Salicylate-induced Tinnitus Research Articles

Involvement of BK Channels and Ryanodine Receptors in Salicylate-induced Tinnitus.

Neural hyperexcitability of the central auditory system is a key pathological characteristic of tinnitus, but its underlying molecular mechanisms remain elusive. The large-conductance Ca2+-activated K+ channel (BK) plays a crucial role in down- or upregulating neuronal activity. This study aims to investigate the role of BK channels in mediating tinnitus-associated neural hyperexcitability and elucidate the mechanisms behind it. Immunofluorescent staining revealed extensive expression of the BK channels on neurons within the central auditory system of rats. After long-term systemic administration of salicylate, a stable tinnitus inducer, we observed a significant change in the expression levels of BKα and β4 subunits in the rat central auditory system. In addition, salicylate was found to enhance the outward potassium currents mediated by the BK channel when exogenously expressed in HEK293 cells. Interestingly, this effect could be blocked by ryanodine, a potent inhibitor of ryanodine receptors (RyRs). Molecular docking identified Gln4020 within the central domain of RyR as a key residue in RyR-salicylate interactions. The results indicated that salicylate might directly activate RyRs leading to Ca2+ release from endoplasmic reticulum, and increased BK currents subsequently. Systemic treatment with paxilline, a potent blocker of BK channel, selectively reversed the increased P4/P1 amplitude ratios in the frequency region of tinnitus perception induced by single-dose salicylate administration. These results suggest that BK channels and ryanodine receptors may play a selective role in salicylate-induced tinnitus.

Exploring the effects of naringenin on cell functioning and energy synthesis in the hippocampus of male Wistar rats with chronic tinnitus, by examining genetic indicators such as Bax, Bcl-2, Tfam, and Pgc-1α

BackgroundThe pivotal factors, including neural plasticity, oxidative stress, neuronal inflammation, and apoptosis, play a significant role in the pathogenesis of tinnitus. The balance between Bax/Bcl-2 genes is an important factor in determining the rate of apoptosis. Pgc-1α and Tfam genes are fundamental regulators of mitochondrial biogenesis. Naringenin possesses significant antioxidant, neuroprotective, anti-inflammatory, anti-apoptotic, and antiviral properties, and its compounds are effective on cell signaling pathways. AimsIn light of the aforementioned information, we endeavored to evaluate the impact of naringenin on the expression levels of Bax, Bcl-2, Pgc-1α, and Tfam genes in the hippocampus of male Wistar rats with chronic tinnitus. Material and MethodsTo demonstrate the existence of tinnitus, all rats were instructed to complete an “active avoidance test” utilizing a conditioning box. The expression levels of genes mentioned above were assessed using real-time PCR. ResultsThe sodium salicylate at a dosage of 350 mg/kg showed an upregulation in the expression level of Bax and a downregulation in the expression level of the Bcl-2 gene (p < 0.001). Furthermore, the sodium salicylate displayed significantly higher expression levels of Tfam and Pgc-1α (p < 0.001) genes. The naringenin, at a dose of 100 mg/kg, led to a decrease in Bax gene expression (p < 0.05) and an increase in Bcl-2 gene expression (p < 0.05). On the other hand, naringenin restored the expression level of both Tfam (p < 0.001) and Pgc-1α (p < 0.01) genes. ConclusionsOur research findings demonstrate that sodium salicylate-induced tinnitus leads to enhanced apoptosis and mitochondrial biogenesis within the hippocampus. Additionally, our evidence recommends that naringenin can reduce apoptosis effectively and maintain a balanced mitochondrial state.

Photobiomodulation therapy in improvement of harmful neural plasticity in sodium salicylate-induced tinnitus.

