Aerobic Exercise Promotes Hippocampal Neurogenesis and Ameliorates Cognitive Dysfunction Induced by Unilateral Labyrinthectomy methods
Aim. Evidence-backed execution summary for Aerobic Exercise Promotes Hippocampal Neurogenesis and Ameliorates Cognitive Dysfunction Induced by Unilateral Labyrinthectomy methods from Aerobic Exercise Promotes Hippocampal Neurogenesis and Ameliorates Cognitive Dysfunction Induced by Unilateral Labyrinthectomy.
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mouse
Subject model for the experiment.
- Use
- confirm full cohort details in the source paper
Contextual and Tone-Cued Fear Conditioning
reagent used in the protocol.
- Use
- The shock-associated training context (Context A) and the analogous non-shock context (Context B) had the same exposed stainless-steel grid floor and ceiling. Context B differed from Context A only in that it included four plastic wall inserts. To maintain cleanliness between trials, a non-al...
Immunofluorescence Staining
reagent used in the protocol.
- Use
- Mice were anesthetized using 1% pentobarbital sodium (35 mg/kg, i.p.) and perfused transcardially with 0.1 M PBS, followed by 4% paraformaldehyde in 0.1 M PBS. The brains were then removed and fixed in 4% paraformaldehyde at 4°C overnight. After dehydration in 30% sucrose, brains were frozen i...
RNA Extraction and Sequencing
reagent used in the protocol.
- Use
- Total RNA was extracted from the DG using the TRIzol reagent protocol (Life Technologies, California, USA). RNA concentration and purity were assessed using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Wilmington, DE), and RNA integrity was assessed using the RNA Nano 6000 Assay Kit on the Agilent Bi...
Quantitative Real-Time PCR (qRT-PCR)
reagent used in the protocol.
- Use
- DG tissue samples were isolated on ice and immediately stored at -80°C. Total RNA was extracted using RNAiso Plus (code 9108, TaKaRa). RNA concentration and purity were assessed using NanoDrop2000C spectrophotometry (Thermo Scientific). One microgram of RNA was reverse transcribed to complementary DNA (cD...
Quantitative Real-Time PCR (qRT-PCR)
reagent used in the protocol.
- Use
- Primer sequences of genes.
The JAK1/JAK2 Inhibitor AZD1480 Attenuates Neuroinflammation
reagent used in the protocol.
- Use
- At 2 weeks after UL induction, mice received daily oral gavage of either AZD1480 (HY-10193, MCE) at 25 mg/kg or a vehicle (VH) control consisting of 0.1% dimethyl sulfoxide (DMSO; HY-Y0320, MCE) [ ]. Depending on the experimental endpoint, mice were either used for behavioral assays or sacrif...
LPS Induces Neuroinflammatory Responses
reagent used in the protocol.
- Use
- Two weeks after UL and running, mice were intraperitoneally injected daily with 0.25 mg/kg lipopolysaccharide (LPS) (L2880, Sigma) or saline as a vehicle control [ ]. Based on analytical requirements, mice were either subjected to behavioral tests or euthanized 2 weeks after administration. Upon sacrific...
Inhibition of DG Neurogenesis in Mice at 28 Days After UL
reagent used in the protocol.
- Use
- In alignment with the observed cognitive deficits, UL mice exhibited a significant reduction in neural stem cell proliferation within the DG, as demonstrated by quantitative analysis of BrdU incorporation (Figure ), as well as Ki67- and DCX-positive cells (Figure ). To fully characterize the...
Open-Field Test ( OFT )
Mice were placed at the center of a rectangular chamber (45 × 45 × 45 cm), and their movement was monitored for 5 min using an automated video tracking system (Supermaze 2.0, Softmaze, China). To mitigate the impact of olfactory cues, the apparatus was cleaned with 75% a...
- Use
- Mice were placed at the center of a rectangular chamber (45 × 45 × 45 cm), and their movement was monitored for 5 min using an automated video tracking system (Supermaze 2.0, Softmaze, China). To mitigate the impact of olfactory cues, the apparatus was cleaned with 75% a...
