Dental noise exposed mice display depressive-like phenotypes
© Dong et al. 2016
Received: 25 March 2016
Accepted: 21 April 2016
Published: 10 May 2016
Studies have indicated that depressive disorders are observed frequently in dentists. It’s suggested that dentists encounter numerous sources of stress in their professional career. We noticed that the noises in dental environments are very unpleasant. The animal modeling studies suggested that stressful noise could produce depressive-like phenotypes in rodent animals. We hypothesize that the dental noise may be one of the primary stressors causing depressive disorders in dentists.
We treated C57BL/6 mice with programmatically played wide-spectrum dental noise for 8 h/day at 75 ± 10 dB SPL level for 30 days, and then tested the behaviors. After exposure to dental noise, animals displayed the depressive-like phenotypes, accompanied by inhibition of neurogenesis in hippocampus. These deficits were ameliorated by orally administered with antidepressant fluoxetine.
Our results suggested that dental noise could be one of the primary stressors for the pathogenesis of depressive disorders and the dental noise mouse model could be used in further depression studies.
KeywordsDental noise Depression Sucrose preference Forced swimming Fluoxetine Neurogenesis Weight
Studies have indicated that depressive disorders are observed frequently in dentists than other professional groups [1, 2]. It has been considered that dentists encounter numerous sources of stress including feeling physically or emotionally exhausted, headaches or backaches, coping with difficult or uncooperative patients, heavy workload and financial problems  in their professional career. Consequently, mental disorders such as anxiety, depression and even suicide may result from these stresses. The suicide rate among dentists was much higher than that of other occupations according to the death data of 21 states of USA in late 20th century, and the suicide rate of dentists is 4.45–5.43 times more than general working-age population according to different logistic regression analysis methods . All these evidences revealed that dentists suffer from the stressful work.
Dentists are predisposed to a number of occupational hazards such as viral infection, dental materials, radiation, noise and eyestrain . We noticed that noises in dental environments are very unpleasant. Noise pollution has been criticized by mankind for decades of years and was historically regarded as a large public concern . The inner ear hair cell system of rodents was very similar to that of human beings and other mammals . The animal modeling studies suggest that stressful noise might produce neurogenesis impairment in rodent animals [8–10]. Increasing evidences implied that impaired hippocampal neurogenesis is intimately linked with the onset of depression as discussed in the neurogenic hypothesis of depression , so the decrease of dentate gyrus neurogenesis is considered a causal factor for depression disorder .
Wrap these all together, we hypothesize that the dental noise may be one of the primary stressors causing depressive disorders in dentists. So we attempted to establish a dental noise-exposed mice model to test and verify whether dental noise could produce depressive-like phenotypes in mice.
Adult male C57BL/6 mice (10 weeks of age) were from Charles River Laboratories. The mice were housed in standard cages with the cycle of 12 h light to 12 h dark (light from 7:00 am to 7:00 pm), with 40–50 % relative humidity and temperature 24 ± 2 °C. All animal experiments were approved by Institutional Animal Care and Use Committee (IACUC) of Shanghai Jiao Tong University.
The dental noise
These two types of dental noise from Shanghai Elli Dental Clinic in this study are available in the Additional file 1.
Dental noise mouse model and treatment
Mice were exposed to noise from a dental clinic (Fig. 1a). Dental noise (75 ± 10 dB SPL) lower than the reversible threshold of tissue injury  was played 8 h/d (from 9:00 am to 5:00 pm) for 30 days with a program mode of 1 min looped noise with random intervals from 1 to 60 s (Fig. 1b). The mice were administered orally with 120 mg/L fluoxetine hydrochloride (Flu, Jinhchem, Shanghai, China) in water (Noise/Flu) or water only (Noise/Veh) during 30 days. Fluoxetine should be available and renewed every day to ensure efficacy. Mice with (Con/Flu) or without (Con/Veh) fluoxetine hydrochloride were used as control groups absent from the exposure to dental noise.
Sucrose preference test
Sucrose preference task is commonly used for evaluating the depressive-like phenotypes in animals. Sucrose preference is defined as the ratio of the consumption of sucrose solution and the consumption of both pure water and sucrose solution. All the mice underwent the Sucrose Preference test before and after the noise exposure stage.