Tinnitus is a common annoying symptom without effective and accepted treatment. In this controlled experimental study, photobiomodulation therapy (PBMT), which uses light to modulate and repair target tissue, was used to treat sodium salicylate (SS)-induced tinnitus in a rat animal model. Here, PBMT was performed simultaneously on the peripheral and central regions involved in tinnitus. The results were evaluated using objective tests including gap pre-pulse inhibition of acoustic startle (GPIAS), auditory brainstem response (ABR) and immunohistochemistry (IHC). Harmful neural plasticity induced by tinnitus was detected by doublecortin (DCX) protein expression, a known marker of neural plasticity. PBMT parameters were 808 nm wavelength, 165 mW/cm2 power density, and 99 J/cm2 energy density. In the tinnitus group, the mean gap in noise (GIN) value of GPIAS test was significantly decreased indicated the occurrence of an additional perceived sound like tinnitus and also the mean ABR threshold and brainstem transmission time (BTT) were significantly increased. In addition, a significant increase in DCX expression in the dorsal cochlear nucleus (DCN), dentate gyrus (DG) and the parafloccular lobe (PFL) of cerebellum was observed in the tinnitus group. In PBMT group, a significant increase in the GIN value, a significant decrease in the ABR threshold and BTT, and also significant reduction of DCX expression in the DG were observed. Based on our findings, PBMT has the potential to be used in the management of SS-induced tinnitus.

Characterization of the neural circuitry of the auditory thalamic reticular nucleus and its potential role in salicylate-induced tinnitus.

Subjective tinnitus, the perception of sound without an external acoustic source, is often subsequent to noise-induced hearing loss or ototoxic medications. The condition is believed to result from neuroplastic alterations in the auditory centers, characterized by heightened spontaneous neural activities and increased synchrony due to an imbalance between excitation and inhibition. However, the role of the thalamic reticular nucleus (TRN), a structure composed exclusively of GABAergic neurons involved in thalamocortical oscillations, in the pathogenesis of tinnitus remains largely unexplored. We induced tinnitus in mice using sodium salicylate and assessed tinnitus-like behaviors using the Gap Pre-Pulse Inhibition of the Acoustic Startle (GPIAS) paradigm. We utilized combined viral tracing techniques to identify the neural circuitry involved and employed immunofluorescence and confocal imaging to determine cell types and activated neurons. Salicylate-treated mice exhibited tinnitus-like behaviors. Our tracing clearly delineated the inputs and outputs of the auditory-specific TRN. We discovered that chemogenetic activation of the auditory TRN significantly reduced the salicylate-evoked rise in c-Fos expression in the auditory cortex. This finding posits the TRN as a potential modulatory target for tinnitus treatment. Furthermore, the mapped sensory inputs to the auditory TRN suggest possibilities for employing optogenetic or sensory stimulations to manipulate thalamocortical activities. The precise mapping of the auditory TRN-mediated neural pathways offers a promising avenue for designing targeted interventions to alleviate tinnitus symptoms.

Effectiveness of transcutaneous vagus nerve stimulation (tVNS) on salicylate-induced tinnitus

IntroductionTinnitus is the most common symptom of auditory system disorders. It affects the quality of life of millions of people, but it is still incurable in most cases. Vagus nerve stimulation (VNS) therapy is a potential new treatment for subjective tinnitus. In this study, transcutaneous vagus nerve stimulation (tVNS) combined with tones was utilized to treat salicylate-induced tinnitus since salicylate is a reliable and convenient approach for rapidly inducing tinnitus. MethodsWistar rats were divided into acoustic stimulation alone (AS, n = 6), tVNS alone (n = 6), and tVNS with AS (n = 6) groups for behavioral and electrophysiological tests. They were assessed by auditory brainstem response (ABR), prepulse inhibition (PPI), gap prepulse inhibition of the acoustic startle (GPIAS), social interactions, and aggressive behavior tests at baseline and seven days’ post-salicylate (175 mg/kg, twice a day) injection. ResultsThe inhibition percentage of the GPIAS test was significantly reduced post-salicylate injection in the tVNS and AS alone groups, while it was not significant in the tVNS with AS group. There was no significant difference in the mean percentage of the GPIAS test between the tVNS groups (with or without AS) after salicylate injections. Social interactions were significantly different in the AS alone group pre- and post-salicylate injections, but they were not significant in other groups. Moreover, the results of aggressive behavior tests showed significantly increased post-salicylate injections in the AS alone group, while they were not significant in the tVNS groups (with or without AS). ConclusionsThe current study revealed that the application of tVNS alone produced improved social interaction and mood and alleviated salicylate-induced tinnitus severity. Moreover, combining tVNS with acoustic stimulation can prevent salicylate-induced tinnitus.

