). It loses 94% of its activity after EDTA treatment and can be reactivated by Mn(II), Co(II), and Zn(II). It splits Arg0-enk into equimolar amounts of Arg-Tyr and Gly-Gly-Phe-Met with a Km of 100 microM, and Vmax of 3.8 mumol/mg of protein/min. Dipeptidyl aminopeptidase does not hydrolyze model substrates for dipeptidyl aminopeptidases I, II, III, and IV, aminoacyl beta-naphthylamides, actin, desmin, tubulin, glial fibrillary acidic protein, and cytoskeletal neurofilament proteins. The enzyme is insensitive to puromycin, but is inhibited by several neuropeptides. Angiotensin III is the most potent with a Ki of 0.3 microM. Substrate specificity, pH optimum, molecular weight, activators, and catalytic site demonstrate that this enzyme is distinct from dipeptidyl aminopeptidases previously described.

  18. Capillary reference half-cell

    DOEpatents

    Hall, Stephen H.

    1996-01-01

    The present invention is a reference half-cell electrode wherein intermingling of test fluid with reference fluid does not affect the performance of the reference half-cell over a long time. This intermingling reference half-cell may be used as a single or double junction submersible or surface reference electrode. The intermingling reference half-cell relies on a capillary tube having a first end open to reference fluid and a second end open to test fluid wherein the small diameter of the capillary tube limits free motion of fluid within the capillary to diffusion. The electrode is placed near the first end of the capillary in contact with the reference fluid. The method of operation of the present invention begins with filling the capillary tube with a reference solution. After closing the first end of the capillary, the capillary tube may be fully submerged or partially submerged with the second open end inserted into test fluid. Since the electrode is placed near the first end of the capillary, and since the test fluid may intermingle with the reference fluid through the second open end only by diffusion, this intermingling capillary reference half-cell provides a stable voltage potential for long time periods.

  19. Capillary reference half-cell

    DOEpatents

    Hall, S.H.

    1996-02-13

    The present invention is a reference half-cell electrode wherein intermingling of test fluid with reference fluid does not affect the performance of the reference half-cell over a long time. This intermingling reference half-cell may be used as a single or double junction submersible or surface reference electrode. The intermingling reference half-cell relies on a capillary tube having a first end open to reference fluid and a second end open to test fluid wherein the small diameter of the capillary tube limits free motion of fluid within the capillary to diffusion. The electrode is placed near the first end of the capillary in contact with the reference fluid. The method of operation of the present invention begins with filling the capillary tube with a reference solution. After closing the first end of the capillary, the capillary tube may be fully submerged or partially submerged with the second open end inserted into test fluid. Since the electrode is placed near the first end of the capillary, and since the test fluid may intermingle with the reference fluid through the second open end only by diffusion, this intermingling capillary reference half-cell provides a stable voltage potential for long time periods. 11 figs.

  20. Capillary pericytes regulate cerebral blood flow in health and disease

    PubMed Central

    Sutherland, Brad A.; O’Farrell, Fergus M.; Buchan, Alastair M.; Lauritzen, Martin; Attwell, David

    2014-01-01

    Brain blood flow increases, evoked by neuronal activity, power neural computation and are the basis of BOLD functional imaging. It is controversial whether blood flow is controlled solely by arteriole smooth muscle, or also by capillary pericytes. We demonstrate that neuronal activity and the neurotransmitter glutamate evoke the release of messengers that dilate capillaries by actively relaxing pericytes. Dilation is mediated by prostaglandin E2, but requires nitric oxide release to suppress vasoconstricting 20-HETE synthesis. In vivo, when sensory input increases blood flow, capillaries dilate before arterioles and are estimated to produce 84% of the blood flow increase. In pathology, ischaemia evokes capillary constriction by pericytes. We show that this is followed by pericyte death in rigor, which may irreversibly constrict capillaries and damage the blood-brain barrier. Thus, pericytes are major regulators of cerebral blood flow and initiators of functional imaging signals. Prevention of pericyte constriction and death may reduce the long-lasting blood flow decrease which damages neurons after stroke. PMID:24670647

  1. Fatty Acid-Binding Protein 5 at the Blood-Brain Barrier Regulates Endogenous Brain Docosahexaenoic Acid Levels and Cognitive Function.

    PubMed

    Pan, Yijun; Short, Jennifer L; Choy, Kwok H C; Zeng, Annie X; Marriott, Philip J; Owada, Yuji; Scanlon, Martin J; Porter, Christopher J H; Nicolazzo, Joseph A

    2016-11-16

    Fatty acid-binding protein 5 (FABP5) at the blood-brain barrier contributes to the brain uptake of docosahexaenoic acid (DHA), a blood-derived polyunsaturated fatty acid essential for maintenance of cognitive function. Given the importance of DHA in cognition, the aim of this study was to investigate whether deletion of FABP5 results in cognitive dysfunction and whether this is associated with reduced brain endothelial cell uptake of exogenous DHA and subsequent attenuation in the brain levels of endogenous DHA. Cognitive function was assessed in male and female FABP5(+/+) and FABP5(-/-) mice using a battery of memory paradigms. FABP5(-/-) mice exhibited impaired working memory and short-term memory, and these cognitive deficits were associated with a 14.7 ± 5.7% reduction in endogenous brain DHA levels. The role of FABP5 in the blood-brain barrier transport of DHA was assessed by measuring (14)C-DHA uptake into brain endothelial cells and capillaries isolated from FABP5(+/+) and FABP5(-/-) mice. In line with a crucial role of FABP5 in the brain uptake of DHA, (14)C-DHA uptake into brain endothelial cells and brain capillaries of FABP5(-/-) mice was reduced by 48.4 ± 14.5% and 14.0 ± 4.2%, respectively, relative to those of FABP5(+/+) mice. These results strongly support the hypothesis that FABP5 is essential for maintaining brain endothelial cell uptake of DHA, and that cognitive deficits observed in FABP5(-/-) mice are associated with reduced CNS access of DHA. Genetic deletion of fatty acid-binding protein 5 (FABP5) in mice reduces uptake of exogenous docosahexaenoic acid (DHA) into brain endothelial cells and brain capillaries and reduces brain parenchymal levels of endogenous DHA. Therefore, FABP5 in the brain endothelial cell is a crucial contributor to the brain levels of DHA. Critically, lowered brain DHA levels in FABP5(-/-) mice occurred in tandem with cognitive deficits in a battery of memory paradigms. This study provides evidence of a critical role

  2. Functional interactions within the parahippocampal region revealed by voltage-sensitive dye imaging in the isolated guinea pig brain.

    PubMed

    Biella, Gerardo; Spaiardi, Paolo; Toselli, Mauro; de Curtis, Marco; Gnatkovsky, Vadym

    2010-02-01

    The massive transfer of information from the neocortex to the entorhinal cortex (and vice versa) is hindered by a powerful inhibitory control generated in the perirhinal cortex. In vivo and in vitro experiments performed in rodents and cats support this conclusion, further extended in the present study to the analysis of the interaction between the entorhinal cortex and other parahippocampal areas, such as the postrhinal and the retrosplenial cortices. The experiments were performed in the in vitro isolated guinea pig brain by a combined approach based on electrophysiological recordings and fast imaging of optical signals generated by voltage-sensitive dyes applied to the entire brain by arterial perfusion. Local stimuli delivered in different portions of the perirhinal, postrhinal, and retrosplenial cortex evoked local responses that did not propagate to the entorhinal cortex. Neither high- and low-frequency-patterned stimulation nor paired associative stimuli facilitated the propagation of activity to the entorhinal region. Similar stimulations performed during cholinergic neuromodulation with carbachol were also ineffective in overcoming the inhibitory network that controls propagation to the entorhinal cortex. The pharmacological inactivation of GABAergic transmission by local application of bicuculline (1 mM) in area 36 of the perirhinal cortex facilitated the longitudinal (rostrocaudal) propagation of activity into the perirhinal/postrhinal cortices but did not cause propagation into the entorhinal cortex. Bicuculline injection in both area 35 and medial entorhinal cortex released the inhibitory control and allowed the propagation of the neural activity to the entorhinal cortex. These results demonstrate that, as for the perirhinal-entorhinal reciprocal interactions, also the connections between the postrhinal/retrosplenial cortices and the entorhinal region are subject to a powerful inhibitory control.

  3. Does isolated traumatic subarachnoid hemorrhage merit a lower intensity level of observation than other traumatic brain injury?

    PubMed

    Phelan, Herb A; Richter, Adam A; Scott, William W; Pruitt, Jeffrey H; Madden, Christopher J; Rickert, Kim L; Wolf, Steven E

    2014-10-15

    Evidence is emerging that isolated traumatic subarachnoid hemorrhage (ITSAH) may be a milder form of traumatic brain injury (TBI). If true, ITSAH may not benefit from intensive care unit (ICU) admission, which would, in turn, decrease resource utilization. We conducted a retrospective review of all TBI admissions to our institution between February 2010 and November 2012 to compare the presentation and clinical course of subjects with ITSAH to all other TBI. We then performed descriptive statistics on the subset of ITSAH subjects presenting with a Glasgow Coma Score (GCS) of 13-15. Of 698 subjects, 102 had ITSAH and 596 had any other intracranial hemorrhage pattern. Compared to all other TBI, ITSAH had significantly lower injury severity scores (p<0.0001), lower head abbreviated injury scores (p<0.0001), higher emergency department GCS (p<0.0001), shorter ICU stays (p=0.007), higher discharge GCS (p=0.005), lower mortality (p=0.003), and significantly fewer head computed tomography scans (p<0.0001). Of those ITSAH subjects presenting with a GCS of 13-15 (n=77), none underwent placement of an intracranial monitor or craniotomy. One subject (1.3%) demonstrated a change in exam (worsened headache and dizziness) concomitant with a progression of his intracranial injury. His symptoms resolved with readmission to the ICU and continued observation. Our results suggest that ITSAH are less-severe brain injuries than other TBI. ITSAH patients with GCS scores of 13-15 demonstrate low rates of clinical progression, and when progression occurs, it resolves without further intervention. This subset of TBI patients does not appear to benefit from ICU admission.

  4. Studies of isolated global brain ischaemia: II. Controlled reperfusion provides complete neurologic recovery following 30 min of warm ischaemia - the importance of perfusion pressure.

    PubMed

    Allen, Bradley S; Ko, Yoshihiro; Buckberg, Gerald D; Tan, Zhong

    2012-05-01

    Neurologic injury after sudden death is likely due to a reperfusion injury following prolonged brain ischaemia, and remains problematic, especially if the cardiac arrest is unwitnessed. This study applies a newly developed isolated model of global brain ischaemia (simulating unwitnessed sudden death) for 30 min to determine if controlled reperfusion permits neurologic recovery. Among the 17 pigs undergoing 30 min of normothermic global brain ischaemia, 6 received uncontrolled reperfusion with regular blood (n = 6), and 11 were reperfused for 20 min with a warm controlled blood reperfusate containing hypocalcaemia, hyper-magnesemia, alkalosis, hyperosmolarty and other constituents that were passed through a white blood cell filter and delivered at flow rates of 350 cc/min (n = 3), 550 cc/min (n = 2) or 750 cc/min (n = 6). Neurologic deficit score (NDS) evaluated brain function (score 0 = normal, 500 = brain death) 24 h post-reperfusion and 2,3,5-triphenyltetrazolium chloride (TTC) staining determined brain infarction. Regular blood (uncontrolled) reperfusion caused negligible brain O(2) uptake by IN Vivo Optical Spectroscopy (INVOS) (<10-15% O(2) extraction), oxidant damage demonstrated by raised conjugated diene (CD) levels (1.78 ± 0.13 A233 mn), multiple seizures, 1 early death from brain herniation, high NDS (249 ± 39) in survivors, brain oedema (84.4 ± 0.6%) and extensive cerebral infarctions. Conversely, controlled reperfusion restored surface brain oxygen saturation by INVOS to normal (55-70%), but the extent of neurologic recovery was determined by the brain reperfusion pressure. Low pressure reperfusion (independent of flow) produced the same adverse functional, metabolic and anatomic changes that followed uncontrolled reperfusion in seven pigs (three at 350 cc/min, two at 550 and two at 750 cc/min). Conversely, higher reperfusion pressure in four pigs (all at 750 cc/min) resulted in NDS of 0-70* indicating complete (n = 2) or near complete (n = 2

  5. Does early beta-blockade in isolated severe traumatic brain injury reduce the risk of post traumatic depression?

    PubMed

    Ahl, Rebecka; Sjolin, Gabriel; Mohseni, Shahin

    2017-01-01

    Depressive symptoms occur in approximately half of trauma patients, negatively impacting on functional outcome and quality of life following severe head injury. Pontine noradrenaline has been shown to increase upon trauma and associated β-adrenergic receptor activation appears to consolidate memory formation of traumatic events. Blocking adrenergic activity reduces physiological stress responses during recall of traumatic memories and impairs memory, implying a potential therapeutic role of β-blockers. This study examines the effect of pre-admission β-blockade on post-traumatic depression. All adult trauma patients (≥18 years) with severe, isolated traumatic brain injury (intracranial Abbreviated Injury Scale score (AIS) ≥3 and extracranial AIS <3) were recruited from the trauma registry of an urban university hospital between 2007 and 2011. Exclusion criteria were in-hospital deaths and prescription of antidepressants up to one year prior to admission. Pre- and post-admission β-blocker and antidepressant therapy data was requested from the national drugs registry. Post-traumatic depression was defined as the prescription of antidepressants within one year of trauma. Patients with and without pre-admission β-blockers were matched 1:1 by age, gender, Glasgow Coma Scale, Injury Severity Score and head AIS. Analysis was carried out using McNemar's and Student's t-test for categorical and continuous data, respectively. A total of 545 patients met the study criteria. Of these, 15% (n=80) were prescribed β-blockers. After propensity matching, 80 matched pairs were analyzed. 33% (n=26) of non β-blocked patients developed post-traumatic depression, compared to only 18% (n=14) in the β-blocked group (p=0.04). There were no significant differences in ICU (mean days: 5.8 (SD 10.5) vs. 5.6 (SD 7.2), p=0.85) or hospital length of stay (mean days: 21 (SD 21) vs. 21 (SD 20), p=0.94) between cohorts. β-blockade appears to act prophylactically and significantly

  6. Capillary rogue waves.

    PubMed

    Shats, M; Punzmann, H; Xia, H

    2010-03-12

    We report the first observation of extreme wave events (rogue waves) in parametrically driven capillary waves. Rogue waves are observed above a certain threshold in forcing. Above this threshold, frequency spectra broaden and develop exponential tails. For the first time we present evidence of strong four-wave coupling in nonlinear waves (high tricoherence), which points to modulation instability as the main mechanism in rogue waves. The generation of rogue waves is identified as the onset of a distinct tail in the probability density function of the wave heights. Their probability is higher than expected from the measured wave background.

  7. Stability of capillary surfaces

    NASA Technical Reports Server (NTRS)

    Weislogel, Mark

    1991-01-01

    An extensive data set quantifying the stability limits of capillary surfaces for a wide range of fluid properties, container geometry, and input disturbance and orientation does not currently exist. To begin to provide such a data set an experimental apparatus has been designed for which the dynamics and stability of fluid interfaces will be investigated. The apparatus consists primarily of a programmable shaker table and a high speed motion picture and video camera for viewing the fluid surface as drop tower tests will be performed to note the stability of a variety of surface/vessel configurations.

  8. Aggressive operative treatment of isolated blunt traumatic brain injury in the elderly is associated with favourable outcome.

    PubMed

    Wutzler, Sebastian; Lefering, Rolf; Wafaisade, Arasch; Maegele, Marc; Lustenberger, Thomas; Walcher, Felix; Marzi, Ingo; Laurer, Helmut

    2015-09-01

    Outcome after traumatic brain injury (TBI) in the elderly has not been fully elucidated. The present retrospective observational study investigates the age-dependent outcome of patients suffering from severe isolated TBI with regard to operative and non-operative treatment. Data were prospectively collected in the TraumaRegister DGU. Anonymous datasets of 8629 patients with isolated severe blunt TBI (AISHead≥3, AISBody≤1) documented from 2002 to 2011 were analysed. Patients were grouped according to age: 1-17, 18-59, 60-69, 70-79 and ≥80 years. Cranial fractures (44.8%) and subdural haematomas (42.6%) were the most common TBIs. Independent from the type of TBI the group of patients with operative treatment declined with rising age. Subgroup analysis of patients with critical TBI (AISHead=5) revealed standardised mortality ratios (SMRs) of 0.81 (95% CI 0.75-0.87) in case of operative treatment (n=1201) and 1.13 (95% CI 1.09-1.18) in case of non-operative treatment (n=1096). All age groups ≥60 years showed significantly reduced SMRs in case of operative treatment. Across all age groups the group of patients with low/moderate disability according to the GOS (4 or 5 points) was higher in case of operative treatment. Results of this retrospective observational study have to be interpreted cautiously. However, good outcome after TBI with severe space-occupying haemorrhage is more frequent in patients with operative treatment across all age groups. Age alone should not be the reason for limited care or denial of operative intervention.

  9. In vivo and ex vivo regulation of breast cancer resistant protein (Bcrp) by peroxisome proliferator-activated receptor alpha (Pparα) at the blood-brain barrier.

    PubMed

    Hoque, Md Tozammel; Shah, Arpit; More, Vijay; Miller, David S; Bendayan, Reina

    2015-12-01

    Breast cancer resistance protein (Bcrp/Abcg2) localized at the blood-brain barrier (BBB) limits permeability into the brain of many xenobiotics, including pharmacological agents. Peroxisome proliferator-activated receptor α (Pparα), a ligand-activated transcription factor, primarily involved in lipid metabolism, has been shown to regulate the functional expression of Bcrp in human cerebral microvascular endothelial cells (hCMEC/D3). The aim of this study was to investigate ex vivo and in vivo, the regulation of Bcrp by Pparα in an intact BBB. Ex vivo quantitative real-time PCR and immunoblot analyses showed significant up-regulation of Abcg2/Bcrp mRNA and protein levels in CD-1 mouse brain capillaries incubated with clofibrate, a Pparα ligand. Fluorescence-based transport assays in CD-1 and C57BL/6 brain capillaries showed that exposure to clofibrate significantly increased Bcrp transport activity. This increase was not observed in capillaries isolated from Pparα knockout mice. In vivo, we found: i) significant Bcrp protein up-regulation in clofibrate-dosed CD-1 and C57BL/6 capillary lysates, but no effect in Pparα knockout capillary lysates, and ii) significantly increased Bcrp transport activity in capillaries isolated from clofibrate-treated mice. These results demonstrate an increase in Bcrp functional expression by Pparα in brain capillaries, and suggest that Pparα is another nuclear receptor that can contribute to the regulation of membrane efflux transporters and drug permeability at the BBB. We propose the involvement of the following pathways in clofibrate-mediated induction of the drug transporter Abcg2/Bcrp mRNA, protein expression and function by the nuclear receptor Pparα, in mouse brain capillary endothelial cells. Upon activation with clofibrate (Pparα, ligand), Pparα complex translocates from the cytoplasm into the nucleus and further recruits coactivators and transcription machinery which induce the transcription of Abcg2 gene and

  10. Capillary suspensions: Particle networks formed through the capillary force

    PubMed Central

    Koos, Erin

    2014-01-01

    The addition of small amounts of a secondary fluid to a suspension can, through the attractive capillary force, lead to particle bridging and network formation. The capillary bridging phenomenon can be used to stabilize particle suspensions and precisely tune their rheological properties. This effect can even occur when the secondary fluid wets the particles less well than the bulk fluid. These materials, so-called capillary suspensions, have been the subject of recent research studying the mechanism for network formation, the properties of these suspensions, and how the material properties can be modified. Recent work in colloidal clusters is summarized and the relationship to capillary suspensions is discussed. Capillary suspensions can also be used as a pathway for new material design and some of these applications are highlighted. Results obtained to date are summarized and central questions that remain to be answered are proposed in this review. PMID:25729316

  11. Tapered capillary optics

    DOEpatents

    Hirsch, Gregory

    1998-01-01

    A metal or glass wire is etched with great precision into a very narrowly tapering cone which has the shape of the desired final capillary-optics bore. By controlling the rate of removal of the wire from an etchant bath, a carefully controlled taper is produced. A sensor measures the diameter of the wire as it leaves the surface of the etchant. This signal is used for feedback control of the withdrawal speed. The etched wire undergoes a treatment to produce an extremely low surface-roughness. The etched and smoothed wire is coated with the material of choice for optimizing the reflectivity of the radiation being focused. This could be a vacuum evaporation, sputtering, CVD or aqueous chemical process. The coated wire is either electroplated, built up with electroless plating, or encapsulated in a polymer cylinder such as epoxy to increase the diameter of the wire for easier handling and greater robustness. During this process, the wire is vertically oriented and tensioned to assure that the wire is absolutely straight. The coated and electroformed wire is bonded to a flat, rigid substrate and is then periodically segmented by cutting or etching a series of narrow slits or grooves into the wire. The wire is vertically oriented and tensioned during the bonding process to assure that it is straight. The original wire material is then chemically etched away through the slits or otherwise withdrawn to leave the hollow internal bore of the final tapered-capillary optical element.

  12. Capillary electrochromatography of cannabinoids.

    PubMed

    Lurie, I S; Meyers, R P; Conver, T S

    1998-08-01

    The applicability of capillary electrochromatography (CEC) with photodiode array UV detection for the analysis of cannabinoids is presented. Baseline separation of seven cannabinoids (cannabigerol, cannabidiol, cannabinol, delta-9-tetrahydrocannabinol, delta-8-tetrahydrocannabinol, cannabichromene, delta-9-tetrahydrocannabinolic acid) is obtained using a 3-micron CEC Hypersil C18 capillary with an acetonitrile/phosphate (pH 2.57) mobile phase. The effects of acetonitrile concentration, buffer concentration, voltage, temperature, stationary phase, and column length on the separation of the cannabinoids were investigated. Good short- and long-term precision in retention times are observed, with significant improvement obtained using relative retention times with cannabinol as reference compound. Although short- and long-term peak area precisions are poor, satisfactory reproducibility is obtained using relative peak areas with cannabinol as reference compound. The applicability of the CEC methodology to drug seizures was demonstrated on marijuana and hashish. Using a high-sensitivity UV flow cell with an extended path length of 1.2 mm, concentration sensitivities approaching HPLC were obtained.

  13. Microbiota-Gut-Brain Axis: Yeast Species Isolated from Stool Samples of Children with Suspected or Diagnosed Autism Spectrum Disorders and In Vitro Susceptibility Against Nystatin and Fluconazole.

    PubMed

    Kantarcioglu, A Serda; Kiraz, Nuri; Aydin, Ahmet

    2016-02-01

    Autism spectrum disorder (ASD) is a general term for a group of complex neurodevelopmental disorders of brain development that limits a person's ability to function normally. Etiology has not been clearly defined up to date. However, gut microbiota and the bidirectional communication between the gastrointestinal tract and brain, the so-called microbiota-gut-brain axis, are hypothesized, which may be involved in the etiology of several mental disorders. Recent reports suggest that Candida, particularly Candida albicans, growth in intestines may cause lower absorption of carbohydrates and minerals and higher toxin levels which are thought to contribute autistic behaviors. The aim of this study was to identify the 3-year deposited yeasts isolated from stool samples of children with diagnosed or suspected ASD and to determine in vitro activity of nystatin and fluconazole against these isolates using Clinical Laboratory Standards Institute M27-A3 guidelines. A 17-year retrospective assessment was also done using our laboratory records. Among the species identified, intrinsically fluconazole-resistant Candida krusei (19.8 %) and Candida glabrata (14.8 %) with elevated MICs were remarkable. Overall, C. albicans (57.4 %) was the most commonly isolated species in 17 years. The species identification and/or antifungal susceptibility tests have to be performed using the strain isolated from stool sample, to select the appropriate antifungal agent, if antimycotic therapy is needed.

  14. Diesel exhaust particles induce oxidative stress, proinflammatory signaling, and P-glycoprotein up-regulation at the blood-brain barrier.

    PubMed

    Hartz, Anika M S; Bauer, Björn; Block, Michelle L; Hong, Jau-Shyong; Miller, David S

    2008-08-01

    Here, we report that diesel exhaust particles (DEPs), a major constituent of urban air pollution, affect blood-brain barrier function at the tissue, cellular, and molecular levels. Isolated rat brain capillaries exposed to DEPs showed increased expression and transport activity of the key drug efflux transporter, P-glycoprotein (6 h EC(50) was approximately 5 microg/ml). Up-regulation of P-glycoprotein was abolished by blocking transcription or protein synthesis. Inhibition of NADPH oxidase or pretreatment of capillaries with radical scavengers ameliorated DEP-induced P-glycoprotein up-regulation, indicating a role for reactive oxygen species in signaling. DEP exposure also increased brain capillary tumor necrosis factor-alpha (TNF-alpha) levels. DEP-induced P-glycoprotein up-regulation was abolished when TNF-receptor 1 (TNF-R1) was blocked and was not evident in experiments with capillaries from TNF-R1 knockout mice. Inhibition of JNK, but not NF-kappaB, blocked DEP-induced P-glycoprotein up-regulation, indicating a role for AP-1 in the signaling pathway. Consistent with this, DEPs increased phosphorylation of c-jun. Together, our results show for the first time that a component of air pollution, DEPs, alters blood-brain barrier function through oxidative stress and proinflammatory cytokine production. These experiments disclose a novel blood-brain barrier signaling pathway, with clear implications for environmental toxicology, CNS pathology, and the pharmacotherapy of CNS disorders.

  15. Isolation and pharmacological analysis of two ionic forms of the GABA sub A /benzodiazepine receptor from rat brain

    SciTech Connect

    Moffett, J.R.

    1988-01-01

    The objective of this dissertation research was to investigate the ionic characteristics of solubilized, partially purified GABA{sub A} receptors, and to determine if physical variability within this receptor population was associated with ligand binding heterogeneity at the GABA and BZ binding sites. Utilizing BZ affinity chromatography, the GABA{sub A} receptor was partially purified from Triton X-100 solubilized rat brain membrane preparations. Anion exchange high performance liquid chromatography was employed to isolate two distinct ionic forms of the receptor, which remained stable and did not interconvert. Equilibrium binding assays with {sup 3}H-flunitrazepam, a BZ binding site agonist, and {sup 3}H-muscimol, a GABA binding site agonist, revealed no significant binding differences between these two ionic forms of the GABA{sub A} receptor. Type I and Type II GABA{sub A} receptor-have been distinguished by virtue of the differential sensitivity of their BZ binding site to the trizolopyridazine, CL 218,872. Competition of 3H-flunitrazepam binding by CL 218,872 in assays of the separated ionic forms of the GABA{sub A} receptor indicate that both high and low affinity CL 218,872 binding sites were present in both forms.

