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A ) Schematic of wild-type GPR75 gene <t>construct</t> <t>and</t> <t>GPR75-GFP-Ires-Cre</t> gene construct. B ) Gel electrophoresis confirmation of Cre insertion, 100kB DNA ladder (lane 1; L), homozygous GPR75-GFP-Ires-Cre genotype (lane 2), Wild-type genotype (lane 3), heterozygous GPR75-GFP-Ires-Cre genotype (lane 4). C-F) In situ hybridization of GPR75 (red), EGFP (green), iCre (blue), and DAPI (white) for ( C ) wild-type, ( D ) heterozygous GPR75-GFP-Ires-Cre, ( E ) homozygous GPR75-GFP-Ires-Cre, and (F) total body deletion of GPR75 in the ventromedial hypothalamus. D’-E’ ). A yellow triangle points to colocalization of GPR75, EGFP, and iCre. C-F ) Images taken at 20x magnification, ( C’-F’ ) taken at 20x magnification with 5x zoom. Scale bar is 50µm in all images.
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A ) Schematic of wild-type GPR75 gene construct and GPR75-GFP-Ires-Cre gene construct. B ) Gel electrophoresis confirmation of Cre insertion, 100kB DNA ladder (lane 1; L), homozygous GPR75-GFP-Ires-Cre genotype (lane 2), Wild-type genotype (lane 3), heterozygous GPR75-GFP-Ires-Cre genotype (lane 4). C-F) In situ hybridization of GPR75 (red), EGFP (green), iCre (blue), and DAPI (white) for ( C ) wild-type, ( D ) heterozygous GPR75-GFP-Ires-Cre, ( E ) homozygous GPR75-GFP-Ires-Cre, and (F) total body deletion of GPR75 in the ventromedial hypothalamus. D’-E’ ). A yellow triangle points to colocalization of GPR75, EGFP, and iCre. C-F ) Images taken at 20x magnification, ( C’-F’ ) taken at 20x magnification with 5x zoom. Scale bar is 50µm in all images.

Journal: bioRxiv

Article Title: A genetic tool targeting brain GPR75

doi: 10.64898/2026.05.29.728898

Figure Lengend Snippet: A ) Schematic of wild-type GPR75 gene construct and GPR75-GFP-Ires-Cre gene construct. B ) Gel electrophoresis confirmation of Cre insertion, 100kB DNA ladder (lane 1; L), homozygous GPR75-GFP-Ires-Cre genotype (lane 2), Wild-type genotype (lane 3), heterozygous GPR75-GFP-Ires-Cre genotype (lane 4). C-F) In situ hybridization of GPR75 (red), EGFP (green), iCre (blue), and DAPI (white) for ( C ) wild-type, ( D ) heterozygous GPR75-GFP-Ires-Cre, ( E ) homozygous GPR75-GFP-Ires-Cre, and (F) total body deletion of GPR75 in the ventromedial hypothalamus. D’-E’ ). A yellow triangle points to colocalization of GPR75, EGFP, and iCre. C-F ) Images taken at 20x magnification, ( C’-F’ ) taken at 20x magnification with 5x zoom. Scale bar is 50µm in all images.

Article Snippet: In situ hybridization of brain sections from wild-type C57BL6, GPR75-KO, and GPR75-GFP-Ires-Cre mice was performed using the RNAscope Multiplex Fluorescent V2 Assay kit (323100, ACDBio) according to the manufacturer’s guidelines.

Techniques: Construct, Nucleic Acid Electrophoresis, In Situ Hybridization

A-C ) Body weight and fat data for GPR75-GFP-Ires-Cre mice on HFD. A ) Overall body weight gain on high fat diet in homozygous, heterozygous, and wild-type GPR75-GFP-Ires-Cre mice over 10 weeks (Two-Way Repeated Measures ANOVA, Week; F (1.220, 31.72) = 356.9, p < 0.0001, Genotype; F (2, 26) = 0.4474, p = 0.6441). B ) Absolute body weight gain on HFD (Two-Way Repeated Measures ANOVA, Week; F (1.220, 31.72) = 356.9, p < 0.0001, Genotype; F (2, 23.96) = 0.4542, p = 0.6399). C ) Percentage of fat mass to total mass after 10 weeks of HFD (One-Way ANOVA; F (x,x) = x, P = x) . D-F ) A separate cohort of mice, GPR75-KO and wild-type control were also fed HFD. D ) Overall body weight gain on HFD in wild-type and GPR75-KO mice (Two-Way Repeated Measures ANOVA, Week; F (1.272, 25.43) = 173.9, p < 0.0001, Genotype; F (1, 20) = 36.55, p < 0.0001). E ) Absolute body weight gain on HFD (Two-Way Repeated Measures ANOVA, Week; F (1.231, 24.61) = 173.9, p < 0.0001, Genotype; F (1, 20) = 17.27, p = 0.0005). F ) Percentage of fat mass to total mass after 10 weeks of HFD (unpaired two-tailed t-test Welch Corrected; t (16.58) = 2.356, P = 0.0311).