Open field test
The Open field test was utilized to examine locomotor activity and anxious behavior. Every mouse was placed in a square plexiglass box (27.5 cm L× 27.5 cm W × 18 cm H) and was allowed to explore the arena freely for 20 min. The total distance of movement was recorded by software of Med Associates inc.
Elevated plus-maze test
The apparatus consisted of four arms (29 cm L × 8 cm W) at 90° angles to each other. At each trial, the mouse was placed in the center with its nose directed toward the closed arm and was allowed to explore the maze freely for 5 min. The entry frequency and stay time in the open arms and closed arms was calculated, respectively.
The apparatus consisted of a 3-chambered plexiglass box (60 cm L × 40 cm W × 50 cm H) divided by plexiglass walls with openings allowing animals to move between chambers. Small cages (8 cm diameter × 10 cm H) were placed in the two outer chambers for snout contact but not fighting between animals. Every mouse was first released in the central chamber and was allowed to freely explore the three chambers for a 10 min habituation period. A male wild-type stimulus mouse about 4–5 weeks old was then placed in one of the small cages and the experimental mouse was allowed to explore the apparatus for an additional 5 min. Time spent in each chamber and in sniffing of each cages was recorded.
Forced swimming test
Forced Swimming test is an effective detection method for depression, and it is widely used for screening anti-depression drugs and testing drug efficacy . Each mouse was individually placed in a plexiglass cylinder (20 cm H × 14 cm diameter) filled with 15 cm depth of water at 24 ± 1 °C for 5 min. Take mice out of water and clean them up after test. Record the latency to the first bout of immobility and calculate the total duration of immobility of the last 3 min. It could be judged immobility when all active movements stopped for over 2 s such as struggling and swimming, only floating or making minimal movements .
Prepulse inhibition test
Contextual fear conditioning test
Contextual Fear Conditioning test was performed in test boxes from Med Associates inc. This test consisted of a training phase and a testing phase. During the training phase, mice were individually placed in test boxes and were allowed to explore for 5 min with three 0.75 mA electric foot shock delivered from the floor of each box. Mice were then returned to their home cages. Twenty-four hours after training, mice were placed back to test boxes for another 5 min and the freezing time was recorded. Freezing was defined as the absence of all movement except for respiration.
BrdU assay for neurogenesis
Mice were injected with 5′-Bromo-2-deoxyuridine (BrdU) (10 mg/ml; Sigma) of 100 mg/kg body weight in distilled water every 2 h for three times. Two hours after the last injection, mice were sacrificed and fresh brains were perfused and fixed in 4 % paraformaldehyde (Sigma) at 4 °C overnight. Transfer the brains into 30 % sucrose (Sigma) in phosphate buffered saline (PBS) until sunken. Cryosections at 40 μm for immunofluorescence were obtained with a cryostat CM 3080S (Leica).
After being incubated with blocking solution containing 5 % goat serum (Millipore) and 0.3 % Triton X-100 (Sigma) for 1 h at room temperature, sections were incubated in the primary antibody, rat anti-BrdU (Abcam) diluted 1:500, overnight at 4 °C in a humidified box. The sections were then rinsed in PBS and incubated with the secondary antibody, goat anti-rat IgG (Life Technologies) diluted 1:500, for 2 h at room temperature. At last, nuclei were labeled with fluorescent dye 4′-6-diamidino-2-phenylindole (DAPI) (10 μg/ml; Sigma). The number of BrdU-positive neurons in the granule cell layer (GCL) and the subgranular zone (SGZ) of dentate gyrus was counted under a a Leica confocal microscope.
Statistical processing methods
We used unpaired two-tailed t-test and ANOVAs to compare different groups. Interactions between conditions (noise and control) and treatment (Flu and vehicle) were analyzed by two-way ANOVA followed by appropriate post-hoc tests using StatView Software. P < 0.05 indicates statistical significance between groups (*p < 0.05, **p < 0.01, ***p < 0.001), and all results are presented as mean ± SEM. Graphs were drawn by GraphPad Prism Software.