  16. Acute alcohol intoxication, diffuse axonal injury and intraventricular bleeding in patients with isolated blunt traumatic brain injury.

    PubMed

    Matsukawa, Hidetoshi; Shinoda, Masaki; Fujii, Motoharu; Takahashi, Osamu; Murakata, Atsushi; Yamamoto, Daisuke

    2013-01-01

    The influence of blood alcohol level (BAL) on outcome remains unclear. This study investigated the relationships between BAL, type and number of diffuse axonal injury (DAI), intraventricular bleeding (IVB) and 6-month outcome. This study reviewed 419 patients with isolated blunt traumatic brain injury. First, it compared clinical and radiological characteristics between patients with good recovery and disability. Second, it compared BAL among DAI lesions. Third, it evaluated the correlation between the BAL and severity of IVB, number of DAI and corpus callosum injury lesions. Regardless of BAL, older age, male gender, severe Glasgow Coma Scale score (<9), abnormal pupil, IVB and lesion on genu of corpus callosum were significantly related to disability. There were no significant differences between the BAL and lesions of DAI. Simple regression analysis revealed that there were no significant correlation between BAL and severity of IVB, number of DAI and corpus callosum injury lesions. Acute alcohol intoxication was not associated with type and number of DAI lesion, IVB and disability. This study suggested that a specific type of traumatic lesion, specifically lesion on genu of corpus callosum and IVB, might be more vital for outcome.

  17. Diagnostic value of brain chronic black holes on T1-weighted MR images in clinically isolated syndromes.

    PubMed

    Mitjana, Raquel; Tintoré, Mar; Rocca, Maria A; Auger, Cristina; Barkhof, Frederik; Filippi, Massimo; Polman, Chris; Fazekas, Franz; Huerga, Elena; Montalban, Xavier; Rovira, Alex

    2014-10-01

    Non-enhancing black holes (neBHs) are more common in multiple sclerosis (MS) patients with longer disease durations and progressive disease subtypes. Our aim was to analyse the added value of neBHs in patients with clinically isolated syndromes (CISs) for predicting conversion to clinically definite MS (CDMS). Patients were classified based on the presence or absence of neBHs and on the number of Barkhof-Tintoré (B-T) criteria fulfilled. Dissemination in space (DIS) was defined as the presence of at least three of the four B-T criteria. Dissemination in time (DIT)1 was defined by simultaneous presence of enhancing and non-enhancing lesions. DIT2 was defined by simultaneous presence of neBHs and T2 lesions not apparent on T1-weighted images. Focal T2-hyperintense brain lesions were identified in 87.7% of the 520 CIS patients, and 41.4% of them presented at least one neBH. Patients meeting DIS, DIT1, and DIT2 had a significantly higher rate of conversion to CDMS. After adjusting for DIS, only patients who fulfilled DIT1 preserved a significant increase in CDMS conversion. Non-enhancing black holes in CIS patients are associated with a higher risk of conversion to CDMS. However, the predictive value of this finding is lost when added to the DIS criteria. © The Author(s) 2014.

  18. On Capillary Rise and Nucleation

    ERIC Educational Resources Information Center

    Prasad, R.

    2008-01-01

    A comparison of capillary rise and nucleation is presented. It is shown that both phenomena result from a balance between two competing energy factors: a volume energy and a surface energy. Such a comparison may help to introduce nucleation with a topic familiar to the students, capillary rise. (Contains 1 table and 3 figures.)

  19. On Capillary Rise and Nucleation

    ERIC Educational Resources Information Center

    Prasad, R.

    2008-01-01

    A comparison of capillary rise and nucleation is presented. It is shown that both phenomena result from a balance between two competing energy factors: a volume energy and a surface energy. Such a comparison may help to introduce nucleation with a topic familiar to the students, capillary rise. (Contains 1 table and 3 figures.)

  20. Noise suppressing capillary separation system

    DOEpatents

    Yeung, E.S.; Xue, Y.

    1996-07-30

    A noise-suppressing capillary separation system for detecting the real-time presence or concentration of an analyte in a sample is provided. The system contains a capillary separation means through which the analyte is moved, a coherent light source that generates a beam which is split into a reference beam and a sample beam that irradiate the capillary, and a detector for detecting the reference beam and the sample beam light that transmits through the capillary. The laser beam is of a wavelength effective to be absorbed by a chromophore in the capillary. The system includes a noise suppressing system to improve performance and accuracy without signal averaging or multiple scans. 13 figs.

  1. Capillary optics for radiation focusing

    SciTech Connect

    Peurrung, A.J.; Reeder, P.L.; Bliss, M.; Craig, R.A.; Lepel, E.A.; Stromswold, D.C.; Stoffels, J.; Sunberg, D.S.; Tenny, H.

    1996-11-01

    Capillary lens technology may ultimately bring benefits to neutron and x-ray-based science like conventional lenses with visible light. Although the technology is not yet 10 years old, these lenses have already had a significant impact in engineering, science, and medicine. Capillary lenses are advantageous when it is desirable to increase the radiation flux at a location without regard to its angular divergence. PNNL has worked to improve the technology in several ways. A single, optimally tapered capillary was manufactured, which allows intensity gains of a factor of 270 for an initially parallel, incident x-ray beam. Feasibility of constructing neutron lenses using {sup 58}Ni (particularly effective at reflecting neutrons) has been explored. Three applications for capillary optics have been identified and studied: neutron telescope, Gandolphi x-ray diffractometry, and neutron radiotherapy. A brief guide is given for determining which potential applications are likely to be helped by capillary optics.

  2. Geometry-induced capillary emptying.

    PubMed

    Rascón, Carlos; Parry, Andrew O; Aarts, Dirk G A L

    2016-10-24

    When a capillary is half-filled with liquid and turned to the horizontal, the liquid may flow out of the capillary or remain in it. For lack of a better criterion, the standard assumption is that the liquid will remain in a capillary of narrow cross-section, and will flow out otherwise. Here, we present a precise mathematical criterion that determines which of the two outcomes occurs for capillaries of arbitrary cross-sectional shape, and show that the standard assumption fails for certain simple geometries, leading to very rich and counterintuitive behavior. This opens the possibility of creating very sensitive microfluidic devices that respond readily to small physical changes, for instance, by triggering the sudden displacement of fluid along a capillary without the need of any external pumping.

  3. Biomedical applications of capillary electrophoresis

    NASA Astrophysics Data System (ADS)

    Kartsova, L. A.; Bessonova, E. A.

    2015-08-01

    The review deals with modern analytical approaches used in capillary electrophoresis for solving medical and biological problems: search for biomarkers of various diseases and rapid diagnosis based on characteristic profiles of biologically active compounds by capillary electrophoresis with mass spectrometric detection; monitoring of the residual drugs in biological fluids for evaluating the efficiency of drug therapy; testing of the enantiomeric purity of pharmaceutical products; the use of novel materials as components of stationary and pseudo-stationary phases in capillary electrophoresis and capillary electrochromatography to increase the selectivity of separation of components of complex matrices; and identification of various on-line preconcentration techniques to reduce the detection limits of biologically active analytes. A topical trend in capillary electrophoresis required in clinical practice, viz., the design of microfluidic systems, is discussed. The bibliography includes 173 references.

  4. Geometry-induced capillary emptying

    PubMed Central

    Parry, Andrew O.; Aarts, Dirk G. A. L.

    2016-01-01

    When a capillary is half-filled with liquid and turned to the horizontal, the liquid may flow out of the capillary or remain in it. For lack of a better criterion, the standard assumption is that the liquid will remain in a capillary of narrow cross-section, and will flow out otherwise. Here, we present a precise mathematical criterion that determines which of the two outcomes occurs for capillaries of arbitrary cross-sectional shape, and show that the standard assumption fails for certain simple geometries, leading to very rich and counterintuitive behavior. This opens the possibility of creating very sensitive microfluidic devices that respond readily to small physical changes, for instance, by triggering the sudden displacement of fluid along a capillary without the need of any external pumping. PMID:27791079

  5. Physiological factors influencing capillary growth.

    PubMed

    Egginton, S

    2011-07-01

    (1) Angiogenesis (growth of new capillaries from an existing capillary bed) may result from a mismatch in microvascular supply and metabolic demand (metabolic error signal). Krogh examined the distribution and number of capillaries to explore the correlation between O(2) delivery and O(2) consumption. Subsequently, the heterogeneity in angiogenic response within a muscle has been shown to reflect either differences in fibre type composition or mechanical load. However, local control leads to targetted angiogenesis in the vicinity of glycolytic fibre types following muscle stimulation, or oxidative fibres following endurance training, while heterogeneity of capillary spacing is maintained during ontogenetic growth. (2) Despite limited microscopy resolution and lack of specific markers, Krogh's interest in the structure of the capillary wall paved the way for understanding the mechanisms of capillary growth. Angiogenesis may be influenced by the response of perivascular or stromal cells (fibroblasts, macrophages and pericytes) to altered activity, likely acting as a source for chemical signals modulating capillary growth such as vascular endothelial growth factor. In addition, haemodynamic factors such as shear stress and muscle stretch play a significant role in adaptive remodelling of the microcirculation. (3) Most indices of capillarity are highly dependent on fibre size, resulting in possible bias because of scaling. To examine the consequences of capillary distribution, it is therefore helpful to quantify the area of tissue supplied by individual capillaries. This allows the spatial limitations inherent in most models of tissue oxygenation to be overcome generating an alternative approach to Krogh's tissue cylinder, the capillary domain, to improve descriptions of intracellular oxygen diffusion. © 2010 The Author. Acta Physiologica © 2010 Scandinavian Physiological Society.

  6. Capillary electrophoresis of carbohydrates.

    PubMed

    Oefner, P J; Chiesa, C

    1994-08-01

    Capillary electrophoresis has emerged as a highly promising technique for the analysis of mono- and oligosaccharides. The approaches developed for overcoming the lack of chromophoric and fluorophoric functions in most carbohydrates involve the use of indirect photometric detection, amperometry, mass spectrometry, and precolumn derivatization with various tags. The merits and drawbacks of the derivatizing agents, including 2-aminopyridine, 4-amino-benzoic acid and its analogues, which for the first time permitted the reproducible determination of aldoses, uronic acids and even ketoses in the low femtomole range by means of readily available UV detection, and other agents such as 8-aminonaphthalene-1,3,6-trisulphonic acid, 1-phenyl-3-methyl-5-pyrazolone and 3-(4-carboxybenzoyl)-2-quinoline-carboxaldehyde, are discussed in detail. Means to secure electromigration of the usually neutral carbohydrates are: (i) ionization of hydroxyl groups at high pH; (ii) complexation of vicinal or alternate hydroxyl groups with borate or other charged compounds such as alkaline earth metal ions; (iii) derivatization with a reagent possessing ionizable functions; and (iv) partitioning into a pseudostationary phase such as sodium dodecyl sulphate micelles. Each alternative has its own analytical rewards, and combinations of the above mechanisms allow the two-dimensional and perhaps even three-dimensional mapping of oligosaccharides. Pyridylaminated oligosaccharides, for instance, have been separated both according to size by exploiting differences in the charge-to-mass ratio, with the charge being identical for each oligomer under acidic conditions due to protonation of the imino group incorporated by precolumn derivatization, as well as on the basis of structural differences, as a consequence of differences in the ease of borate complexation of the peripheral monosaccharide residues. It is also shown that the 4-aminobenzonitrile derivatives of mono- and disaccharides can be separated

  7. Fc receptor density, MHC antigen expression and superoxide production are increased in interferon-gamma-treated microglia isolated from adult rat brain.

    PubMed Central

    Woodroofe, M N; Hayes, G M; Cuzner, M L

    1989-01-01

    Treatment of microglia isolated from adult rat brain with interferon-gamma (IFN-gamma) at a concentration of 1 U/ml resulted in enhanced expression of Fc receptors and major histocompatibility complex (MHC) antigens and increased production of superoxide anions. Neonatal microglia and peritoneal macrophages, isolated and cultured in the same manner, displayed functional properties very similar to those of adult microglia, indicating a common origin for different macrophage populations. The Fc binding capacity of microglia was found to be significantly greater than that of peritoneal cells, thus underlining the potential role of microglia in immune-mediated demyelination. PMID:2556346

  8. Multidimensional capillary electrophoresis.

    PubMed

    Grochocki, Wojciech; Markuszewski, Michał J; Quirino, Joselito P

    2015-01-01

    Multidimensional separation where two or more orthogonal displacement mechanisms are combined is a promising approach to increase peak capacity in CE. The combinations allow dramatic improvement of analytical performance since the total peak capacity is given by a product of the peak capacities of all methods. The initial reports were concentrated on the construction of effective connections between capillaries for 2D analysis. Today, 2D and 3D CE systems are now able to separate real complex biological or environmental mixtures with good repeatability, improved resolution with minimal loss of sample. This review will present the developments in the field of multidimensional CE during the last 15 years. The endeavors in this specific field were on the development of interfaces, interface-free techniques including integrated separations, microdevices, and on-line sample concentration techniques to improve detection sensitivity.

  9. Capillary electrophoresis of DNA.

    PubMed

    Smith, Alan; Nelson, Robert J

    2003-08-01

    Capillary electrophoresis (CE) is an alternative to conventional slab gel electrophoresis for the separation of DNA fragments. CE offers a number of advantages over slab gel separations in terms of speed, resolution, sensitivity, and data handling. Separation times are generally only a few minutes and the DNA is detected either by UV absorption or by fluorescent labeling. The quantity of DNA required for separation is in the nanogram range. Single-base resolution can be obtained on fragments up to several hundred base pairs. In the presence of appropriate standards, fragments can be accurately sized based on relative electrophoretic mobility. A protocol for the analysis of synthetic oligonucleotides in a flowable matrix is described in this unit.

  10. A capillary Archimedes' screw

    NASA Astrophysics Data System (ADS)

    Darbois Texier, Baptiste; Dorbolo, Stephane

    2014-11-01

    As used by Egyptians for irrigation and reported by Archimedes, a screw turning inside a hollow pipe can pull out a fluid againt gravity. At a centimetric scale, an analagous system can be found with a drop pending on a rotating spiral which is tilted toward the horizontal. The ascent of the drop to the top of the spiral is considered and a theoretical model based on geometrical considerations is proposed. The climb of the drop is limited by the fluid deposition on the screw at high capillary number and by a centrifugation phenomenon. We find out the range of fluid proprities and spiral characteristics for which an ascending motion of the drop is possible. Finally we discuss the efficiency of such system to extract a fluid from a bath at a centrimetric scale.

  11. Steady Capillary Driven Flow

    NASA Technical Reports Server (NTRS)

    Weislogel, Mark M.

    1996-01-01

    A steady capillary driven flow is developed for a liquid index in a circular tube which is partially coated with a surface modifier to produce a discontinuous wetting condition from one side of the tube to the other. The bulk flow is novel in that it is truly steady, and controlled solely by the physics associated with dynamic wetting. The influence of gravity on the flow is minimized through the use of small diameter tubes approximately O(1 mm) tested horizontally in a laboratory and larger tubes approximately O(10 mm) tested in the low gravity environment of a drop tower. Average steady velocities are predicted and compared against a large experimental data set which includes the effects of tube dimensions and fluid properties. The sensitivity of the velocity to surface cleanliness is dramatic and the advantages of experimentation in a microgravity environment are discussed.

  12. Capillaries for use in a multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, Edward S.; Chang, Huan-Tsang; Fung, Eliza N.

    1997-12-09

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification ("base calling") is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations.

  13. Capillaries for use in a multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, E.S.; Chang, H.T.; Fung, E.N.

    1997-12-09

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification (``base calling``) is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations. 19 figs.

  14. Abnormal Capillary Vasodynamics Contribute to Ictal Neurodegeneration in Epilepsy.

    PubMed

    Leal-Campanario, Rocio; Alarcon-Martinez, Luis; Rieiro, Hector; Martinez-Conde, Susana; Alarcon-Martinez, Tugba; Zhao, Xiuli; LaMee, Jonathan; Popp, Pamela J Osborn; Calhoun, Michael E; Arribas, Juan I; Schlegel, Alexander A; Stasi, Leandro L Di; Rho, Jong M; Inge, Landon; Otero-Millan, Jorge; Treiman, David M; Macknik, Stephen L

    2017-02-27

    Seizure-driven brain damage in epilepsy accumulates over time, especially in the hippocampus, which can lead to sclerosis, cognitive decline, and death. Excitotoxicity is the prevalent model to explain ictal neurodegeneration. Current labeling technologies cannot distinguish between excitotoxicity and hypoxia, however, because they share common molecular mechanisms. This leaves open the possibility that undetected ischemic hypoxia, due to ictal blood flow restriction, could contribute to neurodegeneration previously ascribed to excitotoxicity. We tested this possibility with Confocal Laser Endomicroscopy (CLE) and novel stereological analyses in several models of epileptic mice. We found a higher number and magnitude of NG2+ mural-cell mediated capillary constrictions in the hippocampus of epileptic mice than in that of normal mice, in addition to spatial coupling between capillary constrictions and oxidative stressed neurons and neurodegeneration. These results reveal a role for hypoxia driven by capillary blood flow restriction in ictal neurodegeneration.

  15. Abnormal Capillary Vasodynamics Contribute to Ictal Neurodegeneration in Epilepsy

    PubMed Central

    Leal-Campanario, Rocio; Alarcon-Martinez, Luis; Rieiro, Hector; Martinez-Conde, Susana; Alarcon-Martinez, Tugba; Zhao, Xiuli; LaMee, Jonathan; Popp, Pamela J. Osborn; Calhoun, Michael E.; Arribas, Juan I.; Schlegel, Alexander A.; Stasi, Leandro L. Di; Rho, Jong M.; Inge, Landon; Otero-Millan, Jorge; Treiman, David M.; Macknik, Stephen L.

    2017-01-01

    Seizure-driven brain damage in epilepsy accumulates over time, especially in the hippocampus, which can lead to sclerosis, cognitive decline, and death. Excitotoxicity is the prevalent model to explain ictal neurodegeneration. Current labeling technologies cannot distinguish between excitotoxicity and hypoxia, however, because they share common molecular mechanisms. This leaves open the possibility that undetected ischemic hypoxia, due to ictal blood flow restriction, could contribute to neurodegeneration previously ascribed to excitotoxicity. We tested this possibility with Confocal Laser Endomicroscopy (CLE) and novel stereological analyses in several models of epileptic mice. We found a higher number and magnitude of NG2+ mural-cell mediated capillary constrictions in the hippocampus of epileptic mice than in that of normal mice, in addition to spatial coupling between capillary constrictions and oxidative stressed neurons and neurodegeneration. These results reveal a role for hypoxia driven by capillary blood flow restriction in ictal neurodegeneration. PMID:28240297

  16. Exercise-induced changes of the capillaries in the cortex of middle-aged rats.

    PubMed

    Huang, C-X; Qiu, X; Wang, S; Wu, H; Xia, L; Li, C; Gao, Y; Zhang, L; Xiu, Y; Chao, F; Tang, Y

    2013-03-13

    Previous studies have shown that running exercise could increase regional cerebral blood flow. There have been previous studies investigating the effects of running exercise on capillary density in the brain and showing that running exercise could induce brain angiogenesis. However, there have been no studies investigating the effects of running exercise on the total volume, total length and total surface area of the capillaries in the cortex. Moreover, sex differences in the effects of running exercise on the capillaries of the cortex have not previously been investigated. The current study was designed to investigate the effects of running exercise on the capillaries in the cortex of middle-aged rats using the new unbiased stereological methods. The present study found that the total length and total surface area of the capillaries in the cortex of running middle-aged female rats were significantly increased, compared to control rats. Our results also reveal that there are sex differences in the effects of running exercise on the capillaries in the cortex of middle-aged rats. These results demonstrate that exercise-induced increases of the capillaries in the female rat cortex might be one of the structural bases for the exercise-induced improvement in the spatial learning capacity of middle-aged female rats. These results provide a baseline for further studies that search for strategies to delay the deleterious effects of brain aging.

  17. Inertial Rise in Short Capillaries

    NASA Astrophysics Data System (ADS)

    Shardt, Orest; Waghmare, Prashant; Mitra, Sushanta; Derksen, Jos

    2013-11-01

    We investigate the primarily inertial rise of liquid in vertical glass capillaries that are shorter than the equilibrium rise height (Jurin height). We focus on the behavior of the liquid upon reaching the top of the capillary and use high-speed imaging to observe the motion of the liquid-air interface with high spatial and temporal resolution. We examine the dependence of the interface behavior on the meniscus speed and capillary height and describe a new phenomenon. Upon reaching the upper edge of a sufficiently short capillary, the meniscus inverts, rises upward, and bulges out radially. The bulging liquid then wets the external surface of the capillary and slides down. The meniscus inside the capillary retracts, falling below the upper edge, and then oscillates vertically with decaying amplitude, inverting several times before reaching a steady shape. A theoretical analysis is used to interpret the conditions required for this phenomenon to occur. A key assumption in the analysis is that the transient flow is inertial and therefore the capillary driving force is balanced by the weight and inertia of the rising liquid column while viscous forces are comparatively small. The analysis points to the possibility of obtaining previously-unseen behavior under reduced gravity.

  18. Instability of the capillary bridge

    NASA Astrophysics Data System (ADS)

    Pare, Gounseti; Hoepffner, Jerome

    2014-11-01

    Capillary adhesion is a physical mechanism that maintains two bodies in contact by capillarity through a liquid ligament. The capillary bridge is an idealization of this capillary adhesion. In this study we first focus on the classical case of the stability of the capillary bridge. Secondly we study a slightly more complex configuration, imagining a flow in the capillary bridge as in the case of the dynamics of the neck of a liquid ligament, in its withdrawal under the effect of capillarity. Inspired by the experiments on soap films of Plateau, the configuration analyzed consists of an initially axisymmetric, mass of fluid held by surface tension forces between two parallel, coaxial, solid pipes of the same diameter. The results presented are obtained by numerical simulations using the free software, Gerris Flow Solver. We first focus on the capillary Venturi. In the static configuration the stability diagram of the capillary bridge obtained is in perfect agreement with the results of Lev A. Slobozhanin. In the dynamic case we develop a matlab code based on the one dimensional equations of Eggers and Dupont. The comparison of the bifurcation diagram obtained and the numerical simulations shows a good agreement.

  19. Detection of free radicals by isolated perfusion of the rat brain following hemorrhagic stroke: a novel approach to cerebrovascular biomarker research.

    PubMed

    Grienberger, Hubert J; Pillai, Deepu R; Schlachetzki, Felix; Gruber, Michael; Dittmar, Michael S

    2010-10-01

    Blood-borne biomarkers are a mainstay of diagnosis and follow-up in many diseases. For stroke, however, no reliable biomarkers have thus far been identified. To remedy this situation, we investigated the usefulness of a modified in situ isolated brain perfusion (IBP) technique for screening potential biomarker candidates. As a proof of concept, the production of reactive oxygen species (ROS) was estimated in a rat model of experimental intracerebral hemorrhage (ICH). After stereotactic infusion of whole blood into the rat striatum, we initiated IBP without intracranial manipulation or discontinuation of cerebral blood flow. To detect ROS, we employed the salicylate trapping method, which involves the hydroxylation of salicylic acid during oxidative stress into dihydroxybenzoic acid (DHBA), and quantification of the latter in venous eluate by using high-performance liquid chromatography. Venous eluate was collected separately from both injured and healthy hemispheres (n=10). Control groups consisted of sham-injured (n=4) and healthy animals (n=3). In animals subjected to ICH (n=10), 50% more 2,5-DHBA was detected in venous eluate on the injured side than in eluate on the contralateral side. Hemorrhagic hemispheres produced more 2,5-DHBA than hemispheres in sham-injured and healthy animals (72 and 110% more 2,5-DHBA, respectively). Isolated brain perfusion combined with salicylate trapping produced data indicating an elevation in the formation of ROS subsequent to ICH. Our findings suggest that isolated in situ brain perfusion is a promising approach to detecting biomarkers of cerebrovascular pathologic conditions.

  20. Acute traumatic coagulopathy in the setting of isolated traumatic brain injury: a systematic review and meta-analysis.

    PubMed

    Epstein, Daniel S; Mitra, Biswadev; O'Reilly, Gerard; Rosenfeld, Jeffrey V; Cameron, Peter A

    2014-05-01

    Acute traumatic coagulopathy (ATC) has been reported in the setting of isolated traumatic brain injury (iTBI) and associated with high mortality and poor outcomes. The aim of this systematic review was to examine the incidence and outcome of patients with ATC in the setting of iTBI. We conducted a search of the MEDLINE database and Cochrane library, focused on subject headings and keywords involving coagulopathy and TBI. Design and results of each study were described. Studies were assessed for heterogeneity and the pooled incidence of ATC in the setting of iTBI determined. Reported outcomes were described. There were 22 studies selected for analysis. A statistically significant heterogeneity among the studies was observed (p<0.01). Using the random effects model the pooled proportion of patients with ATC in the setting of iTBI was 35.2% (95% CI: 29.0-41.4). Mortality of patients with ATC and iTBI ranged between 17% and 86%. Higher blood transfusion rates, longer hospital stays, longer ICU stays, decreased ventilator free days, higher rates of single and multiple organ failure and higher incidence of delayed injury and disability at discharge were reported among patients with ATC. ATC is commonly associated with iTBI and almost uniformly associated with worse outcomes. Any disorder of coagulation above the normal range appears to be associated with worse outcomes and therefore a clinically important target for management. Earlier identification of patients with ATC and iTBI, for recruitment into prospective trials, presents avenues for further research. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Endothelial cell heterogeneity of blood-brain barrier gene expression along the cerebral microvasculature.

    PubMed

    Macdonald, Jennifer A; Murugesan, Nivetha; Pachter, Joel S

    2010-05-15

    The blood-brain barrier (BBB) refers to the network of microvessels that selectively restricts the passage of substances between the circulation and the central nervous system (CNS). This microvascular network is comprised of arterioles, capillaries and venules, yet the respective contribution of each of these to the BBB awaits clarification. In this regard, it has been postulated that brain microvascular endothelial cells (BMEC) from these different tributaries might exhibit considerable heterogeneity in form and function, with such diversity underlying unique roles in physiological and pathophysiological processes. Means to begin exploring such endothelial differences in situ, free from caveats associated with cell isolation and culturing procedures, are crucial to comprehending the nature and treatment of CNS diseases with vascular involvement. Here, the recently validated approach of immuno-laser capture microdissection (immuno-LCM) coupled to quantitative real-time PCR (qRT-PCR) was used to analyze gene expression patterns of BMEC retrieved in situ from either capillaries or venules. From profiling 87 genes known to play a role in BBB function and/or be enriched in isolated brain microvessels, results imply that most BBB properties reside in both segments, but that capillaries preferentially express some genes related to solute transport, while venules tend toward higher expression of an assortment of genes involved in inflammatory-related tasks. Fuller appreciation of such heterogeneity will be critical for efficient therapeutic targeting of the endothelium and the management of CNS disease.

