gad Search Results


gad1 expression construct  (ATCC)
94
ATCC gad1 expression construct
Generation and validation of the Tg ( <t>Npy-eGFP/miRNA:Gad1</t> ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.
Gad1 Expression Construct, supplied by ATCC, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/gad/pmc03011211-53-17-10?v=ATCC
Average 94 stars, based on 1 article reviews
gad1 expression construct - by Bioz Stars, 2026-08
94/100 stars
  Buy from Supplier
mouse anti gad 65  (Developmental Studies Hybridoma Bank)
94
Developmental Studies Hybridoma Bank mouse anti gad 65
Generation and validation of the Tg ( <t>Npy-eGFP/miRNA:Gad1</t> ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.
Mouse Anti Gad 65, supplied by Developmental Studies Hybridoma Bank, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/gad/pmc06554622-195-34-37?v=Developmental+Studies+Hybridoma+Bank
Average 94 stars, based on 1 article reviews
mouse anti gad 65 - by Bioz Stars, 2026-08
94/100 stars
  Buy from Supplier
mouse anti gad67  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology mouse anti gad67
Generation and validation of the Tg ( <t>Npy-eGFP/miRNA:Gad1</t> ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.
Mouse Anti Gad67, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/gad/10__1213_slash_ane__0000000000000729-84-23-26?v=Santa+Cruz+Biotechnology
Average 93 stars, based on 1 article reviews
mouse anti gad67 - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
polyclonal antibodies for gad1  (Proteintech)
94
Proteintech polyclonal antibodies for gad1
Generation and validation of the Tg ( <t>Npy-eGFP/miRNA:Gad1</t> ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.
Polyclonal Antibodies For Gad1, supplied by Proteintech, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/gad/pm41611658-91-4-20?v=Proteintech
Average 94 stars, based on 1 article reviews
polyclonal antibodies for gad1 - by Bioz Stars, 2026-08
94/100 stars
  Buy from Supplier
anti glutamate decarboxylase 1 gad1 mouse monoclonal antiserum  (Proteintech)
93
Proteintech anti glutamate decarboxylase 1 gad1 mouse monoclonal antiserum
Generation and validation of the Tg ( <t>Npy-eGFP/miRNA:Gad1</t> ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.
Anti Glutamate Decarboxylase 1 Gad1 Mouse Monoclonal Antiserum, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/gad/pm41820376-283-40-47?v=Proteintech
Average 93 stars, based on 1 article reviews
anti glutamate decarboxylase 1 gad1 mouse monoclonal antiserum - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
mouse santa cruz biotech  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology mouse santa cruz biotech
Generation and validation of the Tg ( <t>Npy-eGFP/miRNA:Gad1</t> ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.
Mouse Santa Cruz Biotech, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/gad/pmc12158774__41586_2025_8985_MOESM1_ESM-12-147-148?v=Santa+Cruz+Biotechnology
Average 93 stars, based on 1 article reviews
mouse santa cruz biotech - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
gad6  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology gad6
Characterization of the SOXB2 expression in the developing dorsal midbrain at E12.5. Coronal sections of SOX21 GFPn midbrains stained with BrdU (A) , MASH1 (B) , LHX2 and SOX14 (C) and GATA3 (D) antibodies. Coronal sections of SOX14 +/− midbrains were stained with BHLHB5 and SOX21 (E) , BHLHB5 and SOX2 (F) , SOX21 and GATA2 (G) , BHLHB5 and GATA2 (H) , BrdU and LHX2 (I) , GATA3 and LHX2 (K) and <t>GAD6</t> (J) antibodies. The results of this analysis are summarized in the schematic (L) . Red, green and blue arrows indicate the corresponding single stained cells (as shown in the label) and yellow and purple arrows denote co-localization between green and red cells and blue and red, respectively. Scale bar, 150 μm.
Gad6, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti pgp9 5  (Boster Bio)
93
Boster Bio anti pgp9 5
Characterization of the SOXB2 expression in the developing dorsal midbrain at E12.5. Coronal sections of SOX21 GFPn midbrains stained with BrdU (A) , MASH1 (B) , LHX2 and SOX14 (C) and GATA3 (D) antibodies. Coronal sections of SOX14 +/− midbrains were stained with BHLHB5 and SOX21 (E) , BHLHB5 and SOX2 (F) , SOX21 and GATA2 (G) , BHLHB5 and GATA2 (H) , BrdU and LHX2 (I) , GATA3 and LHX2 (K) and <t>GAD6</t> (J) antibodies. The results of this analysis are summarized in the schematic (L) . Red, green and blue arrows indicate the corresponding single stained cells (as shown in the label) and yellow and purple arrows denote co-localization between green and red cells and blue and red, respectively. Scale bar, 150 μm.
Anti Pgp9 5, supplied by Boster Bio, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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gad67 activity  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology gad67 activity
Characterization of the SOXB2 expression in the developing dorsal midbrain at E12.5. Coronal sections of SOX21 GFPn midbrains stained with BrdU (A) , MASH1 (B) , LHX2 and SOX14 (C) and GATA3 (D) antibodies. Coronal sections of SOX14 +/− midbrains were stained with BHLHB5 and SOX21 (E) , BHLHB5 and SOX2 (F) , SOX21 and GATA2 (G) , BHLHB5 and GATA2 (H) , BrdU and LHX2 (I) , GATA3 and LHX2 (K) and <t>GAD6</t> (J) antibodies. The results of this analysis are summarized in the schematic (L) . Red, green and blue arrows indicate the corresponding single stained cells (as shown in the label) and yellow and purple arrows denote co-localization between green and red cells and blue and red, respectively. Scale bar, 150 μm.
Gad67 Activity, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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mouse anit β actin  (Proteintech)
