Electrocochleographic Changes Predict an Early Sign of Cochlear Degeneration

ABSTRACT

Objectives

The purpose of this study is to identify the earliest appearing auditory electrophysiological indicators that change with age progression in young adults with normal hearing, and to analyze the frequency distribution patterns of these markers in the cochlear.

Methods

In this study, 74 participants with normal hearing were divided into four groups: aged 18–25, 26–30, 31–34, and 35–40 years for statistical purposes. Electrocochleography (EcochG), transient evoked otoacoustic emissions (TEOAE), and Hearing in Noise Test (HINT) were used.

Results

Our study found: (1) EcochG action potential (AP) and summating potential (SP) amplitude of the left ear decreased after 25 years except at 6 kHz. (2) The strength of EcochG AP amplitude peaks at 4 kHz. (3) The EcochG SP/AP has no statistically significant difference between different age groups, and there is no characteristic distribution of frequencies. (4) TEOAE amplitude, TEOAE contralateral acoustic stimulation (CS) amplitude has no statistically significant difference between different age groups. The strength of TEOAE amplitude peaks at 1–2 kHz. (5) There are no significant differences in the HINT among the various age groups. (6) The auditory function of the right ear declines more slowly than that of the left ear.

Conclusion

The study's findings reveal several key insights: (1) The EcochG AP amplitude is the most sensitive electrophysiological indicator of cochlear degeneration. (2) Cochlear electrophysiologic testing exhibits distinct frequency distribution characteristics. (3) There is an inconsistent rate of electrophysiological change between the two ears.

Summary

Significant Findings of the Study

• The EcochG AP amplitude is a sensitive indicator for predicting the early sign of cochlear degeneration; an inconsistent rate of decay of electrophysiological change in both ears.

What This Study Adds

• The results indicate an early decrease in EcochG AP amplitude, but it is not related to pure‐tone thresholds or speech discrimination ability in noise.

1. Introduction

The World Health Organization (WHO) estimates that approximately half of the global population is susceptible to developing hearing loss as a result of unsafe listening habits [1]. Furthermore, over half of individuals between the ages of 12 and 35 are frequently exposed to harmful sound levels, whether through the use of personal listening devices or participation in social activities that involve loud noise [2].

Indeed, approximately 10% of patients visiting a hearing outpatient clinic report difficulties with speech perception in noise, yet remain untreated. This is because they exhibit no noticeable changes in their hearing thresholds, which makes it challenging to identify the specific nature of their auditory difficulties [2].

Recent studies have demonstrated that cochlear synaptic lesions emerge earlier than hair cell damage, which may represent the earliest signs of cochlear degeneration [3]. The silencing of these synapses impairs auditory processing and may lead to challenges in comprehending speech amidst background noise. This phenomenon has been termed “hidden hearing loss” (HHL) [4].”

Animal studies show that auditory brainstem response (ABR) Wave I is consistent with cochlear afferent synaptopathy [5, 6]; empirical evidence linking HHL and ABR Wave I amplitude changes in humans with normal hearing threshold remains a challenge. Several studies have reported changes in different metrics that appear to support noise‐exposure‐related cochlear synaptopathy [7]. Furthermore, ABR Wave I amplitude can exhibit considerable variation among individuals due to a multitude of factors, including sex, head size, and electrode contact impedance. To mitigate the substantial inter‐subject variability in ABR Wave I amplitudes, the Electrocochleography (EcochG) is often utilized in the clinical diagnosis of certain cochlear pathologies [8]. EcochG is an electrophysiologic testing that measures cochlear summating potential (SP), which arises from hair cells (outer hair cells [OHCs] and inner hair cells [IHCs]), and action potential (AP), which is equivalent to the ABR I wave produced by Type I cochlear nerve [9]. Liberman et al. observed an enhanced SP/AP ratio in the noise exposure group [7], suggesting that the SP/AP can serve as an indicator to assess synaptic damage [10].

Previous studies have primarily concentrated on afferent nerve synapses, while recent research has revealed that efferent nerves also play a role in HHL [11]. Cochlear synaptopathy is defined by the degeneration of connections between cochlear Type I afferent nerve fibers (ANFs) and IHCs, as well as the de‐efferentation between medial olivocochlear complex (MOC) terminals and OHCs. As a result, gaining insight into cochlear synaptopathy can be achieved by evaluating the auditory feedback reflexes, particularly focusing on the MOC efferent reflex (MOCR) [12].

The primary objective of this study is to evaluate hypotheses related to age‐associated cochlear degeneration in humans. The research objectives encompass two key areas: (1) to identify evidence supporting age‐related cochlear degeneration in human subjects and its correlation with speech discrimination ability and (2) to examine the frequency distribution patterns of EcochG and TEOAE.