Methods
Behavioral experiments were conducted to assess the cognitive impairment induced by unilateral labyrinthectomy (UL). Immunofluorescence staining was performed to assess neural stem cell proliferation and newborn neuron maturation in the hippocampal dentate gyrus. Aerobic exercise was applied as an intervention to ex...
- Use
- Behavioral experiments were conducted to assess the cognitive impairment induced by unilateral labyrinthectomy (UL). Immunofluorescence staining was performed to assess neural stem cell proliferation and newborn neuron maturation in the hippocampal dentate gyrus. Aerobic exercise was applied as an intervention to ex...
T-Maze
The apparatus consisted of an opaque acrylic T-maze (50 cm long, 10 cm wide, and 25 cm high), with a camera mounted above and connected to a video tracking system (Supermaze 2.0, Softmaze, China). To minimize stress and promote acclimatization, mice were placed in the testing room 24 h...
- Use
- The apparatus consisted of an opaque acrylic T-maze (50 cm long, 10 cm wide, and 25 cm high), with a camera mounted above and connected to a video tracking system (Supermaze 2.0, Softmaze, China). To minimize stress and promote acclimatization, mice were placed in the testing room 24 h...
Radial 8-Arm Maze ( RAM )
The RAM was composed of eight transparent organic plastic arms equally spaced around an octagonal central platform. Before the training sessions, the animals' body weight was maintained at 85% of their free-feeding weight, with water available ad libitum. To facilitate acclimatization to the RAM, all arms...
- Use
- The RAM was composed of eight transparent organic plastic arms equally spaced around an octagonal central platform. Before the training sessions, the animals' body weight was maintained at 85% of their free-feeding weight, with water available ad libitum. To facilitate acclimatization to the RAM, all arms...
Contextual and Tone-Cued Fear Conditioning
The shock-associated training context (Context A) and the analogous non-shock context (Context B) had the same exposed stainless-steel grid floor and ceiling. Context B differed from Context A only in that it included four plastic wall inserts. To maintain cleanliness between trials, a non-al...
- Use
- The shock-associated training context (Context A) and the analogous non-shock context (Context B) had the same exposed stainless-steel grid floor and ceiling. Context B differed from Context A only in that it included four plastic wall inserts. To maintain cleanliness between trials, a non-al...
Immunofluorescence Staining
Mice were anesthetized using 1% pentobarbital sodium (35 mg/kg, i.p.) and perfused transcardially with 0.1 M PBS, followed by 4% paraformaldehyde in 0.1 M PBS. The brains were then removed and fixed in 4% paraformaldehyde at 4°C overnight. After dehydration in 30% sucrose, brains were frozen i...
- Use
- Mice were anesthetized using 1% pentobarbital sodium (35 mg/kg, i.p.) and perfused transcardially with 0.1 M PBS, followed by 4% paraformaldehyde in 0.1 M PBS. The brains were then removed and fixed in 4% paraformaldehyde at 4°C overnight. After dehydration in 30% sucrose, brains were frozen i...
Dissection of Hippocampal Dentate Gyrus
The DG was isolated as described previously [ ]. Briefly, mice were anesthetized with 1% sodium pentobarbital (35 mg/kg, i.p.), and the brain was removed and immediately placed in ice-cold PBS. The brain was sectioned along the longitudinal fissure in a petri dish containing ice-cold PBS, and the r...
- Use
- The DG was isolated as described previously [ ]. Briefly, mice were anesthetized with 1% sodium pentobarbital (35 mg/kg, i.p.), and the brain was removed and immediately placed in ice-cold PBS. The brain was sectioned along the longitudinal fissure in a petri dish containing ice-cold PBS, and the r...
RNA Extraction and Sequencing
Total RNA was extracted from the DG using the TRIzol reagent protocol (Life Technologies, California, USA). RNA concentration and purity were assessed using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Wilmington, DE), and RNA integrity was assessed using the RNA Nano 6000 Assay Kit on the Agilent Bi...
