synaptotagmin  (Alomone Labs)


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    Structured Review

    Alomone Labs synaptotagmin
    WT1 -expressing neurons terminate in close proximity to populations of commissurally projecting interneurons. A , A 20-μm-thick section of a spinal cord 46 h after PRV-152 injection into the GS muscle on the left side and stained with antibodies to GFP (green) and WT1 (blue). At this time point, no WT1 -expressing neurons on either side of the spinal cord have taken up the tracer. Contralateral region containing WT1 -expressing cells (dashed box) is expanded to the right. B , To identify presumptive synapses on genetically defined interneuronal subtypes, we inspected the <t>synaptotagmin</t> + “halo” (indicated by white arrows) surrounding each labeled nuclei for WT1 + /synaptotagmin + terminals. C–F , The 20-μm-thick sections cut from a P0 WT1 CreER ROSA26 tdTomato spinal cord and stained with antibodies to tdTomato (red), the synaptic marker synatotagmin (blue), as well as (green) a nuclear marker of V2a cells ( Chx10 -, C ), V1 cells ( En1 -, D ), V0 V cells ( Evx1 -, E ), or DMRT3 -expressing dI6 cells ( F ). WT1 -expressing terminals (tdTomato + /synaptotagmin + processes) were rare or absent nearby Chx10 - or En1 -expressing cells but were commonly seen in close proximity to Evx1 + and DMRT3 + neurons. C , D , Dashed boxes are expanded to the right. E , F , Arrow in the low-magnification image indicates the specific WT1 cell of interest in the panel to the right. In magnified images, arrows indicate presumptive WT1 + axon terminals. Double labeling of these processes is confirmed in orthogonal views to the right of E , F . Scale bars: Low-magnification images, 100 μm; High-magnification images: A , C , D , 20 μm; B , E , F , 5 μm.
    Synaptotagmin, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "WT1-Expressing Interneurons Regulate Left–Right Alternation during Mammalian Locomotor Activity"

    Article Title: WT1-Expressing Interneurons Regulate Left–Right Alternation during Mammalian Locomotor Activity

    Journal: The Journal of Neuroscience

    doi: 10.1523/JNEUROSCI.0328-18.2018

    WT1 -expressing neurons terminate in close proximity to populations of commissurally projecting interneurons. A , A 20-μm-thick section of a spinal cord 46 h after PRV-152 injection into the GS muscle on the left side and stained with antibodies to GFP (green) and WT1 (blue). At this time point, no WT1 -expressing neurons on either side of the spinal cord have taken up the tracer. Contralateral region containing WT1 -expressing cells (dashed box) is expanded to the right. B , To identify presumptive synapses on genetically defined interneuronal subtypes, we inspected the synaptotagmin + “halo” (indicated by white arrows) surrounding each labeled nuclei for WT1 + /synaptotagmin + terminals. C–F , The 20-μm-thick sections cut from a P0 WT1 CreER ROSA26 tdTomato spinal cord and stained with antibodies to tdTomato (red), the synaptic marker synatotagmin (blue), as well as (green) a nuclear marker of V2a cells ( Chx10 -, C ), V1 cells ( En1 -, D ), V0 V cells ( Evx1 -, E ), or DMRT3 -expressing dI6 cells ( F ). WT1 -expressing terminals (tdTomato + /synaptotagmin + processes) were rare or absent nearby Chx10 - or En1 -expressing cells but were commonly seen in close proximity to Evx1 + and DMRT3 + neurons. C , D , Dashed boxes are expanded to the right. E , F , Arrow in the low-magnification image indicates the specific WT1 cell of interest in the panel to the right. In magnified images, arrows indicate presumptive WT1 + axon terminals. Double labeling of these processes is confirmed in orthogonal views to the right of E , F . Scale bars: Low-magnification images, 100 μm; High-magnification images: A , C , D , 20 μm; B , E , F , 5 μm.
    Figure Legend Snippet: WT1 -expressing neurons terminate in close proximity to populations of commissurally projecting interneurons. A , A 20-μm-thick section of a spinal cord 46 h after PRV-152 injection into the GS muscle on the left side and stained with antibodies to GFP (green) and WT1 (blue). At this time point, no WT1 -expressing neurons on either side of the spinal cord have taken up the tracer. Contralateral region containing WT1 -expressing cells (dashed box) is expanded to the right. B , To identify presumptive synapses on genetically defined interneuronal subtypes, we inspected the synaptotagmin + “halo” (indicated by white arrows) surrounding each labeled nuclei for WT1 + /synaptotagmin + terminals. C–F , The 20-μm-thick sections cut from a P0 WT1 CreER ROSA26 tdTomato spinal cord and stained with antibodies to tdTomato (red), the synaptic marker synatotagmin (blue), as well as (green) a nuclear marker of V2a cells ( Chx10 -, C ), V1 cells ( En1 -, D ), V0 V cells ( Evx1 -, E ), or DMRT3 -expressing dI6 cells ( F ). WT1 -expressing terminals (tdTomato + /synaptotagmin + processes) were rare or absent nearby Chx10 - or En1 -expressing cells but were commonly seen in close proximity to Evx1 + and DMRT3 + neurons. C , D , Dashed boxes are expanded to the right. E , F , Arrow in the low-magnification image indicates the specific WT1 cell of interest in the panel to the right. In magnified images, arrows indicate presumptive WT1 + axon terminals. Double labeling of these processes is confirmed in orthogonal views to the right of E , F . Scale bars: Low-magnification images, 100 μm; High-magnification images: A , C , D , 20 μm; B , E , F , 5 μm.

