| Objective To establish a group of stable tinnitus animal models through white noise exposure.It is the first time to evaluate a new brain functional imaging model by combining behavioral and biomarker molecular detection,and is expected to provide an objective and effective diagnostic and detection method for tinnitus research.Methods 1.Tinnitus modeling experiment: SD rats with Auditory brainstem response(ABR)threshold ≤25 dB were randomly divided into 5groups with 8 rats in each group.They were exposed to white noise with110 dB sound intensity for 2 hours(N2 group),2.5 hours(N2.5 group),3hours(N3 group),3.5 hours(N3.5 group)and 4 hours(N4 group),respectively.ABR and Gap prepμlse inhibition of the acoustic startle(GPIAS)tests were performed on 1,3,5,7,14,21 and 28 days after noise exposure to determine the animal model of noise-induced tinnitus(NIT).2.Grouping experiment: white noise with 110 dB sound intensity was exposed for 3.5 hours,and the rats were divided into 1 day after exposure group(Day1 group),21 days after exposure group(Day21 group)and Control group,with 4 rats in each group.In Day1 group,audiological and PAI tests were performed one day after exposure,and the samples were sacrificed.In Day21 group,audiological and PAI tests were performed 21 days after exposure,and the samples were sacrificed.In the Control group,audiology and PAI were performed 21 days after exposure to false noise(exposed sound source was not turned on),and the subjects were sacrificed.The protein expression levels of c-fos and TNF-α in the auditory cortex of rats in each group were detected by Western blot.The mRNA expression levels of c-fos and TNF-α in auditory cortex(ACx)were detected by Real-time Quantitativa Polymerase Chain Reaction(RT-qPCR).Results In the modeling experiment,the ABR test results of N3.5group showed temporary threshold shift,and the GPIAS test results of N3.5group showed significantly lower GPIAS values at each time node after exposure(P < 0.01)compared with those before exposure,showing stable tinnitus behavior.In the grouping experiment,the PAI results showed that compared with the control group,the photoacoustic signal values of cerebral cortex in Day1 and Day21 groups were significantly enhanced,with statistically significant differences(P < 0.01).Western blot results showed that the expression levels of c-fos and TNF-α in ACx in Day1 and Day21 groups were higher than those in Control group(P < 0.05).RT-qPCR results showed that the expression levels of c-fos and TNF-αmRNA in ACx in Day1 and Day21 groups were increased compared with that in Control group(P < 0.05).Conclusion 1.A temporary threshold shift and stable tinnitus animal model of SD rats can be established by continuous stimulation of white noise with sound intensity of 110 dB for 3.5 hours.2.After ABR threshold was restored,compared with the control group,the expression levels of c-fos and TNF-α protein and mRNA in the ACx of rats in the tinnitus group were increased,suggesting that tinnitus behavior persisted at the molecular level.3.After the ABR threshold was restored,the PAI value of cerebral cortex of rats in tinnitus group was significantly increased compared with the control group,suggesting that PAI is expected to provide a practical and stable imaging detection method for tinnitus. |