2. Materials and Methods

2.1. Subject Pool and Inclusion Criteria

A total of 74 healthy individuals, aged 18–40 years, were recruited for this study. All participants had no history of hearing issues, no neurological disorders, and normal otoscopic findings. These participants, all with normal hearing, were divided into four age groups: 18–25, 26–30, 31–34, and 35–40 years old [13].

Participants were required to have absolute hearing thresholds no greater than 25 dB HL across all audiometric frequencies. In addition, they were required to have normal otoscopy and tympanometry results, indicating normal middle‐ear function. Tympanometry was performed using the Titan suite from Otometrics, with a probe‐tone frequency of 226 Hz and an ear‐canal pressure range from −300 to +200 daPa in each ear. The study was reviewed and approved by the Institutional Review Board of Shanghai Jiao Tong University.

All auditory electrophysiological examinations were completed in a soundproof booth with a sound insulation level exceeding 50 dB (SL). The EcochG and TEOAE were performed by one audiologist independently, while pure‐tone audiometry and the Hearing in Noise Test (HINT) were carried out by another audiologist. Both audiologists had received standardized training to ensure consistency and accuracy in their assessments.

2.2. Audiometric Thresholds

Audiometric thresholds were obtained using Otometrics Equinox. Pure‐tone air conduction thresholds were measured at standard audiometric frequencies from 0.25 to 8 kHz (Figure 1).

Figure 1.

Audiometric thresholds, TEOAE, and EcochG diagram in both ears. (A) EcochG: The acoustic stimuli were either 100‐µs clicks delivered at 90 dB nHL at a rate of 7.1 s⁻¹, or tone busts stimuli at 2, 3, 4 kHz (90 dB nHL) or 6 kHz (85 dB nHL). SP and AP are measured from baseline to peak. (B) TEOAE diagram: A linear click sound stimulus (with parameters set to linear) was used at a rate of 19.3 times per second, nonlinear clicks of 80‐μs duration, and a peak‐equivalent sound pressure level (peSPL) of 60 dB. In a quiet environment for the contralateral ear, the SNR and OAE intensity (amplitude) at frequencies of 1.0, 2.0, 3.0, and 4.0 kHz were obtained. RESP stands for Response, which refers to the signal or waveform of the OAE. Noise refers to the background noise level detected in the ear canal during the test. (C) Audiometric thresholds: No significant difference in hearing thresholds in both ears, and have absolute thresholds lower (better) than or equal to 25 dB HL at audiometric frequencies.

2.3. TEOAE

2.3.1. TEOAE (Without Contralateral Acoustic Stimulation)

The procedure was conducted in a standard soundproof booth. Participants were seated comfortably with their hands resting naturally on the sides of their body and were instructed to relax as much as possible. Given that TEOAEs are highly susceptible to self‐noise, participants were required to keep their heads still during the test, with no irrelevant movements or speech allowed. The test room was kept free from distractions caused by other people or unrelated events. The tester needed to have a clear view of the participant's behavior, while the participant should not be able to see the tester's actions, test results, or the instrument display.

First, the daily calibration of the OAE instrument was performed using a 1 cm³ calibration cavity. After successful calibration, a properly sized rubber earplug was placed in the subject's external auditory canal, and an insert earphone was placed in the contralateral ear. The probe calibration program was run, and once passed, the test began and OAE were recorded.

Initially, a nonlinear click sound at 80 dB was used to see if TEOAE could be elicited, and the signal‐to‐noise ratio (SNR) was recorded. Subsequently, a linear click sound stimulus (with parameters set to linear) was used at a rate of 19.3 times per second, nonlinear clicks of 80‐μs duration, and a peak‐equivalent sound pressure level (peSPL) of 60 dB. In a quiet environment for the contralateral ear, the SNR and OAE intensity (amplitude) at frequencies of 1.0, 2.0, 3.0, and 4.0 kHz were obtained, with 2080 averages (Otometrics, Denmark). SNR refers to the ratio of the power of the OAE signal to the power of the background noise. In OAE testing, it is used to measure the strength of the detected OAE signal relative to the background noise. It is usually expressed in decibels (dB), and the calculation formula is SNR (dB) = 10log 10 (P _noise P _signal), where P _signal is the power of the OAE signal and P _noise is the power of the background noise. In OAE testing, SNR is a key indicator for determining whether OAE are normal. Generally, an SNR of ≥ 3 dB is considered detectable for OAE (Figure 1). TEOAE SNR and amplitude provide an objective, rapid, and independent measure of cochlear amplifier function [14].