  2. Integrated multiplexed capillary electrophoresis system

    DOEpatents

    Yeung, Edward S.; Tan, Hongdong

    2002-05-14

    The present invention provides an integrated multiplexed capillary electrophoresis system for the analysis of sample analytes. The system integrates and automates multiple components, such as chromatographic columns and separation capillaries, and further provides a detector for the detection of analytes eluting from the separation capillaries. The system employs multiplexed freeze/thaw valves to manage fluid flow and sample movement. The system is computer controlled and is capable of processing samples through reaction, purification, denaturation, pre-concentration, injection, separation and detection in parallel fashion. Methods employing the system of the invention are also provided.

  3. Capillary oscillations on liquid jets

    NASA Astrophysics Data System (ADS)

    Wetsel, Grover C.

    1980-07-01

    Capillary oscillations on modulated liquid jets have been investigated using laser illumination and electronic detection of the magnified jet shadow. The amplitudes of several wave harmonics of a growing spatial instability were measured as a function of distance from the orifice for a range of jet velocities and initial-disturbance amplitudes. The experimentally determined growth rates at the fundamental frequency are compared with theories of capillary-wave propagation. An empirically derived explanation of the suppression of satellite formation is given. Experimental evidence for infinite-wavelength capillary oscillations is reported; a description of these oscillations in terms of the Rayleigh theory is presented.

  4. Electrospraying from nanofluidic capillary slot

    NASA Astrophysics Data System (ADS)

    Arscott, Steve; Troadec, David

    2005-09-01

    We present here an original electrospray emitter tip based on a nanofluidic capillary slot. The nanofabrication involves focused ion beam etching to form the slot which has a cross-section of 50×300nm and a length of 4μm. The liquid is deformed into the nanofluidic capillary slot by capillary action; the electrospray is produced by the application of a low voltage. Using a water-methanol-formic acid solution, we have observed an electrospray onset voltage of 125V. At a measured electrospray current of 1nA, the unforced flow-rate is estimated to be 110pLmin-1.

  5. Analysis of Capillary Rise in Asymmetric Branch-Like Capillary

    NASA Astrophysics Data System (ADS)

    Li, Caoxiong; Shen, Yinghao; Ge, Hongkui; Yang, Zhihui; Su, Shuai; Ren, Kai; Huang, Heyu

    2016-05-01

    Transport in porous media is common in nature, attracting many attentions for a long time. Tree-like network model is often used as a simplification for porous space, expressing the complexity of pore spaces instead of capillary bundle. To investigate spontaneous imbibition characteristics in this network, a dynamic asymmetric branch-like capillary model is used to represent basic network structure, using fractal method to represent tortuosity. This work investigates the influence of parameters on imbibition process in the branch-like capillary model. An analytical equation for the imbibition mass versus time is derived. Parameters from capillary structures to liquid properties are taken into account and analyzed based on the numerical solution of the equation. It is found that the imbibition process in asymmetric branch-like capillary model can be recognized by four sections and brunching tubes are positive for imbibition process. Concomitantly, meniscus arrest event is simulated and discussed. Moreover, the influence of parameters on imbibition process is discussed. These parameters can be classified as static and dynamic. Static parameters mainly change the capillary force, which are related to the ultimate imbibition mass or imbibition ability, while dynamic parameters mainly have influence on resistance of flowing fluid, which are related to the imbibition speed in the imbibition process.

  6. Symptomatic capillary telangiectasia of the pons and intracerebral developmental venous anomaly - A rare association.

    PubMed

    Sarac, Helena; Telarović, Srdana; Markeljević, Jasenka; Perić, Berislav; Pavlisa, Goran; Rados, Marko

    2011-01-01

    Various combinations of vascular malformations of the brain in one lesion have been reported, while others seem to be very rare. In this report, the authors discuss the case of a coexistence of an capillary telangiectasia of the pons and intracerebral venous anomaly. To our knowledge, this is the first report of coexistence of a capillary telangiectasia of the pons and intracerebral venous anomaly apparted from each other. These discrete vascular malformations of the brain raise attention on possible interrelations in the pathogenesis of these entities. We report a case of pontine capillary telangiectasia and intracerebral venous anomaly in a 42-year-old woman with a right side facial palsy. Hight field magnetic resonance imaging suggested presence of a capillary telangiectasia of the pons. Another lesion in the left frontal gyrus was attributable to the venous anomaly. Along with neuroradiological findings, results of the somatosensor evoked potentials, brain stem auditory potentials, laboratory analysis including blood, cerebrospinal fluid and urine investigation are demonstrated. Awareness of the magnetic resonance imaging finding of the capillary telangiectasias and of the venous anomalies may help in defining clinical correlates of this vascular malformations, while the follow up of these malformations might help to asses risk of vascular rupture. We and others previously selects capillary telangiectasia and venous anomaly in two discrete entities. Coexistence of these malformations in the brain apparted from each other appear to be very rare and raise attention on possible interactions in their natural history and pathogenesis.

  7. Aralar mRNA and protein levels in neurons and astrocytes freshly isolated from young and adult mouse brain and in maturing cultured astrocytes.

    PubMed

    Li, Baoman; Hertz, Leif; Peng, Liang

    2012-12-01

    Intense glucose-based energy metabolism and glutamate synthesis by astrocytes require malate-aspartate-shuttle (MAS) activity to regenerate NAD⁺ from NADH formed during glycolysis, since brain lacks significant glycerophosphate shuttle activity. Aralar is a necessary aspartate/glutamate exchanger for MAS function in brain. Based on cytochemical immunoassays the absence of aralar in adult astrocytes was repeatedly reported. This would mean that adult astrocytes must regenerate NAD⁺ by producing lactate from pyruvate, eliminating its use by oxidative and biosynthetic pathways. We alternatively used astrocytes and neurons from adult brain, freshly isolated by fluorescence-activated cell sorting, to determine aralar protein by a specific antibody and its mRNA by real-time PCR. Both protein and mRNA expressions were identical in adult neurons and astrocytes and similar to whole brain levels. The same level of aralar expression was reached in well-differentiated astrocyte cultures, but not until late development, coinciding with the late-maturing brain capability for glutamate formation and degradation. Copyright © 2012 Elsevier Ltd. All rights reserved.

  8. Capillary Fracturing in Granular Media

    NASA Astrophysics Data System (ADS)

    Holtzman, Ran; Szulczewski, Michael L.; Juanes, Ruben

    2012-06-01

    We study the displacement of immiscible fluids in deformable, noncohesive granular media. Experimentally, we inject air into a thin bed of water-saturated glass beads and observe the invasion morphology. The control parameters are the injection rate, the bead size, and the confining stress. We identify three invasion regimes: capillary fingering, viscous fingering, and “capillary fracturing,” where capillary forces overcome frictional resistance and induce the opening of conduits. We derive two dimensionless numbers that govern the transition among the different regimes: a modified capillary number and a fracturing number. The experiments and analysis predict the emergence of fracturing in fine-grained media under low confining stress, a phenomenon that likely plays a fundamental role in many natural processes such as primary oil migration, methane venting from lake sediments, and the formation of desiccation cracks.

  9. Capillary fracturing in granular media.

    PubMed

    Holtzman, Ran; Szulczewski, Michael L; Juanes, Ruben

    2012-06-29

    We study the displacement of immiscible fluids in deformable, noncohesive granular media. Experimentally, we inject air into a thin bed of water-saturated glass beads and observe the invasion morphology. The control parameters are the injection rate, the bead size, and the confining stress. We identify three invasion regimes: capillary fingering, viscous fingering, and "capillary fracturing," where capillary forces overcome frictional resistance and induce the opening of conduits. We derive two dimensionless numbers that govern the transition among the different regimes: a modified capillary number and a fracturing number. The experiments and analysis predict the emergence of fracturing in fine-grained media under low confining stress, a phenomenon that likely plays a fundamental role in many natural processes such as primary oil migration, methane venting from lake sediments, and the formation of desiccation cracks.

  10. Surface Tension and Capillary Rise

    ERIC Educational Resources Information Center

    Walton, Alan J.

    1972-01-01

    Discussion of the shortcomings of textbook explanations of surface tension, distinguishing between concepts of tension and capillary rise. The arguments require only a clear understanding of Newtonian mechanics, notably potential energy. (DF)

  11. DNA typing by capillary electrophoresis

    SciTech Connect

    Zhang, N.

    1997-10-08

    Capillary electrophoresis is becoming more and more important in nucleic acid analysis including DNA sequencing, typing and disease gene measurements. This work summarized the background of DNA typing. The recent development of capillary electrophoresis was also discussed. The second part of the thesis showed the principle of DNA typing based on using the allelic ladder as the absolute standard ladder in capillary electrophoresis system. Future work will be focused on demonstrating DNA typing on multiplex loci and examples of disease diagnosis in the on-line format of PCR-CE. Also capillary array electrophoresis system should allow high throughput, fast speed DNA typing. Only the introduction and conclusions for this report are available here. A reprint was removed for separate processing.

  12. Simultaneous two-photon imaging of cerebral oxygenation and capillary blood flow in atherosclerotic mice

    NASA Astrophysics Data System (ADS)

    Lu, Xuecong; Li, Baoqiang; Moeini, Mohammad; Lesage, Frédéric

    2017-02-01

    Gradual changes in brain microvasculature and cerebral capillary blood flow occurring with atherosclerosis may significantly contribute to cognition decline due to their role in brain tissue oxygenation. However, previous stud- ies of the relationship between cerebral capillary blood flow and brain tissue oxygenation are limited. This study aimed to investigate vascular and concomitant changes in brain tissue pO2 with atherosclerosis. Experiments in young healthy C57B1/6 mice (n=6 , WT), young atherosclerotic mice (n=6 , ATX Y) and old atherosclerotic mice (n=6 , ATX O) were performed imaging on the left sensory-motor cortex at resting state under urethane (1.5 g/kg) anesthesia using two-photon fluorescence microscopy. The results showed that pO2 around capillaries, correlated with red blood cell (RBC) flux, increased with atherosclerosis.

  13. Capillary electrophoresis electrospray ionization mass spectrometry interface

    DOEpatents

    Smith, Richard D.; Severs, Joanne C.

    1999-01-01

    The present invention is an interface between a capillary electrophoresis separation capillary end and an electrospray ionization mass spectrometry emitter capillary end, for transporting an anolyte sample from a capillary electrophoresis separation capillary to a electrospray ionization mass spectrometry emitter capillary. The interface of the present invention has: (a) a charge transfer fitting enclosing both of the capillary electrophoresis capillary end and the electrospray ionization mass spectrometry emitter capillary end; (b) a reservoir containing an electrolyte surrounding the charge transfer fitting; and (c) an electrode immersed into the electrolyte, the electrode closing a capillary electrophoresis circuit and providing charge transfer across the charge transfer fitting while avoiding substantial bulk fluid transfer across the charge transfer fitting. Advantages of the present invention have been demonstrated as effective in providing high sensitivity and efficient analyses.

  14. Capillary gas-solid chromatography

    NASA Astrophysics Data System (ADS)

    Berezkin, Viktor G.

    1996-11-01

    The current state of gas adsorption chromatography on open tubular capillary columns is analysed. The history of the development of this method and its role in gas chromatography are considered. The preparation of open tubular adsorption capillary columns, fundamentals of the theory of retention and of broadening of chromatographic zones, and the use of columns of this type in practical analytical chemistry are discussed. The bibliography includes 148 references.

  15. Effect of hypothermic pulmonary artery flushing on capillary filtration coefficient.

    PubMed

    Andrade, R S; Wangensteen, O D; Jo, J K; Tsai, M Y; Bolman, R M

    2000-07-27

    We previously demonstrated that surfactant dilution and inhibition occur immediately after pulmonary artery flushing with hypothermic modified Euro-Collins solution. Consequently, we speculated that increased capillary permeability contributed to these surfactant changes. To test this hypothesis, we evaluated the effects of hypothermic pulmonary artery flushing on the pulmonary capillary filtration coefficient (Kfc), and additionally performed a biochemical analysis of surfactant. We used a murine isolated, perfused lung model to measure the pulmonary capillary filtration coefficient and hemodynamic parameters, to determine the wet to dry weight ratio, and to evaluate surfactant by biochemical analysis of lung lavage fluid. We defined three study groups. In group I (controls), we harvested lungs without hypothermic pulmonary artery flushing, and measured Kfc immediately. In group II (in situ flush), we harvested lungs after hypothermic pulmonary artery flushing with modified Euro-Collins solution, and then measured Kfc. Experiments in groups I and II were designed to evaluate persistent changes in Kfc after pulmonary artery flushing. In group III (ex vivo flush), we flushed lungs ex vivo to evaluate transient changes in Kfc during hypothermic pulmonary artery flushing. Groups I and II did not differ significantly in capillary filtration coefficient and hemodynamics. Group II showed significant alterations on biochemical surfactant analysis and a significant increase in wet-to-dry weight ratio, when compared with group I. In group III, we observed a significant transient increase in capillary filtration coefficient during pulmonary artery flushing. Hypothermic pulmonary artery flushing transiently increases the capillary filtration coefficient, leads to an increase in the wet to dry weight ratio, and induces biochemical surfactant changes. These findings could be explained by the effects of hypothermic modified Euro-Collins solution on pulmonary capillary

  16. Capillary viscosimetry on ferrofluids.

    PubMed

    Pop, L M; Odenbach, S

    2008-05-21

    Experiments performed for different ferrofluids under shear flow have shown that an increase of the magnetic field strength applied to the sample yields an increase of the fluid's viscosity, the so called magnetoviscous effect. It has been shown that the magnitude of the effect is strongly related to the modification of the microstructure of ferrofluids and can be influenced by varying both the dipole-dipole interaction between the particles and the concentration of large particles within the fluid. This result has been further used to synthesize new ferrofluids which, on one hand, are more compatible for technical applications but, on the other hand, led to difficulties for the experimenters in measuring the viscous behavior in the presence of a magnetic field. To overcome this problem, a specially designed ferrofluid-compatible capillary viscometer has been developed. Within this paper, the experimental setup as well as experimental results concerning the investigation of the magnetoviscous effect in both diluted and concentrated cobalt-based ferrofluids are presented.

  17. Systemic platelet dysfunction is the result of local dysregulated coagulation and platelet activation in the brain in a rat model of isolated traumatic brain injury.

    PubMed

    Ploplis, Victoria A; Donahue, Deborah L; Sandoval-Cooper, Mayra J; MorenoCaffaro, Maria; Sheets, Patrick; Thomas, Scott G; Walsh, Mark; Castellino, Francis J

    2014-10-01

    Coagulopathy after severe traumatic brain injury (TBI) has been extensively reported. Clinical studies have identified a strong relationship between diminished platelet-rich thrombus formation, responsiveness to adenosine diphosphate agonism, and severity of TBI. The mechanisms that lead to platelet dysfunction in the acute response to TBI are poorly understood. The development of a rodent model of TBI that mimics the coagulopathy observed clinically has recently been reported. Using immunohistochemical techniques and thromboelastography platelet mapping, the current study demonstrated that the expression of coagulation (tissue factor and fibrin) and platelet activation (P-selectin) markers in the injured brain paralleled the alteration in systemic platelet responsiveness to the agonists, adenosine diphosphate and arachodonic acid. Results of this study demonstrate that local procoagulant changes in the injured brain have profound effects on systemic platelet function.

  18. Ultrasensitive quantification of cerebral capillary flow networks and dynamics

    NASA Astrophysics Data System (ADS)

    You, Jiang; Park, Ki; Du, Congwu; Pan, Yingtian

    2015-03-01

    Ultra-high resolution optical Doppler coherence tomography (μODT) is a promising tool for brain functional imaging. However, its sensitivity for detecting slow flows in capillary beds may limit its utility in visualizing and quantifying subtle changes in brain microcirculation. To address this limitation, we developed a novel method called contrast-enhanced μODT (c-μODT) in which intralipid is injected into mouse tail vein to enhance μODT detection sensitivity. We demonstrate that after intralipid injection, the flow detection sensitivity of μODT is dramatically enhanced by 230% as quantified by the fill factor (FF) of microvasculature. More importantly, we show that c-μODT preserves the quantitative properties for flow imaging, i.e., showing a comparable change ratio of hypercapnia-induced flow increase in the capillary network before and after injecting intralipid.

  19. Human and mouse brain-derived endothelial cells require high levels of growth factors medium for their isolation, in vitro maintenance and survival

    PubMed Central

    2013-01-01

    Background Brain microvascular endothelial cells (BMVECs) constitute the primary limitation for passage of ions and molecules from the blood into the brain through the blood brain barrier. Numerous multi-step procedures for isolating and culturing BMVECs have been described. However, each one demonstrates major limitations in purity of culture and/or low proliferation rate. Our goal was to study the efficiency of our pending patent medium, Endothelial Proliferation Medium (EndoPM), on the isolation and purification of human and murine BMVECs. Methods BMVECs, cultured in EndoPM were compared to those cultured in a commercial medium EBM. Cultures were characterized by flow cytometric analysis, lineage differentiation, the ability to form tube-like structure, immunofluorescence, molecular analyses and also in an in vivo model assay. Moreover permeability was assayed by monitoring the passage of Dextran-FITC through a tight monolayer of BMVECs grown to confluence in Boyden chambers. One way Anova two-tailed test was utilized for all statistical analyses. Results The properties of ECs in human and murine BMVECs is confirmed by the expression of endothelial markers (CD31, CD105, CD146, Tie-2 and vWF), of representative proangiogenic genes (ICAM1, VCAM1 and integrin ITGAV), of considerable tube-forming ability, with low-density lipoprotein uptake, eNOS and GLUT-1 expression. Furthermore cells are able to express markers of the junctional architecture as VE-cadherin, β-catenin and Claudin-5 and greatly reduce dextran permeability as barrier functional test. Moreover BMVECs spontaneously organize in vascular-like structures and maintain the expression of endothelial markers in an in vivo xenograft model assay. The significant effect of EndoPM is confirmed by the study of proliferation index, survival index and the behaviour of BMVECs and fibroblasts in co-culture conditions. Conclusion Herein we describe a simple and reproducible method for the isolation and expansion of

  20. Seizure-like discharges induced by 4-aminopyridine in the olfactory system of the in vitro isolated guinea pig brain

    PubMed Central

    Uva, Laura; Trombin, Federica; Carriero, Giovanni; Avoli, Massimo; de Curtis, Marco

    2016-01-01

    Summary Purpose The study of the interactions leading to network- or region-specific propagation of seizures is crucial to understand ictogenesis. We have recently found that systemic (arterial) application of the potassium channel blocker, 4-aminopyridine (4AP), induces different and independent seizure activities in olfactory and in limbic structures. Here, we have characterized the network and cellular features that support 4AP-induced seizure-like events in the olfactory cortex. Methods Simultaneous extracellular recordings were performed from the piriform cortex, the entorhinal cortex, the olfactory tubercle, and the amygdala of the in vitro isolated guinea pig brain preparation. Intracellular, sharp electrode recordings were obtained from neurons of different layers of the region of ictal onset, the piriform cortex. Seizure-like discharges were induced by both arterial perfusion and local intracortical injections of 4AP. Key Findings Arterial application of 4AP induces independent seizure activities in limbic and olfactory cortices. Both local applications of 4AP and cortico-cortical disconnections demonstrated that region-specific seizure-like events initiated in the primary olfactory cortex and propagate to anatomically related areas. Seizures induced by arterial administration of 4-AP are preceded by runs of fast activity at circa 30–40 Hz and are independently generated in the hemispheres. Simultaneous extracellular and intracellular recordings in the piriform cortex revealed that the onset of seizure correlates with (1) a gradual amplitude increase of fast activity runs, (2) a large intracellular depolarization with action potential firing of superficial layer neurons, and (3) no firing in a subpopulation of deep layers neurons. During the ictal event, neuronal firing was abolished for 10–30 s in all neurons and gradually restored and synchronized before seizure termination. Significance Our data show that olfactory neuronal networks sustain the

  1. Rotenone exerts similar stimulatory effects on H2O2 production by isolated brain mitochondria from young-adult and old rats.

    PubMed

    Michelini, Luiz G B; Figueira, Tiago R; Siqueira-Santos, Edilene S; Castilho, Roger F

    2015-03-04

    Chronic and systemic treatment of rodents with rotenone, a classical inhibitor of mitochondrial respiratory complex I, results in neurochemical, behavioral, and neuropathological features of Parkinson's disease. The aim of the present study was to evaluate whether brain mitochondria from old rats (24 months old) would be more susceptible to rotenone-induced inhibition of oxygen consumption and increased generation of H2O2 than mitochondria from young-adult rats (3-4 months old). Isolated brain mitochondria were incubated in the presence of different rotenone concentrations (5, 10, and 100nM), and oxygen consumption and H2O2 production were measured during respiratory states 3 (ADP-stimulated respiration) and 4 (resting respiration). Respiratory state 3 and citrate synthase activity were significantly lower in mitochondria from old rats. Mitochondria from young-adult and old rats showed similar sensitivity to rotenone-induced inhibition of oxygen consumption. Similarly, H2O2 production rates by both types of mitochondria were dose-dependently stimulated to the same extent by increasing concentrations of rotenone. We conclude that rotenone exerts similar effects on oxygen consumption and H2O2 production by isolated brain mitochondria from young-adult and old rats. Therefore, aging does not increase the mitochondrial H2O2 generation in response to complex I inhibition. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  2. Human blood-brain barrier insulin-like growth factor receptor

    SciTech Connect

    Duffy, K.R.; Pardridge, W.M.; Rosenfeld, R.G.

    1988-02-01

    Insulin-like growth factor (IGF)-1 and IGF-2, may be important regulatory molecules in the CNS. Possible origins of IGFs in brain include either de novo synthesis or transport of circulating IGFs from blood into brain via receptor mediated transcytosis mechanisms at the brain capillary endothelial wall, ie, the blood-brain barrier (BBB). In the present studies, isolated human brain capillaries are used as an in vitro model system of the human BBB and the characteristics of IGF-1 or IGF-2 binding to this preparation were assessed. The total binding of IGF-2 at 37 degrees C exceeded 130% per mg protein and was threefold greater than the total binding for IGF-1. However, at 37 degrees C nonsaturable binding equaled total binding, suggesting that endocytosis is rate limiting at physiologic temperatures. Binding studies performed at 4 degrees C slowed endocytosis to a greater extent than membrane binding, and specific binding of either IGF-1 or IGF-2 was detectable. Scatchard plots for either peptide were linear and the molar dissociation constant of IGF-1 and IGF-2 binding was 2.1 +/- 0.4 and 1.1 +/- 0.1 nmol/L, respectively. Superphysiologic concentrations of porcine insulin inhibited the binding of both IGF-1 (ED50 = 2 micrograms/mL) and IGF-2 (ED50 = 0.5 microgram/mL). Affinity cross linking of /sup 125/I-IGF-1, /sup 125/I-IGF-2, and /sup 125/I-insulin to isolated human brain capillaries was performed using disuccinimidylsuberate (DSS). These studies revealed a 141 kd binding site for both IGF-1 and IGF-2, and a 133 kd binding site for insulin.

  3. Stimulation and release from neurons via a dual capillary collection device interfaced to mass spectrometry.

    PubMed

    Fan, Yi; Lee, Chang Young; Rubakhin, Stanislav S; Sweedler, Jonathan V

    2013-11-07

    Neuropeptides are cell to cell signaling molecules that modulate a wide range of physiological processes. Neuropeptide release has been studied in sample sizes ranging from single cells and neuronal clusters, to defined brain nuclei and large brain regions. We have developed and optimized cell stimulation and collection approaches for the efficient measurement of neuropeptide release from neuronal samples using a dual capillary system. The defining feature is a capillary that contains octadecyl-modified silica nanoparticles on its inner wall to capture and extract releasates. This collection capillary is inserted into another capillary used to deliver solutions that chemically stimulate the cells, with solution flowing up the inner capillary to facilitate peptide collection. The efficiency of peptide collection was evaluated using six peptide standards mixed in physiological saline. The extracted peptides eluted from these capillaries were characterized via matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) with low femtomole detection limits. Using the capillary collection system in small custom-fabricated culturing chambers, individual cultured neurons and neuronal clusters from the model animal Aplysia californica were stimulated with distinct neuronal secretagogues and the releasates were collected and characterized using MALDI-TOF MS.

  4. Threshold capillary pressure in capillaries with curved sides

    NASA Astrophysics Data System (ADS)

    Lago, Marcelo; Araujo, Mariela

    2003-03-01

    Modeling of fluid flow through permeable media is of great importance in assessing the performance of both hydrocarbon reservoirs and aquifers. In this process, network models based on cylindrical capillaries with circular cross sections are frequently used. This type of capillaries are not able to reproduce interesting physical phenomena observed in the experiments, for example, situations where there is flow by films with the wetting fluid occupying the crevices and wedges of the structure. We present an analysis of the behavior of the capillary pressure of a droplet of non-wetting fluid with an infinite length, inside objects of cylindrical symmetry with curved sides. The calculation is based on a method proposed by Mayer and Stowe and Princen (MS-P). Different capillary geometries are considered, and the behavior of the capillary pressure and transversal fluid saturation as a function of the shape factor is studied. The results found either analytically or numerically, allow to understand the relation between geometry and flow properties, and helps in the building of more realistic pore network models for flow studies at the pore scale.

  5. Pulsatility of Parafoveal Capillary Leukocytes

    PubMed Central

    Martin, Joy A.; Roorda, Austin

    2009-01-01

    The use of adaptive optics (AO) in a confocal scanning laser ophthalmoscope (AOSLO) allows for long-term imaging of parafoveal capillary leukocyte movement and measurement of leukocyte velocity without contrast dyes. We applied the AOSLO to investigate the possible role of the cardiac cycle on capillary leukocyte velocity by directly measuring capillary leukocyte pulsatility. The parafoveal regions of 8 eight normal healthy subjects with clear ocular media were imaged with an AOSLO. All subjects were dilated and cyclopleged. The AOSLO field of view was either 1.4 × 1.5 degrees or 2.35 × 2.5 degrees, the imaging wavelength was 532 nm and the frame rate was 30 fps. A photoplethysmograph was used to record the subject’s pulse synchronously with each AOSLO video. Parafoveal capillary leukocyte velocities and pulsatility were determined for two or three capillaries per subject. Leukocyte velocity and pulsatility were determined for all eight subjects. The mean parafoveal capillary leukocyte velocity for all subjects was Vmean = 1.30 mm/sec (SD = +/− 0.40 mm/sec). There was a statistically significant difference between leukocyte velocities, Vmax and Vmin, over the pulse cycle for each subject (p<0.05). The mean pulsatility was Pmean= 0.45 (+/− 0.09). Parafoveal capillary leukocyte pulsatility can be directly and non-invasively measured without the use of contrast dyes using an AOSLO. A substantial amount of the variation found in leukocyte velocity is due to the pulsatility that is induced by the cardiac cycle. By controlling for the variation in leukocyte velocity caused by the cardiac cycle, we can better detect other changes in retinal leukocyte velocity induced by disease or pharmaceutical agents. PMID:18708051

  6. Brain activation in teenagers with isolated spelling disorder during tasks involving spelling assessment and comparison of pseudowords. fMRI study.