    Techniques Used: Expressing, Injection, Staining, Labeling, Marker

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    Alomone Labs anti nr2b
    Leptin signaling increases pNR2B Y1472 levels and surface expression. ( A ) Representative Western blot of hippocampal neurons treated with leptin (50 nM), PP1 (10 µM), or both for 2 hours. ( B ) Quantification of pNR2B Y1472 intensity normalized to total <t>NR2B</t> intensity (n = 3). ( C ) Representative Western blot of hippocampal protein extracts from P10 wild-type and ob/ob mice pups (wild-type: n = 5; ob/ob : n = 5). ( D ) Quantification of pNR2B Y1472 intensity normalized to total NR2B intensity and total NR2B intensity normalized to the neuronal marker MAP2B intensity (n = 3). ( E ) Representative Western blot of surface biotinylated hippocampal cultures treated with leptin (50 nM, 2 hours). Biotinylated proteins were affinity purified (AP) with streptavidin magnetic beads. ( F ) Quantification of biotinylated NR2B intensity normalized to NR2B intensity in total lysate (n = 3). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P
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    Alomone Labs na k atpase α subunit
    Localization of CLC2- and Na + -K + -ATPase <t>α-subunit</t> (NaKα)-like proteins in rat distal colon. A and A’ : CLC2-like proteins (green) localized predominantly in surface cells (arrows), while minimally expressed in crypt regions (asterisks). B and B’ : NaKα-like proteins (red) localized to both surface (arrows) and crypt (asterisks) cells. C and C’ : nuclei (blue) stained with Hoechst 33342. D : composite of A , B , and C . D’ : composite of A’ , B’ , and C’ . Higher magnification of the boxed region from D and D’ presented in E and E’ , respectively. E : CLC2 and NaKα colocalized (orange) in both lateral (arrows) and basement (arrowheads) membranes of surface cells. E’ : in crypt cells, NaKα (arrow) and CLC2 (arrowhead) localized in the basolateral membranes of crypts. Identical results obtained in different specimens obtained from three different experiments with three different male and female (not shown) rat distal colons.
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    Alomone Labs rabbit anti dat antibody
    Expression of dopamine transporter <t>(DAT),</t> norepinephrine transporter <t>(NET),</t> and serotonin transporter (SERT) in astroglial cultures prepared from Sprague-Dawley newborn cortex (a), (c), (e) or embryonic striatum (b), (d), (f). Confocal laser-scanning microscopic images showing double immunostaining for DAT (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (a) and (b)), NET (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (c) and (d)), or SERT (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (e) and (f)) in cortical or striatal astroglia prepared from Sprague-Dawley rats. Scale bar = 20 µm.
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    Alomone Labs anti gat 3
    The expressions of neurotransmitters and different subunits of GABA A Rs. A‐B, The expression of GABA was decreased in the laparotomy mice, and no difference was found about glutamate (n = 9). C, The different average spectra of selected metabolites (GABA and glutamate). D‐F, The mRNA level of α5 subunit was up‐regulated at 1 day and continued to 10 days after laparotomy. No difference was found about the α1 and β3 subunits (n = 3). G‐I, The expressions of <t>GAT‐3</t> and GAD65 were decreased, and the levels of surface α5GABA A Rs were increased in the laparotomy mice (n = 4). Data are presented as mean ± SEM. * P
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    Image Search Results