- Use
- Total RNA was extracted from the DG using the TRIzol reagent protocol (Life Technologies, California, USA). RNA concentration and purity were assessed using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Wilmington, DE), and RNA integrity was assessed using the RNA Nano 6000 Assay Kit on the Agilent Bi...
Statistical Analysis
Software used for acquisition, scoring, statistics, or reporting.
- Use
- All statistical analyses were performed using GraphPad Prism 8.0 (GraphPad Software, USA). The normality of data distribution was assessed using the Shapiro-Wilk test. Data conforming to a normal distribution were analyzed using the two-tailed Student's T -test, one-way ANOVA followed by Tuke...
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Behavioral Analysis
Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by handling for a minimum of 5 min, twice daily, for three consecutive days. No significant differences in body weight or whisker number were observed between groups.
Materials and Methods
Experimental procedures involving mice were approved by the Animal Welfare Committee of Huazhong University of Science and Technology (No. 3971). All experiments were conducted in strict accordance with the committee guidelines. C57BL/6J mice (8 weeks old) were obtained from the Experimental Animals Center of Tongji Medical College, Huazhong University of Science and Technology. To minimize variability associated with cyclical hormonal fluctuations in females, only male mice were used in this study. Animals were group-housed (≤ 5 per cage) under controlled environmental conditions (temperature: 22°C-24°C; humidity: 55%-80%) with a 12-h light/dark cycle and provided ad libitum access to food and water.
Unilateral Labyrinthectomy
Male mice (22-25 g) were randomly allocated to the UL or sham surgery groups. In the UL group, mice were anesthetized with 1% sodium pentobarbital (35 mg/kg, i.p.). A postauricular incision was made to expose the middle ear structures. The tympanic membrane was then punctured using fine forceps, and the malleus, incus, and stapes were carefully removed. The stapedial artery was then cauterized using an electrocautery device, and the vestibular window was widened with a probe to allow deep puncture of the labyrinth, ensuring complete destruction. Following this, a small amount of anhydrous ethanol was injected to further damage the labyrinth, and a gelatin sponge was placed in the vestibular window. The incision was then sutured closed. In the sham surgery group, the tympanic membrane was perforated, and the ossicles were removed, but the vestibular structures and lab...
Aerobic Exercise
During the first week, mice in the aerobic exercise groups performed adaptive running training in individual treadmill lanes (FT-200, Taimeng, China) at a speed of 5 m/min for 45 min per session. Over the following 3 weeks, the running protocol was adjusted to 5 m/min for 10 min, 8 m/min for 30 min, and 5 m/min for 10 min per session to minimize stress-induced inhibition of hippocampal neurogenesis [ ]. Mice in the static groups were placed on the stationary treadmill for the same duration.
BrdU Injection
Mice were administered 5′-Bromo-2′-Deoxyuridine (BrdU; B5002, Sigma) in saline via intraperitoneal injection at a dose of 100 mg/kg, once daily for five consecutive days. To avoid labeling the acute burst of progenitor proliferation induced by surgery or stimulation, we administered BrdU during the late phase of the intervention to more accurately capture the cell populations most relevant to exercise-responsive neurogenesis. Animals were sacrificed 24 h after the final BrdU injection.
Immunofluorescence Staining
Mice were anesthetized using 1% pentobarbital sodium (35 mg/kg, i.p.) and perfused transcardially with 0.1 M PBS, followed by 4% paraformaldehyde in 0.1 M PBS. The brains were then removed and fixed in 4% paraformaldehyde at 4°C overnight. After dehydration in 30% sucrose, brains were frozen in OCT medium (TissueTek, Sakura, Japan) and sliced into 30 µm coronal sections using a cryostat (HM550, Thermo Scientific). After three rinses with PBS, brain sections were rinsed in blocking buffer (PBS with 0.3% Triton X-100 containing 10% goat serum and 3% BSA) for 60 min at room temperature. Sections were incubated overnight at 4°C in blocking buffer containing rat anti-BrdU antibody (ab6326, Abcam; 1:300 dilution), rabbit anti-Ki67 antibody (ab15580, Abcam; 1:500 dilution), and rabbit anti-DCX antibody (13925-1-...