93
Proteintech mouse anit β actin
a Representative image of AAV-DIO-Gad2 expression in the LS. Scale bar, 500 μm. This pattern of expression and projection was consistently observed across 3 independent biological replicates. b Left: western blot analysis of Gad2 expression in the EGFP (Control) and Gad2 (Gad2 overexpression) groups <t>(β-actin</t> was an internal reference). Right: quantification of Gad2 signal intensity for the EGFP and Gad2 groups ( p < 0.05). c Quantification of GABA expression levels for the EGFP and Gad2 groups ( p < 0.01). d Schematic of the experimental design. e – g Liquid food intake in EGFP vs. Gad2-overexpressing mice. Left: licking behavior; middle: cumulative licks; right: total intake of standard liquid food ( e ), sucrose solution ( f ) and Ensure ( g ) (standard liquid food: p = 0.94; sucrose solution: p < 0.001; Ensure: p < 0.001). h Viral strategy and timeline for overexpression Gad2 in LS GABA neurons. i Quantification of energy intake (left) and body weight (right) for the Chow+EGFP, Chow+Gad2, HFD + EGFP, and HFD+Gad2 groups [Energy intake (virus: F 1,28 = 18.1, p < 0.001; diet: F 1,28 = 29.8; p < 0.001; time: F 3.08,86.25 = 33.3, p < 0.001). Body weight (virus: F 1,28 = 10.1, p < 0.01; diet: F 1,28 = 10.5; p < 0.01; time: F 1.848,51.75 = 219.9, p < 0.001)]. j Quantification of Gad2 expression level (virus: F 1,27 = 6.2, p < 0.05; diet: F 1,27 = 1895; p < 0.001; interaction: F 1,27 = 7.9, p < 0.01). Statistics: ( b , c , e–g ) Mann-Whitney U-test, ( i ) three-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, ( j ) two-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, with detailed statistics provided in Supplementary Data . Sample sizes are indicated in the figures. ns, no significant difference, * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as mean ± SEM. All tests are two-sided. Source data are provided as a Source Data file.
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gad67  (OriGene)
90
OriGene gad67
a Representative image of AAV-DIO-Gad2 expression in the LS. Scale bar, 500 μm. This pattern of expression and projection was consistently observed across 3 independent biological replicates. b Left: western blot analysis of Gad2 expression in the EGFP (Control) and Gad2 (Gad2 overexpression) groups <t>(β-actin</t> was an internal reference). Right: quantification of Gad2 signal intensity for the EGFP and Gad2 groups ( p < 0.05). c Quantification of GABA expression levels for the EGFP and Gad2 groups ( p < 0.01). d Schematic of the experimental design. e – g Liquid food intake in EGFP vs. Gad2-overexpressing mice. Left: licking behavior; middle: cumulative licks; right: total intake of standard liquid food ( e ), sucrose solution ( f ) and Ensure ( g ) (standard liquid food: p = 0.94; sucrose solution: p < 0.001; Ensure: p < 0.001). h Viral strategy and timeline for overexpression Gad2 in LS GABA neurons. i Quantification of energy intake (left) and body weight (right) for the Chow+EGFP, Chow+Gad2, HFD + EGFP, and HFD+Gad2 groups [Energy intake (virus: F 1,28 = 18.1, p < 0.001; diet: F 1,28 = 29.8; p < 0.001; time: F 3.08,86.25 = 33.3, p < 0.001). Body weight (virus: F 1,28 = 10.1, p < 0.01; diet: F 1,28 = 10.5; p < 0.01; time: F 1.848,51.75 = 219.9, p < 0.001)]. j Quantification of Gad2 expression level (virus: F 1,27 = 6.2, p < 0.05; diet: F 1,27 = 1895; p < 0.001; interaction: F 1,27 = 7.9, p < 0.01). Statistics: ( b , c , e–g ) Mann-Whitney U-test, ( i ) three-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, ( j ) two-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, with detailed statistics provided in Supplementary Data . Sample sizes are indicated in the figures. ns, no significant difference, * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as mean ± SEM. All tests are two-sided. Source data are provided as a Source Data file.
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gad1 human shrna lentiviral particles  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology gad1 human shrna lentiviral particles
A <t>GAD1</t> expression of KRAS wild-type (WT) CRC cell lines plotted against their metastatic potential in mice (DepMap). B GAD1 expression of KRAS mutant CRC cell lines plotted against their metastatic potential in mice (DepMap). C Schematic of OOC model of tumor cell lines invading from the top channel into the bottom endothelial channel. D On-chip tumor invasion of GAD1 knockdown HCT116 tumor cells, normalized to control <t>shRNA.</t> Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0005. E On-chip tumor invasion of HCT116 tumor cells treated with GAD1 inhibitor 3-MPA (5 μM), normalized to vehicle control. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0006. F On-chip tumor invasion of HCT116 tumor cells with increasing concentrations of exogenous GABA. Data was analyzed using a one-way ANOVA with Šídák’s multiple comparison test. N = 6–8 chips; **** P < 0.0001. G Intracellular [ 13 C 4 ]GABA or unlabeled GABA was measured via mass spectrometry-based metabolomics in the HCT116 tumor-chips after the addition of exogenous GABA for six days. N = 3 chips. The intensities of the internal standard 13 C 11 Tryptophan (Trp) were plotted as a control. H On-chip GABA staining in the top channel (1) representing non-invaded HCT116 tumor cells, and in the bottom channel (2) capturing invaded HCT116 tumor cells. Scale bars represent 200 μm in the top channel image and 100 μm in the bottom channel images. Individual data points are shown and mean ± SEM are represented. All schematics were made in or are from BioRender.
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Image Search Results


