Introduction

Tinnitus is defined as an auditory perception or sensation without any external stimuli. Tinnitus itself originates from the Latin word, “tinnire” which means “to ring” [1,2]. Patients with tinnitus often described the auditory perception as ringing, buzzing, hissing, or whistling [1]. About 15-20% of people in the world will experience tinnitus in their lifetime, and constant noise perception in the ears can range from mild to very debilitating [1,2]. Tinnitus can also affect children and adolescents with 7.5%-60% prevalence [3]. In children with normal hearing, the majority is around 6%-36%, while in deaf children, approximately 55% [4].

Tinnitus can negatively impact the patients’ psychological and emotional well-being, lowering their quality of life [5]. Some of the most common psychological impacts experienced by tinnitus patients are concentration disturbance, sleep disorder, and mood disorder, which lead to unstable emotions such as getting angry more easily. Patients with tinnitus also have a greater risk of experiencing depression, anxiety, and insomnia, thus affecting their quality of life [5,6].

The etiology of tinnitus is not fully known and is often multifactorial.7 Most of the cases are related to hearing impairment and changes in the inner ear [7,8]. Previous studies showed that one of the significant risk factors for tinnitus is excessive noise exposure. The exposure could be both long-term or a brief loud noise that can result in some changes in the structures of the inner ear. Studies also showed that the prevalence of tinnitus with hearing loss or hearing impairment is higher than those with normal hearing [4,9]. In addition, neurological disorders or head trauma, such as skull fractures, temporomandibular joint disorders, whiplash injuries, and multiple sclerosis, as well as other conditions, such as thyroid disease, hyperlipidemia, and vascular disease, including arterial bruit and arteriovenous malformation can also be related to tinnitus, which contributes to 5-10% of tinnitus cases [7].

The mechanism underlying tinnitus is also not fully understood [10]. Some experts have proposed the central gain control theory, suggesting that tinnitus is related to neural plasticity, an increase in spontaneous neural activity due to loss or reduction of sensory input. Evidence of an increase in auditory neural activity, particularly in fusiform cells, inferior colliculus (IC), and medial geniculate body (MGB), also reorganization of the tonotopic map in the auditory cortex of animals showing evidence of behavioral tinnitus reinforces the theory [11-13]. In their study reported a significant reduction in gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the brain, in the auditory cortex of patients with tinnitus. This loss of inhibition is likely associated with increased neuronal activity in tinnitus [11,14].

Besides the central gain control theory, recent evidence suggests that neuroinflammation also underlies tinnitus. Wang et al. indicate that inflammation of the primary auditory pathway induced by noise-induced hearing loss (NIHL) plays a role in developing tinnitus in mice. Their research showed that mice with NIHL had elevated proinflammatory cytokines and activated microglia, a non-neuronal cell in the primary auditory cortex, the two key features of neuroinflammatory responses. Proinflammatory cytokine such as TNF-α is required for noise-induced neuroinflammation and tinnitus. Experimental rats no longer showed signs of tinnitus after being given TNF-α inhibitors. Furthermore, genetic knockout TNF-α prevented neuroinflammation and ameliorated the behavioral phenotype associated with tinnitus in mice with NIHL [15].

In recent years, the gut microbiome has attracted the attention of researchers. Various studies have shown that microbiota has specific and complex cross-talk with their hosts, particularly with the central nervous system (CNS), creating the “gut-brain axis”. The secretion of various metabolites, bacterial structural components, and signaling molecules were involved in this gut, gut microbiota, and brain’s communication. In addition, imbalances in the gut microbiota composition can modulate the immune system and barrier function of tissues such as the blood-brain barrier (BBB), so impaired gut-brain interactions may give rise to various neurological disturbances [16,17]. Interestingly, recent studies have shown that the gut microbiota can regulate the production of neurotransmitters, such as GABA and serotonin [17-19], and inflammatory mediators, such as TNF-α and IL-6 [20,21]. Furthermore, this altered regulation of gut microbiota could contribute to the changes in the concentration of neurotransmitters and inflammatory mediators which possibly related to tinnitus [15,16,22,23].

So far, tinnitus is still an unsolved clinical problem. There is no cure for tinnitus. Some treatments may help some people, but none work for everyone. Increased knowledge about the pathomechanism of tinnitus can improve tinnitus management. However, recent studies indicating altered neurotransmitter production and activity and increased levels of proinflammatory cytokines in experimental animals exhibiting tinnitus behavior have provided insights into the involvement of the central auditory system and neuroinflammation in tinnitus mechanisms.