Effect of lidocaine on salicylate-induced tinnitus in guinea pigs: A focus on the auditory cortex.

This study aimed to investigate the effects of the intravenous administration of lidocaine in the auditory cortex after the systemic administration of salicylate. Healthy male albino Hartley guinea pigs were divided into two groups. The control group received only lidocaine, whereas the experimental group received lidocaine after checking for the effects of salicylate. Extracellular recordings of spikes in the primary auditory cortex and dorsocaudal areas in healthy albino Hartley guinea pigs were continuously documented (pre- and post-lidocaine, pre- and post-salicylate, and post-salicylate after adding lidocaine to post-salicylate). We recorded 160 single units in the primary auditory cortex from five guinea pigs and 155 single units in the dorsocaudal area from another five guinea pigs to confirm the effects of lidocaine on untreated animals. No significant change was detected in either the threshold or Q10dB value after lidocaine administration in the primary auditory cortex and dorsocaudal areas. Spontaneous firing activity significantly decreased after lidocaine administration in the primary auditory cortex and dorsocaudal areas. Next, we recorded 160 single units in the primary auditory cortex from five guinea pigs and 137 single units in the dorsocaudal area from another five guinea pigs to determine the effects of lidocaine on salicylate-treated animals. The threshold was significantly elevated after salicylate administration; however, no additional change was detected after adding lidocaine to the primary auditory cortex and dorsocaudal areas. Regarding the Q10dB value, lidocaine negated the significant changes induced by salicylate in the primary auditory cortex and dorsocaudal areas. Moreover, lidocaine negated the significant changes in spontaneous firing activities induced by salicylate in the primary auditory cortex and dorsocaudal areas. In conclusion, changes in the Q10dB value and spontaneous firing activities induced by salicylate administration are abolished by lidocaine administration, suggesting that these changes are related to the presence of tinnitus.

Functional Connectivity Alterations and Molecular Characterization of the Anterior Cingulate Cortex in Tinnitus Pathology without Hearing Loss.

Compared with individuals with hearing loss, tinnitus patients without hearing loss have more psychological or emotional problems. Tinnitus is closely associated to abnormal metabolism and function of the limbic system, a key brain region for emotion experience, but the underlying molecular mechanism remains unknown. Using whole-brain microvasculature dynamics imaging, the anterior cingulate cortex (ACC) is identified as a key brain region of limbic system involve in the onset of salicylate-induced tinnitus in mice. In the tinnitus group, there is enhanced purine metabolism, oxidative phosphorylation, and a distinct pattern of phosphorylation in glutamatergic synaptic pathway according to the metabolome profiles, quantitative proteomic, and phosphoproteomic data of mice ACC tissue. Electroencephalogram in tinnitus patients with normal hearing thresholds show that the functional connectivity between pregenual anterior cingulate cortex and the primary auditory cortex is significantly increased for high-gamma frequency band, which is positively correlated with the serum glutamate level. These findings indicate that ACC plays an important role in the pathophysiology of tinnitus by interacting with the primary auditory cortex and provide potential molecular targets in the ACC for tinnitus treatment.

Temporal Changes in Functional and Structural Neuronal Activities in Auditory System in Non-Severe Blast-Induced Tinnitus.