2.3.2. Contralateral Suppression of TEOAE

Without changing the earplug status of the subject, TEOAE recordings were evoked with 60 dB peSPL linear click stimuli at a rate of 19.3 s⁻¹ under two conditions, with or without a 60 dB SPL contralateral white noise suppressor, by inserting earphones (Etymotic Research Model ER‐3A, USA). Responses were averaged to 2048 sweeps. In the study, contralateral acoustic stimulation (CS) amplitude was defined as follows: CS amplitude = TEOAE AP without contralateral acoustic stimulation − TEOAE AP with contralateral acoustic stimulation. The intensity of the contralateral stimulation was less than what was presented to the ear in which OAE were being recorded, and less than what was needed to create the middle‐ear reflex.

2.4. EcochG

Stimuli were generated by Intelligent Hearing System Inc. (IHS4225O.US), stimulus waveforms were transduced via ER‐3A insert earphones, and data acquisition was processed by the Interacoustics Eclipse computer system. EcochG was measured in both ears, and was evoked by clicks acoustic stimuli or tone busts (2, 3, 4, or 6 kHz) in either ear.

The ear canals were prepared by gently scrubbing with a cotton swab. Electrode placement was consistently positioned at the junction of the external auditory canal and the tympanic membrane. Electrode gel was applied to the silver ball electrode before insertion. The ground electrode was placed at the midline of the forehead, with one tiptrode serving as the inverting electrode and the other as the non‐inverting electrode in the contralateral ear. The impedance between pairs of electrodes was < 3 kΩ, and the acoustic stimuli were either 100‐µs clicks delivered at 90 dB nHL at a rate of 7.1 s⁻¹ or tone busts stimuli at 2, 3, 4 kHz (90 dB nHL) or 6 kHz (85 dB nHL). Electrical responses were amplified 100,000× with a 10–3000 Hz passband filter. Up to 512 sweeps were averaged. The SP and AP peaks were defined as the difference between baseline and peak from 1 to 2 ms post onset by two observers blinded to all test results. The time from the onset of the stimulus to the appearance of the peak of the AP wave is called the latency (Figure 1).

2.5. Hearing in Noise Test (HINT)

The HINT is an assessment of speech recognition in noise, designed to mimic real‐life listening scenarios, and is available in multiple languages, including Standard Chinese. The Standard Chinese version of the HINT (MHINT) was evaluated using the BLIMP software system, a collaborative development between Beijing Tongren Hospital affiliated with Capital Medical University in China and the House Ear Institute in the United States. The MHINT consists of 12 lists, each containing 20 sentences. Each sentence within the MHINT comprises 10 monosyllabic words, carefully constructed to be brief, phonetically balanced, comprehensible, and uniformly challenging. The test stimuli were presented at an intensity of 65 dB SPL and were delivered via Sennheiser HD580 headphones in a monaural fashion. Participants were required to repeat each sentence, after which the audiologist would indicate correctness and proceed to the subsequent sentence. The test's intensity automatically adjusts based on the participant's accuracy. As an adaptive test, the MHINT measures the reception threshold for sentences in noise, employing an adaptive protocol to ascertain the SNR at which the participant achieves 100% sentence recognition. Upon completion of the 20 sentences, the BLIMP test software calculates the final SNR. The test commences with an initial SNR set at 0 dB, and includes two preliminary sentences for adaptation before the formal test. Each ear is tested individually, with the constraint that the vocabulary used cannot be repeated across lists.

2.6. Statistics

The outcome measures were ear‐specific, so there are two measures per subject. The following measures were considered as predictors: (1) TEOAE amplitude; (2) amplitude of TEOAE CS; (3) EcochG SP amplitude, AP amplitude, AP latency, and SP/AP ratio; and (4) MHINT.

The differences in each indicator across age groups were analyzed using a two‐way ANOVA. Bonferroni's multiple comparisons test was subsequently applied to correct for multiple testing. This analysis was conducted using SAS software version 9.4. Additionally, the correlations between TEOAE amplitude and the amplitude of TEOAE CS were assessed through linear regression analysis. All figures were produced by Graphpad Prism 7.0. For all statistical analysis, in the figures, the error bar represents the standard error of mean (SEM), NS represents p > 0.05, *p < 0.05, **p < 0.01.

3. Results

3.1. TEOAE

3.1.1. TEOAE Amplitude in Quiet (Without Contralateral Acoustic Stimulation)

The TEOAE amplitude in quiet of both ears did not show a significant decrease with age (p > 0.05, Figure 2), although a decline trend was observed. Specifically, the amplitude was highest in the 18–25‐year‐old group and gradually decreased in the subsequent three age groups. The trend was consistent across frequencies of 1, 2, 3, and 4 kHz, thereby validating the reliability of the results.