On the other hand, the gut microbiota has played an essential role in regulating the neuroendocrine system via the gut-brain axis, maintaining immune homeostasis, and preventing chronic inflammation. A dysbiosis condition of the gut microbiota will disrupt the gut-brain axis and stimulate an inflammatory process, which may cause tinnitus. Hence, this literature review focuses on gut dysbiosis and its effect on neurotransmitter and immune homeostasis, particularly neuroinflammation, and its role in the development of tinnitus, which is expected to become a theoretical framework for further research on the pathomechanism or therapeutic approach for tinnitus.

Methods

This research is done using a qualitative study design with the literature study approach encompassing literature seeking, identifying, analyzing, and interpreting previous studies that discussed factors related to gut microbial dysbiosis, neurotransmitters, neuroinflammation, and its implication in tinnitus. Literature searching was done from October 2021 until January 2022 and the strategy used was based on the Preferred Reporting Items for Systematic Reviews and Meta-analyses Statement (PRISMA). PubMed was the major search engine to collect literature in the full-text original article published between January 2016 and December 2021. In addition, reports from other sources, such as manual searches through the list of found study references that have relevant topics, were also used in this study. Some keywords accounted for the searching process: Gut Microbiome OR Tinnitus AND (“Neurotransmitter” OR Serotonin OR Dopamine OR Tinnitus Signaling); Microbiota-Gut-Brain-Axis AND Neuroinflammation OR Proinflammatory Cytokines OR TNF-α OR IL-6.

The inclusion criteria are literatures (1) in English; (2) published between January 2016 until December 2021; (3) in the form of full text original article; (4) an experimental study, inclusive of in vivo, in vitro, and human studies; (5) discusses about gut microbiome, gut dysbiosis, gut-brainaxis, neurotransmitter, neuroinflammation, and tinnitus signaling. Literature discusses other than neurological symptoms or diseases was excluded.

Results

In this study, we found 650 articles from an initial search on PubMed and other sources, and the literature continued for further selection was 252 original articles. Subsequently, we selected 28 writings by title and abstract. After a complete reading of the text, some literature did not meet the inclusion criteria, resulting in 22 articles relevant to this study (Figure 1).

Figure 1: Brazilian Scale of Hearing and Language Development.

The main focus of this study is the pathophysiology of tinnitus concerning neurotransmitter activity and neuroinflammation associated with gut microbiota dysbiosis. However, some data need to be considered, such as the influence of gut microbiota and gut dysbiosis on neurotransmitters and neuroinflammatory mediators’ production, brain neurotransmitters associated with tinnitus, and neuroinflammatory mediators’ profile in conditions related to tinnitus.

The gut microbiome has attracted the attention of many researchers in recent years. The ability of gut microbiota to communicate with the host via the gut-brain axis by modulating the neurotransmitter production or activity may underlie the neural plasticity involved in generating tinnitus. Evidence showed that several neurotransmitters’ production increase following the presence of gut microbiota, as provided in Table 1 below.

Table 1: Neurotransmitter Modulation by the Gut Microbiota

GABA is one of the major inhibitory neurotransmitters in the brain that is reported to decrease in tinnitus. Table 1 describes several in vitro studies proving that gut microbiota can produce GABA, particularly Bifidobacterium, Bacteroides, Parabacteroides, and Eubacterium. have examined the genome of 83 Bifidobacteria species and found out that 30 out of 83 species contain succinate-semialdehyde dehydrogenase involves in the conversion of succinate to GABA, while 79 out of 83 species possess the genes to produce GABA from glutamate or glutamine. In vitro study by also showed a similar result: Bifidobacterium contains Gad genes involved in GABA production, and Bifidobacterium adolescentis could generate GABA from precursor GMS.

On the other hand, serotonin or 5-HT is a neurotransmitter involved in enhancing the firing rate in the auditory pathway [24]. Studies conducted and Mandic et al. (2019) showed that gut microbiota, namely Bifidobacterium dentium and Clostridium ramosum could regulate the production of 5-HT but not directly from de novo synthesis of 5-HT due to the lack of genes involved in 5-HT production. However, they increase the 5-HT positive enteroendocrine cells, Tph1 and Maoa genes expression to stimulate neurotransmitter production. Moreover, the B. dentium secreted products, including acetate, and C. ramosum cells lysates, can also stimulate enterochromaffin cells to produce 5-HT. Other microbiota, namely Enterococcus faecium, has shown the ability to utilize tyrosine to produce tyramine. In contrast, Lactobacillus Plantarum has been reported to produce acetylcholine in a pantothenic acid-supplied medium [21].