Background and Objectives: Epidemiological data indicate that blast exposure is the most common morbidity responsible for mild TBI among Service Members (SMs) during recent military operations. Blast-induced tinnitus is a comorbidity frequently reported by veterans, and despite its wide prevalence, it is also one of the least understood. Tinnitus arising from blast exposure is usually associated with direct structural damage that results in a conductive and sensorineural impairment in the auditory system. Tinnitus is also believed to be initiated by abnormal neuronal activities and temporal changes in neuroplasticity. Clinically, it is observed that tinnitus is frequently accompanied by sleep disruption as well as increased anxiety. In this study, we elucidated some of the mechanistic aspects of sensorineural injury caused by exposure to both shock waves and impulsive noise. The isolated conductive auditory damage hypothesis was minimized by employing an animal model wherein both ears were protected. Materials and Methods: After the exposure, the animals' hearing circuitry status was evaluated via acoustic startle response (ASR) to distinguish between hearing loss and tinnitus. We also compared the blast-induced tinnitus against the well-established sodium salicylate-induced tinnitus model as the positive control. The state of the sensorineural auditory system was evaluated by auditory brainstem response (ABR), and this test helped examine the neuronal circuits between the cochlea and inferior colliculus. We then further evaluated the role of the excitatory and inhibitory neurotransmitter receptors and neuronal synapses in the auditory cortex (AC) injury after blast exposure. Results: We observed sustained elevated ABR thresholds in animals exposed to blast shock waves, while only transient ABR threshold shifts were observed in the impulsive noise group solely at the acute time point. These changes were in concert with the increased expression of ribbon synapses, which is suggestive of neuroinflammation and cellular energy metabolic disorder. It was also found that the onset of tinnitus was accompanied by anxiety, depression-like symptoms, and altered sleep patterns. By comparing the effects of shock wave exposure and impulsive noise exposure, we unveiled that the shock wave exerted more significant effects on tinnitus induction and sensorineural impairments when compared to impulsive noise. Conclusions: In this study, we systematically studied the auditory system structural and functional changes after blast injury, providing more significant insights into the pathophysiology of blast-induced tinnitus.

Online ascorbate sensing reveals oxidative injury occurrence in inferior colliculus in salicylate-induced tinnitus animal model

Tinnitus is a widespread and serious clinical and social problem. Although oxidative injury has been suggested to be one of pathological mechanisms in auditory cortex, whether this mechanism could be applied to inferior colliculus remains unclear. In this study, we used an online electrochemical system (OECS) integrating in vivo microdialysis with selective electrochemical detector to continuously monitor the dynamics of ascorbate efflux, an index of oxidative injury, in inferior colliculus of living rats during sodium salicylate-induced tinnitus. We found that OECS with a carbon nanotubes (CNTs)-modified electrode as the detector selectively responses to ascorbate, which is free from the interference from sodium salicylate and MK-801 that were used to induce tinnitus animal model and investigate the N-methyl-d-aspartate (NMDA) receptor mediated excitotoxicity, respectively. With the OECS, we found that the extracellular ascorbate level in inferior colliculus significantly increases after salicylate administration and such increase was suppressed by immediate injection of NMDA receptor antagonist MK-801. In addition, we found that salicylate administration significantly increases the spontaneous and sound stimuli evoked neural activity in inferior colliculus and that the increases were inhibited by the injection of MK-801. These results suggest that oxidative injury may occur in inferior colliculus following salicylate-induced tinnitus, which is closely relevant to the NMDA-mediated neuronal excitotoxicity. This information is useful for understanding the neurochemical processes in inferior colliculus involved in tinnitus and its related brain diseases.