Figure 2.

Correlation between TEOAE amplitude, TEOAE SNR, TEOAE CS amplitude, and age. (A) The SNR of TEOAE at the left ear exhibited a declining trend across all age groups, as indicated by the histograms. However, this trend did not reach statistical significance (p > 0.05). The amplitude reached its peak in the 18–25‐year‐old group and subsequently decreased progressively in the following three groups. However, these decreases appeared to plateau, indicating stabilization. Moreover, this trend was consistently observed across all tested frequencies of 1, 2, 3, and 4 kHz. (B) The amplitude of TEOAE at the left ear also demonstrated a downward trend across all age groups, according to the histograms. Nonetheless, this trend was not statistically significant (p > 0.05). The changes in TEOAE amplitude were essentially consistent with those of the SNR. (C) The amplitude of TEOAE CS at the left ear displayed a similar declining trend across all age groups, as shown by the histograms. Despite this, the trend did not achieve statistical significance (p > 0.05). The changes in the amplitude of TEOAE CS were essentially consistent with those of the TEOAE amplitude. (D) The SNR of TEOAE at the right ear revealed a deteriorating trend among all age groups, based on the histograms. However, this trend was not statistically significant (p > 0.05). The amplitude was highest in the 18–25‐year‐old group and showed a gradual downward trend in the subsequent three groups. However, the changes tended to stabilize, and the trends were consistent across 1, 2, 3, and 4 kHz. (E) The amplitude of TEOAE at the right ear showed a similar declining trend across all age groups, according to the histograms. Yet, this trend did not reach statistical significance (p > 0.05). The changes in TEOAE amplitude were essentially consistent with those of the SNR. (F) The amplitude of TEOAE with CS at the right ear also exhibited a downward trend among all age groups, as indicated by the histograms. But, this trend was not statistically significant (p > 0.05). The variations in TEOAE CS amplitude were fundamentally aligned with those observed in TEOAE amplitude.

Additionally, the strength of TEOAE amplitude and SNR peaked at 1–2 kHz, showing significant difference from other higher frequencies (p < 0.05, Figure 3).

Figure 3.

Frequency distribution characteristics of TEOAE amplitude, SNR, TEOAE CS, EcochG AP amplitude, AP latency, SP amplitude, and SA. (A) The amplitude of TEOAE was highest at 1–2 kHz. In both ears, there was a significant difference between 1 kHz and other frequencies. The 2 kHz region was the second‐highest peak for AP amplitude, showing statistically significant differences compared to the other three groups (**p < 0.01; *p < 0.05). (B) The SNR of TEOAE was highest at 1–3 kHz. In both ears, there was a significant difference between 4 kHz and other frequencies (**p < 0.01). (C) The amplitude of TEOAE CS was highest at 1–2 kHz. In both ears, there was a significant difference between 1 and 2 kHz and other frequencies (**p < 0.01; *p < 0.05). (D) The amplitude of EcochG AP was highest at 3–4 kHz. In both ears, significant differences were observed between 4, 2, and 6 kHz (*p < 0.05). (E) The amplitude of EcochG SP showed no significant differences in either ear (p > 0.05). (F) The latency of EcochG AP was longest at 2 kHz. In both ears, there were significant differences between 2 kHz and 3, 4, and 6 kHz (**p < 0.01). (G) The EcochG SA showed no significant differences in either ear (p > 0.05).

3.1.2. TEOAE Contralateral Acoustic Stimulation (CS)

The decline trend of TEOAE CS amplitude is consistent with TEOAE amplitude under the stimulation of 1, 2, 3, and 4 kHz frequencies. Specifically, the amplitude was highest in the 18–25‐year‐old group and showed a gradual downward trend in the subsequent three groups. Notably, the differences between the four age groups were not significant for either ear (p > 0.05, Figure 2).

Consistent with TEOAE amplitude, the strength of TEOAE CS amplitude also peaked at 1–2 kHz, showing significant differences from other higher frequencies (p < 0.05, Figure 3). Additionally, this study also analyzed the correlation between TEOAE CS and TEOAE, for the left ear, TEOAE CS amplitude was linearly related to the TEOAE amplitude at 1 and 3 kHz (p < 0.05, Figure 4). For the right ear, TEOAE CS amplitude was linearly related to the TEOAE amplitude at 1, 2, 3, and 4 kHz (p < 0.05, Figure 4).

Figure 4.