    PubMed

    Borkowska, Aneta Rita; Francuz, Piotr; Soluch, Paweł; Wolak, Tomasz

    2014-10-01

    The present study aimed at defining the specific traits of brain activation in teenagers with isolated spelling disorder in comparison with good spellers. fMRI examination was performed where the subject's task involved taking a decision 1/whether the visually presented words were spelled correctly or not (the orthographic decision task), and 2/whether the two presented letters strings (pseudowords) were identical or not (the visual decision task). Half of the displays showing meaningful words with an orthographic difficulty contained pairs with both words spelled correctly, and half of them contained one misspelled word. Half of the pseudowords were identical, half of them were not. The participants of the study included 15 individuals with isolated spelling disorder and 14 good spellers, aged 13-15. The results demonstrated that the essential differences in brain activation between teenagers with isolated spelling disorder and good spellers were found in the left inferior frontal gyrus, left medial frontal gyrus and right cerebellum posterior lobe, i.e. structures important for language processes, working memory and automaticity of behaviour. Spelling disorder is not only an effect of language dysfunction, it could be a symptom of difficulties in learning and automaticity of motor and visual shapes of written words, rapid information processing as well as automating use of orthographic lexicon. Copyright © 2013 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

  7. A single exposure to social isolation in domestic piglets activates behavioural arousal, neuroendocrine stress hormones, and stress-related gene expression in the brain.

    PubMed

    Kanitz, E; Puppe, B; Tuchscherer, M; Heberer, M; Viergutz, T; Tuchscherer, A

    2009-08-04

    Stressful early life events can have short- and long-term effects on neuroendocrine and behavioural mechanisms of adaptation. Here, we investigated the effects of a single social isolation (4 h) of domestic piglets on both behavioural alterations in open-field tests and modifications in the expression of genes regulating glucocorticoid response in stress-related brain regions at 7, 21 or 35 days of age. The mRNAs of glucocorticoid receptor (GR), mineralocorticoid receptor (MR), 11ss-hydroxysteroid dehydrogenase 1 and 2 (11ss-HSD1 and 11ss-HSD2) and c-fos were analysed by real-time RT-PCR in the hypothalamus, hippocampus and amygdala. The social isolation caused both elevated stress hormone concentrations (e.g. cortisol) and open-field reactivity (e.g. locomotion, vocalisation) compared to control piglets. The enhanced behavioural and neuroendocrine activity was associated with distinct changes in gene expression in the limbic system. The hypothalamic GR, MR and 11ss-HSD1 mRNA expressions and the hippocampal 11ss-HSD1 mRNA was significantly higher in isolated piglets, whereas in the amygdala social isolation caused a significant decrease in MR mRNA expression. Isolated piglets also displayed significantly higher c-fos mRNA expression, an estimate of neuronal activation, in hypothalamus and amygdala. The mRNA alterations as well as the behavioural and hormonal pattern show an effect of social isolation on days 7 and 21, but no effect on day 35. In conclusion, a single social isolation in piglets caused age-dependent neuroendocrine and behavioural changes that indicate increased arousal and experienced distress. The present results also suggest that psychosocial stress effects should be considered for the assessment of livestock handling practices with respect to health and welfare.

  8. Capillary wave measurements on helically-supported capillary channels

    NASA Astrophysics Data System (ADS)

    Chandurwala, Fahim; Thiessen, David

    2010-10-01

    NASA is considering power generation by the Rankine cycle to save weight on long-duration manned missions to the moon or Mars. Phase separation technology is critical to this process in microgravity. Arrays of capillary channels might be useful for filtering liquid drops from a flowing vapor. The efficiency of droplet capture by a helically-supported capillary channel is being studied. A droplet impinging on the channel launches capillary waves that propagate down the channel helping to dissipate some of the drop's kinetic energy. High-speed video of the channel combined with image processing allows for measurement of the amplitude and speed of the wave packets. Increasing the pitch of the support structure decreases the wave speed. An understanding of the dynamic response of the channel to drop impact is a first step in predicting drop-capture efficiency.

  9. Development of a cryogenic capillary pumped loop

    SciTech Connect

    Kroliczek, E.J.; Cullimore, B.

    1996-03-01

    This paper describes the initial development of a promising new cryogenic technology. Room temperature capillary pumped loops (CPLs), a derivative of heat pipe technology, have been under development for almost two decades and are emerging as a design solution for many spacecraft thermal control problems. While cryogenic capillary pumped loops have application to passive spacecraft radiators and to long term storage of cryogenic propellants and open-cycle coolants, their application to the integration of spacecraft cryocoolers has generated the most excitement. Without moving parts or complex controls, they are able to thermally connect redundant cryocoolers to a single remote load, eliminating thermal switches and providing mechanical isolation at the same time. Development of a cryogenic CPL (CCPL) presented some unique challenges including start-up from a super-critical state, the management of parasitic heat leaks and pressure containment at ambient temperatures. These challenges have been overcome with a novel design that requires no additional devices or preconditioning for start-up. This paper describes the design concept and development and results conducted under SBIR Phase I and Phase II. {copyright} {ital 1996 American Institute of Physics.}

  10. Nerve growth cones isolated from fetal rat brain. IV. Preparation of a membrane subfraction and identification of a membrane glycoprotein expressed on sprouting neurons

    PubMed Central

    1985-01-01

    This study describes the preparation of a membrane subfraction from isolated nerve growth cone particles (GCPs) (see Pfenninger, K. H., L. Ellis, M. P. Johnson, L. B. Friedman, and S. Somlo, 1983, Cell, 35:573- 584) and the identification in this fraction of a glycoprotein expressed during neurite growth. While approximately 40 major polypeptides are visible in Coomassie Blue-stained SDS polyacrylamide gels of pelleted (partially disrupted) GCPs, a salt-washed membrane fraction prepared from lysed, detergent-permeabilized GCPs contains only 14% of this protein and has an unusually simple polypeptide pattern of seven major bands. Monoclonal antibodies have been generated to GCP membranes isolated from fetal rat brain. These antibodies have been screened differentially with synaptosomes from adult rat brain in order to identify those which recognize antigens expressed selectively during neurite growth. One such antibody (termed 5B4) recognizes a developmentally regulated membrane glycoprotein that is enriched in GCP membranes and expressed in fetal neurons sprouting in vitro. The 5B4 antigen in fetal brain migrates in SDS polyacrylamide gels as a diffuse band of approximately 185-255 kD, is rich in sialic acid, and consists of a small family of isoelectric variants. Freezing-thawing and neuraminidase digestion result in the cleavage of the native antigen into two new species migrating diffusely around 200 and 160 kD. Prolonged neuraminidase digestion sharpens these bands at about 180 and 135 kD, respectively. In the mature brain, antibody 5B4 recognizes a sparse polypeptide migrating at approximately 140 kD. As shown in the following paper (Wallis, I., L. Ellis, K. Suh, and K. H. Pfenninger, 1985, J. Cell Biol., 101:1990-1998), the fetal antigen is specifically associated with regions of neuronal sprouting and, therefore, can be used as a molecular marker of neurite growth. PMID:3902858

  11. Brain Abscess Associated with Isolated Left Superior Vena Cava Draining into the Left Atrium in the Absence of Coronary Sinus and Atrial Septal Defect

    SciTech Connect

    Erol, Ilknur Cetin, I. Ilker; Alehan, Fuesun; Varan, Birguel; Ozkan, Sueleyman; Agildere, A. Muhtesem; Tokel, Kursad

    2006-06-15

    A previously healthy 12-year-old girl presented with severe headache for 2 weeks. On physical examination, there was finger clubbing without apparent cyanosis. Neurological examination revealed only papiledema without focal neurologic signs. Cerebral magnetic resonance imaging showed the characteristic features of brain abscess in the left frontal lobe. Cardiologic workup to exclude a right-to-left shunt showed an abnormality of the systemic venous drainage: presence of isolated left superior vena cava draining into the left atrium in the absence of coronary sinus and atrial septal defect. This anomaly is rare, because only a few other cases have been reported.

  12. Capillary imbibition in parallel tubes

    NASA Astrophysics Data System (ADS)

    McRae, Oliver; Ramakrishnan, T. S.; Bird, James

    2016-11-01

    In modeling porous media two distinct approaches can be employed; the sample can be examined holistically, using global variables such as porosity, or it can be treated as a network of capillaries connected in series to various intermediate reservoirs. In forced imbibition this series-based description is sufficient to characterize the flow, due to the presence of an externally maintained pressure difference. However, in spontaneous imbibition, flow is driven by an internal capillary pressure, making it unclear whether a series-based model is appropriate. In this talk, we show using numerical simulations the dynamics of spontaneous imbibition in concentrically arranged capillary tubes. This geometry allows both tubes access to a semi-infinite reservoir but with inlets in close enough proximity to allow for interference. We compare and contrast the results of our simulations with theory and previous experiments. Schlumberger-Doll Research.

  13. Decay of capillary wave turbulence.

    PubMed

    Deike, Luc; Berhanu, Michael; Falcon, Eric

    2012-06-01

    We report on the observation of freely decaying capillary wave turbulence on the surface of a fluid. The capillary wave turbulence spectrum decay is found to be self-similar in time with the same power law exponent as the one found in the stationary regime, in agreement with weak turbulence predictions. The amplitude of all Fourier modes are found to decrease exponentially with time at the same damping rate. The longest wavelengths involved in the system are shown to be damped by a viscous surface boundary layer. These long waves play the role of an energy source during the decay that sustains nonlinear interactions to keep capillary waves in a wave turbulent state.

  14. Paramecium swimming in capillary tube

    NASA Astrophysics Data System (ADS)

    Jana, Saikat; Um, Soong Ho; Jung, Sunghwan

    2012-04-01

    Swimming organisms in their natural habitat need to navigate through a wide range of geometries and chemical environments. Interaction with boundaries in such situations is ubiquitous and can significantly modify the swimming characteristics of the organism when compared to ideal laboratory conditions. We study the different patterns of ciliary locomotion in glass capillaries of varying diameter and characterize the effect of the solid boundaries on the velocities of the organism. Experimental observations show that Paramecium executes helical trajectories that slowly transition to straight lines as the diameter of the capillary tubes decreases. We predict the swimming velocity in capillaries by modeling the system as a confined cylinder propagating longitudinal metachronal waves that create a finite pressure gradient. Comparing with experiments, we find that such pressure gradient considerations are necessary for modeling finite sized ciliary organisms in restrictive geometries.

  15. Optothermally actuated capillary burst valve

    NASA Astrophysics Data System (ADS)

    Eriksen, Johan; Bilenberg, Brian; Kristensen, Anders; Marie, Rodolphe

    2017-04-01

    We demonstrate the optothermal actuation of individual capillary burst valves in an all-polymer microfluidic device. The capillary burst valves are realised in a planar design by introducing a fluidic constriction in a microfluidic channel of constant depth. We show that a capillary burst valve can be burst by raising the temperature due to the temperature dependence of the fluid surface tension. We address individual valves by using a local heating platform based on a thin film of near infrared absorber dye embedded in the lid used to seal the microfluidic device [L. H. Thamdrup et al., Nano Lett. 10, 826-832 (2010)]. An individual valve is burst by focusing the laser in its vicinity. We demonstrate the capture of single polystyrene 7 μm beads in the constriction triggered by the bursting of the valve.

  16. Energy Invariance in Capillary Systems.

    PubMed

    Ruiz-Gutiérrez, Élfego; Guan, Jian H; Xu, Ben; McHale, Glen; Wells, Gary G; Ledesma-Aguilar, Rodrigo

    2017-05-26

    We demonstrate the continuous translational invariance of the energy of a capillary surface in contact with reconfigurable solid boundaries. We present a theoretical approach to find the energy-invariant equilibria of spherical capillary surfaces in contact with solid boundaries of arbitrary shape and examine the implications of dynamic frictional forces upon a reconfiguration of the boundaries. Experimentally, we realize our ideas by manipulating the position of a droplet in a wedge geometry using lubricant-impregnated solid surfaces, which eliminate the contact-angle hysteresis and provide a test bed for quantifying dissipative losses out of equilibrium. Our experiments show that dissipative energy losses for an otherwise energy-invariant reconfiguration are relatively small, provided that the actuation time scale is longer than the typical relaxation time scale of the capillary surface. We discuss the wider applicability of our ideas as a pathway for liquid manipulation at no potential energy cost in low-pinning, low-friction situations.

  17. Energy Invariance in Capillary Systems

    NASA Astrophysics Data System (ADS)

    Ruiz-Gutiérrez, Élfego; Guan, Jian H.; Xu, Ben; McHale, Glen; Wells, Gary G.; Ledesma-Aguilar, Rodrigo

    2017-05-01

    We demonstrate the continuous translational invariance of the energy of a capillary surface in contact with reconfigurable solid boundaries. We present a theoretical approach to find the energy-invariant equilibria of spherical capillary surfaces in contact with solid boundaries of arbitrary shape and examine the implications of dynamic frictional forces upon a reconfiguration of the boundaries. Experimentally, we realize our ideas by manipulating the position of a droplet in a wedge geometry using lubricant-impregnated solid surfaces, which eliminate the contact-angle hysteresis and provide a test bed for quantifying dissipative losses out of equilibrium. Our experiments show that dissipative energy losses for an otherwise energy-invariant reconfiguration are relatively small, provided that the actuation time scale is longer than the typical relaxation time scale of the capillary surface. We discuss the wider applicability of our ideas as a pathway for liquid manipulation at no potential energy cost in low-pinning, low-friction situations.

  18. On-column electrochemical detection for microchip capillary electrophoresis.

    PubMed

    Osbourn, Damon M; Lunte, Craig E

    2003-06-01

    The development of a cellulose acetate decoupler for on-column electrochemical detection in microchip capillary electrophoresis is presented. The capillary based laser-etched decoupler is translated to the planar format to isolate the detector circuit from the separation circuit. The decoupler is constructed by aligning a series of 20 30-microm holes through the coverplate of the microchip with the separation channel and casting a thin film of cellulose acetate within the holes. The decoupler shows excellent isolation of the detection circuit for separation currents up to 60 microA, with noise levels at or below 1 pA at a carbon fiber electrode. Detection limits of 25 nM were achieved for dopamine. This decoupler design combines excellent mechanical stability, effective shunting of high separation currents, and ease of manufacture.

  19. Electrical resistance of a capillary endothelium

    PubMed Central

    1981-01-01

    The electrical resistance of consecutive segments of capillaries has been determined by a method in which the microvessels were treated as a leaky, infinite cable. A two-dimensional analytical model to describe the potential field in response to intracapillary current injection was formulated. The model allowed determination of the electrical resistance from four sets of data: the capillary radius, the capillary length constant, the length constant in the mesentery perpendicular to the capillary, and the relative potential drop across the capillary wall. Of particular importance were the mesothelial membranes covering the mesenteric capillaries with resistances several times higher than that of the capillary endothelium. 27 frog mesenteric capillaries were characterized. The average resistance of the endothelium was 1.85 omega cm2, which compares well with earlier determinations of the ionic permeability of such capillaries. However, heterogeneity with respect to resistance was observed, that of 10 arterial capillaries being 3.0 omega cm2 as compared with 0.95 omega cm2 for 17 mid- and venous capillaries. The average in situ length constant was 99 micrometers for the arterial capillaries and 57 micrometers for the mid- and venous capillaries. It is likely that the ions that carry the current must move paracellularly, through junctions that are leaky to small solutes. PMID:7241087

  20. Revised Capillary Breakup Rheometer Method

    NASA Astrophysics Data System (ADS)

    Lu, Louise; Schultz, William; Solomon, Michael

    2014-11-01

    Rather than integrate the one-dimensional equation of motion for a capillary breakup rheometer, we take the axial derivative of that equation. This avoids the determination of the axial force with all of its complications and correction factors. The resulting evolutionary equation that involves either two or four derivatives of the capillary radius as a function of the axial coordinate determines the ratio of elongational viscosity to surface tension coefficient. We examine several silicone and olive oils to show the accuracy of the method for Newtonian fluids. We will discuss our surface tension measurement techniques and briefly describe measurements of viscoelastic materials, including saliva.

  1. Nonsteady Flow in Capillary Tubes

    NASA Astrophysics Data System (ADS)

    Hara, Ayako

    2000-03-01

    Surface phenomena in the field of electron devices and the problem of how long. It takes plants to absorb water during their growth in hydroponic cultivation is attraching the attention of riseachers. However, the related study of non-steady flow in capillary tubes has a number of issues that require investigation. In response to this situation, we made attempted to assess nonsteady fiow in capillary tubes, the liquid rise time and other issues, using a motion equation that takes factors including the friction force of the tube and the surface tension into consideration.

  2. Capillary haemangioma of the testis

    PubMed Central

    Mazal, P; Kratzik, C; Kain, R; Susani, M

    2000-01-01

    A case of testicular capillary haemangioma is reported and the importance of intraoperative examination of this very rare lesion emphasised. Capillary haemangioma of the testis can be similar to malignant testicular tumours on clinical presentation, as well as on ultrasonography and magnetic resonance imaging, and therefore should be included in the intraoperative differential diagnosis. Because of the benign nature of this lesion, conservative surgical treatment by means of tumour enucleation with preservation of the testis is possible, if intraoperative examination of frozen sections of representative tissue can be performed. Key Words: testis • haemangioma PMID:11002773

  3. Modulatory influence of Parkia biglobosa protein isolate on testosterone and biomarkers of oxidative stress in brain and testes of streptozotocin-induced diabetic male rats

    PubMed Central

    Ogunyinka, Bolajoko Idiat; Oyinloye, Babatunji Emmanuel; Osunsanmi, Foluso Oluwagbemiga; Opoku, Andrew Rowland; Kappo, Abidemi Paul

    2016-01-01

    Parkia biglobosa seed an important household spice commonly consumed in Nigeria is believed to possess antioxidant activity that may exert modulatory effects in diabetes and diabetic complications. This study investigated the modulatory potential of Parkia biglobosa protein isolate (PBPi) on serum testosterone (sTT) level as well as its influence on biomarkers of oxidative stress in brain and testes of streptozotocin-induced diabetic male rats. Animals were made diabetic by single intraperitoneal administration of streptozotocin (STZ; 60 mg/kg body weight). PBPi (200 or 400 mg/kg body weight) was given orally by gavage or insulin (5 U/kg, i.p.) was administered daily to STZ-induced diabetic rats for 28 days. The results revealed a significant elevation in thiobarbituric acid reactive substances (TBARS) levels in the brain and testes of diabetic rats. This was closely associated with a concomitant reduction in levels of sTT and reduced testes weight, a noticeable decline in the glutathione-S-transferase (GST), superoxide dismutase (SOD) and catalase (CAT) as well as total glutathione (Total GSH) level in the brain and testes of diabetic rats. Interestingly, treatment with PBPi efficiently prevented the alterations witnessed in the serum sTT and also ameliorated various alterations in the biomarkers of oxidative stress (TBARS, Total GSH, GST, SOD and CAT) in brain and testes of diabetic rats. These results provide evidence that PBPi could protect the brain and testicular tissues against oxidative stress induced by STZ, via modulation of serum testosterone concentration and also by enhancing antioxidant defence system in STZ-diabetic rats. PMID:27785334

  4. Use of non-porous reversed-phase high-performance liquid chromatography for protein profiling and isolation of proteins induced by temperature variations for Siberian permafrost bacteria with identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and capillary electrophoresis-electrospray ionization mass spectrometry.

    PubMed

    Chong, B E; Kim, J; Lubman, D M; Tiedje, J M; Kathariou, S

    2000-10-01

    Non-porous reversed-phase high-performance liquid chromatography (NP-RP-HPLC) has been used to separate and isolate proteins from whole cell lysates of ED 7-3, a bacterium from the buried Siberian permafrost sediment. The proteins collected from the liquid eluent of this separation were then analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and capillary electrophoresis-electrospray ionization mass spectrometry (CE-ESI-MS). In order to study the differences in expression of cold-shock proteins (CSPs) at different growth temperatures, cultures of the ED 7-3 strain were prepared at 4 degrees C and 25 degrees C. The goals of this work were twofold: firstly, to identify the presence of CSPs and other proteins that are highly expressed at 4 degrees C but not at 25 degrees C; and secondly, to isolate these proteins for MALDI-TOF-MS and CE-ESI-MS identification. In this initial work, distinct protein profiles were observed for these cultures as a function of temperature. Fraction collection from the eluent of NP-RP-HPLC of some of the highly expressed proteins was performed and the proteins were mass analyzed for molecular mass. Peptide maps of the proteins were generated by tryptic digestion and were analyzed by CE-ESI-MS and MALDI-TOF-MS for database identification of the expressed proteins.

  5. Specific binding of atrial natriuretic factor in brain microvessels

    SciTech Connect

    Chabrier, P.E.; Roubert, P.; Braquet, P.

    1987-04-01

    Cerebral capillaries constitute the blood-brain barrier. Studies of specific receptors (neurotransmitters or hormones) located on this structure can be performed by means of radioligand-binding techniques on isolated brain microvessels. The authors examined on pure bovine cerebral microvessel preparations the binding of atrial natriuretic factor (ANF), using /sup 125/I-labeled ANF. Saturation and competition experiments demonstrated the presence of a single class of ANF-binding sites with high affinity and with a binding capacity of 58 fmol/mg of protein. The binding of /sup 125/I-labeled ANF to brain microvessels is specific, reversible, and time dependent, as is shown by association-dissociation experiments. The demonstration of specific ANF-binding sites on brain microvessels supposes a physiological role of ANF on brain microvasculature. The coexistence of ANF and angiotensin II receptors on this cerebrovascular tissue suggests that the two circulating peptides may act as mutual antagonists in the regulation of brain microcirculation and/or blood-brain barrier function.

  6. Multiplexed fluorescence detector system for capillary electrophoresis

    DOEpatents

    Yeung, Edward S.; Taylor, John A.

    1996-03-12

    A fluorescence detection system for capillary electrophoresis is provided wherein the detection system can simultaneously excite fluorescence and substantially simultaneously monitor separations in multiple capillaries. This multiplexing approach involves laser irradiation of a sample in a plurality of capillaries through optical fibers that are coupled individually with the capillaries. The array is imaged orthogonally through a microscope onto a charge-coupled device camera for signal analysis.

  7. Multiplexed fluorescence detector system for capillary electrophoresis

    DOEpatents

    Yeung, Edward S.; Taylor, John A.

    1994-06-28

    A fluorescence detection system for capillary electrophoresis is provided wherein the detection system can simultaneously excite fluorescence and substantially simultaneously monitor separations in multiple capillaries. This multiplexing approach involves laser irradiation of a sample in a plurality of capillaries through optical fibers that are coupled individually with the capillaries. The array is imaged orthogonally through a microscope onto a charge-coupled device camera for signal analysis.

  8. Multiplexed fluorescence detector system for capillary electrophoresis

    DOEpatents

    Yeung, E.S.; Taylor, J.A.

    1994-06-28

    A fluorescence detection system for capillary electrophoresis is provided wherein the detection system can simultaneously excite fluorescence and substantially simultaneously monitor separations in multiple capillaries. This multiplexing approach involves laser irradiation of a sample in a plurality of capillaries through optical fibers that are coupled individually with the capillaries. The array is imaged orthogonally through a microscope onto a charge-coupled device camera for signal analysis. 14 figures.

  9. Multiplexed fluorescence detector system for capillary electrophoresis

    DOEpatents

    Yeung, E.S.; Taylor, J.A.

    1996-03-12

    A fluorescence detection system for capillary electrophoresis is provided wherein the detection system can simultaneously excite fluorescence and substantially simultaneously monitor separations in multiple capillaries. This multiplexing approach involves laser irradiation of a sample in a plurality of capillaries through optical fibers that are coupled individually with the capillaries. The array is imaged orthogonally through a microscope onto a charge-coupled device camera for signal analysis. 14 figs.

  10. Toxicity of cuprizone a Cu(2+) chelating agent on isolated mouse brain mitochondria: a justification for demyelination and subsequent behavioral dysfunction.

    PubMed

    Faizi, Mehrdad; Salimi, Ahmad; Seydi, Enayatolla; Naserzadeh, Parvaneh; Kouhnavard, Mehdi; Rahimi, Atena; Pourahmad, Jalal

    2016-05-01

    Multiple Sclerosis (MS) is a complex disease with an unknown etiology and no effective cure, despite decades of extensive research that led to the development of several partially effective treatments. In this study we aimed to investigate brain mitochondrial dysfunction in demyelination induced by cuprizone in mice. Cuprizone was used for induction of demyelination in mice through a diet containing 0.2% w/w cuprizone for 5 weeks. Behavioral tests for proving of MS was performed and then mitochondria from brain of animals were isolated and afterwards parameters of mitochondrial dysfunction examined. Results of mitochondrial dysfunction parameters such as mitochondrial swelling, production ROS, collapse of the membrane potential showed that isolated mitochondria from cuprizone treated mice have been damaged compared to those of untreated control mice. It is likely that demyelination induced mitochondrial damage led to increased mitochondrial ROS formation and progression of oxidative damages in neurons. It is suggested that cuprizone which is a Cu(2+) chelating agent causes impairment of electron transport chain (complex IV) and antioxidant system (SOD) in mitochondria leading to decreased ATP production and increased ROS formation.

  11. Particle trajectories in linear periodic capillary and capillary-gravity water waves.

    PubMed

    Henry, David

    2007-09-15

    Surface tension plays a significant role as a restoration force in the setting of small-amplitude waves, leading to pure capillary and gravity-capillary waves. We show that within the framework of linear theory, the particle paths in a periodic gravity-capillary or pure capillary wave propagating at the surface of water over a flat bed are not closed.

  12. Capillary electrophoresis with online stacking in combination with AgNPs@MCM-41 reinforced hollow fiber solid-liquid phase microextraction for quantitative analysis of Capecitabine and its main metabolite 5-Fluorouracil in plasma samples isolated from cancer patients.

