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Research Article
Erratum Notice
Important: There has been an erratum issued for this article. View Erratum Notice
Retraction Notice
The article Assisted Selection of Biomarkers by Linear Discriminant Analysis Effect Size (LEfSe) in Microbiome Data (10.3791/61715) has been retracted by the journal upon the authors' request due to a conflict regarding the data and methodology. View Retraction Notice
This protocol aims to decode prefrontal alpha-band neural oscillatory reprogramming induced by aerobic exercise in high-trait-anxiety individuals, using EEG-deep learning integration. The developed predictive model (81.82% accuracy) identifies alpha oscillation as the core mechanism for exercise-mediated anxiety alleviation, advancing precision neuromodulation targets for emotional disorders.
Exercise intervention demonstrates unique potential in treating emotional dysregulation, yet the ambiguity of its neuromodulation targets hinders the development of precise exercise prescriptions. This study investigates trait anxiety as a representative emotional disorder in 40 high-trait-anxiety university students, who were randomly assigned to either an exercise intervention group (40 min moderate-intensity aerobic exercise, n = 20) or a non-exercise control group (40 min quiet reading, n = 20), followed by resting EEG data collection. By integrating resting-state electroencephalography (EEG) after exercise with deep learning algorithms, we developed an alpha-band time-frequency predictive model to systematically decode the neural oscillatory reprogramming mechanisms in the prefrontal cortex induced by exercise. The deep learning model exhibited superior classification efficacy (accuracy 83.33%, F1 score 0.83, Kappa coefficient 0.67) in identifying exercise-induced alpha-band power spectral entropy alterations. This study pioneers the identification of prefrontal Alpha excitatory rebalancing through neural oscillation remodeling as the core mechanism underlying exercise-mediated anxiety mitigation.
In contemporary society, the accelerating pace of life and the increasing burden of life pressures have led to a significant upsurge in the prevalence of emotional dysregulation. Among various manifestations of emotional dysregulation, anxiety, a prevalent subtype, poses a great challenge to individuals. Pharmacological therapies have long been regarded as a fundamental approach in the treatment of emotional dysregulation, particularly anxiety. However, research has shown that approximately 30% of individuals with emotional dysregulation do not respond to first-line medications. Moreover, long-term use of these drugs may give rise to various risks, such as metabolic disorders and cognitive impairment1. Psychological interventions, although addressing etiological factors through evidence-based frameworks, are limited by prolonged treatment durations requiring substantial time, effort, and financial resources, alongside the delayed onset of therapeutic effects2,3.
In recent years, exercise intervention has been demonstrating remarkable advantages in the treatment of emotional dysregulation. A multitude of studies have indicated that exercise has the potential to naturally enhance emotional states and alleviate anxiety and depression, achieved through the promotion of endogenous neurotransmitter release and the induction of synaptic changes4. For example, research on exercise-trained mice revealed that their hypoxic burden was reduced by 52%, and a significant enhancement in cognitive function was observed5. Trait anxiety, which represents an individual's relatively stable and long-lasting tendency to experience anxiety across diverse situations6, is a key factor in understanding the underlying mechanisms of emotional dysregulation. It serves as a core feature of chronic anxiety, and studying it can provide valuable insights into the pathophysiology of such emotional dysregulation. By understanding trait anxiety, we can better comprehend why some individuals are more prone to developing anxiety-related mood problems. In our previous work, we elaborated on the main brain regions related to emotional cognitive functions that are impaired in emotional disorders and how exercise intervention can improve these cognitive functions and relevant brain regions7. Additionally, we conducted two electroencephalogram (EEG) experiments to explore in detail how exercise intervention can improve the characteristics of brain activity in attention control ability among individuals with high-trait anxiety8.
While exercise intervention has emerged as a promising non-pharmacological approach in depression treatment, the precise neural biomarkers associated with the positive effects of exercise intervention have not yet been clearly identified9,10. Neural oscillatory rhythms, acting as the "spatiotemporal encoders" of brain information processing, exhibit characteristic dysregulation in anxiety. For example, research has shown that prefrontal Alpha(α) desynchronization is associated with cognitive control deficits commonly observed in anxiety11,12. This dysregulation of neural oscillatory rhythms indicates an underlying disruption in the normal neural communication processes that are crucial for emotional regulation. However, there is a dearth of studies that comprehensively explore how exercise actually reshapes emotional function by modulating cross-regional rhythmic coupling or local field potential dynamics13,14.