Correlation between TEOAE amplitude and TEOAE CS amplitude in the both ears. Left ear: (A, C) The correlation between TEOAE amplitude and TEOAE CS amplitude of 1 and 3 kHz is statistically significant (**p < 0.01). (B, D) The correlation between TEOAE amplitude and TEOAE CS amplitude of 2 and 4 kHz is not statistically significant (p > 0.05). Right ear: (E–H) The correlation between TEOAE amplitude and TEOAE CS amplitude of 1, 2, 3, and 4 kHz is statistically significant (1 and 2 kHz *p < 0.05; 3 and 4 kHz **p < 0.01).

3.2. EcochG

3.2.1. EcochG AP Amplitude

For EcochG AP amplitude in the left ear, there were statistically significant differences between the 18‐25 group and other age groups (p < 0.05, Figure 5), except for 6 kHz. Under click and tone burst stimuli at 2, 3, and 4 kHz, the EcochG AP amplitude was highest in the 18–25‐year‐old group and showed a downward trend in subsequent age groups, although there were fluctuations across the different groups.

Figure 5.

Correlation between EcochG AP amplitude, AP latency, SP amplitude, SA, and age in both ears. (A, C) The EcochG AP and SP amplitude of the left ear decreased after 25 years, and showed a significant correlation with age, except 6 kHz (*p < 0.05; **p < 0.01); under click and tone burst stimuli at 2, 3, and 4 kHz, the EcochG AP amplitude was highest in the 18–25‐year‐old group and showed a downward trend in subsequent age groups, although there were fluctuations across the different groups. (B, D) The correlation between AP latency and SA of the left ear and age is not statistically significant (p > 0.05). (E–H) The correlation between EcochG AP amplitude, AP latency, SP amplitude, and SA in the right ear and age is not statistically significant (p > 0.05).

The EcochG AP amplitude in the right ear exhibited fluctuations across different age groups, generally trending downward. However, these changes did not reach statistical significance (p > 0.05, Figure 5).

The strength of EcochG AP amplitude reached its peak at 4 kHz, with significant differences compared to the 2 and 6 kHz frequencies (p < 0.05, Figure 3). At 2 kHz, the EcochG AP amplitude was the lowest, corresponding to the longest latency (Figure 3).

3.2.2. EcochG AP Latency

The EcochG AP latency remained stable across different age groups. The correlation between latency in both ears and age was not statistically significant (p > 0.05, Figure 5). Notably, the latency peaked at 2 kHz, consistent with the findings for EcochG AP amplitude (**p < 0.01, Figure 3).

3.2.3. EcochG SP Amplitude

For the EcochG SP amplitude in the left ear, a statistically difference was observed between the 18–25 group and other age groups, aligning with the downward trend seen in EcochG AP amplitude (*p < 0.05, Figure 5).

In contrast, EcochG SP amplitude of the right ear exhibited fluctuations across different age groups, yet no significant differences were observed (p > 0.05, Figure 5). For both ears, the frequency distribution characteristics of SP mirrored those of AP, although the correlation did not reach statistical significance (Figure 3).

3.2.4. EcochG SP/AP Ratio

The SP/AP ratio exhibited no statistically significant correlation with age for either ear (p > 0.05, Figure 5). This ratio was uniformly distributed across all frequencies, with no frequency characterization (Figure 3).

3.3. MHINT SNR (dB)

The thresholds are presented in terms of SNR (dB). There are no significant differences in the SNR of the HINT between the left and right ears, nor among the various age groups (p > 0.05, Figure 6).

Figure 6.

Correlation between MHINT SNR and age. There are no differences in the HINT SNR among the groups (p > 0.05).

4. Discussion

This study identified an early decline in the EcochG AP. Both human and animal experiments suggest that the reduction in the amplitude of the EcochG AP and SP indicates a decrease in the number of cochlear afferent nerve synapses [6, 7, 15]. Therefore, it is hypothesized that this finding may imply an early reduction in cochlear synapses in the human population.

The pathological process of cochlear synaptopathy precedes permanent changes in the auditory threshold [16]. The predominant clinical manifestation of cochlear synaptopathy is an impaired ability to comprehend speech in noisy environments [16]. Histopathological studies have demonstrated that cochlear synaptopathy is a prevalent feature in age‐related hearing loss in humans [17]. Aging, noise, and ototoxic drugs are potential etiological factors. In our study, we recruited young, healthy subjects to investigate the frequency distribution characteristics of afferent and efferent neural functions, as well as their age‐related decline. Surprisingly, we observed the onset of functional decline in EcochG AP and SP after the age of 25. Moreover, the frequency distribution observed through electrophysiological examinations differs from the histological patterns observed in humans.