Susceptibility and mechanism of sodium salicylate-induced tinnitus model in low estrogen rats

Objective: The susceptibility of tinnitus rats with low estrogen level induced by sodium salicylate and the changes of tumor necrosis factor α (TNF-α) in serum were observed to investigate the relationship between tinnitus occurrence and estrogen level. Methods: Forty-two healthy female Wistar rats were randomly divided into control group(n=6), normal group(n=6), sham operation group(n=6) and ovariectomized group(n=24). Control group was intraperitoneally injected with normal saline 200 mg/kg for 14 consecutive days. Normal group, sham operation group and ovariectomized group were intraperitoneally injected with sodium salicylate 200 mg/kg for 14 consecutive days. Before and after sodium salicylate induction, the tinnitus behavior of rats in each group was detected by prepulse inhibition (PPI) and gap pre-pulse inhibition of the acoustic startle (GPIAS) test. Before and after sodium salicylate induction, blood samples were collected from eyeballs of rats in each group, and serum levels of estradiol and TNF-α were detected by ELISA. SPSS 25.0 software was used to analyze the data. Results: (1) Following 14 days of sodium salicylate intervention, there was no significant difference in PPI inhibition rate between groups or within groups(all P>0.05). (2)There was no significant difference in the inhibition rate of GPIAS in the four groups before sodium salicylate injection(F=0.217, P>0.05). With sodium salicylate injected for 14 days, the inhibition rate of GPIAS in ovariectomized group (30.88%±15.40%) was significantly lower than that in the other three groups (44.11%±21.06%, 38.27%±10.92%, 51.59%±11.34%), and the difference was statistically significant(F=3.533, P<0.05). The inhibition rate of GPIAS in ovariectomized group with sodium salicylate injected for 14 days was significantly lower than that before injection, and the difference was statistically significant(t=2.977, P<0.05).There was no significant difference in GPIAS inhibition rate between the other three groups before and after sodium salicylate injection(P>0.05). (3)The level of TNF-α in ovariectomized rats was significantly higher than that in the other three groups, the difference was statistically significant(all P<0.05). With sodium salicylate injection for 14 days, TNF-α level in the ovariectomized group increased more significantly than that in the other three groups, the difference was statistically significant(F=8.045, P<0.05). TNF-α levels increased following salicylate injection in normal group, sham operation group and ovariectomized group, and the differences were statistically significant(t value was -4.843, -4.932 and -5.965 respectively, each P<0.05). There was no significant difference in TNF-α levels before and after normal saline injection in control group(all P>0.05). Conclusion: Low estrogen levels increase susceptibility to sodium salicylate-induced tinnitus. Decreased estrogen levels may increase susceptibility to tinnitus through the increased expression of pro-inflammatory factor TNF-α.

Protective Effect of Resveratrol in an Experimental Model of Salicylate-Induced Tinnitus.

To date, the effect of resveratrol on tinnitus has not been reported. The attenuative effects of resveratrol (RSV) on a salicylate-induced tinnitus model were evaluated by in vitro and in vivo experiments. The gene expression of the activity-regulated cytoskeleton-associated protein (ARC), tumor necrosis factor-alpha (TNFα), and NMDA receptor subunit 2B (NR2B) in SH-SY5Y cells was examined using qPCR. Phosphorylated cAMP response element-binding protein (p-CREB), apoptosis markers, and reactive oxygen species (ROS) were evaluated by in vitro experiments. The in vivo experiment evaluated the gap-prepulse inhibition of the acoustic startle reflex (GPIAS) and auditory brainstem response (ABR) level. The NR2B expression in the auditory cortex (AC) was determined by immunohistochemistry. RSV significantly reduced the salicylate-induced expression of NR2B, ARC, and TNFα in neuronal cells; the GPIAS and ABR thresholds altered by salicylate in rats were recovered close to their normal range. RSV also reduced the salicylate-induced NR2B overexpression of the AC. These results confirmed that resveratrol exerted an attenuative effect on salicylate-induced tinnitus and may have a therapeutic potential.