Furthermore, any disruption of the microbiota composition resulting in gut dysbiosis may also affect the neurotransmitter activity or production in the body. Table 2 shows the implications of the dysbiosis gut microbiota induced by several factors, such as a highfat diet, antibiotics, and other neurological conditions, on neurotransmitter activity (Figure 2).

Table 2: Study on Neurotransmitter Alteration in Dysbiosis Gut Microbiome

Figure 2: Brazilian Scale of Hearing and Language Development.

One of the main characteristics of gut dysbiosis is an increase or decrease in abundance, diversity, or operational taxonomic units (OTUs) of some bacteria taxa. Guo et al. identified an increased bacteria OTUs phyla Bacteroidetes, Proteobacteria, Deferribacteres, Cyanobacteria and Actinobacteria following an HSHF diet in mice, while Labban et al. (2020) found that HFD fed rats had a higher abundance of Gram-negative bacteria, Clostridium, and Bacteroides [17,25]. Moreover, another study conducted by Wu et al. (2020) reported a positive correlation between 5-HT levels and bacteria taxa genus Allobaculum and Akkermansia found to be least abundant in depressed mood mice. Different studies observed some changes in the gut microbiota diversity in schizophrenic patients. Gut microbiota in schizophrenic patients tends to be more varied compared to control, with a greater α and β diversity. Furthermore, they also observed some changes in neurotransmitter levels in mice transplanted with S. vestibularis, one of the bacteria found to increase in schizophrenic patients. Interestingly, the concentrations of dopamine and GABA decreased while 5-HT levels increased in the intestinal and serum of mice transplanted with S. vestibularis, but no changes in the brain. Besides, this study also showed a negative correlation between lower serum tryptophan levels and 38 bacterial enriched-schizophrenic patients, and higher levels of kynurenic acid (KYNA) positively correlated with ten bacterial enriched schizophrenics [26].

In addition, gut dysbiosis, also associated with an altered gut barrier, subsequently leads to the impairment of bloodbrain barrier permeability and neuroinflammation, which may contribute to the generation of several neurological diseases [27]. Table 3 describes some findings of the impact of gut dysbiosis on neuroinflammation, mainly through the activation of some proinflammatory mediators and pathways (Figure 3).

Table 3: Profile of Inflammatory Mediator in Dysbiosis Gut Microbiome

Figure 3: Brazilian Scale of Hearing and Language Development.

Numerous critical pro-inflammatory cytokines, namely TNF-α, IFN-γ, IL-6, and IL-12, were increased in gut dysbiosis induced by several factors, including bacterial infection, high fat diet, and medical conditions such as Alzheimer’s and Parkinson’s diseases [25,28,29]. Furthermore, analyzed a positive correlation between plasma concentrations of TNF-α and Bacteroides abundance and between IFN-γ and Verrucomicrobia in Parkinson patients suggesting that microbiota alteration contributes to the systemic inflammation involved in the disease progression [29].

On the other hand, different studies showed increased concentrations of SCFAs in immune-activated schizophrenic patients. SCFAs is a bacterial metabolite that has essential functions in maintaining the gut epithelial barrier and has an anti-inflammatory effect by increasing anti-inflammatory and decreasing the proinflammatory cytokines [30]. However, reported an increase in several proinflammatory cytokines in immune-activated schizophrenic patients and mice model of schizophrenia following SCFAs supplementation [31].

This result may be related to the findings that showed excessive concentration of SCFAs can be less inhibitory, thus can increase the production of pro-inflammatory cytokines, inducing an inflammatory response instead of their known anti-inflammatory effect [31-33].

While there is no firm evidence of gut dysbiosis involvement in tinnitus yet, several studies have described some changes in the neurotransmitters’ activity and neuroinflammatory mediators in the central auditory pathway associated with tinnitus; this may indicate the role of gut dysbiosis in the tinnitus mechanism. One of the neurotransmitters altered in tinnitus is dopamine. As shown in Table 4, the extracellular dopamine levels decreased in mice following salicylate-induced tinnitus [34]. Dopamine is a monoamine neurotransmitter that can exert heterogeneous effects in the auditory system. Dopamine can increase or decrease neural activity, although the most significant impact is reducing its responsiveness [35]. Another study found a decrease in Gabra1 mRNA, which encodes α1 subunits of GABAA receptors after auditory nerve damage. They also noted changes in excitatory/inhibitory indices in the auditory cortex and inferior colliculus of denervated mice that tend towards hyperexcitability [36]. Besides, Deng et al. (2020) have examined the occurrence of tinnitus was prevented by the infusion of M-8324, thus increasing the GABAergic inhibitory responses [37].

