cx43  (Alomone Labs)

Journal: Cells

Article Title: Blockade of Glial Connexin 43 Hemichannels Reduces Food Intake

doi: 10.3390/cells9112387

Figure Lengend Snippet: ( A ): Representative images depicting ethidium bromide (EtBr) uptake via hemichannels in hypothalamic (ARC) slices treated with either artificial cerebrospinal fluid (aCSF), 200 μM carbenoxolone (Cbx) or 100 µM the connexin-43 (Cx43) hemichannel blocker TAT-GAP19 for 15 min, then exposed to EtBr for 10 min. After fixation, slices were submitted to Glial Fibrillary Acidic Protein (GFAP) immunostaining and Hoechst labelling. Scale bar: 15 µm. ( B ): Quantification of EtBr fluorescence measured in GFAP+ astrocytes. n = 5, with each n representing the average of three replicates. ** p

Article Snippet: Once again, the same saturation solution was used before the Cx43 rabbit primary antibody (#ACC-201 Alomone Labs 1/3000, Jerusalem, Israel) and goat anti-rabbit-IgG secondary antibody (1/400) on the same conditions.

Techniques: Immunostaining, Fluorescence

Journal: Cells

Article Title: Blockade of Glial Connexin 43 Hemichannels Reduces Food Intake

doi: 10.3390/cells9112387

Figure Lengend Snippet: ( A , B ): Connexin-43 (Cx43) and Glial Fibrillary Acidic Protein (GFAP) mRNA expression (normalized with Glyceraldehyde-3-phosphate dehydrogenase gene, GAPDH) was assessed by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) from the hypothalamus ( A ) and dorsal vagal complex (DVC) ( B ) of control ad libitum ( n = 14), 24 h fasted ( n = 13), 24 h fasted plus 2 h refeed ( n = 8) or plus 4 h refeed ( n = 8) mice. ( C ): Representative Western blot analysis of phosphorylated Cx43 (pS368) and Cx43 expression in the hypothalamus of control ad libitum ( n = 5) and 24 h fasted ( n = 5) mice. ( D , E ): Cx43 and GFAP mRNA expressions (normalized with GAPDH) were assessed by Reverse Transcription quantitative Polymerase Chain Reaction (RT-qPCR) from the hypothalamus ( D ) and dorsal vagal complex (DVC) ( D ) of Normal Chow (NC) ( n = 9) and High Fat Diet (HFD) fed ( n = 9) mice. ( F ): Representative Western blot analysis and quantification of phosphorylated Cx43 (pS368) and Cx43 expression in the hypothalamus of NC ( n = 9) and HFD-fed ( n = 9) mice. A two-way analysis of variance test was performed between different experimental groups * p

Article Snippet: Once again, the same saturation solution was used before the Cx43 rabbit primary antibody (#ACC-201 Alomone Labs 1/3000, Jerusalem, Israel) and goat anti-rabbit-IgG secondary antibody (1/400) on the same conditions.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Mouse Assay, Western Blot

Journal: Cells

Article Title: Blockade of Glial Connexin 43 Hemichannels Reduces Food Intake

doi: 10.3390/cells9112387

Figure Lengend Snippet: ( A ): Confocal images of connexin-43 (Cx43) (red) and bassoon (green) double labeling immunohistochemistry performed within the nucleus tractus solitarius (NTS). Scale bar: 5 μm. Inset: Arrows show Cx43 labelling in close vicinity of synapses labelled by presynaptic bassoon protein. Scale bar 1 µm. ( B ): An example of a voxel (4 × 4 µm) used for the quantification of Cx43/bassoon appositions. Scale bar: 0.5 µm. ( C ): Twelve z-stack images defining the voxel illustrated in B and allowing the identification of Cx43/bassoon appositions. ( D ): Stereological estimation of Cx43/bassoon appositions in the NTS (85.13%) and arcuate nucleus of the hypothalamus (ARC) (80.46%) nuclei. ( E ): Table gathering the different stereological values acquired or estimated for each studied animal.

Article Snippet: Once again, the same saturation solution was used before the Cx43 rabbit primary antibody (#ACC-201 Alomone Labs 1/3000, Jerusalem, Israel) and goat anti-rabbit-IgG secondary antibody (1/400) on the same conditions.

Techniques: Labeling, Immunohistochemistry

Journal: Cells

Article Title: Blockade of Glial Connexin 43 Hemichannels Reduces Food Intake

doi: 10.3390/cells9112387

Figure Lengend Snippet: ( A ): Cumulative food intake (g), measured over a 24 h period, of mice having received an i.c.v. injection of either vehicle (Sodium Chloride (NaCl) 0.9%) or TAT-GAP19 (12.5 or 25 µg). The TAT-GAP19-I14A, which includes a point mutation making it ineffective on connexin 43 hemichannels (Cx43 HCs) activity was tested on food intake at the dose of 25 µg. ( B ): Graph showing meal size over 24 h after i.c.v. injection of vehicle or TAT-GAP19 25 µg. Note that each bar represents a meal and its width the meal duration. The dark period is represented by the shaded box. ( C – F ): Meal microstructure analysis performed during the dark period for mice that received either vehicle or TAT-GAP19 25 µg allowed quantification of meals number ( C ), meals size (in mg of food, ( D )), meal duration (in min, ( E )) and interprandial period (in min, average intervals between meals, ( F – G )): Measure of energy expenditure (in Kcal/day-1/kg 0.75 ) within 12 h after injection of 0.9% NaCl vehicle ( n = 7) or 50 µg TAT-GAP19 ( n = 7). ( H , I ): Measure of total spontaneous activity (in Arbitrary Unit, UA; ( H ) and cumulative spontaneous activity ( I ) within 12 h for the 2 groups. ns: no significant difference; * p