Inhibition of DG Neurogenesis in Mice at 28 Days After UL
In alignment with the observed cognitive deficits, UL mice exhibited a significant reduction in neural stem cell proliferation within the DG, as demonstrated by quantitative analysis of BrdU incorporation (Figure ), as well as Ki67- and DCX-positive cells (Figure ). To fully characterize the maturation process of neural stem cells in the DG, we used a retroviral labelling assay in which newborn neurons were specifically labeled with pROV-EF1a-EGFP by microinjection into the DG immediately after UL surgery in mice, and the maturation of neural stem cells was assessed 28 days later. Quantitative morphological analysis using the Sholl method revealed a significant reduction in total dendritic branching complexity compared to controls (Figure ). These findings indicate that UL leads to reduced proliferation and maturation of NSCs in the DG,...
Aerobic Exercise Improves Cognitive Function in Mice After UL
Cognitive dysfunction in mice after UL was improved by aerobic exercise (A) Schematic diagram of the experimental design of an exercise intervention in UL mice. (B, C) In the open field test, there were no significant differences in total movement distance and average speed among the four groups of mice: Sham + Static, Sham + UL, Run + Static, and Run + UL (two-way ANOVA: F (1,36) = 0.04837, p = 0.8272 and F (1,36) = 0.0009785, p = 0.9752). (D, E) There were no significant intergroup differences in terms of total immobility time and the duration spent in the central zone of the open field for the Sham + Static, Sham + UL, Run + Static, and Run + UL groups (two-way ANOVA: F (1,36) = 3.000, p = 0.0918 and F (1,36) = 0.2262, p = 0.6372, n = 10/group in OFT). (F, G) Aerobic exercise amelio...
Measurement outputs
What raw and processed outputs should exist?
Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by h...
- Raw artifact
- Field or section images captured from matched samples
- Processed artifact
- Selected representative panels with quantified intensity, counts, or area measurements
- Reported as
- Per-group imaging summaries with representative figures and quantified endpoints
Behavioral experiments were conducted to assess the cognitive impairment induced by unilateral labyrinthectomy (UL). Immunofluorescence staining was performed to assess neural s...
- Raw artifact
- Field or section images captured from matched samples
- Processed artifact
- Selected representative panels with quantified intensity, counts, or area measurements
- Reported as
- Per-group imaging summaries with representative figures and quantified endpoints
QRT-PCR validation of gene expression changes related to cognitive function. (A-E, G, H, K, L) The Lyz2 (Ordinary one-way ANOVA, F (2,27) = 102.0, p R...
- Raw artifact
- Field or section images captured from matched samples
- Processed artifact
- Selected representative panels with quantified intensity, counts, or area measurements
- Reported as
- Per-group imaging summaries with representative figures and quantified endpoints
The shock-associated training context (Context A) and the analogous non-shock context (Context B) had the same exposed stainless-steel grid floor and ceiling....
- Raw artifact
- Field or section images captured from matched samples
- Processed artifact
- Selected representative panels with quantified intensity, counts, or area measurements
- Reported as
- Per-group imaging summaries with representative figures and quantified endpoints
Analysis plan
How should the outputs become interpretable results?
Acquisition
Capture matched images from the relevant tissue region using the same acquisition settings across samples.
inferred from protocolPreprocessing / cleaning
Behavioral analysis was conducted by investigators blinded to the group assignments.
from paperScoring or quantification
Quantify the primary readouts for this experiment: Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by h...; Behavioral experiments were conducted to assess the cognitive impairment induced by unilateral labyrinthectomy (UL). Immunofluorescence staining was performed to assess neural s...; QRT-PCR validation of gene expression changes related to cognitive function. (A-E, G, H, K, L) The Lyz2 (Ordinary one-way ANOVA, F (2,27) = 102.0, p R...; The shock-associated training context (Context A) and the analogous non-shock context (Context B) had the same exposed stainless-steel grid floor and ceiling.....
from paperNormalization
Normalize image-derived measurements against the matched acquisition or segmentation rules before comparing groups.