    Leptin signaling increases pNR2B Y1472 levels and surface expression. ( A ) Representative Western blot of hippocampal neurons treated with leptin (50 nM), PP1 (10 µM), or both for 2 hours. ( B ) Quantification of pNR2B Y1472 intensity normalized to total NR2B intensity (n = 3). ( C ) Representative Western blot of hippocampal protein extracts from P10 wild-type and ob/ob mice pups (wild-type: n = 5; ob/ob : n = 5). ( D ) Quantification of pNR2B Y1472 intensity normalized to total NR2B intensity and total NR2B intensity normalized to the neuronal marker MAP2B intensity (n = 3). ( E ) Representative Western blot of surface biotinylated hippocampal cultures treated with leptin (50 nM, 2 hours). Biotinylated proteins were affinity purified (AP) with streptavidin magnetic beads. ( F ) Quantification of biotinylated NR2B intensity normalized to NR2B intensity in total lysate (n = 3). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Journal: Endocrinology

    Article Title: Leptin Controls Glutamatergic Synaptogenesis and NMDA-Receptor Trafficking via Fyn Kinase Regulation of NR2B

    doi: 10.1210/endocr/bqz030

    Figure Lengend Snippet: Leptin signaling increases pNR2B Y1472 levels and surface expression. ( A ) Representative Western blot of hippocampal neurons treated with leptin (50 nM), PP1 (10 µM), or both for 2 hours. ( B ) Quantification of pNR2B Y1472 intensity normalized to total NR2B intensity (n = 3). ( C ) Representative Western blot of hippocampal protein extracts from P10 wild-type and ob/ob mice pups (wild-type: n = 5; ob/ob : n = 5). ( D ) Quantification of pNR2B Y1472 intensity normalized to total NR2B intensity and total NR2B intensity normalized to the neuronal marker MAP2B intensity (n = 3). ( E ) Representative Western blot of surface biotinylated hippocampal cultures treated with leptin (50 nM, 2 hours). Biotinylated proteins were affinity purified (AP) with streptavidin magnetic beads. ( F ) Quantification of biotinylated NR2B intensity normalized to NR2B intensity in total lysate (n = 3). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Article Snippet: Neurons fixed and incubated with an anti-NR2B (1:100, Alomone Labs) ( ) and anti-Flag (1:250, Sigma Aldrich) ( ) antibody for 1 hour and then incubated with the appropriate Alexa Fluor secondary IgG antibody ( ) for 1 hour at room temperature.

    Techniques: Expressing, Western Blot, Mouse Assay, Marker, Affinity Purification, Magnetic Beads

    Leptin-regulated NR2B Y1472 phosphorylation and surface expression is Fyn dependent. ( A ) Representative Western blot of HEK293T cells transfected with NR2B-V5, NR1, LepRb-myc, and either V5-Fyn or V5-DN Fyn and treated with leptin (50 nM, 2 hours). ( B ) Quantification of pNR2B Y1472 intensity normalized to total NR2B-V5 intensity (n = 3). ( C ) Hippocampal neurons were transfected with Clover and EGFP-NR2B-V5 and either V5-Fyn or V5-DN Fyn ± leptin stimulation (50 nM, 2 hours) and live immunostained for surface EGFP–NR2B. Quantification of immunostained EGFP-integrated signal density (n = 15). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Journal: Endocrinology