Dental noise decreased body weight gain, which was ameliorated by antidepressant fluoxetine
Dental noise induced depressive-like phenotypes, which were reversed by fluoxetine
Dental noise decreased hippocampal neurogenesis, which was reversed by fluoxetine
Higher suicide rate and risk of depressive disorders are observed frequently in dentists. Explanations for the phenomenon were widely discussed, which generally focused on occupational stress or sociodemographic factors, such as gender and divorce . However, there was no study about the impact of dental noise per se. Strong noise was commonly believed to disturb normal life, causing mood irritability and even inducing exacerbate psychiatric disorders such as depression . To assay the effect of dental noise on the depression, we established a dental noise exposed depressive-like mouse model. Dental noise exposure decreased the body weight increase rate, impaired the sucrose preference and prolonged the duration of immobility in Forced Swimming test, which were common features of depression. We did not observe the difference in other behavioral tests, such as Elevated Plus-Maze, Open Field test, Sociability Test, Prepulse inhibition test and Fear Conditioning test. It suggested that dental noise had specificity of inducing depressive-like phenotypes.
Fluoxetine, a selective serotonin (5-HT) reuptake inhibitor, was the most commonly prescribed treatment for depression . Fluoxetine increases the concentration of extracellular 5-HT and makes the desensitization of presynaptic 5-HT1A receptors . The depressive-like phenotypes induced by dental noise were evidently ameliorated with fluoxetine in the Sucrose Preference test and the Forced Swimming test. Fluoxetine administration also led to an increase in body weight in our experiment. Fluoxetine was reported to have anorectic effect as a serotonin reuptake inhibitor [23–25], this fluoxetine-induced weight gain argued that the antidepressant effect of fluoxetine overroded the anorectic effect . In this experiment, fluoxetine was administrated from the beginning of dental noise exposure and it reduced the incidence of depressive-like phenotypes, so it was suggested that fluoxetine had protective effects from the impairment induced by dental noise.
Recently, increased studies suggested that adult hippocampal neurogenesis was associated with mental disorders including depression [27, 28], so we counted new-born neurons in dentate gyrus. The number of BrdU-positive neurons was decreased in Noise/Veh group comparing to Con/Veh, which hinted that dental noise negatively affects adult hippocampal proliferation. After fluoxetine administration, proliferation was significantly increased in the Noise/Flu group comparing to Noise/Veh. However, there was no change in proliferation of Con/Flu group comparing with Con/Veh group. Several studies have shown that chronic fluoxetine treatment increased the survival of BrdU-positive cells in wild type animals [29–31]. And studies also showed no significantly increased cells proliferation in mice after chronic fluoxetine treatment [31–33]. The effects of chronic fluoxetine treatment on survival and proliferation during adult hippocampal neurogenesis may need further studies.
There is still the possibility that our findings might not be restricted to the dental noise. Unpleasant noises from crowded traffic or other workplaces may have analogous influence, which could be studied in the future. Furthermore, sleep disturbances were associated with mental disorders . The 8-h noise exposure might also affect the sleep of mice which might contribute to depressive-like phenotypes. It also needs to be investigated in our future studies.
Nevertheless, this is the first reported depressive mouse model induced by dental noise per se. However, further studies need to be performed to reveal the molecular mechanism of how dental noise affects neurogenesis and depression in our model.
In conclusion, our results provide evidence that mice exposed to dental noise exhibit depressive-like phenotypes in behavior tests and neurogenesis. This is the first report demonstrating that dental noise could be one of the primary stressors for the pathogenesis of depressive disorders. The dental noise mouse model could be used in further depression studies.
Ethics approval and consent to participate
All animal experiments were approved by Institutional Animal Care and Use Committee (IACUC) of Shanghai Jiao Tong University.
Consent for publication
Availability of data and materials
The dental noise used in the experiment is available in Microsoft OneDrive. https://onedrive.live.com/redir?resid=3E89C4A9A8D4332F!460&authkey=!AHySn7wfIE5_uxg&ithint=folder%2cwav. Accessed 23 Mar 2016.
sound pressure level
phosphate buffered saline
granule cell layer
The authors are grateful to all those who participated in this research for their contributions.
This work was supported by the National Major Scientific Instruments Development Project (2012YQ03026007, 2013YQ030923 to WL), the National Nature Science Foundation of China (81271511 to WL, 31300895 to YZ, 81421061 to LH), the Shanghai Municipal Commission of Science and Technology Program (14JC1403700 to WL), “Eastern Scholar” project supported by Shanghai Municipal Education Commission, the fund of Shanghai Jiao Tong University (15JCZZ02 to WL), National Key Laboratory of Human Factors Engineering Open Fund Project (HF2013-K-02 to WL), National Key Laboratory Grant of Human Factors Engineering (SYFD140051801 to XC) and National Basic Research Program of China (2011CB711000 to XC).
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