    PubMed

    Forough, Mehrdad; Farhadi, Khalil; Molaei, Rahim; Khalili, Hedayat; Shakeri, Ramin; Zamani, Asghar; Matin, Amir Abbas

    2017-01-01

    The purpose of this study is the development and validation of a simple, novel, selective and fast off-line microextraction technique combining capillary electrophoresis with in-column field-amplified sample injection (FASI) for the simultaneous determination of capecitabine (CAP) and its active metabolite, 5-Fluorouracil (5-FU), in human plasma. At the moment, there is a lack of using cost-effective CE tool combined with novel miniaturized sample clean-up techniques for analysis of these important anti-cancer agents in plasma samples. This paper intends to fill this gap and describe a simple off-line sample pretreatment by means of AgNPs@MCM-41 reinforced hollow fiber Solid/Liquid phase microextraction (AgNPs@MCM41-HF-SLPME) with subsequent quantitation by FASI-CE. The separation of analytes was performed using a BGE containing 60mM phosphate-Tris buffer (pH 7) with 10% methanol as an organic modifier. Before sample loading, a short water plug (50mbar, 3s) was injected to permit FASI for stacking. Various parameters affecting the off-line microextraction efficiency as well as FASI were optimized. Migration time was found to be 6.6 (±0.1)min for 5-FU and 7.4 (±0.2)min for CAP. The linearity, precision, accuracy, recovery, selectivity, specificity, stability as well as the robustness of the method was evaluated from spiked plasma samples during the course of validation. The results revealed that the presented technique demonstrates acceptable accuracy and precision, miniaturized sample preparation and a reduced need for complicated equipment along with an acceptable analysis time. The validated method was successfully applied to determine CAP and 5-FU in patient's plasma samples.

  13. Microdialysis monitoring of catecholamines and excitatory amino acids in the rat and mouse brain: recent developments based on capillary electrophoresis with laser-induced fluorescence detection--a mini-review.

    PubMed

    Parrot, Sandrine; Bert, Lionel; Mouly-Badina, Laurence; Sauvinet, Valérie; Colussi-Mas, Joyce; Lambás-Señas, Laura; Robert, Frédéric; Bouilloux, Jean-Pierre; Suaud-Chagny, Marie-Françoise; Denoroy, Luc; Renaud, Bernard

    2003-10-01

    1. Although microdialysis is a widely used approach for in vivo monitoring extracellular neurotransmitter concentrations, it has been previously limited in many cases by its poor temporal resolution. It is clear that when 10-30-min sampling is performed, short-lasting changes in extracellular neurotransmitter concentrations can be overlooked. Such a low sampling rate is necessary when combining microdialysis with the conventional analytical methods like high performance liquid chromatography. 2. Since capillary electrophoresis coupled to laser-induced fluorescence detection (CE-LIFD) allows the detection of attomoles of neurotransmitters, the temporal resolution of microdialysis may be significantly improved: high sampling rates, in the range of 5 s to 1 min, have been already reported by our group and others using CE-LIFD for simultaneously analyzing catecholamines and amino acids in microdialysates. 3. The power of combining microdialyis and CE-LIFD is shown, using examples of physiological and pharmacological studies dealing with the dynamics of in vivo efflux processes and/or interactions between neurotransmitters.

  14. Bacterial motion in narrow capillaries

    PubMed Central

    Ping, Liyan; Wasnik, Vaibhav; Emberly, Eldon

    2014-01-01

    Motile bacteria often have to pass through small tortuous pores in soil or tissue of higher organisms. However, their motion in this prevalent type of niche is not fully understood. Here, we modeled it with narrow glass capillaries and identified a critical radius (Rc) for bacterial motion. Near the surface of capillaries narrower than that, the swimming trajectories are helices. In larger capillaries, they swim in distorted circles. Under non-slip condition, the peritrichous Escherichia coli swam in left-handed helices with an Rc of ∼10 μm near glass surface. However, slipping could occur in the fast monotrichous Pseudomonas fluorescens, when a speed threshold was exceeded, and thus both left-handed and right-handed helices were executed in glass capillaries. In the natural non-cylindrical pores, the near-surface trajectories would be spirals and twisted loops. Engaging in such motions reduces the bacterial migration rate. With a given pore size, the run length and the tumbling angle of the bacterium determine the probability and duration of their near-surface motion. Shear flow and chemotaxis potentially enhance it. Based on this observation, the puzzling previous observations on bacterial migration in porous environments can be interpreted. PMID:25764548

  15. Capillary waves and ellipsometry experiments

    NASA Astrophysics Data System (ADS)

    Bonn, D.; Wegdam, G. H.

    1992-09-01

    The inclusion of higher-order terms in the capillary-wave Hamiltonian may reduce the contributions of these fluctuations to the ellipsometric coefficients. We show that the renormalization of capillary waves at a fluid-fluid interface by Sengers and van Leeuwen [Phys. Rev. A 39 (1989) 6346] using the wave vector-dependent surface tension that follows from the coupled mode theory by Meunier [Phys. France 48 (1987)1819] yields a satisfactory agreement with recent ellipsometry measurements by Schmidt [Phys. Rev. A 38 (1988) 567]. The interface is viewed upon as an intrinsic interface broadened by capillary waves. We suppose that the cutoff wave vector q_{max} that follows from mode-coupling theory marks the transition from the short-wavelength bulk-like fluctuations that contribute to the bare surface tension to the long-wavelength capillary wave-like fluctuations that contribute to the full surface tension. This enables us to calculate, without any adjustable parameters, both the ratio of the bare and experimental surface tension and the universal constant for the elliptical thickness of the interface. Both agree remarkably well with experimental values.

  16. Capillary Rise in a Wedge

    ERIC Educational Resources Information Center

    Piva, M.

    2009-01-01

    In introductory-level physics courses, the concept of surface tension is often illustrated using the example of capillary rise in thin tubes. In this paper the author describes experiments conducted using a planar geometry created with two small plates forming a thin wedge. The distribution of the fluid entering the wedge can be studied as a…

  17. Capillary Rise in a Wedge

    ERIC Educational Resources Information Center

    Piva, M.

    2009-01-01

    In introductory-level physics courses, the concept of surface tension is often illustrated using the example of capillary rise in thin tubes. In this paper the author describes experiments conducted using a planar geometry created with two small plates forming a thin wedge. The distribution of the fluid entering the wedge can be studied as a…

  18. Capillary electrophoresis for drug analysis

    NASA Astrophysics Data System (ADS)

    Lurie, Ira S.

    1999-02-01

    Capillary electrophoresis (CE) is a high resolution separation technique which is amenable to a wide variety of solutes, including compounds which are thermally degradable, non-volatile and highly polar, and is therefore well suited for drug analysis. Techniques which have been used in our laboratory include electrokinetic chromatography (ECC), free zone electrophoresis (CZE) and capillary electrochromatography (CEC). ECC, which uses a charged run buffer additive which migrates counter to osmotic flow, is excellent for many applications, including, drug screening and analyses of heroin, cocaine and methamphetamine samples. ECC approaches include the use of micelles and charged cyclodextrins, which allow for the separation of complex mixtures. Simultaneous separation of acidic, neutral and basic solutes and the resolution of optical isomers and positional isomers are possible. CZE has been used for the analysis of small ions (cations and anions) in heroin exhibits. For the ECC and CZE experiments performed in our laboratory, uncoated capillaries were used. In contrast, CEC uses capillaries packed with high performance liquid chromatography stationary phases, and offers both high peak capacities and unique selectivities. Applications include the analysis of cannabinoids and drug screening. Although CE suffers from limited concentration sensitivity, it is still applicable to trace analysis of drug samples, especially when using injection techniques such as stacking, or detection schemes such as laser induced fluorescence and extended pathlength UV.

  19. Using large-scale Granger causality to study changes in brain network properties in the Clinically Isolated Syndrome (CIS) stage of multiple sclerosis

    NASA Astrophysics Data System (ADS)

    Abidin, Anas Z.; Chockanathan, Udaysankar; DSouza, Adora M.; Inglese, Matilde; Wismüller, Axel

    2017-03-01

    Clinically Isolated Syndrome (CIS) is often considered to be the first neurological episode associated with Multiple sclerosis (MS). At an early stage the inflammatory demyelination occurring in the CNS can manifest as a change in neuronal metabolism, with multiple asymptomatic white matter lesions detected in clinical MRI. Such damage may induce topological changes of brain networks, which can be captured by advanced functional MRI (fMRI) analysis techniques. We test this hypothesis by capturing the effective relationships of 90 brain regions, defined in the Automated Anatomic Labeling (AAL) atlas, using a large-scale Granger Causality (lsGC) framework. The resulting networks are then characterized using graph-theoretic measures that quantify various network topology properties at a global as well as at a local level. We study for differences in these properties in network graphs obtained for 18 subjects (10 male and 8 female, 9 with CIS and 9 healthy controls). Global network properties captured trending differences with modularity and clustering coefficient (p<0.1). Additionally, local network properties, such as local efficiency and the strength of connections, captured statistically significant (p<0.01) differences in some regions of the inferior frontal and parietal lobe. We conclude that multivariate analysis of fMRI time-series can reveal interesting information about changes occurring in the brain in early stages of MS.

  20. Strongylocentrotus purpuratus spindle tubulin. II. Characteristics of its sensitivity to Ca++ and the effects of calmodulin isolated from bovine brain and S. purpuratus eggs

    PubMed Central

    1982-01-01

    Tubulin was extracted from spindles isolated from embryos of the sea urchin Strongylocentrotus purpuratus and purified through cycles of temperature-dependent assembly and disassembly. At 37 degrees C, the majority of the cycle-purified spindle tubulin polymer is insensitive to free Ca++ at concentrations below 0.4 mM, requiring free Ca++ concentrations greater than 1 mM for complete depolymerization. However, free Ca++ at concentrations above 1 microM inhibits initiation of polymer formation without significantly inhibiting the rate of elongation onto existing polymer. At 15 degrees C and 18 degrees C, temperatures that are physiological for S. purpuratus embryos, spindle tubulin polymer is sensitive to free Ca++ at micromolar concentrations such that 3-20 microM free Ca++ causes complete depolymerization. Calmodulin purified from either bovine brain or S. purpuratus eggs does not affect the Ca++ sensitivity of the spindle tubulin at 37 degrees C, although both increase the Ca++ sensitivity of cycle-purified bovine brain tubulin. These results indicate that cycle-purified spindle tubulin and cycle-purified bovine brain tubulin differ significantly in their responses to calmodulin and in their Ca++ sensitivities at their physiological temperatures. They also suggest that, in vivo, spindle tubulin may be regulated by physiological levels of intracellular Ca++ in the absence of Ca++-sensitizing factors. PMID:7119003

  1. Independent Epileptiform Discharge Patterns in the Olfactory and Limbic Areas of the In Vitro Isolated Guinea Pig Brain During 4-Aminopyridine Treatment

    PubMed Central

    Carriero, Giovanni; Uva, Laura; Gnatkovsky, Vadym; Avoli, Massimo; de Curtis, Marco

    2016-01-01

    In vitro studies performed on brain slices demonstrate that the potassium channel blocker 4-aminopyridine (4AP, 50 μM) discloses electrographic seizure activity and interictal discharges. These epileptiform patterns have been further analyzed here in a isolated whole guinea pig brain in vitro by using field potential recordings in olfactory and limbic structures. In 8 of 13 experiments runs of fast oscillatory activity (fast runs, FRs) in the piriform cortex (PC) propagated to the lateral entorhinal cortex (EC), hippocampus and occasionally to the medial EC. Early and late FRs were asynchronous in the hemispheres showed different duration [1.78 ± 0.51 and 27.95 ± 4.55 (SD) s, respectively], frequency of occurrence (1.82 ± 0.49 and 34.16 ± 6.03 s) and frequency content (20–40 vs. 40–60 Hz). Preictal spikes independent from the FRs appeared in the hippocampus/EC and developed into ictal-like discharges that did not propagate to the PC. Ictal-like activity consisted of fast activity with onset either in the hippocampus (n = 6) or in the mEC (n = 2), followed by irregular spiking and sequences of diffusely synchronous bursts. Perfusion of the N-methyl-D-aspartate receptor antagonist 2-amino-5-phosphonopentanoic acid (100 μM) did not prevent FRs, increased the duration of limbic ictal-like discharges and favored their propagation to olfactory structures. The AMPA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (50 μM) blocked ictal-like events and reduced FRs. In conclusion, 4AP-induced epileptiform activities are asynchronous and independent in olfactory and hippocampal-entorhinal regions. Epileptiform discharges in the isolated guinea pig brain show different pharmacological properties compared with rodent in vitro slices. PMID:20220076

  2. Isolation of a Drosophila homolog of the vertebrate homeobox gene Rx and its possible role in brain and eye development

    PubMed Central

    Eggert, Tanja; Hauck, Bernd; Hildebrandt, Nicole; Gehring, Walter J.; Walldorf, Uwe

    1998-01-01

    Vertebrate and invertebrate eye development require the activity of several evolutionarily conserved genes. Among these the Pax-6 genes play a major role in the genetic control of eye development. Mutations in Pax-6 genes affect eye development in humans, mice, and Drosophila, and misexpression of Pax-6 genes in Drosophila can induce ectopic eyes. Here we report the identification of a paired-like homeobox gene, DRx, which is also conserved from flies to vertebrates. Highly conserved domains in the Drosophila protein are the octapeptide, the identical homeodomain, the carboxyl-terminal OAR domain, and a newly identified Rx domain. DRx is expressed in the embryo in the procephalic region and in the clypeolabrum from stage 8 on and later in the brain and the central nervous system. Compared with eyeless, the DRx expression in the embryo starts earlier, similar to the pattern in vertebrates, where Rx expression precedes Pax-6 expression. Because the vertebrate Rx genes have a function during brain and eye development, it was proposed that DRx has a similar function. The DRx expression pattern argues for a conserved function at least during brain development, but we could not detect any expression in the embryonic eye primordia or in the larval eye imaginal discs. Therefore DRx could be considered as a homolog of vertebrate Rx genes. The Rx genes might be involved in brain patterning processes and specify eye fields in different phyla. PMID:9482887

  3. Social isolation mediated anxiety like behavior is associated with enhanced expression and regulation of BDNF in the female mouse brain.

    PubMed

    Kumari, Anita; Singh, Padmanabh; Baghel, Meghraj Singh; Thakur, M K

    2016-05-01

    Adverse early life experience is prominent risk factors for numerous psychiatric illnesses, including mood and anxiety disorders. It imposes serious long-term costs on the individual as well as health and social systems. Hence, developing therapies that prevent the long-term consequences of early life stress is of utmost importance, and necessitates a better understanding of the mechanisms by which early life stress triggers long-lasting alterations in gene expression and behavior. Post-weaning isolation rearing of rodents models the behavioral consequences of adverse early life experiences in humans and it is reported to cause anxiety like behavior which is more common in case of females. Therefore, in the present study, we have studied the impact of social isolation of young female mice for 8weeks on the anxiety like behavior and the underlying molecular mechanism. Elevated plus maze and open field test revealed that social isolation caused anxiety like behavior. BDNF, a well-known molecule implicated in the anxiety like behavior, was up-regulated both at the message and protein level in cerebral cortex by social isolation. CREB-1 and CBP, which play a crucial role in BDNF transcription, were up-regulated at mRNA level in cerebral cortex by social isolation. HDAC-2, which negatively regulates BDNF expression, was down-regulated at mRNA and protein level in cerebral cortex by social isolation. Furthermore, BDNF acts in concert with Limk-1, miRNA-132 and miRNA-134 for the regulation of structural and morphological plasticity. Social isolation resulted in up-regulation of Limk-1 mRNA and miRNA-132 expression in the cerebral cortex. MiRNA-134, which inhibits the translation of Limk-1, was decreased in cerebral cortex by social isolation. Taken together, our study suggests that social isolation mediated anxiety like behavior is associated with up-regulation of BDNF expression and concomitant increase in the expression of CBP, CREB-1, Limk-1 and miRNA-132, and decrease

  4. Molecular characterization of individual D3 dopamine receptor-expressing cells isolated from multiple brain regions of a novel mouse model

    PubMed Central

    Li, Ying

    2012-01-01

    Among dopamine receptors, the expression and function of the D3 receptor subtype is not well understood. The receptor has the highest affinity for dopamine and many drugs that target dopamine receptors. In this paper, we examined, at the single cell level, the characteristics of D3 receptor-expressing cells isolated from different brain regions of male and female mice that were either 35 or 70 days old. The brain regions included nucleus accumbens, Islands of Calleja, olfactory tubercle, retrosplenial cortex, dorsal subiculum, mammillary body, amygdala and septum. The expression analysis was done in the drd3-enhanced green fluorescent protein transgenic mice that report the endogenous expression of D3 receptor mRNA. Using single cell reverse transcriptase PCR, we determined if the D3 receptor-expressing fluorescent cells in these mice were neurons or glia and if they were glutamatergic, GABAergic or catecholaminergic. Next, we determined if the fluorescent cells co-expressed the four other dopamine receptor subtypes, adenylate cyclase V (ACV) isoform, and three different isoforms of G protein-coupled inward rectifier potassium (GIRK) channels. The results suggest that D3 receptor is expressed in neurons, with region-specific expression in glutamatergic and GABAergic neurons. The D3 receptor primarily co-expressed with D1 and D2 dopamine receptors with regional, sex and age-dependent differences in the co-expression pattern. The percentage of cells co-expressing D3 receptor and ACV or GIRK channels varied significantly by brain region, sex and age. The molecular characterization of D3 receptor-expressing cells in mouse brain reported here will facilitate the characterization of D3 receptor function in physiology and pathophysiology. PMID:22286951

  5. Porosity, single-phase permeability, and capillary pressure data from preliminary laboratory experiments on selected samples from Marker Bed 139 at the Waste Isolation Pilot Plant. Volume 3 of 3: Appendices C, D, E, and F

    SciTech Connect

    Howarth, S.M.; Christian-Frear, T.

    1997-08-01

    This volume contains the mineralogy, porosity, and permeability results from the Marker Bed 139 anhydrite specimens evaluated by TerraTek, Inc. for the Waste Isolation Pilot Plant. This volume also documents the brine recipe used by RE/SPEC, Inc., the parameter package submitted to Performance Assessment based on all the data, and a memo on the mixed Brooks and Corey two-phase characteristic curves.

  6. Isolation of compound and CNS depressant activities of Mikania scandens Willd with special emphasis to brain biogenic amines in mice.

    PubMed

    Pal, Dilipkumar; Mazumder, Upal Kanti

    2014-12-01

    Mikania scandens, a twining herb that grows as a weed in India and Bangladesh is used as vegetables and is a good source of vitamin A, C, B complex, mikanin, sesquiterpenes, betasitosterin, stigmasterol and friedelin. The present communication reports CNS depressant activities with special emphasis to brain biogenic amines in mice. Ethanol extract of leaves of M. scandens (EEMS) was prepared by Soxhalation and analyzed chemically. EEMS potentiated sleeping time induced by pentobarbitone, diazepam and meprobamate and showed significant reduction in the number of writhes and stretches. EEMS caused significant protection against pentylene tetrazole-induced convulsion and increased catecholamines and brain amino acids level significantly. Results showed that EEMS produced good CNS depressant effects in mice.

  7. Multiple forms of phosphatase from human brain: isolation and partial characterization of affi-gel blue binding phosphatases.

    PubMed

    Cheng, L Y; Wang, J Z; Gong, C X; Pei, J J; Zaidi, T; Grundke-Iqbal, I; Iqbal, K

    2000-01-01

    Implication of protein phosphatases in Alzheimer disease led us to a systemic investigation of the identification of these enzyme activities in human brain. Human brain phosphatases eluted from DEAE-Sephacel with 0.22 M NaCl were resolved into two main groups by affi-gel blue chromatography, namely affi-gel blue-binding phosphatases and affi-gel blue-nonbinding phosphatases. Affi-gel blue-binding phosphatases were further separated into four different phosphatases, designated P1, P2, P3, and P4 by calmodulin-Sepharose 4B and poly-(L-lysine)-agarose chromatographies. These four phosphatases exhibited activities towards nonprotein phosphoester and two of them, P1 and P4, could dephosphorylate phosphoproteins. The activities of the four phosphatases differed in pH optimum, divalent metal ion requirements, sensitivities to various inhibitors and substrate affinities. The apparent molecular masses as estimated by gel-filtration for P1, P2, P3, and P4 were 97, 45, 42, and 125 kDa, respectively. P1 is markedly similar to PP2B from bovine brain and rabbit skeletal muscle. P4 was labeled with anti-PP2A antibody and may represent a new subtype of PP2A. P1 and P4 were also effective in dephosphorylating Alzheimer disease abnormally hyperphosphorylated tau (AD P-tau). The resulting dephosphorylated AD P-tau had its activity restored in promoting assembly of microtubules in vitro. These results suggest that P1 and P4 might be involved in the regulation of phosphorylation of tau in human brain, especially in neurodegenerative conditions like Alzheimer's disease which are characterized by the abnormal hyperphosphorylation of this protein.

  8. Tapered splice technique for capillary optical fiber

    NASA Astrophysics Data System (ADS)

    Zhu, Xiaoliang; Xiang, Huoxing

    2017-07-01

    We propose a simple but effective technique. It is concerned with a tapered splice technique for capillary optical fiber. In order to contrast, we investigate two kinds of capillary optical fiber. One of the capillary optical fiber has the annular core around the air hole and the other one has the embedded annular core around the inner cladding. We demonstrate the tapered splice technique works for both of the capillary optical fiber in experiment. It is the key to improve the coupling efficiency of the capillary optical fiber. We also build a theoretical model to predict the optical power of the capillary optical fiber and it is confirmed by the experimental results. The method provides an insight of the mode conversion characteristics of capillary optical fiber. It should be used as an easy way to realize the fiber-based in-line components and should be more importantly to explore new possibilities with this kind of fiber.

  9. Capillary rise in cellulose sponges

    NASA Astrophysics Data System (ADS)

    Kim, Jungchul; Kim, Ho-Young; Mahadevan, L.

    2011-11-01

    A cellulose sponge, commonly used for clean-up jobs, can absorb and hold a significant amount of water within its pores, whose size ranges from micrometers to millimeters. We investigate the dynamics of capillary rise of water in the sponge using a combination of experiment and theory. We find that the rate of the capillary rise is significantly lower than Washburn's rule that assumes the sponge as a row of adjoined pores and the liquid flow to be driven by the Laplace pressure. We introduce a novel theory to model the flow in the hygroscopic porous media by combining Darcy's law based on the moisture concentration and the modified Young-Laplace equation. The scaling law constructed through this work agrees well with the experimental results.

  10. How Glutamate Is Managed by the Blood–Brain Barrier

    PubMed Central

    Hawkins, Richard A.; Viña, Juan R.

    2016-01-01

    A facilitative transport system exists on the blood–brain barrier (BBB) that has been tacitly assumed to be a path for glutamate entry to the brain. However, glutamate is a non-essential amino acid whose brain content is much greater than plasma, and studies in vivo show that glutamate does not enter the brain in appreciable quantities except in those small regions with fenestrated capillaries (circumventricular organs). The situation became understandable when luminal (blood facing) and abluminal (brain facing) membranes were isolated and studied separately. Facilitative transport of glutamate and glutamine exists only on the luminal membranes, whereas Na+-dependent transport systems for glutamate, glutamine, and some other amino acids are present only on the abluminal membrane. The Na+-dependent cotransporters of the abluminal membrane are in a position to actively transport amino acids from the extracellular fluid (ECF) into the endothelial cells of the BBB. These powerful secondary active transporters couple with the energy of the Na+-gradient to move glutamate and glutamine into endothelial cells, whereupon glutamate can exit to the blood on the luminal facilitative glutamate transporter. Glutamine may also exit the brain via separate facilitative transport system that exists on the luminal membranes, or glutamine can be hydrolyzed to glutamate within the BBB, thereby releasing ammonia that is freely diffusible. The γ-glutamyl cycle participates indirectly by producing oxoproline (pyroglutamate), which stimulates almost all secondary active transporters yet discovered in the abluminal membranes of the BBB. PMID:27740595

  11. PPAR-α, a lipid-sensing transcription factor, regulates blood-brain barrier efflux transporter expression.

    PubMed

    More, Vijay R; Campos, Christopher R; Evans, Rebecca A; Oliver, Keith D; Chan, Gary Ny; Miller, David S; Cannon, Ronald E

    2017-04-01

    Lipid sensor peroxisome proliferator-activated receptor alpha (PPAR- α) is the master regulator of lipid metabolism. Dietary release of endogenous free fatty acids, fibrates, and certain persistent environmental pollutants, e.g. perfluoroalkyl fire-fighting foam components, are peroxisome proliferator-activated receptor alpha ligands. Here, we define a role for peroxisome proliferator-activated receptor alpha in regulating the expression of three ATP-driven drug efflux transporters at the rat and mouse blood-brain barriers: P-glycoprotein (Abcb1), breast cancer resistance protein (Bcrp/Abcg2), and multidrug resistance-associated protein 2 (Mrp2/Abcc2). Exposing isolated rat brain capillaries to linoleic acid, clofibrate, or PKAs increased the transport activity and protein expression of the three ABC transporters. These effects were blocked by the PPAR- α antagonist, GW6471. Dosing rats with 20 mg/kg or 200 mg/kg of clofibrate decreased the brain accumulation of the P-glycoprotein substrate, verapamil, by 50% (in situ brain perfusion; effects blocked by GW6471) and increased P-glycoprotein expression and activity in capillaries ex vivo. Fasting C57Bl/6 wild-type mice for 24 h increased both serum lipids and brain capillary P-glycoprotein transport activity. Fasting did not alter P-glycoprotein activity in PPAR- α knockout mice. These results indicate that hyperlipidemia, lipid-lowering fibrates and exposure to certain fire-fighting foam components activate blood-brain barrier peroxisome proliferator-activated receptor alpha, increase drug efflux transporter expression and reduce drug delivery to the brain.