Table 4: Profile of Neurotransmitter Activity in Central Auditory Pathway

In addition, several studies have been conducted to assess the changes in some neuroinflammatory mediators, particularly following noise exposure and salicylate injection, as provided in Table 5. Furthermore, activation of neuroglia and microglia characterized by the changes in shape from ramified into an activated non-ramified or amoeboid form were also noted in noise-induced and salicylate-induced tinnitus, proving the contribution of neuroinflammation in tinnitus [15,38].

Table 5: Report Study of Inflammation Mediator Profile Following Noise Exposure and Salicylate Injection

Wang et al. found that TNF-α concentrations increased after noise exposure in mice. They also observed alteration in gap detection performance in wild-type, and TNF-α infused mice but not in TNF-α knockout mice. Also demonstrated that mice exposed to noise exposure and TNF- infusion experienced impaired gap detection performance and PPI. However, changes did not occur with exposure to TNF-α or noise alone. In addition, they also noted changes in microglial morphology to non-ramified or activated amoeboid forms after noise exposure and TNF-α infusion, whereas TNF-α knockout mice showed no changes [15,39].

In another study, observed an increase in TNF-α gene expression and NR2A mRNA levels after chronic salicylate injection for seven days continuously in mice, in contrast to IFN-γ gene expression tended to decrease [40]. Meanwhile, explored neuroglial activation in salicylateinduced tinnitus. They found an increased expression of GFAP and Iba1, specific markers for astrocyte and microglia activation, respectively, in cortex A1 and MGB after salicylate treatment. In addition, there was an increase in IL-1β protein levels in the A1 cortex of mice after acute and chronic salicylate injection [38-42].

DISCUSSION

In general, based on the putative mechanisms that have been studied, tinnitus can be divided into cochleartype and central-type. Cochlear-type tinnitus suggests aberrant activity in the peripheral auditory system, which could happen at the pre-cochlear or cochlear nerve level. Spontaneous Otoacoustic Emissions (SOAEs), hair cells damaged induced by noise exposure or ototoxic agents are some factors that contribute to this cochlear mechanism, altering the endocochlear potential to propagate through the auditory pathway to the cortex, generating tinnitus [43]. Meanwhile, the central-type tinnitus hypothesized that tinnitus generation is primarily influenced by the aberrant neural activity in the central auditory system and not by pre-existing cochlear damage [43]. These proposed mechanisms were strengthened by some studies showing that sectioning the auditory nerve bilaterally did not necessarily terminate the noise perception in tinnitus patients. Since then, many studies have been conducted to better understand the central mechanism of tinnitus.

Of this central-type tinnitus, two significant factors have been thought to contribute to the development of tinnitus, specifically the increased central gain about the maladaptive neural plasticity and neuroinflammation. A reduction or loss of auditory input may result in the decrease of inhibitory neurotransmitters. Additionally, the excitatory neurotransmitter increases, causing a shift in excitatoryinhibitory balances [14,44]. Concurrently, neuroinflammation is defined by increased proinflammatory cytokines and microglial activation.

The Microbiota-Gut-Brain Axis and Neurotransmitter Production: The human gut microbiota refers to the community of trillions of commensal and symbiotic microorganisms living in our intestines. Gram-positive bacteria usually dominate the normal gut, and the most dominant bacterial phyla in the mammalian gut are Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria [25,45,46]. Maintaining this gut microbiota balance is essential to the whole body’s health. Numerous studies have shown that gut microbial community alterations may contribute to several diseases pathogenesis; among these are inflammatory bowel disease [47,48], obesity [49], multiple sclerosis [50], and neurodegenerative or neuropsychiatric disorders such as Parkinson’s [29], Alzheimer’s, [28] schizophrenia, 26 autism spectrum disorders [51], and depression [16].

Bidirectional communication exists between gut microbiota and CNS via the gut-brain axis. The bottomup communication is thought to involve mainly the metabolic, immune, endocrine, and neural systems via the autonomic vagus nerve. [32,45,52]. Bacterial metabolites, i.e. short-chain fatty acids (SCFAs), tryptophan metabolites, and secondary bile acids (2BAs), are mediators for this communication. These metabolites propagate signals mainly by interacting with Enterochromaffin Cells (ECCs), Enteroendocrine Cells (EECs), and the mucosal immune system. However, some can cross the intestinal barrier and enter systemic circulation or even cross the bloodbrain border [52,53].

