Journal: Schizophrenia Bulletin

Article Title: Erbb4 Deletion From Inhibitory Interneurons Causes Psychosis-Relevant Neuroimaging Phenotypes

doi: 10.1093/schbul/sbac192

Figure Lengend Snippet: Representative hippocampal regions of interests for quantitative autoradiography. (A) Regions of interests sampled for [ 3 H]Ro15-4513 and [ 3 H]flumazenil. (B) Regions of interests sampled for [ 3 H]UCB-J. dHip CA1: dorsal hippocampus CA1; mHip CA3: middle hippocampus CA3; mHip CA1/2: middle hippocampus CA1/CA2; vHip CA3: ventral hippocampus CA3; dHip DG: dorsal hippocampus dentate gyrus; mHip DG: middle hippocampus dentate gyrus.

Article Snippet: To quantify density of α5GABA A R, sections were incubated for 60 min at room temperature in 2 nM [ 3 H]Ro15-4513 (Perkin Elmer, NET925250UC), or in 2nM [ 3 H]Ro15-4513 with 10 µM bretazenil (Sigma, B6434) for nonspecific binding.

Techniques: Autoradiography

Journal: Schizophrenia Bulletin

Article Title: Erbb4 Deletion From Inhibitory Interneurons Causes Psychosis-Relevant Neuroimaging Phenotypes

doi: 10.1093/schbul/sbac192

Figure Lengend Snippet: SV2A density, but not α5GABA A R or α1-3;5GABA A R, is decreased in Lhx6-Cre;Erbb4 F/F mice across the hippocampus. (A) [ 3 H]UCB-J autoradiography showed a significant decrease in binding in Lhx6-Cre;Erbb4 F/F mice ( n = 11, 8 female, 3 male) compared to control animals ( n = 9, 3 female, 6 male) across all hippocampal ROIs ( P = .02, η 2 = 0.29). (B) [ 3 H]Ro15-4513 ( Erbb4 F/F n = 11, 5 female, 6 male; Lhx6;Erbb4 F/F n = 10, 7 female, 3 male) or (C) [ 3 H]flumazenil ( Erbb4 F/F n = 10, 4 female, 6 male; Lhx6;Erbb4 F/F n = 9, 7 female, 2 male) binding did not significantly differ by genotype. dHip CA1: dorsal hippocampus CA1; mHip CA3: middle hippocampus CA3; mHip CA1/2: middle hippocampus CA1/CA2; vHip CA3: ventral hippocampus CA3; dHip DG: dorsal hippocampus dentate gyrus; mHip DG: dentate gyrus.

Article Snippet: To quantify density of α5GABA A R, sections were incubated for 60 min at room temperature in 2 nM [ 3 H]Ro15-4513 (Perkin Elmer, NET925250UC), or in 2nM [ 3 H]Ro15-4513 with 10 µM bretazenil (Sigma, B6434) for nonspecific binding.

Techniques: Autoradiography, Binding Assay

Journal: bioRxiv

Article Title: Erbb4 deletion from fast-spiking interneurons causes psychosis-relevant neuroimaging phenotypes

doi: 10.1101/2022.03.07.483347

Figure Lengend Snippet: Representative hippocampal regions of interests. (A) Regions of interests sampled for [ 3 H]Ro15-4513 and [ 3 H]flumazenil. (B) Regions of interests sampled for [ 3 H]UCB-J. dHip CA1: dorsal hippocampus CA1; mHip CA3: middle hippocampus CA3; mHip CA1/2: middle hippocampus CA1/CA2; vHip CA3: ventral hippocampus CA3; dHip DG: dorsal hippocampus dentate gyrus; mHip DG: middle hippocampus dentate gyrus.

Article Snippet: To quantify density of α5GABA A R – sections were incubated for 60 minutes at room temperature in 2nM [ 3 H]Ro15-4513 (Perkin Elmer, NET925250UC), or in 2nM [ 3 H]Ro15-4513 with 10 µM bretazenil (Sigma, B6434) for nonspecific binding.