4.1. The EcochG AP Amplitude Decline With Age

In animal studies, cochlear synaptopathy is diagnosed through the observed decline in the supra‐threshold amplitude of ABR Wave I. The cochlear neurons that are most vulnerable to both noise‐induced and aging‐related degeneration are those with high thresholds and low spontaneous rates (SRs) [18]. These low‐SR ANFs play a crucial role in the neural coding of transient stimuli, particularly in noisy environments, and their loss does not impact hearing threshold. In humans, the diagnostic utility of ABR Wave I or EcochG AP for identifying cochlear synaptopathy is a matter of debate [16, 19]. There is a significant inter‐subject variability, attributed to factors such as sex, head size, and tissue conductivity heterogeneity [7]. Additionally, selecting a specific age group (e.g., 18–40 years) can be an effective method to reduce individual differences in EcochG measurements. Age‐related changes in auditory function are known to affect ECochG. By focusing on a relatively narrow age range, such as 18–40 years, we can minimize variability caused by age‐related factors.

To evaluate cochlear neural function, we recorded click and tone‐burst‐evoked potentials using ear‐canal electrodes and subsequently extracted the amplitudes of both the SP and AP. Animal studies have established that the deterioration of AP typically initiates at higher frequencies [20]. In our study, long‐term accumulation of noise damage could account for the significantly diminished AP amplitudes observed at 2, 3, and 4 kHz. However, the correlation between EcochG AP amplitude at 6 kHz and age of both ears was not statistically significant. These findings suggest that aging is not a predominant factor in the decline of AP amplitude among individuals aged 26 and above.

Additionally, the observed group differences were primarily between the 18–25 age group and the older cohorts, with no significant changes in AP amplitude noted after the age 25. This presents an intriguing yet perplexing phenomenon that is not supported by existing literature. Could it be that after the most vulnerable low‐SR ANFs are diminished, the remaining ANFs become less susceptible to damage? Alternatively, might the decline in AP amplitude beyond the age of 26 be attributed to cochlear toughening? These hypotheses await confirmation through future experimental investigations.

4.2. Frequency Distribution Characteristics of EcochG AP Amplitude

In our study, the maximal activation region of Type I ANF peaked in the upper basal turn of cochlear (near 4 kHz) tested by EcochG AP, which aligns with findings from other studies [21]. However, it is inconsistent with histological analyses in humans [22, 23]. In younger individuals, the peripheral axon density of ANFs is highest in the middle of the cochlea (0.5–2 kHz), with lower densities towards both the base and apex. This density declines with age [22, 23], corresponding to a frequency range one to two octaves lower than the ANF strength assessed by EcochG. But in animal studies, the decline of ABR I wave AP matches with the decrease in ANF counts [6, 11, 24]. This discrepancy may be explained by difference in the distributions of lateral olivocochlear (LOC) in humans. Confocal images from human histological analyses suggest an apical–basal gradient of innervation density for the LOC system. The density of LOC terminals peaks in the apical turn, 0.5–2 kHz in younger adults [15]. These terminals are quantified by measuring areas in the choline acetyltransferase (ChAT) channel, which is the primary inhibitory neurotransmitter in LOC. In mammals, such as cats and mice, LOC terminal density is a relatively uniform projection from apex to base [25, 26]. Howerver, this remains speculative, as the LOC is less studied due to its complexity function. Another possible explanation is the EcochG test in humans may not be sufficiently sensitive to maximize the activation of ANF, thus limiting its ability to accurately assess ANF function.

4.3. Frequency Distribution Characteristics of EcochG AP Latency

The AP latency represents the rate of excitatory conduction of ANF. The duration of this latency is closely associated with several factors, including the conduction velocity of the neuron, the distance between the stimulation site and the recording site, the number of neuronal transitions in the conduction pathway, and the synaptic transmission time. Additionally, it is influenced by individual variations such as age, sex, height, stimulus intensity, skin temperature, and other factors. In our study, the frequency‐specific characteristics of the AP latency align with those of the AP amplitude. Specifically, nerve conduction in 2 kHz region is slower compared to other frequencies. However, there was no prolongation of latency with increasing age, suggesting that early aging does not impact the conduction velocity of cochlear nerve. This indicates that the cochlear nerve conduction velocity is less sensitive to age‐related changes than the AP amplitude.