Article Snippet: Once again, the same saturation solution was used before the Cx43 rabbit primary antibody (#ACC-201 Alomone Labs 1/3000, Jerusalem, Israel) and goat anti-rabbit-IgG secondary antibody (1/400) on the same conditions.

Techniques: Mouse Assay, Injection, Mutagenesis, Activity Assay

Journal: Acta Neuropathologica Communications

Article Title: Astrocyte remodeling without gliosis precedes optic nerve Axonopathy

doi: 10.1186/s40478-018-0542-0

Figure Lengend Snippet: Diminished anterograde transport in a sample of DBA/2 J nerves. a Top Left: Coronal section through superior colliculus of DBA/2 J mouse (between dashed white lines) following intravitreal injection of CTB (green). Bottom Left: Corresponding retinotopic map shows nearly depleted anterograde transport of CTB. Top Right: Coronal section through superior colliculus of D2 control mouse prepared as on left. Bottom Right: Corresponding retinotopic map shows a full complement of anterogradely transported CTB. b Transport of CTB from DBA/2 J eyes was near 20% (22.8 ± 5.5), significantly reduced from D2 control which is near 75% (74.896 ± 4.328) ( p  = 0.0015). c Confocal micrographs of proximal (left) and distal (right) DBA/2 J (top) and D2 control (bottom) optic nerves. Connexin 43 (Cx43, blue) and GFAP (red) colocalize, and both are elevated in proximal optic nerve. d . Density of Cx43 (puncta/μm 2 ) in proximal segment of DBA/2 J nerves is significantly elevated compared to both distal DBA/2 J (*; p  = 0.043) and proximal D2 nerves (#; p  = 0.032)

Article Snippet: We labeled 10 μm cryosections with the following antibodies: anti-glial fibrillary acidic protein (GFAP; EMD Millipore, Billerica, MA, 1:500), and anti-Connexin-43 (Cx43; Alomone Labs, Jerusalem, Israel, 1:250).

Techniques: Injection, CtB Assay

Journal: PLoS ONE

Article Title: Beneficial effect of voluntary physical exercise in Plakophilin2 transgenic mice

doi: 10.1371/journal.pone.0252649

Figure Lengend Snippet: Histological analysis of cardiac tissue after long-term training. ( A ) Representative images for sedentary, 8 weeks of running and 16 weeks of running tissue from wt (upper row) and PKP2 +/- mice (lower row) are shown for Trichrome-stained ventricular slices. No change of fibrous tissue could be detected in either genotype or in response to long-term training. ( B ) The oil stain in frozen sections from cardiac tissue with oil red did not reveal an increase in fatty replacements. ( C ) We labeled the gap junctions with a Cx43 specific antibody and analyzed the images by confocal microscopy. The connexins were predominantly located at the intercalated discs as seen in the representative images shown. There was no shift in subcellular distribution due to genotype or long-term training observed. The single data points indicate the number of animals analyzed.

Article Snippet: The sections were labelled with anti-Cx43 (1:400, Alomone Labs, Jerusalem, Israel) and Alexa488 (1:500, Invitrogen) and subsequently analyzed with a confocal microscope.

Techniques: Mouse Assay, Staining, Labeling, Confocal Microscopy

Journal: PLoS ONE

Article Title: Beneficial effect of voluntary physical exercise in Plakophilin2 transgenic mice

doi: 10.1371/journal.pone.0252649

Figure Lengend Snippet: Histological analysis of cardiac tissue after long-term training. ( A ) Representative images for sedentary, 8 weeks of running and 16 weeks of running tissue from wt (upper row) and PKP2 +/- mice (lower row) are shown for Trichrome-stained ventricular slices. No change of fibrous tissue could be detected in either genotype or in response to long-term training. ( B ) The oil stain in frozen sections from cardiac tissue with oil red did not reveal an increase in fatty replacements. ( C ) We labeled the gap junctions with a Cx43 specific antibody and analyzed the images by confocal microscopy. The connexins were predominantly located at the intercalated discs as seen in the representative images shown. There was no shift in subcellular distribution due to genotype or long-term training observed. The single data points indicate the number of animals analyzed.

Article Snippet: A subset of paraffin sections was used for immunohistochemistry to study the distribution of connexin 43 (Cx43) in the ventricular tissue.

Techniques: Mouse Assay, Staining, Labeling, Confocal Microscopy