Techniques:

Journal: bioRxiv

Article Title: Erbb4 deletion from fast-spiking interneurons causes psychosis-relevant neuroimaging phenotypes

doi: 10.1101/2022.03.07.483347

Figure Lengend Snippet: [ 3 H]UCB-J, but not [ 3 H]Ro15-4513 or [ 3 H]flumazenil binding in Lhx6-Cre;Erbb4 F/F mice is decreased across the hippocampus. (A) [ 3 H]UCB-J showed a significant decrease in synaptic density in Lhx6-Cre;Erbb4 F/F mice (n=11) compared to control animals (n=9) across all hippocampal ROIs (mixed effects analysis; main effect of genotype: F (1,18) =7.27, p =0.02, ). (B) [ 3 H]Ro15-4513 (n=10-11/group) and (C) [ 3 H]flumazenil (n=9-10/group) binding did not differ by genotype. dHip CA1: dorsal hippocampus CA1; mHip CA3: middle hippocampus CA3; mHip CA1/2: middle hippocampus CA1/CA2; vHip CA3: ventral hippocampus CA3; dHip DG: dorsal hippocampus dentate gyrus; mHip DG: dentate gyrus.

Article Snippet: To quantify density of α5GABA A R – sections were incubated for 60 minutes at room temperature in 2nM [ 3 H]Ro15-4513 (Perkin Elmer, NET925250UC), or in 2nM [ 3 H]Ro15-4513 with 10 µM bretazenil (Sigma, B6434) for nonspecific binding.

Techniques: Binding Assay

Journal: The Journal of biological chemistry

Article Title: GABA A α 6-Containing Receptors Are Selectively Compromised in Cerebellar Granule Cells of the Ataxic Mouse, Stargazer *

doi: 10.1074/jbc.M700111200

Figure Lengend Snippet: Full saturation [3H]Ro15-4513 binding curves were generated using +/+ and stg cerebellar membranes and a concentration range of 0.3–40 nm [3H]Ro15-4513. Nonspecific binding was determined in the presence of Ro15-1788 (10 μm). [3H]Ro15-4513 binding in the presence of flunitrazepam (10 μm), defined BZ-IS-binding sites, and hence benzodiazepine-sensitive (BZ-S) sites could be determined by subtraction of the BZ-IS-binding sites from total [3H]Ro15-4513 specific binding sites. The bar graph illustrates the difference in Bmax values calculated for [3H]Ro15-4513 binding to +/+ and stg membranes. Results show differences in Bmax for total specific binding (100 ± 5% and 81 ± 7%), BZ-S binding (100 ± 11% and 92 ± 9%), and BZ-IS binding (100 ± 3% and 57 ± 12%) in +/+ and stg membranes, respectively. The table summarizes the estimated Bmax and KD values for [3H]Ro15-4513 binding to +/+ and stg cerebellar membranes. Data are representative of the mean ± S.E. for assays conducted in triplicate on two separate membrane preparations per mouse strain (n = 6). Ten cerebella were used for each preparation per mouse strain.

Article Snippet: [ 3 H]Muscimol and [ 3 H]Ro15-4513 were purchased from PerkinElmer Life Sciences.

Techniques: Binding Assay, Generated, Concentration Assay

Journal: The Journal of biological chemistry

Article Title: GABA A α 6-Containing Receptors Are Selectively Compromised in Cerebellar Granule Cells of the Ataxic Mouse, Stargazer *

