Journal: bioRxiv

Article Title: ARIH1 deficiency impairs spatial learning and memory via GIRK2 upregulation in hippocampal CaMKII-expressing neurons in mice

doi: 10.1101/2025.03.10.625121

Figure Lengend Snippet: ( A ) Workflow of quantitative proteomic analysis. Proteomic data were obtained from mice hippocampus, n =3. ( B ) Heatmaps of differentially expressed (DE) proteins in WT and ARIH1 +/- hippocampal samples. ( C ) Volcano plot for DE proteins (139 upregulated, 66 downregulated) in ARIH1 +/- hippocampal samples compared with WT hippocampus. Red and blue dots indicate statistical significance DE proteins. The hippocampal tissue of WT and ARIH1 +/- mice were dissected and whole lysis was prepared for immunoblot analysis ( D ) and total RNA was prepared for qPCR analysis ( E , F ) as described in the methods. The qPCR was probed by mouse ARIH1 or GIRK2 primers, n =4. The immunoblot was probed by anti-ARIH1 or anti-GIRK2 antibody. The mean intensity of bands was quantified using Image J and normalized to corresponding loading controls, n =4. * p < 0.05, ** p < 0.01, *** p < 0.001, ns . not significant, unpaired Student’s t test ( D , E , F ). Mean ± SEM.

Article Snippet: The Flag-ARIH1 (HG20139-CF) and HA-GIRK2 (HG18578-CY) plasmids were purchased from Sino Biological.

Techniques: Lysis, Western Blot

Journal: bioRxiv

Article Title: ARIH1 deficiency impairs spatial learning and memory via GIRK2 upregulation in hippocampal CaMKII-expressing neurons in mice

doi: 10.1101/2025.03.10.625121

Figure Lengend Snippet: HT-22 cells were infected with lentivirus expressing scramble (NC) or Arih1-shRNA of two different on-target sequence (948, 949). NC and Arih1 knockdown cells were selected and maintained in cell culture media supplemented with puromycin (2.5μg/ml for selection, and 1μg/ml for maintaining) as described in the methods. ( A ) Whole cell lysates were prepared from HT-22 NC and Arih1 knockdown cells for immunoblot analysis probed by anti-Arih1 or anti-Girk2 antibodies. ( B ) Arih1 knockdown cells were transfected with vector or Flag-Arih1 plasmids. After 48hours transfection, whole cell lysates were prepared for immunoblot analysis probed by anti-Girk2, anti-Arih1, or anti-Flag antibodies. ( C ) HT-22 NC and Arih1 knockdown cells were transfected with Flag-Ub and HA-Girk2 plasmids. After 48hours transfection, whole cell lysates were prepared and immunoprecipitation was performed to examine the ubiquitination of Girk2 using anti-Flag or anti-HA antibodies. Showing blots are representative of at least 3 independent experiments. The mean intensity of bands was quantified using Image J and normalized to corresponding loading controls ( A 1 -A 2 , B 1 -B 4 ), or to corresponding input loadings ( C 1 ). ( D , E ) The total RNA was prepared from HT-22 NC and Arih1 knockdown cells for qPCR analysis with mouse ARIH1 or GIRK2 primers, n =5. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns . not significant, one-way ANOVA followed by Tukey’s test, n =3-6. Mean ± SEM.

Article Snippet: The Flag-ARIH1 (HG20139-CF) and HA-GIRK2 (HG18578-CY) plasmids were purchased from Sino Biological.

Techniques: Infection, Expressing, shRNA, Sequencing, Knockdown, Cell Culture, Selection, Western Blot, Transfection, Plasmid Preparation, Immunoprecipitation

Journal: bioRxiv

Article Title: ARIH1 deficiency impairs spatial learning and memory via GIRK2 upregulation in hippocampal CaMKII-expressing neurons in mice

