Six-week-old female C3H/HeN mice and AKR/N mice were purchased from Japan SLC. Animals were maintained under specific pathogen–free conditions. All the experiments were conducted according to the guidelines for the Animal Research Committee of the Research Institute for Microbial Diseases, Osaka University.
C3h hen mice
Manufactured by Japan SLC
Sourced in Japan
C3H/HeN mice are an inbred strain of laboratory mice that are widely used in biomedical research. They are characterized by their susceptibility to various diseases and conditions, making them a valuable model for studying a range of health-related topics. The C3H/HeN strain is known for its specific genetic and physiological characteristics, which can provide insights into the underlying mechanisms of various diseases and conditions.
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6 protocols using c3h hen mice
1
Pathogen-free Murine Experiments
2
Detailed Protocol for Mouse Experiments
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A total of six ddY mice (obtained from Tokyo Laboratory Animals Science Co., Ltd., Tokyo,
Japan), twelve ICR mice (obtained from Japan SLC, Hamamatsu, Japan), and sixty C3H/HeN
mice (Japan SLC) were used in the present study. Thirty 6-week-old female C3H/HeN mice
were used in both Experiments 1 and 2. In Experiment 3, six retired female ddY mice and
three 6-week-old female ICR mice were used. In Experiment 4, nine 6-week-old female ICR
mice were used.
All mice were sorted into groups of 3 to 6 and acclimatized for several days in a
conventional environment. Tap water in a bottle and solid feed (Lab Diet 5001, Japan SLC)
were supplied without restriction. At the termination of the experiments, the mice were
euthanized by exsanguination after deep anesthesia with 4% isoflurane (DS Pharma Animal
Health Co., Ltd., Osaka, Japan). The research described herein was approved (31-45,
R02-76, R02-77) by the Animal Care and Use Committee at Tokyo University of Agriculture
and Technology (TUAT) and the research was performed according to the guidelines for
animal experiments at TUAT.
Japan), twelve ICR mice (obtained from Japan SLC, Hamamatsu, Japan), and sixty C3H/HeN
mice (Japan SLC) were used in the present study. Thirty 6-week-old female C3H/HeN mice
were used in both Experiments 1 and 2. In Experiment 3, six retired female ddY mice and
three 6-week-old female ICR mice were used. In Experiment 4, nine 6-week-old female ICR
mice were used.
All mice were sorted into groups of 3 to 6 and acclimatized for several days in a
conventional environment. Tap water in a bottle and solid feed (Lab Diet 5001, Japan SLC)
were supplied without restriction. At the termination of the experiments, the mice were
euthanized by exsanguination after deep anesthesia with 4% isoflurane (DS Pharma Animal
Health Co., Ltd., Osaka, Japan). The research described herein was approved (31-45,
R02-76, R02-77) by the Animal Care and Use Committee at Tokyo University of Agriculture
and Technology (TUAT) and the research was performed according to the guidelines for
animal experiments at TUAT.
3
Murine Bone Marrow Macrophage Differentiation
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Femoral and tibial bone marrows of 12‐ to 16‐week‐old nondiabetic C3H/HeN mice (Japan SLC Inc.) were collected in 1% FCS‐supplemented low‐glucose DMEM, and erythrocytes were lysed in RBC lysis buffer. Bone marrow cells were washed in 1% FCS DMEM and cultured 5–7 days in 10% FCS DMEM supplemented with 40 ng/ml recombinant human macrophage colony‐stimulating factor (M‐CSF) (BioLegend). Non‐adhesive cells were washed out with PBS. Differentiated macrophages were collected by scraping in ice‐cold PBS with 1% FCS/ 5 mM EDTA, washed, counted, diluted to 2.0 × 105 cells/ml in 1% FCS DMEM, and cultured in 96‐well plates overnight at 37°C under 5% CO2 atmosphere.