Recent advances in EEG-based deep learning research have provided novel paradigms for understanding pathological mechanisms and developing precision treatments for mental disorders such as depression and anxiety15. Notably, studies using dynamic functional connectivity (DFC) of resting-state EEG combined with hidden Markov models (HMMs) have revealed significant differences in Delta ( δ), Theta (θ), Alpha (α), and Gamma (γ) band network dynamics among non-psychotic depression, psychotic depression, and schizophrenia16,17,18. A DFC-based binary classification model achieved 73.1% accuracy in distinguishing these three conditions, outperforming traditional static analyses. Key biomarkers included θ-band DMN-SN synchronization, γ-band FPCN-limbic system synchronization, and HMM state transition probabilities, establishing a new framework for precision psychiatric classification19 employed graph theoretical analysis to demonstrate that baseline brain network features predict deep brain stimulation (DBS) efficacy in treatment-resistant depression. A random forest model using network metrics achieved 81.2% accuracy in predicting DBS response, surpassing clinical scales. Longitudinal data showed DBS reverses network dysfunction by enhancing δ-band global synchronization and reducing sgACC centrality. Additionally, left prefrontal α-wave power predicted antidepressant non-response, with a convolutional neural network (CNN) model achieving 82.3% accuracy based on α-asymmetry20. Everaert et al. (2022) developed an artificial neural network model with feature selection using 460 participants to identify predictive features of emotion regulation strategies. These findings underscore the critical need to identify precise neural targets to optimize exercise prescriptions21.
In the realm of exercise-related neuroscience research, deep learning has emerged as a powerful tool, enabling the extraction of robust neural biomarkers from the complex, high-dimensional, and low-amplitude spatiotemporal neurological data generated by exercise interventions. Multiple studies have demonstrated that physical activity significantly modulates activation patterns in motor-related brain regions and neural oscillatory dynamics across frequency bands22,23,24. A systematic review of 47 studies revealed consistent increases in prefrontal α/β band power following exercise, likely reflecting enhanced neuroplasticity and cortical inhibition25. Both acute exercise and long-term training induced similar trends, though γ band responses showed intensity-dependent heterogeneity (e.g., moderate aerobic vs. high-intensity interval training). Four-month aerobic interventions in healthy young adults produced significant prefrontal α wave (9-12 Hz) augmentation, positively correlated with aerobic fitness gains. While behavioral improvements in reaction time or accuracy were absent, neural oscillation metrics indicated dynamic optimization of visual attention networks, suggesting α waves may serve as biomarkers for exercise efficacy26. High-level sport experts exhibited elevated sensorimotor rhythm (SMR, 12-15 Hz) power during aiming tasks, concurrent with reduced prefrontal-temporal coherence, indicating automated motor skill execution and network efficiency enhancement27. Notably, table tennis athletes showed reduced activation in exercise-related brain regions compared to non-athletes, suggesting long-term training builds specialized, energy-efficient neural networks28.
This study focuses on trait anxiety as a specific research subject, employing electroencephalography (EEG) to collect neural data and explore its neural biomarkers, thereby providing novel insights for identifying precise neural targets. Previous research indicates that alpha waves in the prefrontal region are closely associated with emotional regulation, cognitive control, and emotional recognition (Harmon-Jones et al., 2010), playing a pivotal role in processes such as decoding external emotional cues (e.g., facial expressions, vocal tones) and modulating emotional responses. Studies suggest that alterations in prefrontal alpha activity may serve as physiological markers of emotional dysregulation, particularly in anxiety and negative emotional states29,30,31. Resting-state electroencephalography (EEG) serves as a default experimental condition in neuroscience for investigating the dynamic properties of the brain, requiring participants to remain awake without performing any cognitive tasks32. Experimental conditions may include eyes-closed or eyes-open states.Empirical evidence indicates that changes in prefrontal alpha oscillations could function as biomarkers for impaired emotion regulation, especially in conditions characterized by anxiety and a predominance of negative affect33,34. Its power spectral density and functional connectivity patterns can reveal the intrinsic activity characteristics of the brain, and are applicable to detecting pathological markers in neurodegenerative diseases (e.g., Alzheimer's disease), developmental disorders (e.g., developmental dyslexia)35,36, as well as mental and emotional disorders (e.g., depression and anxiety)37. Among these, the alpha rhythm under the eyes-open condition is commonly utilized in studies on emotional disorders38,39. Consequently, this study investigates the classification performance of alpha oscillations in prefrontal regions before and after exercise interventions for trait anxiety. Building on EEG data, this research employs EEGNet to identify neural targets associated with exercise interventions for individuals with high trait anxiety. EEGNet is specifically designed for EEG signal classification and offers several key advantages over traditional and other deep learning methods, making it particularly suitable for investigating EEG patterns with limited data40.