Moreover, recent studies have shown that gut microbiota also plays a role in producing neuroactive molecules, either by directly producing them from the precursor or indirectly by stimulating the cells to produce neurotransmitters, thereby contributing to gut-brain signaling. Bifidobacterium, Bacteroides, Parabacteroides, and Eubacterium showed the ability to produce neurotransmitter GABA from the precursor glutamate or succinate. Besides, Lactobacillus have been shown to produce acetylcholine and tyramine [18-21] Additionally, Bifidobacterium and Clostridium can stimulate the 5-HT production by enterochromaffin cells, and it is estimated that around 90% of the serotonin needed for CNS function is produced by gut microbiota [22,23,46].

Gut Microbiota Dysbiosis and Neuroinflammation: One of the most important pathways for gut-brain communication is the neuroimmune system. There is increasing evidence that gut microbiota can modulate brain immune function through the production of inflammatory cytokines, reactive oxygen species, and microglia maturation and activation [54]. A study on germfree (GF) mice and specific-pathogen free (SPF) mice showed that the microglia mRNA genes profiles and the CNS histology in GF and SPF mice were markedly different. GF mice demonstrated a lower level of genes needed for cell activation and type I IFN receptor signaling. Furthermore, microglia in GF mice failed to evolve to their activated shape when induced by LPS injection, and the innate immune response was also found to be impaired in GF mice compared to SPF mice [55].

Another study showed a correlation between the abundance of bacteria genus Bacteroides and Verrucomicrobia. They increased plasma levels of TNF-α and IFN-γ in humans with Parkinson’s disease (PD) compared to the healthy control [29]. A Parkinson’s disease mice model also showed the gut microbiota involvement in modulating inflammatory cytokines, particularly IL-6 and microglia activation. Mice given PD-derived microbial donors developed an increased motor dysfunction and microglial activation compared to GF mice. Furthermore, bacterial metabolite SCFAs also showed the same results [56]. Hence, these results demonstrated the importance of gut microbiota in neuroinflammation and neurodegenerative disease.

In addition, numerous studies have provided evidence of the effect of intestinal microbiota dysbiosis on the integrity of the host epithelial barrier. For example, environmental factors such as infections, high-fat diet and antibiotics can cause dysbiosis characterized by reduced symbiotic microbiota and loss of essential metabolites; these can induce a pro-inflammatory response and intestinal leakage due to impaired intestinal barrier integrity [57,58]. Another study showed that the BBB permeability of mice lacking a normal gut microbiota was increased. This may be caused by the decreased expression of occludin and claudin-5, which is the transmembrane tight junction protein. Moreover, recolonization with more complex microbiota and SCFAs producing bacteria improved the BBB integrity [54,59]. Taken together, this gut dysbiosis, intestinal epithelial barrier disruption, neuroinflammation, and loss of blood-brain barrier integrity may contribute to the pathogenesis of several neurological diseases.

The Potential Role of Gut Dysbiosis in Inducing Tinnitus:

The implications of gut dysbiosis on neurotransmitters production and neuroinflammation described above may also indicate the role of gut microbiota dysbiosis in inducing tinnitus. Environmental factors e.g. diet, infections and antibiotics may alter the beneficial microbiota needed for neurotransmitter production, causing the shift in neurotransmitter balance [25,32]. It has been previously described that the presence of tinnitus is related to the decrease in inhibitory neurotransmitters such as GABA and an increase in excitatory neurotransmitters44. Besides, the increased GABAergic inhibitory neurons and decreased excitatory responses have been successfully reported to prevent tinnitus, proving the role of neurotransmitter modulation in tinnitus [37].

In addition, any factors triggering the gut dysbiosis may also induce epithelial barrier dysfunction and microbial adhesion, which further elicits proinflammatory responses [57]. This increased proinflammatory cytokines and activation of microglia, the brain defense mechanism, together with the presence of other risk factors such as noise exposure and excess salicylate administration, then causing the behavioral evidence of tinnitus [15,39,40].

Conclusion

In conclusion, neurotransmitter production and neuroinflammatory modulatory responses are likely to be critical factors in the development of tinnitus. Meanwhile, the gut microbiota’s ability to build a complex cross-talk with their hosts to create the gut-brain axis is crucial for regulating the neuroendocrine system and maintaining immune homeostasis. However, microbial composition changes or dysbiosis can interfere with neurotransmitter production, causing hyperexcitability; and modulating neuroinflammatory responses through activation of microglia and several proinflammatory cytokines. Therefore, gut dysbiosis is likely to occur in tinnitus, inducing sound perception by altering those factors. Disruption of the gut barrier and BBB integrity may also contribute to this process, thus facilitating some bacterial metabolites and neuroactive molecules to reach the brain, altering its function and causing tinnitus.