    Article Title: Leptin Controls Glutamatergic Synaptogenesis and NMDA-Receptor Trafficking via Fyn Kinase Regulation of NR2B

    doi: 10.1210/endocr/bqz030

    Figure Lengend Snippet: Leptin-regulated NR2B Y1472 phosphorylation and surface expression is Fyn dependent. ( A ) Representative Western blot of HEK293T cells transfected with NR2B-V5, NR1, LepRb-myc, and either V5-Fyn or V5-DN Fyn and treated with leptin (50 nM, 2 hours). ( B ) Quantification of pNR2B Y1472 intensity normalized to total NR2B-V5 intensity (n = 3). ( C ) Hippocampal neurons were transfected with Clover and EGFP-NR2B-V5 and either V5-Fyn or V5-DN Fyn ± leptin stimulation (50 nM, 2 hours) and live immunostained for surface EGFP–NR2B. Quantification of immunostained EGFP-integrated signal density (n = 15). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Article Snippet: Neurons fixed and incubated with an anti-NR2B (1:100, Alomone Labs) ( ) and anti-Flag (1:250, Sigma Aldrich) ( ) antibody for 1 hour and then incubated with the appropriate Alexa Fluor secondary IgG antibody ( ) for 1 hour at room temperature.

    Techniques: Expressing, Western Blot, Transfection

    LepRb directly interacts with NR2B. ( A ) Schematic of LepRb–BioID experiment with representative Western blot of NR2B-V5 immunoprecipitated from HEK293T cells expressing the designated BioID constructs and NR2B-V5 and NR1-Clover to the right. ( B ) Quantification of IP biotinylated NR2B-V5 intensity normalized to total NR2B-V5 intensity in the same lane (n = 3). ( C ) Schematic of NR2B–BioID experiment with representative Western blot of LepRb-V5 immunoprecipitated from HEK293T cells expressing designated BioID constructs and LepRb-V5 and NR1-Clover. ( D ) Representative Western blot of LepRb-myc immunoprecipitated from HEK293T cells stimulated with leptin (50 nM, 2 hours) and expressing LepRb-myc, NR2B-V5, and NR1-Clover. ( E ) Quantification of coimmunoprecipitated NR2B-V5 intensity normalized to immunoprecipitated LepRb-myc intensity from the same lane (n = 3). ( F ) Representative fluorescent images of hippocampal cultures expressing Flag-LepRb and Clover. Surface Flag-LepRb and endogenous surface NR2B were live immunostained after stimulation with leptin (50 nM, 2 hours). ( G ) Quantification of NR2B/Flag-LepRb puncta colocalization compared to total NR2B puncta. Colocalization experiments were repeated in 2 independent hippocampal culture preparations. All BioID experiments were stimulated with biotin (50 µM) at the time of transfection. All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Journal: Endocrinology

    Article Title: Leptin Controls Glutamatergic Synaptogenesis and NMDA-Receptor Trafficking via Fyn Kinase Regulation of NR2B