The resting-state EEG data were collected using a 64-channel system (Brain Products, Germany) following the 10-20 international standard, with a sampling rate of 1000 Hz and bandpass filtering (0.1-100 Hz). To ensure signal quality, electrode impedance was maintained below 5 kΩ, and ocular artifacts were removed via Independent Component Analysis (ICA). Participants were instructed to remain awake with eyes open while fixating on a cross, minimizing movement-related noise.
Key inclusion criteria for high-trait-anxiety participants were: (1) Trait Anxiety Inventory scores ≥ 55, (2) limited high-intensity exercise (< 3 days/week) to control for pre-existing fitness effects, and (3) total weekly physical activity < 600 MET-min. These criteria aimed to homogenize the sample while reflecting real-world sedentary populations. A limitation is the potential variability in resting-state EEG dynamics due to individual differences in baseline arousal or undetected subclinical conditions, which future studies could address with larger samples and multimodal assessments (e.g., fMRI or behavioral tasks).
We hypothesize that prefrontal alpha activity can effectively classify the EEG data of exercise and control. In summary, this study aims to leverage AI technologies to analyze the benefits of exercise interventions for emotional disorders, using trait anxiety as a model. Through its methodology and findings, this work seeks to enhance understanding of current developments and challenges in the field, offering guidance and insights for future research.
This study was approved by the Institutional Research Ethics Committee of Wuhan Sports University (2023016).
1. Study participants
2. Task instruction
3. Data collection
4. Offline data analysis
5. Model analysis
NOTE: This convolutional neural network (CNN) achieves the learning of time-frequency features of EEG signals through a multi-scale two-dimensional convolution operation46. The process of the CNN model is shown in Figure 1B.
EEG data processing and statistical analysis
The raw EEG data were segmented into 2 s epochs centered on the event onset, consistent with standard practices in time-frequency analysis to capture transient neural dynamics while minimizing edge artifacts. Each epoch underwent continuous wavelet transformation (CWT) using a complex Morlet wavelet with 3 cycles, which optimally balances temporal and frequency resolution for detecting oscillatory activity in the theta to gamma bands.
The left panel of Figure 2 represents the exercise group, and the right panel represents the Control Group. (1) Data processing quality: Both spectra exhibit smooth curves and the characteristic neurophysiological "1/f" decay pattern (high power at low frequencies decreasing exponentially with frequency). The highly overlapping trajectories indicate effective data preprocessing (e.g., denoising, filtering) and high baseline data quality with good signal fidelity in the frequency domain. (2) Subtle inter-group differences: Within the alpha band (8-12 Hz, shaded gray area for illustration), the control group (right) shows slightly lower power values compared to the exercise group (left), suggesting that a single bout of acute exercise may have induced a mild modulatory effect on the alpha rhythm of resting-state brain oscillations.
For statistical inference, we performed pointwise nonparametric permutation tests (5,000 iterations) across all time-frequency points. This approach controls for multiple comparisons by clustering adjacent significant points (cluster-forming threshold p < 0.05, cluster-level FDR correction), to address the non-Gaussian distribution of wavelet coefficients.
Significant differences in prefrontal electrode activity were observed within the 7-13 Hz frequency band between the exercise and reading groups, as shown in Figure 3.
Validation of CNN model classification performance
In the investigation of the impact of exercise intervention on individuals with high trait anxiety, the classification performance of the Convolutional Neural Network (CNN) model using prefrontal alpha band feature data is a crucial aspect. This analysis aims to determine whether the model can effectively distinguish between the Read group and the Exercise group, thereby providing evidence for the neural-level differences associated with exercise.
The CNN model showed high classification performance when using prefrontal alpha band feature data to discriminate between the Read and Exercise groups, with an accuracy of 83.33%, and achieved an average F1 score of 0.83 and a Kappa coefficient of 0.63. To better understand the model's performance, we turn to the binary classification confusion matrix presented in Figure 3C. In this matrix, a well-structured tool for evaluating classification models, each row represents the true category of the data, and each column represents the category predicted by the model. This layout allows for a detailed assessment of the model's ability to correctly classify different data instances. The model exhibited a relatively good classification performance for both types of data. This high recognition rate implies that the model was able to accurately identify a large proportion of the data belonging to the exercise group. In other words, the neural patterns in the prefrontal alpha band associated with exercise were distinct enough for the model to recognize them with a high degree of certainty. These results from the confusion matrix further support the overall accuracy of the CNN model.