Small molecule modulation of the large-conductance calcium-activated potassium channel suppresses salicylate-induced tinnitus in mice

Tinnitus is the phantom perception of sound that has no external source. A neurological signature of tinnitus, and the frequently associated hyperacusis, is an imbalance between excitatory and inhibitory activity in the central auditory system (CAS), leading to dysregulated network excitability. The large conductance, calcium-activated potassium (BK) channel is a key player in pre- and post-synaptic excitability through its mediation of K+ currents. Changes in BK channel activity are associated with aberrant network activity in sensory regions of the CNS, raising the possibility that BK channel modulation could regulate activity associated with tinnitus and hyperacusis. To test whether BK channel openers are able to suppress biomarkers of drug-induced tinnitus and hyperacusis, the 1,3,4 oxadiazole BMS-191011 was given to young adult CBA mice that had been administered 250 mg/kg sodium salicylate (SS). Systemic treatment with BMS-191011 reduced behavioral manifestations of SS-induced tinnitus, but not hyperacusis, probed via the gap-in-noise startle response method. Systemic BMS-191011 treatment did not influence SS-induced increases in auditory brainstem response functions, but local application at the inferior colliculus did reverse SS-suppressed spontaneous activity, particularly in the frequency region of the tinnitus percept. Thus, action of BMS-191011 in the inferior colliculus may contribute to the reduction in behaviorally measured tinnitus. Together, these findings support the utility of BK channel openers in reducing central auditory processing changes associated with the formation of the tinnitus percept.

The Auditory Brainstem Response (ABR) Test, Supplementary to Behavioral Tests for Evaluation of the Salicylate-Induced Tinnitus.

The Auditory Brainstem Response (ABR) Test, Supplementary to Behavioral Tests for Evaluation of the Salicylate-Induced Tinnitus.

Nitric oxide signalling underlies salicylate-induced increases in neuronal firing in the inferior colliculus: A central mechanism of tinnitus?

The anti-inflammatory drug salicylate induces tinnitus in animals and man. Salicylate reduces cochlear output but causes hyperactivity in higher auditory centres, including the inferior colliculus (the auditory midbrain). Using multi-electrode recording in anaesthetised guinea pigs (Cavia porcellus), we addressed the hypothesis that salicylate-induced hyperactivity in the inferior colliculus involves nitric oxide signalling secondary to increased ascending excitatory input.Systemic salicylate (200 mg/kg i.p., 0 h) markedly increased spontaneous and sound-driven neuronal firing in the inferior colliculus (3-6 h post drug), with both onset and sustained responses to pure tones being massively increased. Reverse microdialysis of increasing concentrations of salicylate directly into the inferior colliculus (100 µM-10 mM, from 0 h) failed to mimic systemic salicylate. In contrast, it caused a small, transient, increase in sound-driven firing (1 h), followed by a larger sustained decrease in both spontaneous and sound-driven firing (2-5 h). When salicylate was given systemically, reverse microdialysis of the neuronal nitric oxide synthase inhibitor L-methyl arginine into the inferior colliculus (500 mM, 2-6 h) completely blocked the salicylate-induced increase in spontaneous and sound-driven neuronal firing.Our data indicate that systemic salicylate induces neuronal hyperactivity in the auditory midbrain via a mechanism outside the inferior colliculus, presumably upstream in the auditory pathway; and that the mechanism is ultimately dependent on nitric oxide signalling within the inferior colliculus.Given that nitric oxide is known to mediate NMDA receptor signalling in the inferior colliculus, we propose that salicylate activates an ascending glutamatergic input to the inferior colliculus and that this is an important mechanism underlying salicylate-induced tinnitus.

Behavioral and Immunohistochemical Evidence for Suppressive Effects of Goshajinkigan on Salicylate-Induced Tinnitus in Rats.