    doi: 10.1210/endocr/bqz030

    Figure Lengend Snippet: LepRb directly interacts with NR2B. ( A ) Schematic of LepRb–BioID experiment with representative Western blot of NR2B-V5 immunoprecipitated from HEK293T cells expressing the designated BioID constructs and NR2B-V5 and NR1-Clover to the right. ( B ) Quantification of IP biotinylated NR2B-V5 intensity normalized to total NR2B-V5 intensity in the same lane (n = 3). ( C ) Schematic of NR2B–BioID experiment with representative Western blot of LepRb-V5 immunoprecipitated from HEK293T cells expressing designated BioID constructs and LepRb-V5 and NR1-Clover. ( D ) Representative Western blot of LepRb-myc immunoprecipitated from HEK293T cells stimulated with leptin (50 nM, 2 hours) and expressing LepRb-myc, NR2B-V5, and NR1-Clover. ( E ) Quantification of coimmunoprecipitated NR2B-V5 intensity normalized to immunoprecipitated LepRb-myc intensity from the same lane (n = 3). ( F ) Representative fluorescent images of hippocampal cultures expressing Flag-LepRb and Clover. Surface Flag-LepRb and endogenous surface NR2B were live immunostained after stimulation with leptin (50 nM, 2 hours). ( G ) Quantification of NR2B/Flag-LepRb puncta colocalization compared to total NR2B puncta. Colocalization experiments were repeated in 2 independent hippocampal culture preparations. All BioID experiments were stimulated with biotin (50 µM) at the time of transfection. All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Article Snippet: Neurons fixed and incubated with an anti-NR2B (1:100, Alomone Labs) ( ) and anti-Flag (1:250, Sigma Aldrich) ( ) antibody for 1 hour and then incubated with the appropriate Alexa Fluor secondary IgG antibody ( ) for 1 hour at room temperature.

    Techniques: Western Blot, Immunoprecipitation, Expressing, Construct, Transfection

    pNR2B Y1472 is necessary for leptin-stimulated spine formation. ( A ) Representative fluorescent images of hippocampal neurons expressing Clover and EGFP-NR2B-V5 or EGFP-NR2B Y1472F -V5 ± leptin stimulation (50 nM, 2 hours) and live immunostained for surface EGFP-NR2B. White bar = 20 µm. ( B ) Quantification of immunostained EGFP-integrated signal density (n = 23). ( C-E ) Hippocampal neurons that were transfected with a fluorescent Clover-βactin and EGFP-NR2B Y1472F -V5. Neurons were stimulated with leptin (50 nM) on DIV8, and on DIV11 to 12 spine density was measured by hand using ImageJ with the NeuronJ plugin ( C,D ), or electrophysiological recordings were performed ( E ). White bar = 5 µm. ( D ) Quantification of dendritic spine density from a minimum of 2 to 3 dendritic segments from 15 neurons. ( E ) Quantification of mEPSC frequency, amplitude, and decay time normalized to control condition (control: n = 32; control + leptin: n = 34; NR2B Y1472F : n = 33; NR2B Y1472F + leptin: n = 33). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Journal: Endocrinology

    Article Title: Leptin Controls Glutamatergic Synaptogenesis and NMDA-Receptor Trafficking via Fyn Kinase Regulation of NR2B

    doi: 10.1210/endocr/bqz030

    Figure Lengend Snippet: pNR2B Y1472 is necessary for leptin-stimulated spine formation. ( A ) Representative fluorescent images of hippocampal neurons expressing Clover and EGFP-NR2B-V5 or EGFP-NR2B Y1472F -V5 ± leptin stimulation (50 nM, 2 hours) and live immunostained for surface EGFP-NR2B. White bar = 20 µm. ( B ) Quantification of immunostained EGFP-integrated signal density (n = 23). ( C-E ) Hippocampal neurons that were transfected with a fluorescent Clover-βactin and EGFP-NR2B Y1472F -V5. Neurons were stimulated with leptin (50 nM) on DIV8, and on DIV11 to 12 spine density was measured by hand using ImageJ with the NeuronJ plugin ( C,D ), or electrophysiological recordings were performed ( E ). White bar = 5 µm. ( D ) Quantification of dendritic spine density from a minimum of 2 to 3 dendritic segments from 15 neurons. ( E ) Quantification of mEPSC frequency, amplitude, and decay time normalized to control condition (control: n = 32; control + leptin: n = 34; NR2B Y1472F : n = 33; NR2B Y1472F + leptin: n = 33). All experiments were repeated in 3 independent culture preparations and expressed as the mean ± SEM, * P

    Article Snippet: Neurons fixed and incubated with an anti-NR2B (1:100, Alomone Labs) ( ) and anti-Flag (1:250, Sigma Aldrich) ( ) antibody for 1 hour and then incubated with the appropriate Alexa Fluor secondary IgG antibody ( ) for 1 hour at room temperature.