4.4. The EcochG SP Amplitude Decline With Age

As is known, the EcochG SP is primarily composed of pre‐synaptic potentials from hair cells. The SP, which is observed as a shoulder on the leading edge of the Wave I peak, is predominantly influenced by contributions from the IHC receptor potentials. This represents the capacity for mechanoelectric transduction by the hair cells [27]. Recent electrophysiologic studies have identified the SP amplitude as a significant predictor of CS [28, 29]. In this study, the SP amplitude of the left ear in the 18–25 group was substantially higher compared to other three age groups, with statistically significant difference. For the right ear, the SP amplitude in the 18–25 age group was slightly higher than in the other three age groups, although this difference was not statistically significant. For both ears, the SP amplitude varied across the subsequent three groups, which may still be related to individual differences [30]. Previous animal and human studies have noted the stability of SP amplitude despite the decline of AP [6, 7, 31]. In our study, the SP amplitude exhibited the same frequency distribution characteristics as the AP wave, but without significant differences. It is probably because the generators of SP and AP are physically close, but the SP amplitude is smaller [29].

4.5. No Significant Change in EcochG SP/AP

In our study, individual SP/AP ratios were also computed. There were no significant differences in the SP/AP ratios across the four tested frequencies, and no significant change with age progression. SP/AP eliminates the frequency characterization of AP amplitude. The result proposes the stability of SA, and is more suitable for the diagnosis of pathological changes of cochlear, such as endolymphatic hydrops [8]. The stability of SA suggests that it is less susceptible to electrode position and individual differences interference, making it a better indicator for cochlear lesion evaluation [7].

Additionally, our study is constrained by a limited sample size, attributable to the extended duration required for each testing procedure. The absence of significant correlations between the SP/AP ratio and other variables, such as age and frequency, in our study could potentially be attributed to the small sample size or the youthful demographic of our subjects.

4.6. Frequency Distribution Characteristics of TEOAE and TEOAE CS Amplitude

The mammalian cochlea is innervated by two cholinergic feedback systems, known as the MOC and LOC pathways, which transmit regulatory signals from the brainstem back to the OHCs and auditory‐nerve fibers. These MOC and LOC systems play a crucial role in protecting the ear from acoustic overstimulation [11], combating the effects of aging [32], and are essential for normal cochlear development [33].

In animals and humans, the MOC reflex can be evoked by sound. Thus, its strength can be assessed by measuring the suppression of ipsilateral thresholds by a contralateral sound [34]. In human studies, contralateral sound suppression typically peaks at 1–2.8 kHz [9], which is two octaves lower than the peak MOC density (4 kHz) in human temporal bones [15]. In our study, the strength of the MOC reflex peaked at 1–2 kHz, which is consistent with previous reports. Liberman et al. once suggested the routine testing of MOC reflexes in human may not be able to activate the maximal reflex activation of MOC [15]. In cats and guinea pigs, MOC innervation peaks in the 5–10 kHz region [35, 36], when MOC activity is evoked by electrophysiological stimulation of the OC bundle. Maximal effects of MOC occur at 5–10 kHz [37]. Liberman has proposed that full activation of the peak density requires descending control from higher centers, superimposed on the brainstem reflex [15, 38].

We propose that there may be additional alternative explanation. In our study, the peak of TEOAE AP was also observed in the lower frequencies band (1–2 kHz), aligning with the distribution of TEOAE CS AP. The correlation between TEOAE CS AP and TEOAE AP was found to be significant in both ears. Others have also reported the consistency in the frequency distributions of both [9, 39]. Our results suggest that the strength of the MOC reflex is contingent upon the functionality of the OHC.

4.7. No Significant Change in TEOAE and TEOAE CS Amplitude Across Different Age Groups

Although TEOAE amplitude and TEOAE CS amplitude showed a downward trend across different age groups, no significant differences were observed. The result suggests the decompensation of afferent synapses might occur before efferent synapses, or an alternative explanation is needed, that the EcochG test is more sensitive than TEOAE. Expanding the sample size in subsequent studies may reveal statistically significant differences.

4.8. The Discrepancy Between Left and Right Ear

The issue of asymmetrical hearing loss remains a subject of debate and is not fully understood. Previous studies have found the average hearing thresholds across 1–2–3 kHz are significantly poorer in the left ear [40]. Prior research has established that noise‐induced hearing loss (NIHL) can induce or exacerbate asymmetry between the right and left ears, with the left ear often exhibiting higher air‐conduction thresholds [41]. Factors such as the head‐shadow effect and the lateralization of sound sources may contribute to this asymmetry, as observed in drivers who drive with open windows [42, 43]. Anatomical factors also play a role. Previous studies have found evidence of functional lateralization in the human brain. The left hemisphere predominantly handles language processing and production in most right‐handed people, while the right hemisphere is more involved in spatial and non‐verbal information processing [44, 45]. Research has consistently demonstrated a strong correlation between the right‐ear advantage and the ability to recognize speech in noisy environments among adults [43]. Furthermore, studies have demonstrated that the right ear possesses a stronger medial olivocochlear (MOC) efferent system compared to the left ear [40], which may confer greater resistance to noise [42, 43]. It has been observed that the MOC is more effective in the right ear than in the left for right‐handed individuals, whereas it functions symmetrically in left‐handed individuals aged between 18 and 34 years [46]. Consequently, the right ear has traditionally been considered the dominant auditory pathway [44]. In this study, all participants were right‐handed, which may account for the lack of significant differences in the amplitude decline of the EcochG AP and SP in the right ear.