Generation and validation of the Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.

Journal: Molecular Psychiatry

Article Title: Novel animal models for studying complex brain disorders: BAC-driven mi RNA-mediated in vivo silencing of gene expression

doi: 10.1038/mp.2010.1

Figure Lengend Snippet: Generation and validation of the Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM mice. ( a ) Schematic linear representation of the construct. Flanking regions of the construct are derived from the driver NPY BAC that provides interneuron subtype-specific expression of the construct. The 5′ region of the driver BAC (NPY promoter region) ensures specific expression of eGFP, and the SV40-pA ensures proper polyadenylation. The construct, containing a non-functional (not translated) part of β-globin exons 1 and 2, and intron 1 in its entirety, is spliced by the cellular machinery, liberating the intron that contains the 70–100 nucleotide long double-stranded miRNA directed against GAD1. The spliced intron- miRNA:Gad1 is processed through DROSHA and exported from the nucleus. In the cytoplasm the miRNA binds to and degrades the endogenous GAD1 mRNA through an RNA-induced silencing complex (RISC). , The eGFP mRNA is translated, thus fluorescently labeling cells of interest. The presence of LoxP sites facilitates generation of animals lacking the silencing part of the miRNA:Gad1 construct, but still expresses eGFP, to serve as controls. ( b ) miRNA-mediated downregulation of GAD1 in vitro . CHO cells co-transfected with GFP reporter vector alone (−), two different intermediate constructs (GFP reporter vectors containing two different synthetic miRNAs directed against GAD1—nos. 1 and 2), and GAD1 expression vector. The cells were harvested at 24 and 48 h after transfection. Western blot analysis confirmed that the two different miRNAs against GAD1 were correctly processed from the β-globin intron and that both miRNAs strongly downregulated GAD1 expression. Similar results have been obtained in HEK293 cells with stable expression of GAD1 that were engineered in our laboratory (data not shown). The same cells transfected with CMV-eGFP,miRNA:Gad1 show high levels of eGFP expression, confirming that the construct performs as expected. In addition, the processing of the miRNA from the β-globin intron in the construct does not interfere with eGFP protein translation. ( c ) Identification of founder animals by PCR-based genotyping of founder animals using eGFP primers with genomic DNA as template. The first lane represents size marker, and the last lane corresponds to positive control, with the remaining lanes containing eGFP amplification products from the genomic DNA of individual animals. The 550-nt product on a 1% agarose gel indicates construct incorporation into the genome of four founder animals (NPY 1, 3, 4 and 5). Similar results were obtained by Southern hybridization (data not sown). ( d ) Quantitative PCR (qPCR) amplification plot from frontal cortex of founder animals using eGFP construct-specific primers. The y axis denotes PCR product accumulation, and the x axis denotes amplification cycle number. Note that two (NPY1 and NPY4) of the four founder lines that incorporated the NPY-BAC/GAD1-miRNA construct reported functional eGFP expression. These lines were used for further characterization. ( e ) The NPY-BAC/GAD1-miRNA construct showed the expected tissue distribution in the brain. Micrographs depict the fidelity of co-localized eGFP and NPY in adult transgenic animals from line Tg ( Npy-eGFP/miRNA:Gad1 ) 1KM . The left column micrographs denote eGFP immunostaining, middle column micrographs represent sections labeled with anti-NPY and the right column micrographs illustrate pseudocolored composite of eGFP-NPY co-localization in the same tissue sections. In the cortex, roman numerals denote cortical laminae. Hippocampus abbreviations: gcl, granule cell layer; h, hilus; ml, molecular layer. Note that all GFP+ neurons are also NPY+, and all NPY+ cells are GFP+, suggesting that the construct is specifically and exclusively expressed in the phenotypically appropriate target cell population. Calibration bar=100 μm. ( f , g ) Tg ( Npy-eGFP/ miRNA:Gad1 ) 1KM animals show undetectable GAD1 levels in the frontal cortex and hippocampus of NPY+ cells compared with NPY+ neurons in control animals. eGFP-GAD1 double-immunohistochemistry (eGFP, green; GAD1, red) was performed from a coronal section through the frontal cortex and hippocampus of a transgenic animal. Wild-type control littermate was double stained against NPY and GAD1. Same cells are denoted by arrowheads. In the cortex, roman numerals denote cortical laminae. Hippocampal abbreviations: f, hippocampal fissure; gcl, ganglion cell layer; h, hilus; ml, molecular layer; slm, stratum lacunosum moleculare; sp, stratum pyramidale; sr, stratum radiatum. Note that all NPY+ cells (white arrows) are GAD1+ in control animals, whereas none of the eGFP+ (and thus NPY+) neurons (white arrows) show GAD1 staining in the transgenic mice. Moreover, note the large number of single-labeled GAD1+ neurons that are eGFP– in the transgenic line. These data indicate selective, miRNA-mediated, cell type-specific downregulation of GAD1 in NPY+ interneurons. Calibration bars=60 μm (overview figures) and 100 μm.

Article Snippet: Intermediate construct DNA (CMV-eGFP, miRNA:Gad1) was co-transfected into HEK-293 cells (American Type Culture Collection) along with a GAD1 expression construct (IMAGE clone 5358787) in a 3:1 ratio using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) according to the manufacturer's instructions.

Techniques: Biomarker Discovery, Construct, Derivative Assay, Expressing, Functional Assay, Labeling, In Vitro, Transfection, Plasmid Preparation, Western Blot, Marker, Positive Control, Amplification, Agarose Gel Electrophoresis, Hybridization, Real-time Polymerase Chain Reaction, Transgenic Assay, Immunostaining, Control, Immunohistochemistry, Staining

Characterization of the SOXB2 expression in the developing dorsal midbrain at E12.5. Coronal sections of SOX21 GFPn midbrains stained with BrdU (A) , MASH1 (B) , LHX2 and SOX14 (C) and GATA3 (D) antibodies. Coronal sections of SOX14 +/− midbrains were stained with BHLHB5 and SOX21 (E) , BHLHB5 and SOX2 (F) , SOX21 and GATA2 (G) , BHLHB5 and GATA2 (H) , BrdU and LHX2 (I) , GATA3 and LHX2 (K) and GAD6 (J) antibodies. The results of this analysis are summarized in the schematic (L) . Red, green and blue arrows indicate the corresponding single stained cells (as shown in the label) and yellow and purple arrows denote co-localization between green and red cells and blue and red, respectively. Scale bar, 150 μm.