    Techniques: Expressing, Transfection

    Localization of CLC2- and Na + -K + -ATPase α-subunit (NaKα)-like proteins in rat distal colon. A and A’ : CLC2-like proteins (green) localized predominantly in surface cells (arrows), while minimally expressed in crypt regions (asterisks). B and B’ : NaKα-like proteins (red) localized to both surface (arrows) and crypt (asterisks) cells. C and C’ : nuclei (blue) stained with Hoechst 33342. D : composite of A , B , and C . D’ : composite of A’ , B’ , and C’ . Higher magnification of the boxed region from D and D’ presented in E and E’ , respectively. E : CLC2 and NaKα colocalized (orange) in both lateral (arrows) and basement (arrowheads) membranes of surface cells. E’ : in crypt cells, NaKα (arrow) and CLC2 (arrowhead) localized in the basolateral membranes of crypts. Identical results obtained in different specimens obtained from three different experiments with three different male and female (not shown) rat distal colons.

    Journal: American Journal of Physiology - Gastrointestinal and Liver Physiology

    Article Title: Parallel intermediate conductance K+ and Cl− channel activity mediates electroneutral K+ exit across basolateral membranes in rat distal colon

    doi: 10.1152/ajpgi.00011.2020

    Figure Lengend Snippet: Localization of CLC2- and Na + -K + -ATPase α-subunit (NaKα)-like proteins in rat distal colon. A and A’ : CLC2-like proteins (green) localized predominantly in surface cells (arrows), while minimally expressed in crypt regions (asterisks). B and B’ : NaKα-like proteins (red) localized to both surface (arrows) and crypt (asterisks) cells. C and C’ : nuclei (blue) stained with Hoechst 33342. D : composite of A , B , and C . D’ : composite of A’ , B’ , and C’ . Higher magnification of the boxed region from D and D’ presented in E and E’ , respectively. E : CLC2 and NaKα colocalized (orange) in both lateral (arrows) and basement (arrowheads) membranes of surface cells. E’ : in crypt cells, NaKα (arrow) and CLC2 (arrowhead) localized in the basolateral membranes of crypts. Identical results obtained in different specimens obtained from three different experiments with three different male and female (not shown) rat distal colons.

    Article Snippet: The NaKα1 monoclonal antibody has been used extensively and has been shown to recognize newly synthesized Na+ -K+ -ATPase α-subunit that binds the N-azidobenzoyl derivative of ouabain used as substrate during immunoprecipitation in Madin-Darby canine kidney (MDCK) cells ( ).

    Techniques: Staining

    Expression of dopamine transporter (DAT), norepinephrine transporter (NET), and serotonin transporter (SERT) in astroglial cultures prepared from Sprague-Dawley newborn cortex (a), (c), (e) or embryonic striatum (b), (d), (f). Confocal laser-scanning microscopic images showing double immunostaining for DAT (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (a) and (b)), NET (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (c) and (d)), or SERT (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (e) and (f)) in cortical or striatal astroglia prepared from Sprague-Dawley rats. Scale bar = 20 µm.

    Journal: ASN NEURO

    Article Title: Neuroprotective Role of Astroglia in Parkinson Disease by Reducing Oxidative Stress Through Dopamine-Induced Activation of Pentose-Phosphate Pathway

    doi: 10.1177/1759091418775562

    Figure Lengend Snippet: Expression of dopamine transporter (DAT), norepinephrine transporter (NET), and serotonin transporter (SERT) in astroglial cultures prepared from Sprague-Dawley newborn cortex (a), (c), (e) or embryonic striatum (b), (d), (f). Confocal laser-scanning microscopic images showing double immunostaining for DAT (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (a) and (b)), NET (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (c) and (d)), or SERT (fluorescein-isothiocyanate, green) and GFAP (rhodamine, red) with nuclear staining (4′,6-diamino-2-phenylindole, blue; (e) and (f)) in cortical or striatal astroglia prepared from Sprague-Dawley rats. Scale bar = 20 µm.