  12. Glial Cell Calcium Signaling Mediates Capillary Regulation of Blood Flow in the Retina

    PubMed Central

    Biesecker, Kyle R.; Srienc, Anja I.; Shimoda, Angela M.; Agarwal, Amit; Bergles, Dwight E.; Kofuji, Paulo

    2016-01-01

    The brain is critically dependent on the regulation of blood flow to nourish active neurons. One widely held hypothesis of blood flow regulation holds that active neurons stimulate Ca2+ increases in glial cells, triggering glial release of vasodilating agents. This hypothesis has been challenged, as arteriole dilation can occur in the absence of glial Ca2+ signaling. We address this controversy by imaging glial Ca2+ signaling and vessel dilation in the mouse retina. We find that sensory stimulation results in Ca2+ increases in the glial endfeet contacting capillaries, but not arterioles, and that capillary dilations often follow spontaneous Ca2+ signaling. In IP3R2−/− mice, where glial Ca2+ signaling is reduced, light-evoked capillary, but not arteriole, dilation is abolished. The results show that, independent of arterioles, capillaries actively dilate and regulate blood flow. Furthermore, the results demonstrate that glial Ca2+ signaling regulates capillary but not arteriole blood flow. SIGNIFICANCE STATEMENT We show that a Ca2+-dependent glial cell signaling mechanism is responsible for regulating capillary but not arteriole diameter. This finding resolves a long-standing controversy regarding the role of glial cells in regulating blood flow, demonstrating that glial Ca2+ signaling is both necessary and sufficient to dilate capillaries. While the relative contributions of capillaries and arterioles to blood flow regulation remain unclear, elucidating the mechanisms that regulate capillary blood flow may ultimately lead to the development of therapies for treating diseases where blood flow regulation is disrupted, including Alzheimer's disease, stroke, and diabetic retinopathy. This finding may also aid in revealing the underlying neuronal activity that generates BOLD fMRI signals. PMID:27605617

  13. Spontaneous oscillations of capillary blood flow in artificial microvascular networks.

    PubMed

    Forouzan, Omid; Yang, Xiaoxi; Sosa, Jose M; Burns, Jennie M; Shevkoplyas, Sergey S

    2012-09-01

    Previous computational studies have suggested that the capillary blood flow oscillations frequently observed in vivo can originate spontaneously from the non-linear rheological properties of blood, without any regulatory input. Testing this hypothesis definitively in experiments involving real microvasculature has been difficult because in vivo the blood flow in capillaries is always actively controlled by the host. The objective of this study was to test the hypothesis experimentally and to investigate the relative contribution of different blood cells to the capillary blood flow dynamics under static boundary conditions and in complete isolation from the active regulatory mechanisms mediated by the blood vessels in vivo. To accomplish this objective, we passed whole blood and re-constituted blood samples (purified red blood cells suspended in buffer or in autologous plasma) through an artificial microvascular network (AMVN) comprising completely inert, microfabricated vessels with the architecture inspired by the real microvasculature. We found that the flow of blood in capillaries of the AMVN indeed oscillates with characteristic frequencies in the range of 0-0.6 Hz, which is in a very good agreement with previous computational studies and in vivo observations. We also found that the traffic of leukocytes through the network (typically neglected in computational modeling) plays an important role in generating the oscillations. This study represents the key piece of experimental evidence in support of the hypothesis that spontaneous, self-sustained oscillations of capillary blood flow can be generated solely by the non-linear rheological properties of blood flowing through microvascular networks, and provides an insight into the mechanism of this fundamentally important microcirculatory phenomenon. Copyright © 2012 Elsevier Inc. All rights reserved.

  14. Role of the hippocampal-entorhinal loop in temporal lobe epilepsy: extra- and intracellular study in the isolated guinea pig brain in vitro.

    PubMed

    Paré, D; deCurtis, M; Llinás, R

    1992-05-01

    This article introduces a new experimental paradigm for the study of temporal lobe epilepsy. This approach utilizes the isolated guinea pig brain in vitro preparation, which generates a pattern of hypersynchronous neuronal activity similar to the peculiar 8-30 Hz rhythm characterizing stereoelectroencephalographic hippocampal recordings in human temporal lobe epilepsy. The present report describes an attempt to identify the functional events underlying the epileptiform activities observed in this preparation. Rhythmic epileptiform discharges (EDs), here defined as population spikes (PSs) recorded from somata or dendritic layers, were induced in the hippocampal formation of the isolated guinea pig brain maintained in vitro by tetanic stimulation of the entorhinal cortex (EC). Two patterns of EDs were distinguished by performing simultaneous field potential recordings along the dentate gyrus (DG), EC, CA1, and CA3. During stage 1, the first self-sustained EDs were isolated PSs occurring at a frequency of 2-3 Hz at all levels of the entorhinal-hippocampal loop, the only exception being the DG, where no signs of synchronized neuronal discharge could be found. Over the next 30-50 sec, the temporal organization of these EDs changed dramatically. During stage 2, at all levels of the entorhinal-hippocampal loop, EDs occurred in 0.3-0.5 sec trains of 16-25 Hz population spikes interrupted by 0.7-1.3 sec silent periods. The transition between stages 1 and 2 coincided with the occurrence of population spikes in the DG. Laminar analyses and multiple simultaneous field potential recordings revealed that the trains of EDs observed in stage 2 resulted from the repetitive, sequential activation of the hippocampal-entorhinal loop. In the transverse axis, the earliest event usually occurred in the CA3 region. Thereafter, population spikes occurred sequentially in the CA1 region, EC, DG, and back to the CA3 region. Intracellular recordings confirmed that the EDs recorded

  15. Step-gradient capillary electrochromatography.

    PubMed

    Euerby, M R; Gilligan, D; Johnson, C M; Bartle, K D

    1997-10-01

    The analytical benefits of using a step-gradient in capillary electrochromatography (CEC) are demonstrated. The application of step-gradient CEC to the analysis of six diuretics of widely differing lipophilicities was evaluated and shown to result in a marked reduction in the analysis time and an improvement in the peak shape for later-eluting lipophilic components. When the step-gradient approach was performed in an automated mode, the retention time RSD for repeated injections was below 1%.

  16. Capillary electrophoresis systems and methods

    DOEpatents

    Dorairaj, Rathissh [Hillsboro, OR; Keynton, Robert S [Louisville, KY; Roussel, Thomas J [Louisville, KY; Crain, Mark M [Georgetown, IN; Jackson, Douglas J [New Albany, IN; Walsh, Kevin M [Louisville, KY; Naber, John F [Goshen, KY; Baldwin, Richard P [Louisville, KY; Franco, Danielle B [Mount Washington, KY

    2011-08-02

    An embodiment of the invention is directed to a capillary electrophoresis apparatus comprising a plurality of separation micro-channels. A sample loading channel communicates with each of the plurality of separation channels. A driver circuit comprising a plurality of electrodes is configured to induce an electric field across each of the plurality of separation channels sufficient to cause analytes in the samples to migrate along each of the channels. The system further comprises a plurality of detectors configured to detect the analytes.

  17. Capillary stretching of elastic fibers

    NASA Astrophysics Data System (ADS)

    Protiere, Suzie; Stone, Howard A.; Duprat, Camille

    2014-11-01

    Fibrous media consisting of constrained flexible fibers can be found in many engineered systems (membranes in filters, woven textile, matted paper). When such materials interact with a liquid, the presence of liquid/air interfaces induces capillary forces that deform the fibers. To model this interaction we study the behaviour of a finite volume of liquid deposited on two parallel flexible fibers clamped at both ends. A tension along the fibers is imposed and may be varied. We show that the system undergoes various morphological changes as the interfiber distance, the elasticity and the tension of the fibers are varied. For a certain range of parameters, the liquid spreads along the fibers and pulls them together, leading to the ``zipping'' of the fibers. This capillary adhesion can then be enhanced or reduced by changing the tension within the fibers. We will show that balancing stretching and capillary forces allows the prediction of this transition as well as the conditions for which detachment of the fibers occurs. These results may be used to prevent the clogging of fibrous membranes or to optimize the capture of liquids.

  18. Capillary Electrophoresis - Optical Detection Systems

    SciTech Connect

    Sepaniak, M. J.

    2001-08-06

    Molecular recognition systems are developed via molecular modeling and synthesis to enhance separation performance in capillary electrophoresis and optical detection methods for capillary electrophoresis. The underpinning theme of our work is the rational design and development of molecular recognition systems in chemical separations and analysis. There have been, however, some subtle and exciting shifts in our research paradigm during this period. Specifically, we have moved from mostly separations research to a good balance between separations and spectroscopic detection for separations. This shift is based on our perception that the pressing research challenges and needs in capillary electrophoresis and electrokinetic chromatography relate to the persistent detection and flow rate reproducibility limitations of these techniques (see page 1 of the accompanying Renewal Application for further discussion). In most of our work molecular recognition reagents are employed to provide selectivity and enhance performance. Also, an emerging trend is the use of these reagents with specially-prepared nano-scale materials. Although not part of our DOE BES-supported work, the modeling and synthesis of new receptors has indirectly supported the development of novel microcantilevers-based MEMS for the sensing of vapor and liquid phase analytes. This fortuitous overlap is briefly covered in this report. Several of the more significant publications that have resulted from our work are appended. To facilitate brevity we refer to these publications liberally in this progress report. Reference is also made to very recent work in the Background and Preliminary Studies Section of the Renewal Application.

  19. Capillary-driven automatic packaging.

    PubMed

    Ding, Yuzhe; Hong, Lingfei; Nie, Baoqing; Lam, Kit S; Pan, Tingrui

    2011-04-21

    Packaging continues to be one of the most challenging steps in micro-nanofabrication, as many emerging techniques (e.g., soft lithography) are incompatible with the standard high-precision alignment and bonding equipment. In this paper, we present a simple-to-operate, easy-to-adapt packaging strategy, referred to as Capillary-driven Automatic Packaging (CAP), to achieve automatic packaging process, including the desired features of spontaneous alignment and bonding, wide applicability to various materials, potential scalability, and direct incorporation in the layout. Specifically, self-alignment and self-engagement of the CAP process induced by the interfacial capillary interactions between a liquid capillary bridge and the top and bottom substrates have been experimentally characterized and theoretically analyzed with scalable implications. High-precision alignment (of less than 10 µm) and outstanding bonding performance (up to 300 kPa) has been reliably obtained. In addition, a 3D microfluidic network, aligned and bonded by the CAP technique, has been devised to demonstrate the applicability of this facile yet robust packaging technique for emerging microfluidic and bioengineering applications.

  20. Isolation, purification, and partial characterization of a membrane-bound Cl(-)/HCO3(-)-activated ATPase complex from rat brain with sensitivity to GABAAergic ligands.

    PubMed

    Menzikov, Sergey A

    2017-02-07

    This study describes the isolation and purification of a protein complex with [Formula: see text]-ATPase activity and sensitivity to GABAAergic ligands from rat brain plasma membranes. The ATPase complex was enriched using size-exclusion, affinity, and ion-exchange chromatography. The fractions obtained at each purification step were subjected to SDS-polyacrylamide gel electrophoresis (SDS-PAGE), which revealed four subunits with molecular mass ∼48, 52, 56, and 59 kDa; these were retained at all stages of the purification process. Autoradiography revealed that the ∼52 and 56 kDa subunits could bind [(3)H]muscimol. The [Formula: see text]-ATPase activity of this enriched protein complex was regulated by GABAAergic ligands but was not sensitive to blockers of the NKCC or KCC cotransporters.

  1. Baratin, a nonamidated neurostimulating neuropeptide, isolated from cockroach brain: distribution and actions in the cockroach and locust nervous systems.

    PubMed

    Nässel, D R; Persson, M G; Muren, J E

    2000-06-26

    During the purification of tachykinin-related peptides from the brain of the cockroach Leucophaea maderae, a few other peptides were collected in adjacent high-performance liquid chromatography fractions. Edman degradation, mass spectrometry, and chemical synthesis revealed that one of these peptides had the sequence DNSQWGGFA. This nonamidated nonapeptide was designated baratin and appears not to be related to any known insect peptide. Baratin was not found to be bioactive in the L. maderae hindgut or oviduct muscle contraction assay. (Both synthetic nonamidated and amidated baratin were tested.) To screen for possible sites of action, we raised a rabbit antiserum to baratin. We found baratin-immunoreactive (BAR-IR) interneurons throughout the cockroach central nervous system. Some prominent brain neuropils were supplied by BAR-IR neuron processes: the central body, the calyx, and lobes of the mushroom bodies, parts of the optic lobe, and the tritocerebral neuropil. Additionally we found BAR-IR neurosecretory cells in the median neurosecretory cell group with processes supplying the storage lobe of the corpora cardiaca. In each of the thoracic and abdominal ganglia processes of BAR-IR projection neurons and local neurons were seen. The baratin antiserum also labeled neurons in the brain of the locust Locusta migratoria, some of which are similar to those of the cockroach. A prominent system of interganglionic BAR-IR processes was found in the locust subesophageal, thoracic, and abdominal ganglia. This was formed by four large projection neurons with cell bodies in the abdominal ganglia A1-2. The processes of these BAR-IR neurons are distributed dorsally and laterally in each of the ventral nerve cord ganglia. When baratin (10(-6)-10(-4) M) was applied to desheathed abdominal ganglia of locusts and cockroaches, we could monitor bursts of action potentials in neurons with axons in the anterior abdominal nerve (nerve 1), but not in the posterior nerve (nerve 2). In

  2. Targeted unlabeled multiple reaction monitoring analysis of cell markers for the study of sample heterogeneity in isolated rat brain cortical microvessels.

    PubMed

    Gomez-Zepeda, David; Chaves, Catarina; Taghi, Méryam; Sergent, Philippe; Liu, Wang-Qing; Chhuon, Cérina; Vidal, Michel; Picard, Martin; Thioulouse, Elizabeth; Broutin, Isabelle; Guerrera, Ida-Chiara; Scherrmann, Jean-Michel; Parmentier, Yannick; Decleves, Xavier; Menet, Marie-Claude

    2017-08-01

    Liquid chromatography coupled to tandem mass spectrometry-based targeted absolute protein quantification (in fmol of the analyte protein per μg of total protein) is employed for the molecular characterization of the blood-brain barrier using isolated brain microvessels. Nevertheless, the heterogeneity of the sample regarding the levels of different cells co-isolated within the microvessels and bovine serum albumin (BSA) contamination (from buffers) are not always evaluated. We developed an unlabeled targeted liquid chromatography coupled to tandem mass spectrometry method to survey the levels of endothelial cells (ECs), astrocytes, and pericytes, as well as BSA contaminant in rat cortical microvessels. Peptide peak identities were evaluated using a spectral library and chromatographic parameters. Sprague-Dawley rat microvessels obtained on three different days were analyzed with this method complemented by an absolute quantification multiple reaction monitoring method for transporter proteins P-gp, Bcrp, and Na(+) /K(+) ATPase pump using stable isotope labeled peptides as internal standard. Inter-day differences in the cell markers and BSA contamination were observed. Levels of cell markers correlated positively between each other. Then, the correlation between cell marker proteins and transporter proteins was evaluated to choose the best EC marker protein for protein quantification normalization. The membrane protein Pecam-1 showed a very high correlation with the EC-specific transporter P-gp (Pearson product-moment correlation coefficient (r) > 0.89) and moderate to high with Bcrp (r ≥ 0.77), that can be found also in pericytes and astrocytes. Therefore, Pecam-1 was selected as a marker for the normalization of the quantification of the proteins of endothelial cells. © 2017 International Society for Neurochemistry.

  3. Fluid Delivery System For Capillary Electrophoretic Applications.

    DOEpatents

    Li, Qingbo; Liu, Changsheng; Kane, Thomas E.; Kernan, John R.; Sonnenschein, Bernard; Sharer, Michael V.

    2002-04-23

    An automated electrophoretic system is disclosed. The system employs a capillary cartridge having a plurality of capillary tubes. The cartridge has a first array of capillary ends projecting from one side of a plate. The first array of capillary ends are spaced apart in substantially the same manner as the wells of a microtitre tray of standard size. This allows one to simultaneously perform capillary electrophoresis on samples present in each of the wells of the tray. The system includes a stacked, dual carrousel arrangement to eliminate cross-contamination resulting from reuse of the same buffer tray on consecutive executions from electrophoresis. The system also has a gel delivery module containing a gel syringe/a stepper motor or a high pressure chamber with a pump to quickly and uniformly deliver gel through the capillary tubes. The system further includes a multi-wavelength beam generator to generate a laser beam which produces a beam with a wide range of wavelengths. An off-line capillary reconditioner thoroughly cleans a capillary cartridge to enable simultaneous execution of electrophoresis with another capillary cartridge. The streamlined nature of the off-line capillary reconditioner offers the advantage of increased system throughput with a minimal increase in system cost.

  4. Progression of Diabetic Capillary Occlusion: A Model

    PubMed Central

    Gens, John Scott; Glazier, James A.; Burns, Stephen A.; Gast, Thomas J.

    2016-01-01

    An explanatory computational model is developed of the contiguous areas of retinal capillary loss which play a large role in diabetic maculapathy and diabetic retinal neovascularization. Strictly random leukocyte mediated capillary occlusion cannot explain the occurrence of large contiguous areas of retinal ischemia. Therefore occlusion of an individual capillary must increase the probability of occlusion of surrounding capillaries. A retinal perifoveal vascular sector as well as a peripheral retinal capillary network and a deleted hexagonal capillary network are modelled using Compucell3D. The perifoveal modelling produces a pattern of spreading capillary loss with associated macular edema. In the peripheral network, spreading ischemia results from the progressive loss of the ladder capillaries which connect peripheral arterioles and venules. System blood flow was elevated in the macular model before a later reduction in flow in cases with progression of capillary occlusions. Simulations differing only in initial vascular network structures but with identical dynamics for oxygen, growth factors and vascular occlusions, replicate key clinical observations of ischemia and macular edema in the posterior pole and ischemia in the retinal periphery. The simulation results also seem consistent with quantitative data on macular blood flow and qualitative data on venous oxygenation. One computational model applied to distinct capillary networks in different retinal regions yielded results comparable to clinical observations in those regions. PMID:27300722

  5. Quantifying the cleanliness of glass capillaries.

    PubMed

    Bowman, C L

    1998-01-01

    I used capillary rise methods to investigate the lumenal surface properties of quartz (fused silica, Amersil T-08), borosilicate (Corning 7800), and high-lead glass (Corning 0010) capillaries commonly used to make patch pipets. I calculated the capillary rise and contact angle for water and methanol from weight measurements. The capillary rise was compared with the theoretical maximum value calculated by assuming each fluid perfectly wetted the lumenal surface of the glass (i.e., zero contact angle, which reflects the absence of surface contamination). For borosilicate, high-lead, and quartz capillaries, the rise for water was substantially less than the theoretical maximum rise. Exposure of the borosilicate, lead, and quartz capillaries to several cleaning methods resulted in substantially better--but not perfect--agreement between the theoretical maximum rise and calculated capillary rise. By contrast, the capillary rise for methanol was almost identical in untreated and cleaned capillaries, but less than its theoretical maximum rise. The residual discrepancy between the observed and theoretical rise for water could not be improved on by trying a variety of cleaning procedures, but some cleaning methods were superior to others. The water solubility of the surface contaminants, deduced from the effectiveness of repeated rinsing, was different for each of the three types of capillaries examined: Corning 7800 > quartz > Corning 0010. A surface film was also detected in quatz tubing with an internal filament. I conclude that these borosilicate, quartz, and high-lead glass capillaries have a film on the lumenal surface, which can be removed using appropriate cleaning methods. The surface contaminants may be unique to each type of capillary and may also be hydrophobic. Two simple methods are presented to quantitate the cleanliness of glass capillary tubing commonly used to make pipets for studies of biological membranes. It is not known if the surface film is of

  6. A new dynamic in vitro modular capillaries-venules modular system: Cerebrovascular physiology in a box

    PubMed Central

    2013-01-01

    Background The study of the cerebrovascular physiology is crucial to understand the pathogenesis of neurological disease and the pharmacokinetic of drugs. Appropriate models in vitro often fail to represent in vivo physiology. To address these issues we propose the use of a novel artificial vascular system that closely mimics capillary and venous segments of human cerebrovasculature while also allowing for an extensive control of the experimental variables and their manipulation. Results Using hollow fiber technology, we modified an existing dynamic artificial model of the blood–brain barrier (BBB) (DIV-capillary) to encompass the distal post-capillary (DIV-venules) segments of the brain circulatory system. This artificial brain vascular system is comprised of a BBB module serially connected to a venule segment. A pump generates a pulsatile flow with arterial pressure feeding the system. The perfusate of the capillary module achieves levels of shear stress, pressure, and flow rate comparable to what observed in situ. Endothelial cell exposure to flow and abluminal astrocytic stimuli allowed for the formation of a highly selective capillary BBB with a trans-endothelial electrical resistance (TEER; >700 ohm cm2) and sucrose permeability (< 1X10-u cm/sec) comparable to in vivo. The venule module, which attempted to reproduce features of the hemodynamic microenvironment of venules, was perfused by media resulting in shear stress and intraluminal pressure levels lower than those found in capillaries. Because of altered cellular and hemodynamic factors, venule segments present a less stringent vascular bed (TEER <250 Ohm cm2; Psucrose > 1X10-4 cm/sec) than that of the BBB. Abluminal human brain vascular smooth muscle cells were used to reproduce the venular abluminal cell composition. Conclusion The unique characteristics afforded by the DIV-BBB in combination with a venule segment will realistically expand our ability to dissect and study the physiological and

  7. Multiple forms of phosphatase from human brain: isolation and partial characterization of affi-gel blue nonbinding phosphatase activities.

    PubMed

    Cheng, L Y; Wang, J Z; Gong, C X; Pei, J J; Zaidi, T; Grundke-Iqbal, I; Iqbal, K

    2001-04-01

    Phosphatases extracted from a human brain were resolved into two main groups, namely affi-gel blue-binding phosphatases and affi-gel blue-nonbinding phosphatases. Affi-gel blue binding phosphatases were further separated into four different phosphatase activities, designated P1-P4, and described previously. In the present study we describe the affi-gel blue-nonbinding phosphatases which were separated into seven different phosphatase activities, designated P5-P11 by poly-(L-lysine)-agarose and aminohexyl Sepharose 4B chromatographies. These seven phosphatase activities were active toward nonprotein phosphoester. P7-P11 and to some extent P5 could also dephosphorylate a phosphoprotein. They displayed different enzyme kinetics. On the basis of activity peak, the apparent molecular mass as estimated by Sephadex G-200 column chromatography for P5 was 49 kDa; P6, 32 kDa; P7, 150 kDa; P8, 250 kDa; P9, 165 kDa; P10, 90 kDa and P11, 165 kDa. Immunoblot analysis indicated that P8-P11 may belong to PP2B family, whereas P7 may associate with PP2A. The phosphatases P7-P11 were found to be effective in the dephosphorylation of Alzheimer's disease abnormally hyperphosphorylated tau. The resulting dephosphorylated tau regained its activity in promoting the microtubule assembly, suggesting that P7-P11 might regulate the phosphorylation of tau protein in the brain.

  8. Isolation of the human genomic brain-2/N-Oct 3 gene (POUF3) and assignment to chromosome 6q16

    SciTech Connect

    Atanasoski, S.; Malipiero, U.; Shreiber, E.

    1995-03-20

    N-Oct 3 is a human POU domain transcription factor that binds to the octamer sequence ATGCAAAT. The protein is expressed in the central nervous system during development and in adult brain. We have isolated and characterized genomic clones encoding the human N-Oct 3 gene (HGMW-approved symbol POUF3). Comparison of the structure of these clones with the N-Oct 3 cDNA revealed that POUF3 is an intronless gene. Sequencing of 650 bp of the promoter region showed 84% sequence identity of POUF3 with its murine homologue, the brain-2 (designated brn-2) gene. Whereas both POUF3 and brn-2 lack a TATA box, consensus sequences for AP-2, GCF, and SP1 transcription factors were identified within the highly conserved 5{prime}- flanking region. These sequences may play a crucial role for the tissue-specific transcription activation of the POUF3 gene. Southern blotting and in situ hybridization localized the human POUF3 gene to chromosome 6q16. 61 refs., 5 figs., 1 tab.

  9. Isolation of the human genomic brain-2/N-Oct 3 gene (POUF3) and assignment to chromosome 6q16.

    PubMed

    Atanasoski, S; Toldo, S S; Malipiero, U; Schreiber, E; Fries, R; Fontana, A

    1995-03-20

    N-Oct 3 is a human POU domain transcription factor that binds to the octamer sequence ATGCAAAT. The protein is expressed in the central nervous system during development and in adult brain. We have isolated and characterized genomic clones encoding the human N-Oct 3 gene (HGMW-approved symbol POUF3). Comparison of the structure of these clones with the N-Oct 3 cDNA revealed that POUF3 is an intronless gene. Sequencing of 650 bp of the promoter region showed 84% sequence identity of POUF3 with its murine homologue, the brain-2 (designated brn-2) gene. Whereas both POUF3 and brn-2 lack a TATA box, consensus sequences for AP-2, GCF, and SP1 transcription factors were identified within the highly conserved 5'-flanking region. These sequences may play a crucial role for the tissue-specific transcription activation of the POUF3 gene. Southern blotting and in situ hybridization localized the human POUF3 gene to chromosome 6q16.

  10. Impact of capillary flow hydrodynamics on carrier-mediated transport of opioid derivatives at the blood-brain barrier, based on pH-dependent Michaelis-Menten and Crone-Renkin analyses.

    PubMed

    Yusof, Siti R; Abbott, N Joan; Avdeef, Alex

    2017-08-30

    Most studies of blood-brain barrier (BBB) permeability and transport are conducted at a single pH, but more detailed information can be revealed by using multiple pH values. A pH-dependent biophysical model was applied to the mechanistic analysis of published pH-dependent BBB luminal uptake data from three opioid derivatives in rat: pentazocine (Suzuki et al., 2002a, 2002b), naloxone (Suzuki et al., 2010a), and oxycodone (Okura et al., 2008). Two types of data were processed: in situ brain perfusion (ISBP) and brain uptake index (BUI). The published perfusion data were converted to apparent luminal permeability values, Papp, and analyzed by the pCEL-X program (Yusof et al., 2014), using the pH-dependent Crone-Renkin equation (pH-CRE) to determine the impact of cerebrovascular flow on the Michaelis-Menten transport parameters (Avdeef and Sun, 2011). For oxycodone, the ISBP data had been measured at pH7.4 and 8.4. The present analysis indicates a 7-fold lower value of the cerebrovascular flow velocity, Fpf, than that expected in the original study. From the pyrilamine-inhibited data, the flow-corrected passive intrinsic permeability value was determined to be P0=398×10(-6)cm·s(-1). The uptake data indicate that the neutral form of oxycodone is affected by a transporter at pH8.4. The extent of the cation uptake was less certain from the available data. For pentazocine, the brain uptake by the BUI method had been measured at pH5.5, 6.5, and 7.4, in a concentration range 0.1-40mM. Under similar conditions, ISBP data were also available. The pH-CRE determined values of Fpf from both methods were nearly the same, and were smaller than the expected value in the original publication. The transport of the cationic pentazocine was not fully saturated at pH5.5 at 40mM. The transport of the neutral species at pH7.4 appeared to reach saturation at 40mM pentazocine concentration, but not at 12mM. In the case of naloxone, a pH-dependent Michaelis-Menten equation (p

  11. Joule heating in packed capillaries used in capillary electrochromatography.

    PubMed

    Rathore, Anurag S; Reynolds, Kimberly J; Colón, Luis A

    2002-09-01

    Effective heat dissipation is critical for reproducible and efficient separations in electrically driven separation systems. Flow rate, retention kinetics, and analyte diffusion rates are some of the characteristics that are affected by variation in the temperature of the mobile phase inside the column. In this study, we examine the issue of Joule heating in packed capillary columns used in capillary electrochromatography (CEC). As almost all commonly used CEC packings are poor thermal conductors, it is assumed that the packing particles do not conduct heat and heat transfer is solely through the mobile phase flowing through the system. The electrical conductivity of various mobile phases was measured at different temperatures by a conductivity meter and the temperature coefficient for each mobile phase was calculated. This was followed by measurement of the electrical current at several applied voltages to calculate the conductivity of the solution within the column as a function of the applied voltage. An overall increase in the conductivity is attributed to Joule heating within the column, while a constant conductivity means good heat dissipation. A plot of conductivity versus applied voltage was used as the indicator of poor heat dissipation. Using theories that have been proposed earlier for modeling of Joule heating effects in capillary electrophoresis (CE), we estimated the temperature within CEC columns. Under mobile and stationary phase conditions typically used in CEC, heat dissipation was found to be not always efficient. Elevated temperatures within the columns in excess of 23 degrees C above ambient temperature were calculated for packed columns, and about 35 degrees C for an open column, under a given set of conditions. The results agree with recently published experimental findings with nuclear magnetic resonance (NMR) thermometry, and Raman spectroscopic measurements.