Many people are affected by tinnitus, a sensation of ringing in the ear despite the absence of external sound. Goshajinkigan (GJG) is one of the formulations of Japanese traditional herbal medicine and is prescribed for the palliative treatment of patients with tinnitus. Although GJG is clinically effective in these patients, its behavioral effects and the underlying neuroanatomical substrate have not been modeled in animals. We modeled tinnitus using salicylate-treated rats, demonstrated the effectiveness of GJG on tinnitus, and examined the underlying neuronal substrate with c-Fos expression. Intraperitoneal injection of sodium salicylate (400 mg/kg) into rats for three consecutive days significantly increased false positive scores, which were used to assess tinnitus behavior. When GJG was orally administered one hour after each salicylate injection, the increase in tinnitus behavior was suppressed. The analysis of c-Fos expression in auditory-related brain areas revealed that GJG significantly reduced the salicylate-induced increase in the number of c-Fos-expressing cells in the auditory cortices, inferior colliculus, and dorsal cochlear nucleus. These results suggest a suppressive effect of GJG on salicylate-induced tinnitus in animal models.

Neuroprotective Effect of Valproic Acid on Salicylate-Induced Tinnitus.

High-dose salicylate induces temporary moderate hearing loss and the perception of a high-pitched tinnitus in humans and animals. Previous studies demonstrated that high doses of salicylate increase N-methyl-d-aspartate (NMDA) receptor levels, resulting in a rise in Ca2+ influx and induction of excitotoxicity. Glutamate excitotoxicity is associated with failure in the maintenance of calcium homeostasis, mitochondrial dysfunction, and production of reactive oxygen species (ROS). Valproic acid (VPA) is widely used for the management of bipolar disorder, epilepsy, and migraine headaches, and is known to regulate NMDA receptor activity. In this study, we examined the beneficial effects of VPA in a salicylate-induced tinnitus model in vitro and in vivo. Cells were pretreated with VPA followed by salicylate treatment. The expression levels of NMDA receptor subunit NR2B, phosphorylated cAMP response element-binding protein—an apoptosis marker, and intracellular levels of ROS were measured using several biochemical techniques. We observed increased expression of NR2B and its related genes TNFα and ARC, increased intracellular ROS levels, and induced expression of cleaved caspase-3. These salicylate-induced changes were attenuated in the neuronal cell line SH-SY5Y and rat cortical neurons after VPA pretreatment. Together, these results provide evidence of the beneficial effects of VPA in a salicylate-induced temporary hearing loss and tinnitus model.

Salicylate-Induced Changes in Hearing Thresholds in Mongolian Gerbils Are Correlated With Tinnitus Frequency but Not With Tinnitus Strength.

Tinnitus is an auditory phantom percept without external sound sources. Despite the high prevalence and tinnitus-associated distress of affected patients, the pathophysiology of tinnitus remains largely unknown, making prevention and treatments difficult to develop. In order to elucidate the pathophysiology of tinnitus, animal models are used where tinnitus is induced either permanently by noise trauma or transiently by the application of salicylate. In a model of trauma-induced tinnitus, we have suggested a central origin of tinnitus-related development of neuronal hyperactivity based on stochastic resonance (SR). SR refers to the physiological phenomenon that weak subthreshold signals for given sensors (or synapses) can still be detected and transmitted if appropriate noise is added to the input of the sensor. The main objective of this study was to characterize the neurophysiological and behavioral effects during salicylate-induced tinnitus and compare these to the conditions within the trauma model. Our data show, in line with the pharmacokinetics, that hearing thresholds generally increase 2 h after salicylate injections. This increase was significantly stronger within the region of best hearing compared to other frequencies. Furthermore, animals showed behavioral signs of tinnitus during that time window and frequency range as assessed by gap prepulse inhibition of the acoustic startle reflex (GPIAS). In contrast to animals with noise trauma-induced tinnitus, salicylate-induced tinnitus animals showed no correlation between hearing thresholds and behavioral signs of tinnitus, indicating that the development of tinnitus after salicylate injection is not based on SR as proposed for the trauma model. In other words, salicylate-induced tinnitus and noise trauma-induced tinnitus are not based on the same neurophysiological mechanism.

Participation of the Anterior Cingulate Cortex in Sodium Salicylate-induced Tinnitus.