4
Prenatal Arsenic Exposure and Offspring Anxiety
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Pregnant C3H/HeN mice (F0, total n = 14) were purchased from JAPAN SLC (Shizuoka, Japan) and these mice (n = 7) were given arsenic exposure (sodium arsenite, NaAsO2, 85 ppm (85 mg/L)) in the drinking water starting from the gestational day (GD) 8 to 18. Control mice (n = 7) were given tap water only. The experimental dose for gestational arsenic exposure was explained in our previous studies [33 (link), 34 (link), 38 ]. The experimental design of this study was illustrated in the Fig. 1 . F1 female offspring (n = 12 per each group) delivered from arsenic exposed F0 or control F0 were randomly collected for the experiments. At the age of 74-week, the control female mice (n = 7) and arsenic-exposed female mice (n = 7) were used for anxiety-related behavioral test and molecular analyses. Then, the control F1 female mice (n = 5) and arsenite F1 female mice (n = 5) were used for immunohistochemical analyses. This study was approved by the Ethics Committee of the Animal Care and Experimentation Council of the National Institute for Environmental Studies (NIES), Japan (Ethics approval code number: AE-22-12).
5
Chronic Stress-Induced Wheel-Running Behavior
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Five-week-old male C3H/HeN mice (Japan SLC Inc., Hamamatsu, Japan) were individually maintained in plastic cages containing paper chip bedding and running wheels (SW-15; Melquest Y.K, Toyama, Japan) were provided free access to a standard diet (AIN-93G; Oriental Yeast, Tokyo) under a 12-h light/dark cycle (LD 12:12; lights on at Zeitgeber time [ZT] 0). A white fluorescent lamp placed at cage level provided light (350 lx) for 2 weeks until daily wheel-running activity reached a plateau (Miyazaki et al., Citation2013) .
The mice were assigned to the following groups (n = 5 per group): control (not stressed) at ZT0 and ZT12, and CSD at ZT0 and ZT12. The CSD mice were created by exposing them to psychophysiological stress for 1 week as described (Miyazaki et al., Citation2013) . Briefly, we replaced paper-chip bedding in the cages with 1.5 cm of water, which caused the mice to remain on wheels 24 h per day for 7 days. The paradigm of CSD model shows minimal effect on body weight gain in mice (Oishi et al., Citation2014) . Wheel-running activity was continuously recorded at 1-min intervals using a Chronobiology Kit ® (Stanford Software Systems, Stanford, CA, USA). The Animal Care and Use Committee at the National Institute of Advanced Industrial Science and Technology (AIST) approved all animal experiments (Permission #2021-0338).
The mice were assigned to the following groups (n = 5 per group): control (not stressed) at ZT0 and ZT12, and CSD at ZT0 and ZT12. The CSD mice were created by exposing them to psychophysiological stress for 1 week as described (Miyazaki et al., Citation2013) . Briefly, we replaced paper-chip bedding in the cages with 1.5 cm of water, which caused the mice to remain on wheels 24 h per day for 7 days. The paradigm of CSD model shows minimal effect on body weight gain in mice (Oishi et al., Citation2014) . Wheel-running activity was continuously recorded at 1-min intervals using a Chronobiology Kit ® (Stanford Software Systems, Stanford, CA, USA). The Animal Care and Use Committee at the National Institute of Advanced Industrial Science and Technology (AIST) approved all animal experiments (Permission #2021-0338).
6
Chronic Stress-Induced Wheel-Running Behavior
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Five-week-old male C3H/HeN mice (Japan SLC Inc., Hamamatsu, Japan) were individually housed in plastic cages containing paper-chip bedding and running wheels (SW-15; Melquest Y.K, Toyama, Japan). The mice had free access to a standard diet (AIN-93G: Oriental Yeast Co. Ltd., Tokyo, Japan) and tap water under a 12 h light-12 h dark cycle (lights on at 08:00) for four weeks until daily wheel-running activity reached a plateau. The CSD mice were then exposed to psychophysiological stress for one week to induce CSD23 (link). Briefly, paper-chip bedding was replaced with water to a depth of 1.5 cm, which caused the mice to remain on the wheels throughout every day of the study. Wheel-running activity was continuously recorded at 1-min intervals using Chronobiology Kits (Stanford Software Systems, Stanford, CA, USA) and activity data are displayed as actograms. All protocols complied with the guidelines for animal experiments published by the National Institute of Advanced Industrial Science and Technology (AIST) and the ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines. The AIST Animal Care and Use Committee approved all experimental protocols (Permission No: #2021-338).
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