Figure 1: Resting-state EEG acquisition and CNN-based classification workflow. (A) Left side: The recording process of resting-state electroencephalogram (EEG). Right side: The EEG waveforms and the distribution of scalp electrodes. (B) The workflow of using a Convolutional Neural Network (CNN) to classify the alpha waves of two groups. Please click here to view a larger version of this figure.

Figure 2: Power Spectral Density Comparison Between Exercise and Control Groups. Left panel: the Exercise Group; Right panel: the Control Group. Please click here to view a larger version of this figure.

Figure 3: Neural dynamics and CNN classification of exercise vs. reading groups. (A) Significant differences between groups identified by point-to-point t-tests, highlighting time-frequency clusters (p < 0.05, FDR-corrected). (B) Topographic maps of grand-averaged alpha-band (7-13 Hz) power. Maps depict the spatial distribution of neural oscillatory activity for the reading group (left) and exercise group (right). (C) Classification performance of prefrontal Alpha activity using a CNN model (accuracy: 83.3%). Please click here to view a larger version of this figure.
| Stage | Criteria/Process | Number | Outcome | Location in Protocol |
| Initial Recruitment | Non-sports majors from Wuhan Sports University | 550 | Eligible for pre-screening | Section 1.1 |
| Anxiety screening | STAI Trait Anxiety score ≥55 | 120 | Meet anxiety threshold | Section 1.2 |
| Activity Screening | Exercise frequency <3 days/week (high-intensity); Total MET-minutes <600/week | 40 | Qualified for final allocation | Section 1.3 |
| Final Groups | Exercise intervention (n=20): Moderate cycling; Control (n=20): Quiet reading | 40 | EEG and CNN | Section 2 |
Table 1: Participant recruitment and screening criteria.
In the realm of mental health research, understanding the underlying neural mechanisms of interventions for individuals with high trait anxiety is of paramount importance. This study was designed with the explicit aim of exploring the neural biomarkers associated with exercise intervention in such individuals through the utilization of artificial intelligence models. The adoption of advanced deep neural networks, including models like EEGNet, has revolutionized electroencephalogram (EEG) signal processing by enabling the precise analysis and decoding of brain activity with unprecedented accuracy47,48. Thus, this study used deep learning models to classify EEG data for exercise intervention, aiming to identify neural biomarkers of high trait anxiety and explore the intervention's impact on these markers. Participants with high trait anxiety were divided into an experimental group and a control group; their brain electrical activities were compared between exercise and reading. The results of the study were highly significant. Specifically, the prefrontal region, and particularly its α-wave activity, demonstrated remarkable accuracy in the classification tasks. This finding strongly suggests that prefrontal α waves may play a pivotal role in the regulatory effect of exercise intervention on high trait anxiety.
In the exploration of the neural mechanisms underlying emotion recognition, especially in the context of anxiety, the prefrontal α-wave activity (8-12 Hz) emerges as a key factor. This specific neural oscillatory pattern has a significant association with an individual's performance in emotion recognition tasks, and understanding its role is crucial for elucidating how emotions are processed and regulated. α Waves, reflecting brain relaxation and anxiety reduction, are crucial for emotion recognition. Their activity relates to attentional resource allocation, emotional information processing depth, and emotional response inhibition. Anxious individuals exhibit lower prefrontal α waves during emotion recognition, especially at rest. This reduction may link to their negative interpretations and hypervigilance. Studies found that the weakened prefrontal α Waves in anxious individuals are correlated with difficulties in recognizing emotional cues49,50. Anxious individuals often overreact to threat-related information, which can interfere with the normal neural processes involved in emotion recognition. Excessive sensitivity to emotional cues (e.g., facial expressions) further impairs recognition accuracy and adaptability51.
The prefrontal α is vital for emotion regulation, particularly of negative emotions. Individuals with high trait anxiety show excessive negative emotional responses, and reduced prefrontal α waves may indicate inadequate regulation. α-wave reduction reflects overreaction to emotional stimuli and weakened inhibitory neural mechanisms, serving as a neural basis for regulation difficulties. Lee52 proposed that increased left prefrontal α waves correlate with better emotion regulation and fewer negative responses, while reduced right prefrontal α waves associate with poor emotion inhibition and heightened negative emotions. Chen53 noted that reduced prefrontal α waves in depression relate to emotion regulation difficulties and impaired cognitive control. Depressed individuals struggle to regulate emotions, with reduced prefrontal activity exacerbating symptoms. Reduced left prefrontal α waves may reflect overreaction to negative emotions, highlighting the role of prefrontal α waves in emotion recognition and regulation for depression patients.