doi: 10.1101/2025.03.10.625121

Figure Lengend Snippet: Schematic of the AAV-ARIH1 shRNA ( A ) and brain injection site ( B ) in mice. ( C ) The MWM experiment is composed of 3 sections which are adaptation (day 1), training (day 2 to 7), and testing (day 8). The latency on day 1 as well as the average latency (4 quadrants) on day 2 to 7 of AAV-Scramble or AAV-ARIH1 shRNA injected mice were calculated as described in the methods. During test on day 8, the latency ( D ) as well as times crossing the platform area ( E ) of AAV-Scramble or AAV-ARIH1 shRNA injected mice were recorded as described in the methods, n =10-12. ( F ) Representative image for showing the site of injection. Scale bar, 100μm. ( G ) The dorsal hippocampal tissue of AAV-Scr and AAV-ARIH1 shRNA injected mice were dissected and whole cell lysates were prepared for immunoblot analysis probed by anti-ARIH1 antibody. The mean intensity of bands was quantified using Image J and normalized to corresponding loading controls, n =3. ( H ) Representative image for showing co-expression of AAV-GFP with CaMKII, but not with GFAP or IB1. Scale bar, 20μm. ( I ) Representative image of immunostaining of Girk2 in mice with dorsal hippocampal injection of AAV-Scramble or AAV-ARIH1 shRNA. Scale bar, 100μm. ( J ) Quantification of the intensity of Girk2 staining, n =5. ( K ) The latency on day 1 as well as the average latency (4 quadrants) on day 2 to 7 of AAV-ARIH1 shRNA injected mice, administrated with Saline or Tipepidine ( i.p. , 20mg/kg), were calculated as described in the methods. During test on day 8, the latency ( L ) as well as times crossing the platform area ( M ) of AAV-ARIH1 shRNA injected mice, administrated with Saline or Tipepidine, were recorded as described in the methods, n =10-11. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, unpaired Student’s t test ( D , E , G , J , L , M ). * p < 0.05, two-way ANOVA followed by Bonferroni’s test ( C , K ). Mean ± SEM.

Article Snippet: The Flag-ARIH1 (HG20139-CF) and HA-GIRK2 (HG18578-CY) plasmids were purchased from Sino Biological.

Techniques: shRNA, Injection, Western Blot, Expressing, Immunostaining, Staining, Saline

Journal: The Journal of Neuroscience

Article Title: G-Protein-Gated Potassium (GIRK) Channels Containing the GIRK2 Subunit Are Control Hubs for Pharmacologically Induced Hypothermic Responses

doi: 10.1523/JNEUROSCI.1699-05.2005

Figure Lengend Snippet: Girk2 gene dosage has a significant effect on the amount of hypothermia induced by 8-OH-DPAT. A, Mean ± SEM core body temperature drops elicited by injections of 8-OH-DPAT in wild-type mice (+/+; open squares), heterozygous Girk2 KO mice (-/+; shaded squares), and homozygous Girk2 KO mice (-/-; filled triangles) (n = 12 for each group of animals) plotted against time. Time = 0, the time immediately before drug injection. B, Dose-response relationships for the maximal drops ± SEM in core temperature produced by 8-OH-DPAT. Curves represent best fit variable slope sigmoidal dose-response curves. *p < 0.05; **p < 0.01; ***p < 0.001.

Article Snippet: Finally, we should note that, because the GIRK2 gene is located in human chromosome 21 (and its mouse ortholog, Girk2 , is in mouse chromosome 16) (The Jackson Laboratory, 2005 ), our findings might be relevant to the study of neurotransmission and thermoregulation in persons with Down syndrome and aneuploid mouse models for this genetic disorder (for review, see Patterson and Costa, 2005 ).

Techniques: Injection, Produced

Journal: The Journal of Neuroscience

Article Title: G-Protein-Gated Potassium (GIRK) Channels Containing the GIRK2 Subunit Are Control Hubs for Pharmacologically Induced Hypothermic Responses

doi: 10.1523/JNEUROSCI.1699-05.2005

Figure Lengend Snippet: The Girk2 gene plays a significant role in the hypothermia induced by various GPCR agonists. A, Maximal core temperature drops ± SEM elicited by 8-OH-DPAT, baclofen, oxotremorine, R-PIA, loperamide, and ethanol in wild-type mice (+/+; open bars) and homozygous Girk2 KO mice (-/-; filled bars) (n = 12 for each group of animals). B, Schematic model showing GIRK2-containing channels functioning as “molecular control hubs” for the hypothermia-inducing action of many GPCRs. *p < 0.05; **p < 0.01.

Article Snippet: Finally, we should note that, because the GIRK2 gene is located in human chromosome 21 (and its mouse ortholog, Girk2 , is in mouse chromosome 16) (The Jackson Laboratory, 2005 ), our findings might be relevant to the study of neurotransmission and thermoregulation in persons with Down syndrome and aneuploid mouse models for this genetic disorder (for review, see Patterson and Costa, 2005 ).

Techniques: Control