Journal: Frontiers in Molecular Neuroscience

Article Title: Sequential Role of SOXB2 Factors in GABAergic Neuron Specification of the Dorsal Midbrain

doi: 10.3389/fnmol.2018.00152

Figure Lengend Snippet: Characterization of the SOXB2 expression in the developing dorsal midbrain at E12.5. Coronal sections of SOX21 GFPn midbrains stained with BrdU (A) , MASH1 (B) , LHX2 and SOX14 (C) and GATA3 (D) antibodies. Coronal sections of SOX14 +/− midbrains were stained with BHLHB5 and SOX21 (E) , BHLHB5 and SOX2 (F) , SOX21 and GATA2 (G) , BHLHB5 and GATA2 (H) , BrdU and LHX2 (I) , GATA3 and LHX2 (K) and GAD6 (J) antibodies. The results of this analysis are summarized in the schematic (L) . Red, green and blue arrows indicate the corresponding single stained cells (as shown in the label) and yellow and purple arrows denote co-localization between green and red cells and blue and red, respectively. Scale bar, 150 μm.

Article Snippet: Immunohistochemistry was performed according to standard procedures using the following primary antibodies: FLAG (fusion protein tag, mouse IgG, Sigma Aldrich, 1:100), SOX14 (guinea pig IgG, Gift from Tom Jessell, 1:2000), GATA3 (lineage-specific marker, rat IgG, gift from Frank Rosvelt, 1:50), DBX1 (lineage-determining factor, rabbit IgG, gift from Alessandra Pieraani, 1:1000), SOX2 (stem cell marker, rabbit IgG, gift from Eumorphia Remboutsika, 1:500), MASH1 (lineage-determining factor, guinea pig IgG, gift from Francois Guillemot, 1:2000), SOX21 (goat IgG, Neuromics USA, 1:200), NKX2.2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), NKX6.1 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GAD6 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GATA2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), PAX6 (lineage-specific marker, rabbit IgG, Millipore USA, 1:200), BRN3A (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), LHX2 (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), BHLHB5 (lineage-specific marker, rabbit IgG, Santa Cruz Biotechnology USA, 1:500), CyclinD1 (proliferation marker, mouse IgG, Santa Cruz Biotechnology USA, 1:50), P27 (cell cycle arrest marker, rabbit IgG, Abcam UK, 1:500), Ki67 (proliferation marker, rabbit IgG, Abcam UK, 1:500) and BrdU (proliferation marker, rat IgG, Abcam UK, 1:100).

Techniques: Expressing, Staining

Analysis of the GABAergic and glutamatergic lineages in SOX21 −/− embryos at E12.5. The immunostaining of the GABAergic marker GATA3 (A,A’) was shown to be reduced in the knockout embryos ( ii ) in comparison to the wild type ( i ). However, the expression of SOX14 (B,B’) and GAD6 (C,C’) as well as the glutamatergic marker LHX2 (D,D’) were not affected in the absence of SOX21. All nuclei were counted dorsally to the ventrolateral midbrain sulcus (indicated by white arrow). These data were confirmed by RT-Q-PCR analysis of embryos at the same age (E) . Data are represented as mean ± SEM. * n = 10 (5 pairs of mice), P = 0.006, two-tailed Mann-Whitney test. ** n = 6 (3 pairs of mice), p = 0.042, DF = 1, Kruskal Wallis test. Scale bar, 150 μm.

Journal: Frontiers in Molecular Neuroscience

Article Title: Sequential Role of SOXB2 Factors in GABAergic Neuron Specification of the Dorsal Midbrain

doi: 10.3389/fnmol.2018.00152

Figure Lengend Snippet: Analysis of the GABAergic and glutamatergic lineages in SOX21 −/− embryos at E12.5. The immunostaining of the GABAergic marker GATA3 (A,A’) was shown to be reduced in the knockout embryos ( ii ) in comparison to the wild type ( i ). However, the expression of SOX14 (B,B’) and GAD6 (C,C’) as well as the glutamatergic marker LHX2 (D,D’) were not affected in the absence of SOX21. All nuclei were counted dorsally to the ventrolateral midbrain sulcus (indicated by white arrow). These data were confirmed by RT-Q-PCR analysis of embryos at the same age (E) . Data are represented as mean ± SEM. * n = 10 (5 pairs of mice), P = 0.006, two-tailed Mann-Whitney test. ** n = 6 (3 pairs of mice), p = 0.042, DF = 1, Kruskal Wallis test. Scale bar, 150 μm.

Article Snippet: Immunohistochemistry was performed according to standard procedures using the following primary antibodies: FLAG (fusion protein tag, mouse IgG, Sigma Aldrich, 1:100), SOX14 (guinea pig IgG, Gift from Tom Jessell, 1:2000), GATA3 (lineage-specific marker, rat IgG, gift from Frank Rosvelt, 1:50), DBX1 (lineage-determining factor, rabbit IgG, gift from Alessandra Pieraani, 1:1000), SOX2 (stem cell marker, rabbit IgG, gift from Eumorphia Remboutsika, 1:500), MASH1 (lineage-determining factor, guinea pig IgG, gift from Francois Guillemot, 1:2000), SOX21 (goat IgG, Neuromics USA, 1:200), NKX2.2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), NKX6.1 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GAD6 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GATA2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), PAX6 (lineage-specific marker, rabbit IgG, Millipore USA, 1:200), BRN3A (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), LHX2 (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), BHLHB5 (lineage-specific marker, rabbit IgG, Santa Cruz Biotechnology USA, 1:500), CyclinD1 (proliferation marker, mouse IgG, Santa Cruz Biotechnology USA, 1:50), P27 (cell cycle arrest marker, rabbit IgG, Abcam UK, 1:500), Ki67 (proliferation marker, rabbit IgG, Abcam UK, 1:500) and BrdU (proliferation marker, rat IgG, Abcam UK, 1:100).