Exercise intervention may improve anxiety management and emotion regulation by enhancing prefrontal α waves, supporting exercise as an effective intervention for high-trait-anxiety populations52. Exercise intervention may enhance emotion regulation and reduce anxiety by strengthening prefrontal α waves. Reduced prefrontal α waves in anxious individuals link to overreaction to negative emotions and deficient recognition/regulation. Weakened prefrontal α waves correlate with emotion regulation difficulties and impaired cognitive control54. This study provides new insights into exercise intervention's neurobiological mechanisms for high trait anxiety and theoretical support for its clinical application as a non-pharmacological method.This finding also aligns with and corroborates the effectiveness of single, acute aerobic exercise sessions as established in previous research. Exercise improves neurobiochemical levels, inhibits inflammation, regulates the neuroendocrine system, and enhances neural plasticity, alleviating depression55. As A single session of acute aerobic exercise enhances activity and network connectivity within the hippocampus and parietal lobe. These findings align with Li (2018), who demonstrated that physical exercise alleviates depression and anxiety in college students, noting sports dance as most effective for depression and badminton for anxiety (Li, 2018)56 Acute aerobic exercise shows significant benefits in treating emotional disorders.
The effectiveness of deep learning models as a testing tool is also supported by prior research. Qi & Zhang (2024)57 highlighted that deep learning models offer new approaches to aphasia's neurocomputational mechanisms. Breedlove et al. (2020)58 used deep generative networks to simulate the brain's top-down imagery mechanism, predicting brain responses to imagery. Other studies confirm deep learning's application in neuroimaging. For example, Jiang et al. (2022) used deep convolutional neural networks (CNNs) to identify brain activity patterns in cognitive tasks59. Amin (2023) analyzed fMRI data via deep learning, revealing functional connections during emotion regulation and providing new biomarkers for emotional disorder treatment60.
These studies demonstrate deep learning's power in neurocomputation, offering tools for cognitive neuroscience, especially in emotion, memory, and disorder treatment. This study used the CNN model to analyze exercise intervention data, finding that CNN distinguishes brain electrical activity changes. Prefrontal α-wave data showed higher accuracy in differentiating exercise intervention vs reading states than resting-state EEG data, suggesting prefrontal α waves play a key role in exercise-induced brain activity changes, especially in emotion regulation and cognitive control. Thus, prefrontal α waves may serve as neural markers to monitor exercise intervention's impact on brain function, providing neurobiological evidence for EEG-based personalized treatment.
The study's sample size (e.g., N = 40) may limit the statistical power to detect subtle but clinically meaningful effects, particularly in subgroup analyses (e.g., age- or pathology-specific responses). Small samples increase the risk of Type II errors (false negatives) and reduce the reliability of effect size estimates, as random variability can disproportionately influence results61. While our cross-validation approach mitigates some risks, larger cohorts are needed to validate findings across diverse populations (e.g., clinical vs. healthy, different age groups) and account for inter-subject variability in EEG dynamics62.
The convolutional neural network (CNN) architecture, though effective for feature extraction, is susceptible to overfitting given the limited EEG dataset size. Overfitting may lead to inflated performance metrics during training that fail to generalize to independent datasets, as CNNs can memorize noise or subject-specific artifacts rather than biologically relevant patterns63. Although we employed PCA for dimensionality reduction (retaining 95% variance) and dropout regularization (p = 0.5), the model's performance on external datasets remains uncertain. Hybrid approaches (e.g., wavelet-CNN fusion with synthetic EEG augmentation) could improve robustness in future work64.
The study's focus on eyes-open resting-state EEG may limit direct applicability to other paradigms (e.g., task-based or eyes-closed recordings). For instance, alpha-band suppression in eyes-open conditions could obscure biomarkers typically observed in eyes-closed protocols (e.g., default mode network activity)65. Additionally, participant demographics (e.g., young adults) and controlled laboratory settings may not fully represent real-world scenarios or clinical populations with comorbidities66. External validation in multi-center studies, using standardized acquisition protocols, is essential to confirm the method's broader utility
The authors declare no conflicts of interest.
None
| BrainAmp SN | Brain Products | AMP12081737 Standard | Acquisition of electroencephalogram (EEG) signals |
| Eprime Professional | PSYCHOLOGY SOFTWARE TOOLS | 2.0.10.92 | Psychology Experiment Software |
| motion cycle 600 | emotion fitness GmbH & Co. KG | F-EF-MC-650 | Bicycle Ergometer |
| DCU(Deep Computing Unit) | HYGON | HYGON Z100L | Model analysis |
| Python | Python Software Foundation | Python 3.8 | Model analysis |