  12. Isolation and characterization of a factor from rat liver which inhibits /sup 3/H-nicotine binding to brain nicotine receptors

    SciTech Connect

    Noggle, H.D.

    1986-01-01

    In studies of /sup 3/H-nicotine binding sites in rat brain membranes, it was observed that crude rat liver homogenates are capable of inhibiting this binding. The purpose of this work was to isolate from the liver homogenate the factor (or factors) responsible for this nicotinelike activity and to identify it. Isolation procedures, including heat denaturation, ultrafiltration, and reverse phase high pressure liquid chromatography, resulted in an incompletely purified substance whose chemical properties were compatible with those of dimethylaminoethyl (DMAE) carbamate, as reflected in infrared, ultraviolet, NMR, and mass spectra; in HPLC elution characteristics into mobile phases; and in IC/sub 50/'s with respect to /sup 3/H-nicotine binding to neural membranes, although additional purification and characterization will be necessary to confirm or reject this identification. This compound has not previously been described in mammalian tissue, although its two component functional moieties, dimethylaminoethanol and carbamic acid, are present in the liver. The binding properties of DMAE carbamate are compared with those of structurally and pharmacological related compounds to assess its potential as an endogenous cholinergic ligand.

  13. Usefulness of plasma brain natriuretic peptide measurement and tissue Doppler imaging in identifying isolated left ventricular diastolic dysfunction without heart failure.

    PubMed

    Goto, Toshihiko; Ohte, Nobuyuki; Wakami, Kazuaki; Asada, Kaoru; Fukuta, Hidekatsu; Mukai, Seiji; Tani, Tomomitsu; Kimura, Genjiro

    2010-07-01

    Left ventricular (LV) diastolic dysfunction carries a substantial risk for the subsequent development of heart failure and reduced survival, even when it is asymptomatic. Plasma brain natriuretic peptide (BNP) level and tissue Doppler imaging indexes provide powerful incremental assessment of LV diastolic function. Accordingly, the aim of this study was to clarify whether these methodologies could identify LV diastolic dysfunction without heart failure in 280 patients with preserved LV ejection fractions (> or =50%) who underwent echocardiography and cardiac catheterization for the evaluation of coronary artery disease. Patients were classified into 2 groups, those with diastolic dysfunction (tau > or =48 ms; n = 91) and those with normal diastolic function (tau <48 ms; n = 189). Plasma BNP > or =22.4 pg/ml, an unexpectedly low value, had sensitivity of 74.7% and specificity of 60.8% for identifying isolated LV diastolic dysfunction; the combined use of BNP > or =22.4 pg/mL and mitral annular velocity during early diastole <7.4 cm/s had relatively low sensitivity of 44.0% but high specificity of 86.8%. In conclusion, using plasma BNP level and with the combination of BNP level and mitral annular velocity during early diastole, invasively proved isolated LV diastolic dysfunction without heart failure could be identified in patients with coronary artery disease.

  14. Histogenesis of hippocampus and neocortex isolated from postnatal rats in organotypic roller tube cultures of floating brain sections.

    PubMed

    Aleksandrova, O P; Lyzhin, A A; Viktorov, I V

    2001-09-01

    Floating sections of the hippocampus and neocortex isolated from 6-10-day-old rats retained their spatial cell organization after 2-week roller-tube culturing. Cell structure in CA1, Ca2, and CA3 fields, polymorphic layer of the fascia dentata and its medial and lateral limbs were revealed in sections of the dorsal hippocampus. In neocortical sections, cortical neurons and subcortical structure were preserved, however, some cortical fragments changed their configuration and formed spherical structures, where cortical neurons were located in the external layer without forming typical of neocortex 6-layer structure.

  15. Capillary pumped loop body heat exchanger

    NASA Technical Reports Server (NTRS)

    Swanson, Theodore D. (Inventor); Wren, deceased, Paul (Inventor)

    1998-01-01

    A capillary pumped loop for transferring heat from one body part to another body part, the capillary pumped loop comprising a capillary evaporator for vaporizing a liquid refrigerant by absorbing heat from a warm body part, a condenser for turning a vaporized refrigerant into a liquid by transferring heat from the vaporized liquid to a cool body part, a first tube section connecting an output port of the capillary evaporator to an input of the condenser, and a second tube section connecting an output of the condenser to an input port of the capillary evaporator. A wick may be provided within the condenser. A pump may be provided between the second tube section and the input port of the capillary evaporator. Additionally, an esternal heat source or heat sink may be utilized.

  16. Microfluidic PMMA interfaces for rectangular glass capillaries

    NASA Astrophysics Data System (ADS)

    Evander, Mikael; Tenje, Maria

    2014-02-01

    We present the design and fabrication of a polymeric capillary fluidic interface fabricated by micro-milling. The design enables the use of glass capillaries with any kind of cross-section in complex microfluidic setups. We demonstrate two different designs of the interface; a double-inlet interface for hydrodynamic focusing and a capillary interface with integrated pneumatic valves. Both capillary interfaces are presented together with examples of practical applications. This communication shows the design optimization and presents details of the fabrication process. The capillary interface opens up for the use of complex microfluidic systems in single-use glass capillaries. They also enable simple fabrication of glass/polymer hybrid devices that can be beneficial in many research fields where a pure polymer chip negatively affects the device's performance, e.g. acoustofluidics.

  17. Copper transport to the brain by the blood-brain barrier and blood-CSF barrier.

    PubMed

    Choi, Byung-Sun; Zheng, Wei

    2009-01-12

    The mechanism of copper (Cu) transport into the brain is unclear. This study evaluated the main species and route of Cu transport into the brain using in situ brain perfusion technique, and assessed the levels of mRNA encoding Cu transporters using real time RT-PCR. Free (64)Cu uptake in rat choroid plexus (CP), where the blood-cerebrospinal fluid barrier (BCB) is primarily located, is about 50 and 1000 times higher than (64)Cu-albumin and (64)Cu-ceruloplasmin uptake, respectively. The unidirectional transport rate constants (K(in)) for Cu in the CP and brain capillaries of the blood-brain barrier (BBB) were 1034 and 319 microl/s/g, respectively, while K(in) in CSF and capillary-depleted parenchyma were much reduced, 0.8 and 112 microl/s/g, respectively. The K(in) in cerebellum was significantly lower than that in hippocampus. The mRNAs encoding Cu transporter-1 (Ctr1) and ATP7A were higher in the CP than those in brain capillaries and parenchyma, whereas ATP7B mRNA was higher in brain capillaries than those in the CP and brain parenchyma. Taken together, these data suggest that the expression of Cu transporters is higher in brain barriers than in brain parenchyma; the Cu transport into the brain is mainly achieved through the BBB as a free Cu ion and the BCB may serve as a main regulatory site of Cu in the CSF.

  18. [Ultrastructural changes in the MP3 neuron of the mollusk Lymnaea stagnalis after cryopreservation of the isolated brain].

    PubMed

    Dmitrieva, E V; Moshkov, D A; Gakhova, E N

    2006-01-01

    Investigation of a possibility of long-term storage of frozen (-196 degrees C) viable neurons and nervous tissue is one of the central present day problems. In this study ultrastructural changes in neurons of frozen-thawed snail brain were examined as a function of time. We studied the influence of cryopreservation, cryoprotectant (Me2SO), cooling to 4-6 degrees C, and a prolonged incubation in physiological solution at 4-6 degrees C on dictyosomes of Golgi apparatus, endoplasmic reticulum (ER) cisternae and mitochondria. It has been found that responses of these intracellular structures of cryopreserved neurons to the above influences are similar: dissociation of Golgi dictyosomes, swelling of endoplasmic reticulum cisternae and mitochondrial cristae. Both freezing-thawing and cryoprotectant were seen to cause an increase in the number of lysosomes, liposomes, myelin-like structures, and to form large vacuoles. The structural changes in molluscan neurons caused by cryopreservation with Me2SO (2 M) were reversible.

  19. DNA Sequencing Using capillary Electrophoresis

    SciTech Connect

    Dr. Barry Karger

    2011-05-09

    The overall goal of this program was to develop capillary electrophoresis as the tool to be used to sequence for the first time the Human Genome. Our program was part of the Human Genome Project. In this work, we were highly successful and the replaceable polymer we developed, linear polyacrylamide, was used by the DOE sequencing lab in California to sequence a significant portion of the human genome using the MegaBase multiple capillary array electrophoresis instrument. In this final report, we summarize our efforts and success. We began our work by separating by capillary electrophoresis double strand oligonucleotides using cross-linked polyacrylamide gels in fused silica capillaries. This work showed the potential of the methodology. However, preparation of such cross-linked gel capillaries was difficult with poor reproducibility, and even more important, the columns were not very stable. We improved stability by using non-cross linked linear polyacrylamide. Here, the entangled linear chains could move when osmotic pressure (e.g. sample injection) was imposed on the polymer matrix. This relaxation of the polymer dissipated the stress in the column. Our next advance was to use significantly lower concentrations of the linear polyacrylamide that the polymer could be automatically blown out after each run and replaced with fresh linear polymer solution. In this way, a new column was available for each analytical run. Finally, while testing many linear polymers, we selected linear polyacrylamide as the best matrix as it was the most hydrophilic polymer available. Under our DOE program, we demonstrated initially the success of the linear polyacrylamide to separate double strand DNA. We note that the method is used even today to assay purity of double stranded DNA fragments. Our focus, of course, was on the separation of single stranded DNA for sequencing purposes. In one paper, we demonstrated the success of our approach in sequencing up to 500 bases. Other

  20. Equilibrium capillary forces with atomic force microscopy.

    PubMed

    Sprakel, J; Besseling, N A M; Leermakers, F A M; Cohen Stuart, M A

    2007-09-07

    We present measurements of equilibrium forces resulting from capillary condensation. The results give access to the ultralow interfacial tensions between the capillary bridge and the coexisting bulk phase. We demonstrate this with solutions of associative polymers and an aqueous mixture of gelatin and dextran, with interfacial tensions around 10 microN/m. The equilibrium nature of the capillary forces is attributed to the combination of a low interfacial tension and a microscopic confinement geometry, based on nucleation and growth arguments.

  1. Heuristic approach to capillary pressures averaging

    SciTech Connect

    Coca, B.P.

    1980-10-01

    Several methods are available to average capillary pressure curves. Among these are the J-curve and regression equations of the wetting-fluid saturation in porosity and permeability (capillary pressure held constant). While the regression equation seem completely empiric, the J-curve method seems to be theoretically sound due to its expression based on a relation between the average capillary radius and the permeability-porosity ratio. An analysis is given of each of these methods.

  2. Human blood-brain barrier insulin receptor.

    PubMed

    Pardridge, W M; Eisenberg, J; Yang, J

    1985-06-01

    A new model system for characterizing the human brain capillary, which makes up the blood-brain barrier (BBB) in vivo, is described in these studies and is applied initially to the investigation of the human BBB insulin receptor. Autopsy brains were obtained from the pathologist between 22-36 h postmortem and were used to isolate human brain microvessels which appeared intact on both light and phase microscopy. The microvessels were positive for human factor 8 and for a BBB-specific enzyme marker, gamma-glutamyl transpeptidase. The microvessels avidly bound insulin with a high-affinity dissociation constant, KD = 1.2 +/- 0.5 nM. The human brain microvessels internalized insulin based on acid-wash assay, and 75% of insulin was internalized at 37 degrees C. The microvessels transported insulin to the medium at 37 degrees C with a t1/2 = approximately 70 min. Little of the 125I-insulin was metabolized by the microvessels under these conditions based on the elution profile of the medium extract over a Sephadex G-50 column. Plasma membranes were obtained from the human brain microvessels and these membranes were enriched in membrane markers such as gamma-glutamyl transpeptidase or alkaline phosphatase. The plasma membranes bound 125I-insulin with and ED50 = 10 ng/ml, which was identical to the 50% binding point in intact microvessels. The human BBB plasma membranes were solubilized in Triton X-100 and were adsorbed to a wheat germ agglutinin Sepharose affinity column, indicating the BBB insulin receptor is a glycoprotein. Affinity cross-linking of insulin to the plasma membranes revealed a 127K protein that specifically binds insulin.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Capillaroscopy and the measurement of capillary pressure

    PubMed Central

    Shore, Angela C

    2000-01-01

    Capillaries play a critical role in cardiovascular function as the point of exchange of nutrients and waste products between the tissues and circulation. Studies of capillary function in man are limited by access to the vascular bed. However, skin capillaries can readily be studied by the technique of capillaroscopy which enables the investigator to assess morphology, density and blood flow velocity. It is also possible to estimate capillary pressure by direct cannulation using glass micropipettes. This review will describe the techniques used to make these assessments and will outline some of the changes that are seen in health and disease. PMID:11136289

  4. Oxygen exchange in silicone rubber capillaries.

    PubMed

    Heineken, F G; Predecki, P K; Filley, G F

    1978-06-01

    Capillaries of 7 and 12.5 mu diameter have been fabricated in silicone rubber. Whole blood treated with heparin has been perfused through these capillaries. Under flowing conditions, no clotting or other clumping effects have been observed and red cells appear to maintain a constant velocity. Oxygen transfer data to and from saline perfusing the 12.5 mu diameter capillaries have been obtained in order to determine how rapidly O2 will permeate the silicone rubber film. The data indicate that the capillaries simulate lung tissue oxygen exchange and will allow for the first time the experimental determination of oxygen exchange kinetics in flowing whole blood.

  5. Transient studies of capillary-induced flow

    NASA Technical Reports Server (NTRS)

    Reagan, M. K.; Bowman, W. J.

    1993-01-01

    This paper presents the numerical and experimental results of a study performed on the transient rise of fluid in a capillary tube. The capillary tube problem provides an excellent mechanism from which to launch an investigation into the transient flow of a fluid in a porous wick structure where capillary forces must balance both adverse gravitational effects and frictional losses. For the study, a capillary tube, initially charged with a small volume of water, was lowered into a pool of water. The behavior of the column of fluid during the transient that followed as more water entered the tube from the pool was both numerically and experimentally studied.

  6. Capillary electrophoresis and nanomaterials - Part I: Capillary electrophoresis of nanomaterials.

    PubMed

    Adam, Vojtech; Vaculovicova, Marketa

    2017-10-01

    Nanomaterials are in analytical science used for a broad range of purposes, covering the area of sample pretreatment as well as separation, detection, and identification of target molecules. This part of the review covers capillary electrophoresis (CE) of nanomaterials and focuses on the application of CE as a method for characterization used during nanomaterial synthesis and modification as well as the monitoring of their properties and interactions with other molecules. The heterogeneity of the nanomaterial family is extremely large. Depending on different definitions of the term Nanomaterial/Nanoparticle, the group may cover metal and polymeric nanoparticles, carbon nanomaterials, liposomes and even dendrimers. Moreover, these nanomaterials are usually subjected to some kind of surface modification or functionalization, which broadens the diversity even more. Not only for purposes of verification of nanomaterial synthesis and batch-to-batch quality check, but also for determination the polydispersity and for functionality characterization on the nanoparticle surface, has CE offered very beneficial capabilities. Finally, the monitoring of interactions between nanomaterials and other (bio)molecules is easily performed by some kind of capillary electromigration technique. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. High pressure pulsed capillary viscometry

    NASA Technical Reports Server (NTRS)

    Smith, R. L.; Walowitt, J. A.; Pan, C. H. T.

    1972-01-01

    An analytical and test program was conducted in order to establish the feasibility of a multichamber pulsed-capillary viscometer. The initial design incorporated a piston, ram, and seals which produced measured pulses up to 30,000 psi in the closed chamber system. Pressure pulses from one to ten milliseconds were investigated in a system volume of 1 cuin. Four test fluids: a MIL-L-7808, a 5P4E polyphenyl ether, a MIL-L-23699A, and a synthetic hydrocarbon were examined in the test pressure assembly. The pressure-viscosity coefficient and viscosity delay time were determined for the MIL-L-7808 lubricant tested.

  8. Capillary wave spectroscopy on ferrofluids

    NASA Astrophysics Data System (ADS)

    Patzke, J.; Rathke, B.; Will, S.

    2007-12-01

    We investigate the magnetoviscous effect in ferrofluids by Capillary Wave Spectroscopy (CWS, Surface Light Scattering). This technique probes a specific mode of thermally excited surface waves giving information on surface tension and viscosity. In ferrofluids we detect a transition from propagating surface modes to overdamped ones depending on the particle concentration and strength and the orientation of an externally applied magnetic field. We interprete this effect as caused by an increase of the liquid viscosity with an increasing particle concentration and field-strength. Changing the relative orientation of the scattering vector and magnetic field shows that the viscous properties of ferrofluids in a magnetic field are anisotropic. Figs 8, Refs 12.

  9. Horizontal microscopy in square capillaries

    NASA Astrophysics Data System (ADS)

    Moroz, Pavel E.

    1992-07-01

    Intracellular protoplasmic movements may, due to gravity, have a vertical component greater or different from the horizontal one. This makes horizontal microscopy indispensable in the search for the cellular sensor of gravity. The possibility of the latter being a cell organelle assigns special significance to high-resolution microscopy. A horizontal suction device for picking up a cell and its high-resolution horizontal microscopy in a rectangular capillary may be helpful for detection of gravity-related shifts of cellular organelles in vivo.

  10. Capillary Bridges between Soft Substrates

    NASA Astrophysics Data System (ADS)

    Wexler, Jason S.; Heard, Tiara M.; Stone, Howard A.

    2014-02-01

    A wetting droplet trapped in the thin gap between two elastic bodies will deflect the bodies towards one another. The deformation increases the total capillary adhesion force by increasing the contact radius and narrowing the gap height. For flat droplets, with a large ratio of radius to gap height, the Laplace pressure causes surface deformations that are orders of magnitude larger than those induced by a sessile droplet of the same radius. We present experiments, scalings, and closed-form solutions that describe the deformation. Using variational techniques, we also show that the problem exhibits a bifurcation, where the gap spontaneously closes due to an incremental increase in drop volume.

  11. Isolation of cosmids and fetal brain cDNAs from the proximal long arm of human chromosome 22.

    PubMed

    Lamour, V; Lévy, N; Desmaze, C; Baude, V; Lécluse, Y; Delattre, O; Bernheim, A; Thomas, G; Aurias, A; Lipinski, M

    1993-05-01

    The proximal portion of human chromosome 22q appears to carry genes implicated in the pathogenesis of various developmental disorders, including the cat eye syndrome (CES) and the DiGeorge syndrome (DGS). A cosmid library was prepared from a radiation hybrid selected for its content in chromosome 22 fragments. A large fraction of cosmids containing human DNA were found to derive from the juxtacentromeric region of chromosome 22, as shown by fluorescence in situ hybridization (FISH) performed using individual cosmids or cosmid pools as probes. Finer mapping was obtained for individual cosmids by hybridization to a somatic cell hybrid mapping panel which splits the long arm of the chromosome into 14 bins numbered 1 to 14 from the centromere to the telomere. Of the 10 cosmids mapped, eight belonged to group 1, the other two to group 14, in agreement with FISH data. Rare endonuclease sites and fragments conserved between species were searched in single cosmids, resulting in the selection of seven cosmid fragments which were used to screen a human fetal brain cDNA library. Three cDNAs were identified, encoded from two chromosome 22 genes which appeared to be novel, as determined from partial end sequence and comparison with the database entries. Fine localization of the 30.9 cDNA indicated that the corresponding gene was located in a segment of proximal 22q overlapping with the critical DGS region.

  12. Brain pericytes from stress-susceptible pigs increase blood-brain barrier permeability in vitro

    PubMed Central

    2012-01-01

    Background The function of pericytes remains questionable but with improved cultured technique and the use of genetically modified animals, it has become increasingly clear that pericytes are an integral part of blood–brain barrier (BBB) function, and the involvement of pericyte dysfunction in certain cerebrovascular diseases is now emerging. The porcine stress syndrome (PSS) is the only confirmed, homologous model of malignant hyperthermia (MH) in veterinary medicine. Affected animals can experience upon slaughter a range of symptoms, including skeletal muscle rigidity, metabolic acidosis, tachycardia and fever, similar to the human syndrome. Symptoms are due to an enhanced calcium release from intracellular stores. These conditions are associated with a point mutation in ryr1/hal gene, encoding the ryanodine receptor, a calcium channel. Important blood vessel wall muscle modifications have been described in PSS, but potential brain vessel changes have never been documented in this syndrome. Methods In the present work, histological and ultrastructural analyses of brain capillaries from wild type and ryr1 mutated pigs were conducted to investigate the potential impairment of pericytes, in this pathology. In addition, brain pericytes were isolated from the three porcine genotypes (wild-type NN pigs; Nn and nn pigs, bearing one or two (n) mutant ryr1/hal alleles, respectively), and tested in vitro for their influence on the permeability of BBB endothelial monolayers. Results Enlarged perivascular spaces were observed in ryr1-mutant samples, corresponding to a partial or total detachment of the astrocytic endfeet. These spaces were electron lucent and sometimes filled with lipid deposits and swollen astrocytic feet. At the ultrastructural level, brain pericytes did not seem to be affected because they showed regular morphology and characteristics, so we aimed to check their ability to maintain BBB properties in vitro. Our results indicated that pericytes from the

  13. Immunoaffinity screening with capillary electrochromatography.

    PubMed

    Mayer, Michael; Muscate-Magnussen, Angelika; Vogel, Horst; Ehrat, Markus; Bruin, Gerard J M

    2002-05-01

    Highly efficient capillary electrochromatographic separations of cardiac glycosides and other steroids are presented. Employing butyl-derivatized silica particles as stationary phase resulted in a nearly three times faster electroosmotic flow (EOF) compared to capillary electrochromatography (CEC) with octadecyl silica particles. On-column focusing with a preconcentration factor of 180 was performed and separation efficiencies of up to 240,000 plates per meter were obtained. Using label-free standard UV absorbance, detection limits of 10-80 nM were reached for all steroids tested. For screening of cardiac glycosides, e.g., digoxin and digitoxin in mixtures of steroids, CEC was combined with immunoaffinity extraction using immobilized polyclonal anti-digoxigenin antibodies and F(ab) fragments. Simply adding small amounts of antibody carrying particles to the samples and comparing chromatograms before and after antibody addition allowed screening for high affinity antigens in mixtures with moderate numbers of compounds. Under conditions of competing antigens, affinity fingerprints of immobilized anti-digoxigenin and anti-digitoxin antibodies were obtained, reflecting the cross-reactivity of eleven steroids. The method provides high selectivity due to the combination of bioaffinity interaction with highly efficient CEC separation and UV detection at several wavelengths in parallel. This selectivity was exploited for the detection of four cardiac glycosides in submicromolar concentrations in an untreated urine sample.

  14. Cryogenic Capillary Screen Heat Entrapment

    NASA Technical Reports Server (NTRS)

    Bolshinskiy, L.G.; Hastings, L.J.; Stathman, G.

    2007-01-01

    Cryogenic liquid acquisition devices (LADs) for space-based propulsion interface directly with the feed system, which can be a significant heat leak source. Further, the accumulation of thermal energy within LAD channels can lead to the loss of sub-cooled propellant conditions and result in feed system cavitation during propellant outflow. Therefore, the fundamental question addressed by this program was: "To what degree is natural convection in a cryogenic liquid constrained by the capillary screen meshes envisioned for LADs.?"Testing was first conducted with water as the test fluid, followed by LN2 tests. In either case, the basic experimental approach was to heat the bottom of a cylindrical column of test fluid to establish stratification patterns measured by temperature sensors located above and below a horizontal screen barrier position. Experimentation was performed without barriers, with screens, and with a solid barrier. The two screen meshes tested were those typically used by LAD designers, "200x1400" and "325x2300", both with Twill Dutch Weave. Upon consideration of both the water and LN2 data it was concluded that heat transfer across the screen meshes was dependent upon barrier thermal conductivity and that the capillary screen meshes were impervious to natural convection currents.

  15. Atomic Force Controlled Capillary Electrophoresis

    NASA Astrophysics Data System (ADS)

    Lewis, Aaron; Yeshua, Talia; Palchan, Mila; Lovsky, Yulia; Taha, Hesham

    2010-03-01

    Lithography based on scanning probe microscopic techniques has considerable potential for accurate & localized deposition of material on the nanometer scale. Controlled deposition of metallic features with high purity and spatial accuracy is of great interest for circuit edit applications in the semiconductor industry, for plasmonics & nanophotonics and for basic research in surface enhanced Raman scattering & nanobiophysics. Within the context of metal deposition we will review the development of fountain pen nanochemistry and its most recent emulation Atomic Force Controlled Capillary Electrophoresis (ACCE). Using this latter development we will demonstrate achievement of unprecedented control of nanoparticle deposition using a three-electrode geometry. Three electrodes are attached: one on the outside of a metal coated glass probe, one on the inside of a hollow probe in a solution containing Au nanoparticles in the capillary, and a third on the surface where the writing takes place. The three electrodes provide electrical pulses for accurate control of deposition and retraction of the liquid from the surface overcoming the lack of control seen in both dip pen lithography & fountain pen nanochemistry when the tip contacts the surface. With this development, we demonstrate depositing a single 1.3 nm Au nanoparticle onto surfaces such as semiconductors.

  16. Evaluation of capillary reinforced composites

    NASA Technical Reports Server (NTRS)

    Cahill, J. E.; Halase, J. F.; South, W. K.; Stoffer, L. J.