Techniques: Immunostaining, Marker, Knock-Out, Comparison, Expressing, Two Tailed Test, MANN-WHITNEY

Analysis of the GABAergic neuron maturation in GATA3 eGFP /SOX21 −/− E12.5 embryos. The immunostaining of the GABAergic marker GATA3 was shown to be reduced in the knockout embryos ( iii,iv ) in comparison to the wild type ( i,ii ) despite no noticeable changes in the expression of GFP (A,A′) . Immunostaining of the sections with CNTN2 illustrated a significant reduction in the co-localization with the GFP +ve axons in the absence of SOX21 (B,B′) as well as a decrease in the thickness of GAD6 +ve axons (C) —white arrows denote the thickness of the axonal bundle, yellow arrows denote GAD6 co-localization.In addition, the arrangement of the GFP +ve cells was investigated in E12.5 clarified midbrains (D) .The dashed lines of the panel ( D iii , D′ iii ) denote the magnified region shown in panel ( D v , D′ v ) and the white arrows indicate the axonal length (R-Rostral, C-Caudal, L-Left, R-Right).Data are represented as mean ± SEM. * n = 10 (5 pairs of mice), P = 0.012, two-tailed Mann-Whitney test. Scale bar, 150 μm.

Journal: Frontiers in Molecular Neuroscience

Article Title: Sequential Role of SOXB2 Factors in GABAergic Neuron Specification of the Dorsal Midbrain

doi: 10.3389/fnmol.2018.00152

Figure Lengend Snippet: Analysis of the GABAergic neuron maturation in GATA3 eGFP /SOX21 −/− E12.5 embryos. The immunostaining of the GABAergic marker GATA3 was shown to be reduced in the knockout embryos ( iii,iv ) in comparison to the wild type ( i,ii ) despite no noticeable changes in the expression of GFP (A,A′) . Immunostaining of the sections with CNTN2 illustrated a significant reduction in the co-localization with the GFP +ve axons in the absence of SOX21 (B,B′) as well as a decrease in the thickness of GAD6 +ve axons (C) —white arrows denote the thickness of the axonal bundle, yellow arrows denote GAD6 co-localization.In addition, the arrangement of the GFP +ve cells was investigated in E12.5 clarified midbrains (D) .The dashed lines of the panel ( D iii , D′ iii ) denote the magnified region shown in panel ( D v , D′ v ) and the white arrows indicate the axonal length (R-Rostral, C-Caudal, L-Left, R-Right).Data are represented as mean ± SEM. * n = 10 (5 pairs of mice), P = 0.012, two-tailed Mann-Whitney test. Scale bar, 150 μm.

Article Snippet: Immunohistochemistry was performed according to standard procedures using the following primary antibodies: FLAG (fusion protein tag, mouse IgG, Sigma Aldrich, 1:100), SOX14 (guinea pig IgG, Gift from Tom Jessell, 1:2000), GATA3 (lineage-specific marker, rat IgG, gift from Frank Rosvelt, 1:50), DBX1 (lineage-determining factor, rabbit IgG, gift from Alessandra Pieraani, 1:1000), SOX2 (stem cell marker, rabbit IgG, gift from Eumorphia Remboutsika, 1:500), MASH1 (lineage-determining factor, guinea pig IgG, gift from Francois Guillemot, 1:2000), SOX21 (goat IgG, Neuromics USA, 1:200), NKX2.2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), NKX6.1 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GAD6 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GATA2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), PAX6 (lineage-specific marker, rabbit IgG, Millipore USA, 1:200), BRN3A (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), LHX2 (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), BHLHB5 (lineage-specific marker, rabbit IgG, Santa Cruz Biotechnology USA, 1:500), CyclinD1 (proliferation marker, mouse IgG, Santa Cruz Biotechnology USA, 1:50), P27 (cell cycle arrest marker, rabbit IgG, Abcam UK, 1:500), Ki67 (proliferation marker, rabbit IgG, Abcam UK, 1:500) and BrdU (proliferation marker, rat IgG, Abcam UK, 1:100).

Techniques: Immunostaining, Marker, Knock-Out, Comparison, Expressing, Two Tailed Test, MANN-WHITNEY

Analysis of the GABAergic and glutamatergic lineages in SOX14 −/− embryos at E12.5. The absence of SOX14 in the knockout embryos was validated by antibody staining (A) .The immunostaining of the GABAergic markers GABA, GAD6 and GATA3 (B,C,E) were shown to be reduced in the knockout embryos ( iii,iv ) in comparison to the heterozygotes ( i,ii ). However, the expression of SOX21 (D) and the glutamatergic marker LHX2 (F) were not affected in the absence of SOX14. These data were confirmed by RT-Q-PCR analysis of embryos at the same age (G) . Data are represented as mean ± SEM. * n = 6 (3 pairs of mice), P = 0.032, DF = 1, Kruskal Wallis test. ** n = 6 (3 pairs of mice), P = 0.034, DF = 1, Kruskal Wallis test. *** n = 6 (3 pairs of mice), p = 0.021, DF = 1, Kruskal Wallis test. Scale bar, 150 μm.