inferred from protocolStatistical comparison
Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by h...; qRT-PCR validation of gene expression changes related to cognitive function. (A-E, G, H, K, L) The Lyz2 (Ordinary one-way ANOVA, F (2,27) = 102.0, p R...; All statistical analyses were performed using GraphPad Prism 8.0 (GraphPad Software, USA). The normality of data distribution was assessed using the Shapiro-Wilk test. Dat...; In alignment with the observed cognitive deficits, UL mice exhibited a significant reduction in neural stem cell proliferation within the DG, as demonstrated by quantitative ana...
from paperReporting output
Report representative outputs alongside summary comparisons for Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by h..., Behavioral experiments were conducted to assess the cognitive impairment induced by unilateral labyrinthectomy (UL). Immunofluorescence staining was performed to assess neural s..., QRT-PCR validation of gene expression changes related to cognitive function. (A-E, G, H, K, L) The Lyz2 (Ordinary one-way ANOVA, F (2,27) = 102.0, p R..., The shock-associated training context (Context A) and the analogous non-shock context (Context B) had the same exposed stainless-steel grid floor and ceiling.....
inferred from protocolStructured statistical methods
Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by h...; qRT-PCR validation of gene expression changes related to cognitive function. (A-E, G, H, K, L) The Lyz2 (Ordinary one-way ANOVA, F (2,27) = 102.0, p R...; All statistical analyses were performed using GraphPad Prism 8.0 (GraphPad Software, USA). The normality of data distribution was assessed using the Shapiro-Wilk test. Dat...; In alignment with the observed cognitive deficits, UL mice exhibited a significant reduction in neural stem cell proliferation within the DG, as demonstrated by quantitative ana...
source structuredSource and audit
What supports the facts on this page?
Evidence quotes (8)
Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by handling for a minimum of 5 min, twice daily, for three consecutive days. No significant differences in body weight or whisker number were observed between groups.
Experimental procedures involving mice were approved by the Animal Welfare Committee of Huazhong University of Science and Technology (No. 3971). All experiments were conducted in strict accordance with the committee guidelines. C57BL/6J mice (8 weeks old) were obtained from the Experimental Animals Center of Tongji Medical College, Huazhong University of Science and Technology. To minimize variability associated with cyclical hormonal fluctuations in females, only male mice were used in this study. Animals were group-housed (≤ 5 per cage) under controlled environmental conditions (temperature: 22°C-24°C; humidity: 55%-80%) with a 12-h light/dark cycle and provided ad libitum access to food and water.
Male mice (22-25 g) were randomly allocated to the UL or sham surgery groups. In the UL group, mice were anesthetized with 1% sodium pentobarbital (35 mg/kg, i.p.). A postauricular incision was made to expose the middle ear structures. The tympanic membrane was then punctured using fine forceps, and the malleus, incus, and stapes were carefully removed. The stapedial artery was then cauterized using an electrocautery device, and the vestibular window was widened with a probe to allow deep puncture of the labyrinth, ensuring complete destruction. Following this, a small amount of anhydrous ethanol was injected to further damage the labyrinth, and a gelatin sponge was placed in the vestibular window. The incision was then sutured closed. In the sham surgery group, the tympanic membrane was perforated, and the ossicles were removed, but the vestibular structures and labyrinth remained intact.
During the first week, mice in the aerobic exercise groups performed adaptive running training in individual treadmill lanes (FT-200, Taimeng, China) at a speed of 5 m/min for 45 min per session. Over the following 3 weeks, the running protocol was adjusted to 5 m/min for 10 min, 8 m/min for 30 min, and 5 m/min for 10 min per session to minimize stress-induced inhibition of hippocampal neurogenesis [ ]. Mice in the static groups were placed on the stationary treadmill for the same duration.
Mice were administered 5′-Bromo-2′-Deoxyuridine (BrdU; B5002, Sigma) in saline via intraperitoneal injection at a dose of 100 mg/kg, once daily for five consecutive days. To avoid labeling the acute burst of progenitor proliferation induced by surgery or stimulation, we administered BrdU during the late phase of the intervention to more accurately capture the cell populations most relevant to exercise-responsive neurogenesis. Animals were sacrificed 24 h after the final BrdU injection.