4.9. Decrease in EcochG AP Amplitude and Its Lack of Effect on HINT

This study suggests that there is no significant change in the MHINT between the ages of 18 and 40, indicating that a decrease in the amplitude of EcochG AP has no apparent effect on the HINT. Our previous research found that the SNR of the MHINT significantly increased after the age of 40 [47]. The decrease in EcochG AP amplitude did not lead to an increase in HINT SNR, suggesting that this reduction is not sufficient to affect speech discrimination ability. We did not find evidence linking EcochG to HINT, as has been reported in other studies [48]. It has been proposed that the suprathreshold effects of synaptopathy may extend to auditory perception. Given that synaptopathy initially affects low‐SR (low‐SR) ANFs, Kujawa and Liberman hypothesized that the loss of these fibers might largely account for the wide variability in speech perception in noise (SPiN) abilities among individuals with normal audiograms [3].

Nevertheless, this reasoning hinges on two critical assumptions: first, that synaptopathy preferentially affects low‐SR fibers in humans; and second, that these low‐SR fibers in humans possess high response thresholds. These assumptions, in particular, may not be well‐founded. For instance, Hickox et al. pointed out that the relationship between low‐SR and high‐threshold fibers observed in the auditory nerve (AN) fibers of mice, gerbils, guinea‐pigs, and cats may not apply to primates [49]. In fact, single‐unit recordings from the AN fibers of macaque monkeys have shown no systematic correlation between SR and threshold [50].

Even assuming this reasoning holds true, one point deserves consideration: whether minor damage to low‐SR nerve fibers is insufficient to impair HINT.

Therefore, it is speculated that a certain accumulation of synaptic pathology might be required to affect speech discrimination ability. Another possible explanation is that HINT may not be sensitive enough to detect minor deficits in SPiN.

4.10. This Study has Some Limitations

Our study is limited by a relative small sample size. The primary reason for the smaller sample size in this study is that EcochG testing is time‐consuming. It takes approximately 4 h to complete all the tests on both ears for each individual. Given the exploratory nature of this study, there is no specific method for calculating the sample size. However, we believe that the chosen sample size is adequate for the exploratory aims of this study. The sample size was determined based on previous studies in the field [51, 52].

Although electrophysiological experiments in animals have shown that the EcochG AP amplitude and TEOAE CS are correlated with cochlear synaptic lesions, it is necessary to consider individual differences between subjects such as sensitivity to sound stimulation in human electrophysiological studies. In subsequent studies, we will further increase the sample size to reduce individual variability and enhance the robustness and credibility of the findings.

5. Conclusion

The results suggest that the early decline in cochlear EcochG AP amplitude differed between the left and right ears. This primary neural degeneration does not affect the audiogram and speech discrimination ability; therefore, therapies designed to regrow peripheral axons could provide clinically meaningful improvement in the aged ear. Early intervention is needed, after all, the decline after 25 years. Early intervention for hearing health primarily involves lifestyle modifications. Limit exposure to high‐noise environments and use hearing protection devices such as earplugs or earmuffs when exposure is unavoidable. For example, take precautions when attending rock concerts. Additionally, minimize exposure to ototoxic medications and chemicals that may damage hearing. Manage conditions such as diabetes and high blood pressure, as these can contribute to hearing problems. If early signs of hearing loss are detected, medications that improve inner ear blood flow, reduce oxidative stress and inflammation, and protect neuronal function may be considered.

Author Contributions

Min‐Fei Qian and Hao Chen wrote the manuscript. Ji‐Ping Li and Zhi‐Wu Huang designed and supervised the whole study. Min‐Fei Qian, Qi‐Xuan Wang, Ya‐Qi Zhou, and Xiao‐Lu Chen performed the experiments.

Ethics Statement

The above manuscript is the authors’ own original work, which has not been previously published and is not in submission elsewhere. The paper reflects the authors’ own research and analysis in a truthful and complete manner and properly credits the meaningful contributions of coauthors and coresearchers.

Conflicts of Interest

The authors declare no conflicts of interest.

Data Availability Statement

Data are available on request from the authors.