Journal: Frontiers in Molecular Neuroscience

Article Title: Sequential Role of SOXB2 Factors in GABAergic Neuron Specification of the Dorsal Midbrain

doi: 10.3389/fnmol.2018.00152

Figure Lengend Snippet: Analysis of the GABAergic and glutamatergic lineages in SOX14 −/− embryos at E12.5. The absence of SOX14 in the knockout embryos was validated by antibody staining (A) .The immunostaining of the GABAergic markers GABA, GAD6 and GATA3 (B,C,E) were shown to be reduced in the knockout embryos ( iii,iv ) in comparison to the heterozygotes ( i,ii ). However, the expression of SOX21 (D) and the glutamatergic marker LHX2 (F) were not affected in the absence of SOX14. These data were confirmed by RT-Q-PCR analysis of embryos at the same age (G) . Data are represented as mean ± SEM. * n = 6 (3 pairs of mice), P = 0.032, DF = 1, Kruskal Wallis test. ** n = 6 (3 pairs of mice), P = 0.034, DF = 1, Kruskal Wallis test. *** n = 6 (3 pairs of mice), p = 0.021, DF = 1, Kruskal Wallis test. Scale bar, 150 μm.

Article Snippet: Immunohistochemistry was performed according to standard procedures using the following primary antibodies: FLAG (fusion protein tag, mouse IgG, Sigma Aldrich, 1:100), SOX14 (guinea pig IgG, Gift from Tom Jessell, 1:2000), GATA3 (lineage-specific marker, rat IgG, gift from Frank Rosvelt, 1:50), DBX1 (lineage-determining factor, rabbit IgG, gift from Alessandra Pieraani, 1:1000), SOX2 (stem cell marker, rabbit IgG, gift from Eumorphia Remboutsika, 1:500), MASH1 (lineage-determining factor, guinea pig IgG, gift from Francois Guillemot, 1:2000), SOX21 (goat IgG, Neuromics USA, 1:200), NKX2.2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), NKX6.1 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GAD6 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), GATA2 (lineage-specific marker, mouse IgG, DSHB USA, 1:50), PAX6 (lineage-specific marker, rabbit IgG, Millipore USA, 1:200), BRN3A (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), LHX2 (lineage-specific marker, goat IgG, Santa Cruz Biotechnology USA, 1:200), BHLHB5 (lineage-specific marker, rabbit IgG, Santa Cruz Biotechnology USA, 1:500), CyclinD1 (proliferation marker, mouse IgG, Santa Cruz Biotechnology USA, 1:50), P27 (cell cycle arrest marker, rabbit IgG, Abcam UK, 1:500), Ki67 (proliferation marker, rabbit IgG, Abcam UK, 1:500) and BrdU (proliferation marker, rat IgG, Abcam UK, 1:100).

Techniques: Knock-Out, Staining, Immunostaining, Comparison, Expressing, Marker

a Representative image of AAV-DIO-Gad2 expression in the LS. Scale bar, 500 μm. This pattern of expression and projection was consistently observed across 3 independent biological replicates. b Left: western blot analysis of Gad2 expression in the EGFP (Control) and Gad2 (Gad2 overexpression) groups (β-actin was an internal reference). Right: quantification of Gad2 signal intensity for the EGFP and Gad2 groups ( p < 0.05). c Quantification of GABA expression levels for the EGFP and Gad2 groups ( p < 0.01). d Schematic of the experimental design. e – g Liquid food intake in EGFP vs. Gad2-overexpressing mice. Left: licking behavior; middle: cumulative licks; right: total intake of standard liquid food ( e ), sucrose solution ( f ) and Ensure ( g ) (standard liquid food: p = 0.94; sucrose solution: p < 0.001; Ensure: p < 0.001). h Viral strategy and timeline for overexpression Gad2 in LS GABA neurons. i Quantification of energy intake (left) and body weight (right) for the Chow+EGFP, Chow+Gad2, HFD + EGFP, and HFD+Gad2 groups [Energy intake (virus: F 1,28 = 18.1, p < 0.001; diet: F 1,28 = 29.8; p < 0.001; time: F 3.08,86.25 = 33.3, p < 0.001). Body weight (virus: F 1,28 = 10.1, p < 0.01; diet: F 1,28 = 10.5; p < 0.01; time: F 1.848,51.75 = 219.9, p < 0.001)]. j Quantification of Gad2 expression level (virus: F 1,27 = 6.2, p < 0.05; diet: F 1,27 = 1895; p < 0.001; interaction: F 1,27 = 7.9, p < 0.01). Statistics: ( b , c , e–g ) Mann-Whitney U-test, ( i ) three-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, ( j ) two-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, with detailed statistics provided in Supplementary Data . Sample sizes are indicated in the figures. ns, no significant difference, * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as mean ± SEM. All tests are two-sided. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: High-fat diet disrupts a septal control on feeding to promote obesity in male mice

doi: 10.1038/s41467-025-68010-x

Figure Lengend Snippet: a Representative image of AAV-DIO-Gad2 expression in the LS. Scale bar, 500 μm. This pattern of expression and projection was consistently observed across 3 independent biological replicates. b Left: western blot analysis of Gad2 expression in the EGFP (Control) and Gad2 (Gad2 overexpression) groups (β-actin was an internal reference). Right: quantification of Gad2 signal intensity for the EGFP and Gad2 groups ( p < 0.05). c Quantification of GABA expression levels for the EGFP and Gad2 groups ( p < 0.01). d Schematic of the experimental design. e – g Liquid food intake in EGFP vs. Gad2-overexpressing mice. Left: licking behavior; middle: cumulative licks; right: total intake of standard liquid food ( e ), sucrose solution ( f ) and Ensure ( g ) (standard liquid food: p = 0.94; sucrose solution: p < 0.001; Ensure: p < 0.001). h Viral strategy and timeline for overexpression Gad2 in LS GABA neurons. i Quantification of energy intake (left) and body weight (right) for the Chow+EGFP, Chow+Gad2, HFD + EGFP, and HFD+Gad2 groups [Energy intake (virus: F 1,28 = 18.1, p < 0.001; diet: F 1,28 = 29.8; p < 0.001; time: F 3.08,86.25 = 33.3, p < 0.001). Body weight (virus: F 1,28 = 10.1, p < 0.01; diet: F 1,28 = 10.5; p < 0.01; time: F 1.848,51.75 = 219.9, p < 0.001)]. j Quantification of Gad2 expression level (virus: F 1,27 = 6.2, p < 0.05; diet: F 1,27 = 1895; p < 0.001; interaction: F 1,27 = 7.9, p < 0.01). Statistics: ( b , c , e–g ) Mann-Whitney U-test, ( i ) three-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, ( j ) two-way ANOVA followed by post hoc test using two-stage step-up method of Benjamini, Krieger and Yekutieli, with detailed statistics provided in Supplementary Data . Sample sizes are indicated in the figures. ns, no significant difference, * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as mean ± SEM. All tests are two-sided. Source data are provided as a Source Data file.