doi: 10.1074/jbc.M700111200

Figure Lengend Snippet: A, total [3H]Ro15-4513 binding. Panel i, in situ autoradiography of total [3H]Ro15-4513 binding (γ2-containing GABARs) to +/+ and stg cerebella using a near-saturating concentration of [3H]Ro15-4513 (20 nm). Nonspecific binding was determined by competitive displacement of Ro15-4513 with the benzodiazepine receptor antagonist, Ro15-1788 (10 μm). The signal obtained under these latter conditions was at the level of film background. Two representative comparable sections per mouse strain are shown. Panel ii, magnified image of cerebellar lobules labeled with [3H]Ro15-4513. Panel iii, histogram illustrating the results of quantitative image analysis of grayscale intensities using image J software to determine the relative amounts of ligand bound in the cerebellar granule cell layers. A small (10 ± 4%) but insignificant (p > 0.05) reduction in total [3H]Ro15-4513 binding in the cerebellar granule cell layer of stg was determined. Data shown are representative of at least three +/+ and three stg brains and a minimum of 10 sections per brain. B, BZ-IS [3H]Ro15-4513 binding. Panel i, in situ autoradiography of flunitrazepam (10 μm)-insensitive [3H]Ro15-4513 (BZ-ISR) binding (α6βγ2, α1α6βγ2 GABARs) in +/+ and stg cerebella using a saturating concentration of [3H]Ro15-4513 (20 nm). Nonspecific binding was determined by competitive displacement of Ro15-4513 with benzodiazepine receptor antagonist, Ro15-1788 (10 μm). The signal obtained under these conditions was at the level of film background. Panel ii, magnified image of cerebellar lobules following autoradiography to identify BZ-ISRs. Panel iii, histogram illustrating the results of quantitative image analysis of grayscale intensities using image J software to determine the relative amounts of radioligand bound in the cerebellar granule cell layer. A small but significant reduction (by 21 ± 7%; p < 0.05) in flunitrazepam-insensitive [3H]Ro15-4513 binding in the cerebellar granule cell layer of stg was determined. Data shown are representative of at least three +/+ and three stg brains and a minimum of 10 sections per brain. C, BZ-S [3H]flunitrazepam binding. Panel i, in situ autoradiography of [3H]flunitrazepam (BZ-SR) binding (e.g. α1βγ2 GABARs) in +/+ and stg cerebella using [3H]flunitrazepam (5 nm). Nonspecific binding was determined by competitive displacement of flunitrazepam with benzodiazepine receptor antagonist, Ro15-1788 (10 μm). The signal obtained under these conditions was at the level of film background. Panel ii, magnified image of cerebellar lobules following autoradiography to identify BZ-SRs. Panel iii, histogram illustrating the results of quantitative image analysis of grayscale intensities using image J software to determine the relative amounts of radioligand bound in the cerebellar granule cell layer. A small, nonsignificant increase (by 13 ± 6%; p > 0.05) in [3H]flunitrazepam binding in the cerebellum of stg was determined.

Article Snippet: [ 3 H]Muscimol and [ 3 H]Ro15-4513 were purchased from PerkinElmer Life Sciences.

Techniques: Binding Assay, In Situ, Autoradiography, Concentration Assay, Labeling, Software

Journal: The Journal of biological chemistry

Article Title: GABA A α 6-Containing Receptors Are Selectively Compromised in Cerebellar Granule Cells of the Ataxic Mouse, Stargazer *

doi: 10.1074/jbc.M700111200

Figure Lengend Snippet: Zolpidem (1 nm to 10 μm) was used to competitively displace BZ-S [3H]Ro15-4513 (5 nm) equilibrium binding from cerebellar membrane homogenates derived from adult +/+ and stg mice. [3H]Ro15-4513 binding displaced by flunitrazepam (10 μm) defined benzodiazepine-sensitive (BZ-S) sites. 100% binding was that obtained in the absence of competitive ligand.

Article Snippet: [ 3 H]Muscimol and [ 3 H]Ro15-4513 were purchased from PerkinElmer Life Sciences.

Techniques: Binding Assay, Derivative Assay

Journal: The Journal of Neuroscience

Article Title: Loss of the Major GABA A Receptor Subtype in the Brain Is Not Lethal in Mice

doi: 10.1523/JNEUROSCI.21-10-03409.2001

Figure Lengend Snippet: Amount (fmol/mg protein) of GABA A receptors and benzodiazepine binding sites in WT, α1−/−, and β2−/− mice determined by nonlinear regression analysis of saturation data sets

Article Snippet: Radioligand binding assays with [ 3 H]Ro15–1788 (87 Ci/mmol; NEN Life Sciences), [ 3 H]Ro15–4513 (21.7 Ci/mmol; NEN Life Sciences) in the presence or absence of 10 μ m diazepam and [ 3 H]muscimol (19.1 Ci/mmol; NEN Life Sciences) were performed on membrane preparations as previously described ( Quirk et al., 1994 ; Sur et al., 1998 , 1999a ).

Techniques: Binding Assay

Journal: The Journal of Neuroscience

Article Title: Loss of the Major GABA A Receptor Subtype in the Brain Is Not Lethal in Mice

doi: 10.1523/JNEUROSCI.21-10-03409.2001

Figure Lengend Snippet: Pharmacological characterization of GABA A receptors in WT, α1 (−/−), and β2 (−/−) mice

Article Snippet: Radioligand binding assays with [ 3 H]Ro15–1788 (87 Ci/mmol; NEN Life Sciences), [ 3 H]Ro15–4513 (21.7 Ci/mmol; NEN Life Sciences) in the presence or absence of 10 μ m diazepam and [ 3 H]muscimol (19.1 Ci/mmol; NEN Life Sciences) were performed on membrane preparations as previously described ( Quirk et al., 1994 ; Sur et al., 1998 , 1999a ).