    Article Snippet: Chemicals Chemicals and materials were obtained from the following sources: 2-deoxy-D-[1-14 C]glucose ([14 C]deoxyglucose; specific activity, 2.13 GBq/mmol), Insta-Fluor Plus, and hyamine hydroxide 10-X were obtained from Perkin-Elmer Life Sciences (Boston, MA, USA); D-[1-14 C]glucose (specific activity, 2.035 GBq/mmol) and D-[6-14 C]glucose (specific activity, 2.035 GBq/mmol) were obtained from American Radiolabeled Chemicals, Inc. (St. Louis, MO, USA); normal goat serum was obtained from Jackson ImmunoResearch (West Grove, PA, USA); rabbit anti-DAT antibody and rabbit anti-NET were obtained from Alomone Labs (Jerusalem, Israel); mouse anti-SERT antibody was obtained from Chemicon International (Temecula, CA, USA); mouse monoclonal anti-glial fibrillary acidic protein (GFAP) was obtained from Sigma (St. Louis, MO, USA); rabbit anti-GFAP antibody was obtained from DAKO (Glostrup, Denmark); rhodamine-conjugated goat anti-mouse (for GFAP), anti-rabbit (for GFAP) IgG antibody, and fluorescein-isothiocyanate-conjugated goat anti-rabbit (for DAT and NET) and anti-mouse (for SERT) antibody were obtained from Santa Cruz Biochemistry (Delaware, CA, USA); Dulbecco’s modified Eagle media (DMEM) with or without glucose, penicillin, and streptomycin were obtained from Life Technologies (Grand Island, NY, USA); defined fetal bovine serum was obtained from HyClone Laboratories (Logan, UT, USA); and 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA) and monochlorobimane (MCB) were obtained from Molecular Probe Inc. (Eugene, OR, USA); and alamarBlue was obtained from Thermo Fisher Scientific (Waltham, MA, USA).

    Techniques: Expressing, Double Immunostaining, Staining

    The expressions of neurotransmitters and different subunits of GABA A Rs. A‐B, The expression of GABA was decreased in the laparotomy mice, and no difference was found about glutamate (n = 9). C, The different average spectra of selected metabolites (GABA and glutamate). D‐F, The mRNA level of α5 subunit was up‐regulated at 1 day and continued to 10 days after laparotomy. No difference was found about the α1 and β3 subunits (n = 3). G‐I, The expressions of GAT‐3 and GAD65 were decreased, and the levels of surface α5GABA A Rs were increased in the laparotomy mice (n = 4). Data are presented as mean ± SEM. * P

    Journal: CNS Neuroscience & Therapeutics

    Article Title: Disruption of the GABAergic system contributes to the development of perioperative neurocognitive disorders after anesthesia and surgery in aged mice, et al. Disruption of the GABAergic system contributes to the development of perioperative neurocognitive disorders after anesthesia and surgery in aged mice

    doi: 10.1111/cns.13388

    Figure Lengend Snippet: The expressions of neurotransmitters and different subunits of GABA A Rs. A‐B, The expression of GABA was decreased in the laparotomy mice, and no difference was found about glutamate (n = 9). C, The different average spectra of selected metabolites (GABA and glutamate). D‐F, The mRNA level of α5 subunit was up‐regulated at 1 day and continued to 10 days after laparotomy. No difference was found about the α1 and β3 subunits (n = 3). G‐I, The expressions of GAT‐3 and GAD65 were decreased, and the levels of surface α5GABA A Rs were increased in the laparotomy mice (n = 4). Data are presented as mean ± SEM. * P

    Article Snippet: The antibodies used in this study include rabbit anti‐α5GABAA receptors, anti‐GAT‐3 (1:500‐1000; Alomone Labs), rabbit anti‐GAD65 (1:1000; Abcam), rabbit anti‐P38, p‐P38, ERK1/2, p‐ERK1/2, JNK1/2, p‐JNK1/2 (1:1000‐2000; Cell Signaling Technology), mouse anti‐GAPDH HRP‐conjugated goat‐anti‐mouse IgG, or anti‐rabbit IgG (1:1000‐5000; Promoter).

    Techniques: Expressing, Mouse Assay