Mice were anesthetized using 1% pentobarbital sodium (35 mg/kg, i.p.) and perfused transcardially with 0.1 M PBS, followed by 4% paraformaldehyde in 0.1 M PBS. The brains were then removed and fixed in 4% paraformaldehyde at 4°C overnight. After dehydration in 30% sucrose, brains were frozen in OCT medium (TissueTek, Sakura, Japan) and sliced into 30 µm coronal sections using a cryostat (HM550, Thermo Scientific). After three rinses with PBS, brain sections were rinsed in blocking buffer (PBS with 0.3% Triton X-100 containing 10% goat serum and 3% BSA) for 60 min at room temperature. Sections were incubated overnight at 4°C in blocking buffer containing rat anti-BrdU antibody (ab6326, Abcam; 1:300 dilution), rabbit anti-Ki67 antibody (ab15580, Abcam; 1:500 dilution), and rabbit anti-DCX antibody (13925-1-AP, Proteintech; 1:300 dilution). After three washes with PBST (PBS containing 0.05% Tween-20), sections were incubated with the appropriate fluorophore-conjugated secondary antibody (goat anti-rabbit IgG conjugated to Alexa Fluor 488, AS039, ABclonal; 1:300 dilution) in blocking b...
In alignment with the observed cognitive deficits, UL mice exhibited a significant reduction in neural stem cell proliferation within the DG, as demonstrated by quantitative analysis of BrdU incorporation (Figure ), as well as Ki67- and DCX-positive cells (Figure ). To fully characterize the maturation process of neural stem cells in the DG, we used a retroviral labelling assay in which newborn neurons were specifically labeled with pROV-EF1a-EGFP by microinjection into the DG immediately after UL surgery in mice, and the maturation of neural stem cells was assessed 28 days later. Quantitative morphological analysis using the Sholl method revealed a significant reduction in total dendritic branching complexity compared to controls (Figure ). These findings indicate that UL leads to reduced proliferation and maturation of NSCs in the DG, suggesting a potential causal relationship between impaired neurogenic processes and cognitive dysfunction.
Cognitive dysfunction in mice after UL was improved by aerobic exercise (A) Schematic diagram of the experimental design of an exercise intervention in UL mice. (B, C) In the open field test, there were no significant differences in total movement distance and average speed among the four groups of mice: Sham + Static, Sham + UL, Run + Static, and Run + UL (two-way ANOVA: F (1,36) = 0.04837, p = 0.8272 and F (1,36) = 0.0009785, p = 0.9752). (D, E) There were no significant intergroup differences in terms of total immobility time and the duration spent in the central zone of the open field for the Sham + Static, Sham + UL, Run + Static, and Run + UL groups (two-way ANOVA: F (1,36) = 3.000, p = 0.0918 and F (1,36) = 0.2262, p = 0.6372, n = 10/group in OFT). (F, G) Aerobic exercise ameliorated the UL-induced working memory impairment, manifested by the total entry to and time spent in the novel arm. Right arm (F) was set as the novel arm (two-way ANOVA: F (1,36) = 1.616, p = 0.2118, Tukey's multiple comparisons test: Static + Sham vs. Static + UL: p &#...
Machine-readable layer
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"description": "Evidence-backed execution summary for Aerobic Exercise Promotes Hippocampal Neurogenesis and Ameliorates Cognitive Dysfunction Induced by Unilateral Labyrinthectomy methods from Aerobic Exercise Promotes Hippocampal Neurogenesis and Ameliorates Cognitive Dysfunction Induced by Unilateral Labyrinthectomy.",
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"name": "Behavioral Analysis",
"text": "Behavioral analysis was conducted by investigators blinded to the group assignments. All tests were conducted during the light phase. Prior to testing, mice were acclimated by handling for a minimum of 5 min, twice daily, for three consecutive days. No significant differences in body weight or whisker number were observed between groups."