Journal: bioRxiv

Article Title: A genetic tool targeting brain GPR75

doi: 10.64898/2026.05.29.728898

Figure Lengend Snippet: A-C ) Body weight and fat data for GPR75-GFP-Ires-Cre mice on HFD. A ) Overall body weight gain on high fat diet in homozygous, heterozygous, and wild-type GPR75-GFP-Ires-Cre mice over 10 weeks (Two-Way Repeated Measures ANOVA, Week; F (1.220, 31.72) = 356.9, p < 0.0001, Genotype; F (2, 26) = 0.4474, p = 0.6441). B ) Absolute body weight gain on HFD (Two-Way Repeated Measures ANOVA, Week; F (1.220, 31.72) = 356.9, p < 0.0001, Genotype; F (2, 23.96) = 0.4542, p = 0.6399). C ) Percentage of fat mass to total mass after 10 weeks of HFD (One-Way ANOVA; F (x,x) = x, P = x) . D-F ) A separate cohort of mice, GPR75-KO and wild-type control were also fed HFD. D ) Overall body weight gain on HFD in wild-type and GPR75-KO mice (Two-Way Repeated Measures ANOVA, Week; F (1.272, 25.43) = 173.9, p < 0.0001, Genotype; F (1, 20) = 36.55, p < 0.0001). E ) Absolute body weight gain on HFD (Two-Way Repeated Measures ANOVA, Week; F (1.231, 24.61) = 173.9, p < 0.0001, Genotype; F (1, 20) = 17.27, p = 0.0005). F ) Percentage of fat mass to total mass after 10 weeks of HFD (unpaired two-tailed t-test Welch Corrected; t (16.58) = 2.356, P = 0.0311).

Article Snippet: In situ hybridization of brain sections from wild-type C57BL6, GPR75-KO, and GPR75-GFP-Ires-Cre mice was performed using the RNAscope Multiplex Fluorescent V2 Assay kit (323100, ACDBio) according to the manufacturer’s guidelines.

Techniques: Control, Two Tailed Test

A-B) Maximum intensity projection images of DAPI and GFP immunostaining acquired from an 11.02µm Z-stack (0.92µm Z-step) in C57BL6 ( A-A’ ) and from a 12.04µm Z-stack (0.93µm Z-step) in GPR75-GFP-Ires-Cre ( B-B’ ) mouse thalamic brain tissue taken at 20x magnification with a 3x zoom, scale bar is 50µm. C-C’ ) Maximum intensity projection image of GFP and DAPI immunostaining in GPR75-GFP-Ires-Cre mouse thalamic tissue acquired from a 16.44µm Z-stack (0.2µm Z-step) taken at 60x magnification with a 2x zoom, scale bar is 25µm. D ) Representative 3D rendering of DAPI and GFP in GPR75-GFP-Ires-Cre mouse thalamic tissue. 3D rendering was compiled from a 16.44µm Z-stack (0.2µm Z-step), scale bar is 25µm.

Journal: bioRxiv

Article Title: A genetic tool targeting brain GPR75

doi: 10.64898/2026.05.29.728898

Figure Lengend Snippet: A-B) Maximum intensity projection images of DAPI and GFP immunostaining acquired from an 11.02µm Z-stack (0.92µm Z-step) in C57BL6 ( A-A’ ) and from a 12.04µm Z-stack (0.93µm Z-step) in GPR75-GFP-Ires-Cre ( B-B’ ) mouse thalamic brain tissue taken at 20x magnification with a 3x zoom, scale bar is 50µm. C-C’ ) Maximum intensity projection image of GFP and DAPI immunostaining in GPR75-GFP-Ires-Cre mouse thalamic tissue acquired from a 16.44µm Z-stack (0.2µm Z-step) taken at 60x magnification with a 2x zoom, scale bar is 25µm. D ) Representative 3D rendering of DAPI and GFP in GPR75-GFP-Ires-Cre mouse thalamic tissue. 3D rendering was compiled from a 16.44µm Z-stack (0.2µm Z-step), scale bar is 25µm.

Article Snippet: In situ hybridization of brain sections from wild-type C57BL6, GPR75-KO, and GPR75-GFP-Ires-Cre mice was performed using the RNAscope Multiplex Fluorescent V2 Assay kit (323100, ACDBio) according to the manufacturer’s guidelines.

Techniques: Immunostaining