The anterior cingulate cortex (ACC) participates in sodium salicylate (SS)-induced tinnitus through alteration of the disordered neural activity and modulates the neuronal changes in the auditory cortex (AC). Although the mechanism underlying tinnitus remains unclear, the crucial roles of the auditory center and limbic system in this process have been elucidated. Recent reports suggest that dysfunction of the ACC, an important component of the limbic system that regulates and controls the conduction of multiple sensations, is involved in tinnitus. Although altered functional connectivity between the ACC and the auditory system has been observed in humans with tinnitus, the underlying neuronal mechanism remains unexplored. SS (350 mg/kg, 10%, i.p.) was used to yield tinnitus model in rats, followed by comparison of the alteration in the spontaneous firing rate (SFR), local field potential (LFP), and extracellular glutamic acid in the ACC. The responses of neurons in the AC to electrical stimulation from the ACC were also observed. We determined significant increases in the neuronal SFR and extracellular glutamate level in the ACC after SS injection (p < 0.05). These effects were accompanied by decreased alpha band activity and increased beta and gamma band activity (p < 0.05). In the majority of AC neurons, the SFR decreased in response to ACC stimulation (p < 0.05). Our results demonstrated that disordered neural activity in the ACC contributes to SS-induced tinnitus and that ACC activation can modulate AC activity.

Role of the caudate-putamen nucleus in sensory gating in induced tinnitus in rats.

Tinnitus can be described as the conscious perception of sound without external stimulation, and it is often accompanied by anxiety, depression, and insomnia. Current clinical treatments for tinnitus are ineffective. Although recent studies have indicated that the caudate-putamen nucleus may be a sensory gating area involved in noise elimination in tinnitus, the underlying mechanisms of this disorder are yet to be determined. To investigate the potential role of the caudate-putamen nucleus in experimentally induced tinnitus, we created a rat model of tinnitus induced by intraperitoneal administration of 350 mg/kg sodium salicylate. Our results revealed that the mean spontaneous firing rate of the caudate-putamen nucleus was increased by sodium salicylate treatment, while dopamine levels were decreased. In addition, electrical stimulation of the caudate-putamen nucleus markedly reduced the spontaneous firing rate of neurons in the primary auditory cortex. These findings suggest that the caudate-putamen nucleus plays a sensory gating role in sodium salicylate-induced tinnitus. This study was approved by the Institutional Animal Care and Use Committee of Peking University Health Science Center (approval No. A2010031) on December 6, 2017.

Aberrant expression of Nav1.6 in the cochlear nucleus correlates with salicylate-induced tinnitus in rats

Hyperactivity in cochlear nucleus (CN) is one of the major neural correlates for tinnitus induction, yet the molecular factors that participate in the neuronal hyperexcitability remain unclear. The present study showed that acute and chronic administrations of salicylate were both capable of inducing reversible tinnitus in rats. The number of GAD 65/67-immunoreactive neurons in the AVCN and DCN was decreased, while the number of VGLUT 1/2-immunoreactive neurons in the AVCN and DCN was increased when rats were experiencing tinnitus, providing evidence for excitatory-inhibitory imbalance in CN is correlated with tinnitus. Interestingly, the expression level of Nav1.6, an important subtype of voltage-gated sodium channels was significantly increased in the DCN and AVCN of rats experiencing tinnitus, the up-regulation of Nav1.6 was returned to normal level following the disappearance of tinnitus. Double-labeling experiments revealed that Nav1.6 expression was down-regulated in the GAD 65/67-positive neurons in the DCN and AVCN of rats experiencing tinnitus. Notably, the percentage of co-localization of Nav1.6 and NeuN-labeling fusiform neurons was markedly increased in the DCN during tinnitus. These findings uncover the tinnitus-associated alteration in Nav1.6, a potential key contributor that can lead to hyperexcitability in CN and contribute to salicylate-induced tinnitus.