Article Snippet: The primary antibodies used were rabbit anti-Gad65 (1:3000, Proteintech, Cat# 20746-1-AP) and mouse anit-β-actin (1:3000, Proteintech, Cat# 66009), The secondary antibodies used were goat anti-rabbit (1:2000, ThermoFisher, Cat# 32460) or goat anti-mouse (1:2000, ThermoFisher, Cat# 62-6520).

Techniques: Expressing, Western Blot, Control, Over Expression, Virus, MANN-WHITNEY

A GAD1 expression of KRAS wild-type (WT) CRC cell lines plotted against their metastatic potential in mice (DepMap). B GAD1 expression of KRAS mutant CRC cell lines plotted against their metastatic potential in mice (DepMap). C Schematic of OOC model of tumor cell lines invading from the top channel into the bottom endothelial channel. D On-chip tumor invasion of GAD1 knockdown HCT116 tumor cells, normalized to control shRNA. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0005. E On-chip tumor invasion of HCT116 tumor cells treated with GAD1 inhibitor 3-MPA (5 μM), normalized to vehicle control. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0006. F On-chip tumor invasion of HCT116 tumor cells with increasing concentrations of exogenous GABA. Data was analyzed using a one-way ANOVA with Šídák’s multiple comparison test. N = 6–8 chips; **** P < 0.0001. G Intracellular [ 13 C 4 ]GABA or unlabeled GABA was measured via mass spectrometry-based metabolomics in the HCT116 tumor-chips after the addition of exogenous GABA for six days. N = 3 chips. The intensities of the internal standard 13 C 11 Tryptophan (Trp) were plotted as a control. H On-chip GABA staining in the top channel (1) representing non-invaded HCT116 tumor cells, and in the bottom channel (2) capturing invaded HCT116 tumor cells. Scale bars represent 200 μm in the top channel image and 100 μm in the bottom channel images. Individual data points are shown and mean ± SEM are represented. All schematics were made in or are from BioRender.

Journal: Oncogene

Article Title: GABAergic signaling contributes to tumor cell invasion and poor overall survival in colorectal cancer

doi: 10.1038/s41388-025-03546-2

Figure Lengend Snippet: A GAD1 expression of KRAS wild-type (WT) CRC cell lines plotted against their metastatic potential in mice (DepMap). B GAD1 expression of KRAS mutant CRC cell lines plotted against their metastatic potential in mice (DepMap). C Schematic of OOC model of tumor cell lines invading from the top channel into the bottom endothelial channel. D On-chip tumor invasion of GAD1 knockdown HCT116 tumor cells, normalized to control shRNA. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0005. E On-chip tumor invasion of HCT116 tumor cells treated with GAD1 inhibitor 3-MPA (5 μM), normalized to vehicle control. Data was analyzed using an unpaired t-test. N = 6 chips; *** P = 0.0006. F On-chip tumor invasion of HCT116 tumor cells with increasing concentrations of exogenous GABA. Data was analyzed using a one-way ANOVA with Šídák’s multiple comparison test. N = 6–8 chips; **** P < 0.0001. G Intracellular [ 13 C 4 ]GABA or unlabeled GABA was measured via mass spectrometry-based metabolomics in the HCT116 tumor-chips after the addition of exogenous GABA for six days. N = 3 chips. The intensities of the internal standard 13 C 11 Tryptophan (Trp) were plotted as a control. H On-chip GABA staining in the top channel (1) representing non-invaded HCT116 tumor cells, and in the bottom channel (2) capturing invaded HCT116 tumor cells. Scale bars represent 200 μm in the top channel image and 100 μm in the bottom channel images. Individual data points are shown and mean ± SEM are represented. All schematics were made in or are from BioRender.

Article Snippet: GAD1 human shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc- sc-35435-V) and control shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc-108080) were used to produce HCT116 H2B-GFP GAD1 knock down cells at a multiplicity of infection (MOI) of 2 and 2 mg/mL of puromycin (Santa Cruz Biotechnology, #sc-108071) was used to select for knocked down cells.

Techniques: Expressing, Mutagenesis, Knockdown, Control, shRNA, Comparison, Mass Spectrometry, Staining

Forest plots show the association of GAD1 tumor expression as a continuous variable with PFS ( A ) and OS ( B ) across treatment arms; Forest plots show the association of ABAT tumor expression with PFS ( C ) and OS ( D ) across treatment arms. Kaplan-Meier curves show PFS ( E ) and OS ( F ) stratified by GAD1 tumor expression tertiles (T1 low, T2 middle, and T3 high) in the overall population of the CALGB/SWOG 80405 trial. The statistical comparison is performed for T3 vs T1. Kaplan-Meier curves show PFS ( G ) and OS ( H ) stratified by ABAT tumor expression tertiles in the overall population of the CALGB/SWOG 80405 trial. The statistical comparison is performed for T3 vs T1. P values are based on log-rank test for PFS and OS in the univariate categorical analysis, and likelihood ratio test in the univariate continuous and multivariable Cox proportional hazards regression model. bev bevacizumab, cet cetuximab, CI confidence interval, HR hazard ratio, OS overall survival, PFS progression-free survival.