Techniques:

Journal: The Journal of Neuroscience

Article Title: Loss of the Major GABA A Receptor Subtype in the Brain Is Not Lethal in Mice

doi: 10.1523/JNEUROSCI.21-10-03409.2001

Figure Lengend Snippet: Generation-dependent upregulation of α2 and α3 subunits in cerebellum and forebrain of α1−/− mice.a, The loss of [3H]Ro15–1788 binding sites increased in cerebellum (left panel) and forebrain (right panel) of α1−/− mice from generation F2 to F5 to finally reach the proportion of [3H]Ro15–1788 sites immunoprecipitated by a selective α1 antibody from wild-type membrane (filled bars). ***p < 0.001; Student's t test. b, Evidence for an upregulation of α3 subunit in the cerebellum of α1−/− mice. Western blot showing an increase in α3 subunit expression in knock-out mice compared with wild-type (left panel). Quantitative immunoprecipitation with a α3 antibody demonstrated that α3 subunit-containing GABAAreceptors account for 14 ± 3% (mean ± SEM;n = 3) and 92 ± 8% (mean ± SEM;n = 4) of total [3H]Ro15–1788 binding sites in the cerebellum of wild-type and α1−/− mice, respectively (right panel). c, Quantitative immunoprecipitation demonstrated a 42% increase in the expression of α2 and α3 subunit-sensitive [3H]Ro15–1788 binding sites in the forebrain of α1−/− compared with wild-type mice.

Article Snippet: Radioligand binding assays with [ 3 H]Ro15–1788 (87 Ci/mmol; NEN Life Sciences), [ 3 H]Ro15–4513 (21.7 Ci/mmol; NEN Life Sciences) in the presence or absence of 10 μ m diazepam and [ 3 H]muscimol (19.1 Ci/mmol; NEN Life Sciences) were performed on membrane preparations as previously described ( Quirk et al., 1994 ; Sur et al., 1998 , 1999a ).

Techniques: Binding Assay, Immunoprecipitation, Western Blot, Expressing, Knock-Out

Journal:

Article Title: Mice devoid of ?-aminobutyrate type A receptor ?3 subunit have epilepsy, cleft palate, and hypersensitive behavior

doi:

Figure Lengend Snippet: Binding of [ 3 H]muscimol and [ 3 H]Ro15-4513 to whole brain homogenates prepared from postnatal day 1 (pups) or 8- to 12-week-old (adult) mice

Article Snippet: The benzodiazepine sites were labeled by [ 3 H]Ro15-4513 (10 nM; DuPont/NEN) ( 17 ) with and without zolpidem (Synthelabo Recherche, Bagneux, France) during a 60-min incubation at 0–4°C.

Techniques: Binding Assay

Journal:

Article Title: Mice devoid of ?-aminobutyrate type A receptor ?3 subunit have epilepsy, cleft palate, and hypersensitive behavior

doi:

Figure Lengend Snippet: Autoradiographic binding of a GABA site ligand ([3H]muscimol) and a benzodiazepine site ligand ([3H]Ro15-4513) to representative horizontal brain sections from newborn wild-type (+/+), heterozygous (+/−), and homozygous (−/−) mice. (A) Total binding of 5 nM [3H]Ro15-4513. (B) Binding of [3H]Ro15-4513 in the presence of 30 μM zolpidem. (C) Total binding of 20 nM [3H]muscimol. All images represent specific binding since [3H]Ro15-4513 and [3H]muscimol failed to label any sites in the presence of 10 μM flumazenil and 100 μM GABA, respectively.

Article Snippet: The benzodiazepine sites were labeled by [ 3 H]Ro15-4513 (10 nM; DuPont/NEN) ( 17 ) with and without zolpidem (Synthelabo Recherche, Bagneux, France) during a 60-min incubation at 0–4°C.

Techniques: Binding Assay

Journal:

Article Title: Mice devoid of ?-aminobutyrate type A receptor ?3 subunit have epilepsy, cleft palate, and hypersensitive behavior

doi:

Figure Lengend Snippet: Binding of [ 3 H]muscimol and [ 3 H]Ro15-4513 to postnatal day 1 mouse brain sections as revealed by autoradiography

Article Snippet: The benzodiazepine sites were labeled by [ 3 H]Ro15-4513 (10 nM; DuPont/NEN) ( 17 ) with and without zolpidem (Synthelabo Recherche, Bagneux, France) during a 60-min incubation at 0–4°C.

Techniques: Binding Assay