    1985-01-01

    Anti-icing of the inlet of jet engines is generally performed with high pressure heated air that is directed forward from the compressor through a series of pipes to various manifolds located near the structures to be anti-iced. From these manifolds, the air is directed to all flowpath surfaces that may be susceptible to ice formation. There the anti-icing function may be performed by either heat conduction or film heating. Unfortunately, the prospect of utilizing lighweight, high strength composites for inlet structures of jet engines has been frustrated by the low transverse thermal conductivity of such materials. It was the objective of this program to develop an advanced materials and design concept for anti-icing composite structures. The concept that was evaluated used capillary glass tubes embedded on the surface of a composite structure with heated air ducted through the tubes. An analytical computer program was developed to predict the anti-icing performance of such tubes and a test program was conducted to demonstrate actual performance of this system. Test data and analytical code results were in excellent agreement. Both indicate feasibility of using capillary tubes for surface heating as a means for composite engine structures to combat ice accumulation.

  17. Capillary Rise in Porous Media.

    PubMed

    Lago, Marcelo; Araujo, Mariela

    2001-02-01

    Capillary rise experiments were performed in columns filled with glass beads and Berea sandstones, using visual methods to register the advance of the water front. For the glass bead filled columns, early time data are well fitted by the Washburn equation. However, in the experiments, the advancing front exceeded the predicted equilibrium height. For large times, an algebraic behavior of the velocity of the front is observed (T. Delker et al., Phys. Rev. Lett. 76, 2902 (1996)). A model for studying the capillary pressure evolution in a regular assembly of spheres is proposed and developed. It is based on a quasi-static advance of the meniscus with a piston-like motion and allows us to estimate the hydraulic equilibrium height, with values very close to those obtained by fitting early time data to a Washburn equation. The change of regime is explained as a transition in the mechanism of advance of the meniscus. On the other hand, only the Washburn regime was observed for the sandstones. The front velocity was fitted to an algebraical form with an exponent close to 0.5, a value expected from the asymptotic limit of the Washburn equation. Copyright 2001 Academic Press.

  18. Capillary electrophoresis analysis of orange juice pectinesterases.

    PubMed

    Braddock, R J; Bryan, C R; Burns, J K

    2001-02-01

    Pectinesterase (PE) was extracted from orange juice and pulp with 1 M NaCl, desalted, and separated using capillary electrophoresis (CE) gel procedures (CE-SDS-CGE) and isoelectric focusing (CE-IEF). PE resolved as a single peak using noncoated fused silica columns with CE-SDS-CGE. CE-IEF separation of PE required acryloylaminoethoxyethanol-coated columns, which had limited stability. Thermal stability of PE extracts before and after heating at 75 degrees C for 30 min and at 95 degrees C for 5 min established heat labile and heat stabile fractions with identical PE migration times by CE-SDS-CGE or CE-IEF. Peak magnitude decreased to a constant value as heating time increased at 75 degrees C. Regression analysis of CE-SDS-CGE peak migration times of molecular weight (MW) standards estimated both heat labile and heat stable PE at MW approximately 36 900. Traditional SDS-PAGE gel separation of MW standards and active PE isolated by IEF allowed estimation of MW approximately 36 000. CE-SDS-CGE allowed presumptive, but not quantitative, detection of active PE in fresh juice.

  19. Ultra-small superparamagnetic iron oxide enhancement is associated with higher loss of brain tissue structure in clinically isolated syndrome.

    PubMed

    Maarouf, Adil; Ferré, Jean-Christophe; Zaaraoui, Wafaa; Le Troter, Arnaud; Bannier, Elise; Berry, Isabelle; Guye, Maxime; Pierot, Laurent; Barillot, Christian; Pelletier, Jean; Tourbah, Ayman; Edan, Gilles; Audoin, Bertrand; Ranjeva, Jean-Philippe

    2016-07-01

    Macrophages are important components of inflammatory processes in multiple sclerosis, closely linked to axonal loss, and can now be observed in vivo using ultra-small superparamagnetic iron oxide (USPIO). In the present 1-year longitudinal study, we aimed to determine the prevalence and the impact on tissue injury of macrophage infiltration in patients after the first clinical event of multiple sclerosis. Thirty-five patients, 32 years mean age, were imaged in a mean of 66 days after their first event using conventional magnetic resonance imaging, gadolinium (Gd) to probe blood-brain barrier integrity, USPIO to study macrophage infiltration and magnetization transfer ratio (MTR) to assess tissue structure integrity. Statistics were performed using two-group repeated-measures ANOVA. Any patient received treatment at baseline. At baseline, patients showed 17 USPIO-positive lesions reflecting infiltration of macrophages present from the onset. This infiltration was associated with local higher loss of tissue structure as emphasized by significant lower MTRnorm values (p<0.03) in USPIO(+)/Gd(+) lesions (n=16; MTRnormUSPIO(+)/Gd(+)=0.78 at baseline, MTRnormUSPIO(+)/Gd(+)=0.81 at M12) relative to USPIO(-)/Gd(+) lesions (n=67; MTRnormUSPIO(-)/Gd(+)=0.82 at baseline, MTRnormUSPIO(-)/Gd(+)=0.85 at M12). No interaction in MTR values was observed during the 12 months follow-up (lesion type × time). Infiltration of activated macrophages evidenced by USPIO enhancement, is present at the onset of multiple sclerosis and is associated with higher and persistent local loss of tissue structure. Macrophage infiltration affects more tissue structure while tissue recovery during the following year has a similar pattern for USPIO and Gd-enhanced lesions, leading to relative higher persistent local loss of tissue structure in lesions showing USPIO enhancement at baseline. © The Author(s), 2015.

  20. Isolation and characterization of patient-derived, toxic, high mass amyloid beta-protein (Abeta) assembly from Alzheimer disease brains.

    PubMed

    Noguchi, Akihiko; Matsumura, Satoko; Dezawa, Mari; Tada, Mari; Yanazawa, Masako; Ito, Akane; Akioka, Manami; Kikuchi, Satoru; Sato, Michio; Ideno, Shouji; Noda, Munehiro; Fukunari, Atsushi; Muramatsu, Shin-ichi; Itokazu, Yutaka; Sato, Kazuki; Takahashi, Hitoshi; Teplow, David B; Nabeshima, Yo-ichi; Kakita, Akiyoshi; Imahori, Kazutomo; Hoshi, Minako

    2009-11-20

    Amyloid beta-protein (Abeta) assemblies are thought to play primary roles in Alzheimer disease (AD). They are considered to acquire surface tertiary structures, not present in physiologic monomers, that are responsible for exerting toxicity, probably through abnormal interactions with their target(s). Therefore, Abeta assemblies having distinct surface tertiary structures should cause neurotoxicity through distinct mechanisms. Aiming to clarify the molecular basis of neuronal loss, which is a central phenotype in neurodegenerative diseases such as AD, we report here the selective immunoisolation of neurotoxic 10-15-nm spherical Abeta assemblies termed native amylospheroids (native ASPDs) from AD and dementia with Lewy bodies brains, using ASPD tertiary structure-dependent antibodies. In AD patients, the amount of native ASPDs was correlated with the pathologic severity of disease. Native ASPDs are anti-pan oligomer A11 antibody-negative, high mass (>100 kDa) assemblies that induce degeneration particularly of mature neurons, including those of human origin, in vitro. Importantly, their immunospecificity strongly suggests that native ASPDs have a distinct surface tertiary structure from other reported assemblies such as dimers, Abeta-derived diffusible ligands, and A11-positive assemblies. Only ASPD tertiary structure-dependent antibodies could block ASPD-induced neurodegeneration. ASPDs bind presynaptic target(s) on mature neurons and have a mode of toxicity different from those of other assemblies, which have been reported to exert their toxicity through binding postsynaptic targets and probably perturbing glutamatergic synaptic transmission. Thus, our findings indicate that native ASPDs with a distinct toxic surface induce neuronal loss through a different mechanism from other Abeta assemblies.

  1. Use of 3h-. gamma. -aminobutyric acid for transport studies with isolated nerve-terminals from rat brain

    SciTech Connect

    Halvarsson, G.B.; Karlsson, I.; Sellstroem, A.

    1985-07-22

    Isolated synaptosomes were used to study the problem of net accumulation of neurotransmitters. The time-course and the kinetics of exogenous and endogenous GABA transport were studied by liquid-scintillation counting and HPLC-amino acid analysis respectively. Different pools of GABA were suggested by a 6-fold difference in tissue-to-medium-ratio of endogenous vs. exogenous GABA. Net accumulation, exchange and net efflux of GABA was found to be a function of the GABA concentration in the incubation medium. The K/sub m/s for net accumulation and for /sup 3/H-GABA accumulation were 2.68 +/- 1.16 and 6.19 +/- 1.26 ..mu..M respectively, whereas the V/sub max/s were 5.9 +/- 4.9 and 134 +/- 13 pmol/mg w.w min respectively. This means that the transport studies which use exogenous substances (e.g. /sup 3/H-GABA) considerably overestimate the transport by overlooking the magnitude of the counter transport. 22 references, 5 figures, 2 tables.

  2. Impaired coagulation is a risk factor for clinical and radiologic deterioration in patients with traumatic brain injury and isolated traumatic subarachnoid hemorrhage.

    PubMed

    von der Brelie, Christian; Schneegans, Insa; van den Boom, Leander; Meier, Ullrich; Hedderich, Juergen; Lemcke, Johannes

    2015-08-01

    Isolated traumatic subarachnoid hemorrhage (itSAH) is found in approximately 25% of all patients with mild traumatic brain injury (TBI). The aim of this study was to analyze the clinical course and identify risk factors for potential clinical and radiologic deterioration in consideration of impaired coagulation in patients with itSAH. A retrospective analysis of 735 patients with TBI resulting in a pathologic computer-assisted tomography (CAT) was performed. Only those patients with itSAH and Glasgow Coma Scale (GCS) of greater than 8 points and follow-up CAT scan were included. Patients with hemorrhage in any other brain compartment (subdural, epidural, and intracerebral) were excluded. Impaired coagulation was operationally defined. Of the 735 patients, 89 met the inclusion criteria. The majority of these patients experienced mild TBI. The rate of radiologic expansion or conversion of the SAH was 28.1%. The rate of clinical deterioration was 6.7%. Neither the initial pattern of itSAH on different intracranial localizations nor the number of sulci involved in the itSAH was associated with clinical worsening. The rate of patients with impaired coagulation was 38%; 17.9% of all patients showed elevated international normalized ratio (INR). Radiologic and clinical deterioration was significantly associated with elevated INR. INR was shown to be independent of age in a logistic regression analysis. TBI patients with itSAH and impaired coagulation especially those who showed elevated INR are at risk of clinical and radiologic deterioration. Despite coagulation status, routine repetition of cranial CAT scan is advised in patients with itSAH to detect potential radiologic worsening, which if occurring should result in close clinical monitoring. Therapeutic study, level IV; prognostic study, level III.

  3. Ca2+ Handling in Isolated Brain Mitochondria and Cultured Neurons Derived from the YAC128 Mouse Model of Huntington’s Disease

    PubMed Central

    Pellman, Jessica J.; Hamilton, James; Brustovetsky, Tatiana; Brustovetsky, Nickolay

    2015-01-01

    We investigated Ca2+ handling in isolated brain synaptic and nonsynaptic mitochondria and in cultured striatal neurons from the YAC128 mouse model of Huntington’s disease (HD). Both synaptic and nonsynaptic mitochondria from 2- and 12-month-old YAC128 mice had larger Ca2+ uptake capacity than mitochondria from YAC18 and wild-type FVB/NJ mice. Synaptic mitochondria from 12-month-old YAC128 mice had further augmented Ca2+ capacity compared with mitochondria from 2-month-old YAC128 mice and age-matched YAC18 and FVB/NJ mice. This increase in Ca2+ uptake capacity correlated with an increase in the amount of mutant huntingtin protein (mHtt) associated with mitochondria from 12-month-old YAC128 mice. We speculate that this may happen due to mHtt-mediated sequestration of free fatty acids thereby increasing resistance of mitochondria to Ca2+-induced damage. In experiments with striatal neurons from YAC128 and FVB/NJ mice, brief exposure to 25 or 100μM glutamate produced transient elevations in cytosolic Ca2+ followed by recovery to near resting levels. Following recovery of cytosolic Ca2+, mitochondrial depolarization with FCCP produced comparable elevations in cytosolic Ca2+, suggesting similar Ca2+ release and, consequently, Ca2+ loads in neuronal mitochondria from YAC128 and FVB/NJ mice. Together, our data argue against a detrimental effect of mHtt on Ca2+ handling in brain mitochondria of YAC128 mice. PMID:25963273

  4. Extracellular Vesicles Isolated from the Brains of rTg4510 Mice Seed Tau Protein Aggregation in a Threshold-dependent Manner*

    PubMed Central

    Polanco, Juan Carlos; Scicluna, Benjamin James; Hill, Andrew Francis

    2016-01-01

    The microtubule-associated protein tau has a critical role in Alzheimer disease and related tauopathies. There is accumulating evidence that tau aggregates spread and replicate in a prion-like manner, with the uptake of pathological tau seeds causing misfolding and aggregation of monomeric tau in recipient cells. Here we focused on small extracellular vesicles enriched for exosomes that were isolated from the brains of tau transgenic rTg4510 and control mice. We found that these extracellular vesicles contained tau, although the levels were significantly higher in transgenic mice that have a pronounced tau pathology. Tau in the vesicles was differentially phosphorylated, although to a lower degree than in the brain cells from which they were derived. Several phospho-epitopes (AT8, AT100, and AT180) thought to be critical for tau pathology were undetected in extracellular vesicles. Despite this, when assayed with FRET tau biosensor cells, extracellular vesicles derived from transgenic mice were capable of seeding tau aggregation in a threshold-dependent manner. We also observed that the dye used to label extracellular vesicle membranes was still present during nucleation and formation of tau inclusions, suggesting either a role for membranes in the seeding or in the process of degradation. Together, we clearly demonstrate that extracellular vesicles can transmit tau pathology. This indicates a role for extracellular vesicles in the transmission and spreading of tau pathology. The characteristics of tau in extracellular vesicles and the seeding threshold we identified may explain why tau pathology develops very slowly in neurodegenerative diseases such as Alzheimer disease. PMID:27030011

  5. Extracellular Vesicles Isolated from the Brains of rTg4510 Mice Seed Tau Protein Aggregation in a Threshold-dependent Manner.

    PubMed

    Polanco, Juan Carlos; Scicluna, Benjamin James; Hill, Andrew Francis; Götz, Jürgen

    2016-06-10

    The microtubule-associated protein tau has a critical role in Alzheimer disease and related tauopathies. There is accumulating evidence that tau aggregates spread and replicate in a prion-like manner, with the uptake of pathological tau seeds causing misfolding and aggregation of monomeric tau in recipient cells. Here we focused on small extracellular vesicles enriched for exosomes that were isolated from the brains of tau transgenic rTg4510 and control mice. We found that these extracellular vesicles contained tau, although the levels were significantly higher in transgenic mice that have a pronounced tau pathology. Tau in the vesicles was differentially phosphorylated, although to a lower degree than in the brain cells from which they were derived. Several phospho-epitopes (AT8, AT100, and AT180) thought to be critical for tau pathology were undetected in extracellular vesicles. Despite this, when assayed with FRET tau biosensor cells, extracellular vesicles derived from transgenic mice were capable of seeding tau aggregation in a threshold-dependent manner. We also observed that the dye used to label extracellular vesicle membranes was still present during nucleation and formation of tau inclusions, suggesting either a role for membranes in the seeding or in the process of degradation. Together, we clearly demonstrate that extracellular vesicles can transmit tau pathology. This indicates a role for extracellular vesicles in the transmission and spreading of tau pathology. The characteristics of tau in extracellular vesicles and the seeding threshold we identified may explain why tau pathology develops very slowly in neurodegenerative diseases such as Alzheimer disease.

  6. The role of the cerebral capillaries in acute ischemic stroke: the extended penumbra model

    PubMed Central

    Østergaard, Leif; Jespersen, Sune Nørhøj; Mouridsen, Kim; Mikkelsen, Irene Klærke; Jonsdottír, Kristjana Ýr; Tietze, Anna; Blicher, Jakob Udby; Aamand, Rasmus; Hjort, Niels; Iversen, Nina Kerting; Cai, Changsi; Hougaard, Kristina Dupont; Simonsen, Claus Z; Von Weitzel-Mudersbach, Paul; Modrau, Boris; Nagenthiraja, Kartheeban; Riisgaard Ribe, Lars; Hansen, Mikkel Bo; Bekke, Susanne Lise; Dahlman, Martin Gervais; Puig, Josep; Pedraza, Salvador; Serena, Joaquín; Cho, Tae-Hee; Siemonsen, Susanne; Thomalla, Götz; Fiehler, Jens; Nighoghossian, Norbert; Andersen, Grethe

    2013-01-01

    The pathophysiology of cerebral ischemia is traditionally understood in relation to reductions in cerebral blood flow (CBF). However, a recent reanalysis of the flow-diffusion equation shows that increased capillary transit time heterogeneity (CTTH) can reduce the oxygen extraction efficacy in brain tissue for a given CBF. Changes in capillary morphology are typical of conditions predisposing to stroke and of experimental ischemia. Changes in capillary flow patterns have been observed by direct microscopy in animal models of ischemia and by indirect methods in humans stroke, but their metabolic significance remain unclear. We modeled the effects of progressive increases in CTTH on the way in which brain tissue can secure sufficient oxygen to meet its metabolic needs. Our analysis predicts that as CTTH increases, CBF responses to functional activation and to vasodilators must be suppressed to maintain sufficient tissue oxygenation. Reductions in CBF, increases in CTTH, and combinations thereof can seemingly trigger a critical lack of oxygen in brain tissue, and the restoration of capillary perfusion patterns therefore appears to be crucial for the restoration of the tissue oxygenation after ischemic episodes. In this review, we discuss the possible implications of these findings for the prevention, diagnosis, and treatment of acute stroke. PMID:23443173

  7. The role of the cerebral capillaries in acute ischemic stroke: the extended penumbra model.

    PubMed

    Østergaard, Leif; Jespersen, Sune Nørhøj; Mouridsen, Kim; Mikkelsen, Irene Klærke; Jonsdottír, Kristjana Ýr; Tietze, Anna; Blicher, Jakob Udby; Aamand, Rasmus; Hjort, Niels; Iversen, Nina Kerting; Cai, Changsi; Hougaard, Kristina Dupont; Simonsen, Claus Z; Von Weitzel-Mudersbach, Paul; Modrau, Boris; Nagenthiraja, Kartheeban; Riisgaard Ribe, Lars; Hansen, Mikkel Bo; Bekke, Susanne Lise; Dahlman, Martin Gervais; Puig, Josep; Pedraza, Salvador; Serena, Joaquín; Cho, Tae-Hee; Siemonsen, Susanne; Thomalla, Götz; Fiehler, Jens; Nighoghossian, Norbert; Andersen, Grethe

    2013-05-01

    The pathophysiology of cerebral ischemia is traditionally understood in relation to reductions in cerebral blood flow (CBF). However, a recent reanalysis of the flow-diffusion equation shows that increased capillary transit time heterogeneity (CTTH) can reduce the oxygen extraction efficacy in brain tissue for a given CBF. Changes in capillary morphology are typical of conditions predisposing to stroke and of experimental ischemia. Changes in capillary flow patterns have been observed by direct microscopy in animal models of ischemia and by indirect methods in humans stroke, but their metabolic significance remain unclear. We modeled the effects of progressive increases in CTTH on the way in which brain tissue can secure sufficient oxygen to meet its metabolic needs. Our analysis predicts that as CTTH increases, CBF responses to functional activation and to vasodilators must be suppressed to maintain sufficient tissue oxygenation. Reductions in CBF, increases in CTTH, and combinations thereof can seemingly trigger a critical lack of oxygen in brain tissue, and the restoration of capillary perfusion patterns therefore appears to be crucial for the restoration of the tissue oxygenation after ischemic episodes. In this review, we discuss the possible implications of these findings for the prevention, diagnosis, and treatment of acute stroke.

  8. Capillary waveguide optrodes for Medical applications

    NASA Astrophysics Data System (ADS)

    Kieslinger, Dietmar; Weigl, Bernhard H.; Draxler, Sonja; Lippitsch, Max E.

    1997-01-01

    Glass capillaries with a chemically sensitive coating on the inner surface are used as optical sensors for medical diagnostics. The capillary simultaneously serves as a sample compartment, a sensor element, and an inhomogeneous optical waveguide. Different optical setups have been investigated and compared regarding its waveguiding properties.

  9. Internal capillary insulation for cryogenic tanks

    NASA Technical Reports Server (NTRS)

    Mcgrew, J. L.

    1972-01-01

    Capillary-type insulation was devised for installation on inside of liquid methane fuel tanks for future aircraft. Insulation consists of honeycomb core of fiberglass cloth impregnated with polyimide resin which is bonded onto metal tank wall using polyimide adhesive. Capillary holes in each honeycomb cell admit methane which provides static pressure in cell.

  10. Sheathless interface for coupling capillary electrophoresis with mass spectrometry

    DOEpatents

    Wang, Chenchen; Tang, Keqi; Smith, Richard D.

    2014-06-17

    A sheathless interface for coupling capillary electrophoresis (CE) with mass spectrometry is disclosed. The sheathless interface includes a separation capillary for performing CE separation and an emitter capillary for electrospray ionization. A portion of the emitter capillary is porous or, alternatively, is coated to form an electrically conductive surface. A section of the emitter capillary is disposed within the separation capillary, forming a joint. A metal tube, containing a conductive liquid, encloses the joint.

  11. A covalent capillary coating of diazoresin and polyglycerol dendrimer for protein analysis using capillary electrophoresis.

    PubMed

    Yu, Bing; Wang, Minghong; Cong, Hailin; Li, Guoling

    2017-08-29

    Overcoming proteins adsorption on the inner surface of capillary has attracted increasing attention recently. By using the unique photochemistry reaction of diazoresin (DR), a new covalent capillary coating was prepared on the fused-silica capillary through layer-by-layer self-assembly of DR with polyglycerol (PG) dendrimer. The separation performance of covalently DR/PG-dendrimer coated capillary noticeably exceeded the bare capillary and the noncovalently linked DR/PG-dendrimer capillary. A baseline separation of lysozyme, myoglobin, bovine serum albumin, and ribonuclease A was achieved using CE within 20 min. Besides, the covalently linked DR/PG-dendrimer coating has the remarkable stability and reproducibility. Especially, compared with the traditional method which use highly toxic and moisture-sensitive silane coupling agent, this method seems to be a simple and environmental friendly way to prepare the covalently coated capillaries for CE. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Activating PKC-β1 at the blood-brain barrier reverses induction of P-glycoprotein activity by dioxin and restores drug delivery to the CNS.

    PubMed

    Wang, Xueqian; Hawkins, Brian T; Miller, David S

    2011-06-01

    Upregulation of blood-brain barrier (BBB) P-glycoprotein expression causes central nervous system (CNS) pharmacoresistance. However, activation of BBB protein kinase C-β1 (PKC-β1) rapidly reduces basal P-glycoprotein transport activity. We tested whether PKC-β1 activation would reverse CNS drug resistance caused by dioxin acting through aryl hydrocarbon receptor. A selective PKC-β1 agonist abolished the increase in P-glycoprotein activity induced by dioxin in isolated rat brain capillaries and reversed the effect of dioxin on brain uptake of verapamil in dioxin-dosed rats. Thus, targeting BBB PKC-β1 may be an effective strategy to improve drug delivery to the brain, even in drug-resistant individuals.

  13. Effect of capillary waves on surface tension

    NASA Technical Reports Server (NTRS)

    Kayser, R. F.

    1986-01-01

    The present study is concerned with the effect which a cutting off of the capillary waves has on surface tension, taking into account a calculation based on capillary-wave theory. For simplicity, three-dimensional systems are considered, and capillary-wave theory is used to calculate sigma-k, the surface tension of an interface where only those modes with at least one wave-vector component greater than k are allowed. Attention is given to a review of capillary-wave theory, the calculation of surface tensions, a determination of the range of validity of capillary-wave theory, and some numerical examples. The quantitative behavior of sigma-k and its relation to the surface tension of a finite-size system are considered. The most surprising result is that sigma-k can be significantly larger than the unconstrained surface tension.

  14. Study of Capillary-Based Gaseous Detectors

    NASA Astrophysics Data System (ADS)

    Iacobaeus, C.; Francke, T.; Danielsson, M.; Ostling, J.; Peskov, V.

    2004-06-01

    We have studied gain vs. voltage characteristics and position resolutions of multistep capillary plates (two or three capillary plates operating in a cascade), as well as capillary plates operating in a mode when the main amplification occurs between plates or between the capillary plate and the readout plate (parallel plate amplification mode). Results of these studies demonstrated that in the parallel-plate amplification mode one can reach both high gains (>100000) and good position resolutions (~100 micro meter) even with a single step arrangement. It offers a compact amplification structure, which can be used in many applications. For example, in preliminary tests we succeeded to combine it with a photocathode and use it as a position sensitive gaseous photomultiplier. CsI coated capillary plates could also be used as a high position resolution and high rate X-ray converter.

  15. Oximetry of retinal capillaries by multicomponent analysis.

    PubMed

    Furukawa, Hiromitsu; Arimoto, Hidenobu; Shirai, Tomohiro; Ooto, Sotaro; Hangai, Masanori; Yoshimura, Nagahisa

    2012-08-01

    Retinal oximetry of capillaries was performed for early detection of retinal vascular abnormalities, which are caused predominantly by complications of systemic circulatory diseases. As the conventional method for determining absorbance is not applicable to capillaries, multicomponent analysis was used to estimate the absorbance spectra of the retinal blood vessels. In this analysis, the capillary spectrum was classified as intermediate between those of the retinal arteries and veins, enabling relative estimation of oxygen saturation in the capillaries. This method could be useful for early recognition of disturbances in the peripheral circulation. Furthermore, a spectroscopic ophthalmoscope system based on the proposed method was developed to examine the human retina. A clinical trial of this system demonstrated that oximetry of the retinal capillaries may be an improvement over the present diagnosis for patients of malignant hypertension.

  16. Citalopram Ameliorates Synaptic Plasticity Deficits in Different Cognition-Associated Brain Regions Induced by Social Isolation in Middle-Aged Rats.

    PubMed

    Gong, Wei-Gang; Wang, Yan-Juan; Zhou, Hong; Li, Xiao-Li; Bai, Feng; Ren, Qing-Guo; Zhang, Zhi-Jun

    2017-04-01

    Our previous experiments demonstrated that social isolation (SI) caused AD-like tau hyperphosphorylation and spatial memory deficits in middle-aged rats. However, the underlying mechanisms of SI-induced spatial memory deficits remain elusive. Middle-aged rats (10 months) were group or isolation reared for 8 weeks. Following the initial 4-week period of rearing, citalopram (10 mg/kg i.p.) was administered for 28 days. Then, pathophysiological changes were assessed by performing behavioral, biochemical, and pathological analyses. We found that SI could cause cognitive dysfunction and decrease synaptic protein (synaptophysin or PSD93) expression in different brain regions associated with cognition, such as the prefrontal cortex, dorsal hipp