Journal: Oncogene

Article Title: GABAergic signaling contributes to tumor cell invasion and poor overall survival in colorectal cancer

doi: 10.1038/s41388-025-03546-2

Figure Lengend Snippet: Forest plots show the association of GAD1 tumor expression as a continuous variable with PFS ( A ) and OS ( B ) across treatment arms; Forest plots show the association of ABAT tumor expression with PFS ( C ) and OS ( D ) across treatment arms. Kaplan-Meier curves show PFS ( E ) and OS ( F ) stratified by GAD1 tumor expression tertiles (T1 low, T2 middle, and T3 high) in the overall population of the CALGB/SWOG 80405 trial. The statistical comparison is performed for T3 vs T1. Kaplan-Meier curves show PFS ( G ) and OS ( H ) stratified by ABAT tumor expression tertiles in the overall population of the CALGB/SWOG 80405 trial. The statistical comparison is performed for T3 vs T1. P values are based on log-rank test for PFS and OS in the univariate categorical analysis, and likelihood ratio test in the univariate continuous and multivariable Cox proportional hazards regression model. bev bevacizumab, cet cetuximab, CI confidence interval, HR hazard ratio, OS overall survival, PFS progression-free survival.

Article Snippet: GAD1 human shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc- sc-35435-V) and control shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc-108080) were used to produce HCT116 H2B-GFP GAD1 knock down cells at a multiplicity of infection (MOI) of 2 and 2 mg/mL of puromycin (Santa Cruz Biotechnology, #sc-108071) was used to select for knocked down cells.

Techniques: Expressing, Comparison

A GAD1 mRNA expression from TCGA in primary colon cancer tumors. N = 196 patients with KRAS , NRAS , or BRAF mutant tumors; N = 201 patients with KRAS , NRAS , or BRAF wildtype tumors. Individual data points are shown and the median with interquartile range is represented. Data was analyzed with an unpaired t-test; **** P < 0.0001. B GAD1 mRNA expression from the Gene Expression Omnibus (GEO), accession GSE39582 , in primary colon cancer tumors. N = 220 patients with KRAS or BRAF mutant tumors; N = 276 patients with KRAS or BRAF wildtype tumors. Individual data points are shown and the median with interquartile range is represented. Data was analyzed with an unpaired t-test; **** P < 0.0001. C Kaplan-Meier curve with univariate analysis of the survival of patients with KRAS , NRAS , or BRAF mutated CRC tumors based on high versus low expression of GAD1 (defined as above or below the median GAD1 mRNA expression z-score of 0.3). Data was extracted from the TCGA. N = 254 patients. Data was analyzed using a log-rank (Mantel-Cox test) and a hazard ratio (HR) with 95% confidence interval (CI) is reported. D Kaplan-Meier curve with univariate analysis of the survival of patients with KRAS or BRAF mutated CRC tumors based on high versus low expression of GAD1 (defined as above or below the median GAD1 mRNA expression of 3.4). Data was extracted from the GEO, accession GSE39582 . N = 217 patients. Data was analyzed using a log-rank (Mantel-Cox test) and a hazard ratio (HR) with 95% confidence interval (CI) is reported. All schematics were made in or are from BioRender.

Journal: Oncogene

Article Title: GABAergic signaling contributes to tumor cell invasion and poor overall survival in colorectal cancer

doi: 10.1038/s41388-025-03546-2

Figure Lengend Snippet: A GAD1 mRNA expression from TCGA in primary colon cancer tumors. N = 196 patients with KRAS , NRAS , or BRAF mutant tumors; N = 201 patients with KRAS , NRAS , or BRAF wildtype tumors. Individual data points are shown and the median with interquartile range is represented. Data was analyzed with an unpaired t-test; **** P < 0.0001. B GAD1 mRNA expression from the Gene Expression Omnibus (GEO), accession GSE39582 , in primary colon cancer tumors. N = 220 patients with KRAS or BRAF mutant tumors; N = 276 patients with KRAS or BRAF wildtype tumors. Individual data points are shown and the median with interquartile range is represented. Data was analyzed with an unpaired t-test; **** P < 0.0001. C Kaplan-Meier curve with univariate analysis of the survival of patients with KRAS , NRAS , or BRAF mutated CRC tumors based on high versus low expression of GAD1 (defined as above or below the median GAD1 mRNA expression z-score of 0.3). Data was extracted from the TCGA. N = 254 patients. Data was analyzed using a log-rank (Mantel-Cox test) and a hazard ratio (HR) with 95% confidence interval (CI) is reported. D Kaplan-Meier curve with univariate analysis of the survival of patients with KRAS or BRAF mutated CRC tumors based on high versus low expression of GAD1 (defined as above or below the median GAD1 mRNA expression of 3.4). Data was extracted from the GEO, accession GSE39582 . N = 217 patients. Data was analyzed using a log-rank (Mantel-Cox test) and a hazard ratio (HR) with 95% confidence interval (CI) is reported. All schematics were made in or are from BioRender.

Article Snippet: GAD1 human shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc- sc-35435-V) and control shRNA lentiviral particles (Santa Cruz Biotechnology, Inc., #sc-108080) were used to produce HCT116 H2B-GFP GAD1 knock down cells at a multiplicity of infection (MOI) of 2 and 2 mg/mL of puromycin (Santa Cruz Biotechnology, #sc-108071) was used to select for knocked down cells.

Techniques: Expressing, Mutagenesis, Gene Expression