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"text": "Experimental procedures involving mice were approved by the Animal Welfare Committee of Huazhong University of Science and Technology (No. 3971). All experiments were conducted in strict accordance with the committee guidelines. C57BL/6J mice (8 weeks old) were obtained from the Experimental Animals Center of Tongji Medical College, Huazhong University of Science and Technology. To minimize variability associated with cyclical hormonal fluctuations in females, only male mice were used in this study. Animals were group-housed (≤ 5 per cage) under controlled environmental conditions (temperature: 22°C-24°C; humidity: 55%-80%) with a 12-h light/dark cycle and provided ad libitum access to food and water."
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"name": "Unilateral Labyrinthectomy",
"text": "Male mice (22-25 g) were randomly allocated to the UL or sham surgery groups. In the UL group, mice were anesthetized with 1% sodium pentobarbital (35 mg/kg, i.p.). A postauricular incision was made to expose the middle ear structures. The tympanic membrane was then punctured using fine forceps, and the malleus, incus, and stapes were carefully removed. The stapedial artery was then cauterized using an electrocautery device, and the vestibular window was widened with a probe to allow deep puncture of the labyrinth, ensuring complete destruction. Following this, a small amount of anhydrous ethanol was injected to further damage the labyrinth, and a gelatin sponge was placed in the vestibular window. The incision was then sutured closed. In the sham surgery group, the tympanic membrane was perforated, and the ossicles were removed, but the vestibular structures and lab..."
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"name": "Immunofluorescence Staining",
"text": "Mice were anesthetized using 1% pentobarbital sodium (35 mg/kg, i.p.) and perfused transcardially with 0.1 M PBS, followed by 4% paraformaldehyde in 0.1 M PBS. The brains were then removed and fixed in 4% paraformaldehyde at 4°C overnight. After dehydration in 30% sucrose, brains were frozen in OCT medium (TissueTek, Sakura, Japan) and sliced into 30 µm coronal sections using a cryostat (HM550, Thermo Scientific). After three rinses with PBS, brain sections were rinsed in blocking buffer (PBS with 0.3% Triton X-100 containing 10% goat serum and 3% BSA) for 60 min at room temperature. Sections were incubated overnight at 4°C in blocking buffer containing rat anti-BrdU antibody (ab6326, Abcam; 1:300 dilution), rabbit anti-Ki67 antibody (ab15580, Abcam; 1:500 dilution), and rabbit anti-DCX antibody (13925-1-..."
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"@type": "HowToStep",
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"name": "Inhibition of DG Neurogenesis in Mice at 28 Days After UL",
"text": "In alignment with the observed cognitive deficits, UL mice exhibited a significant reduction in neural stem cell proliferation within the DG, as demonstrated by quantitative analysis of BrdU incorporation (Figure ), as well as Ki67- and DCX-positive cells (Figure ). To fully characterize the maturation process of neural stem cells in the DG, we used a retroviral labelling assay in which newborn neurons were specifically labeled with pROV-EF1a-EGFP by microinjection into the DG immediately after UL surgery in mice, and the maturation of neural stem cells was assessed 28 days later. Quantitative morphological analysis using the Sholl method revealed a significant reduction in total dendritic branching complexity compared to controls (Figure ). These findings indicate that UL leads to reduced proliferation and maturation of NSCs in the DG,..."
},
{
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"name": "Aerobic Exercise Improves Cognitive Function in Mice After UL",
"text": "Cognitive dysfunction in mice after UL was improved by aerobic exercise (A) Schematic diagram of the experimental design of an exercise intervention in UL mice. (B, C) In the open field test, there were no significant differences in total movement distance and average speed among the four groups of mice: Sham + Static, Sham + UL, Run + Static, and Run + UL (two-way ANOVA: F (1,36) = 0.04837, p = 0.8272 and F (1,36) = 0.0009785, p = 0.9752). (D, E) There were no significant intergroup differences in terms of total immobility time and the duration spent in the central zone of the open field for the Sham + Static, Sham + UL, Run + Static, and Run + UL groups (two-way ANOVA: F (1,36) = 3.000, p = 0.0918 and F (1,36) = 0.2262, p = 0.6372, n = 10/group in OFT). (F, G) Aerobic exercise amelio..."
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