High-salt intake alters autonomic maturation trajectories assessed by an HRV-based ANS Age Index in rats

Excessive dietary salt intake is known to affect autonomic and cardiovascular regulation, but its longitudinal impact on autonomic maturation remains poorly understood. This study developed a heart rate variability (HRV)-based autonomic nervous system (ANS) Age Index to quantify deviations from the normative trajectory of autonomic maturation in rats exposed to different levels of salt loading. Eighteen rats were assigned to control, 4% NaCl, or 8% NaCl diet groups and followed over an 11-week observation period. Three-lead electrocardiograms were recorded twice weekly for 5 min, and HRV-derived time-frequency representations were generated using short-time Fourier transform. Baseline-referenced HRV matrices were segmented using a 30-s sliding window with a 10-s shift, and each HRV segment was combined with zero-padded baseline-referenced body weight-change information to construct a 407 × 300 input for a VGG16-based regression model. The model was trained exclusively on control-group data to learn the normative autonomic maturation trajectory and was then validated using leave-one-animal-out cross-validation to reduce the risk of information leakage from non-independent augmented segments. In this animal-level validation, all augmented segments from one control animal were held out during model training in each fold. After averaging the 28 segment-level predictions within each ECG recording, the model achieved a recording-level root mean square error (RMSE) of 0.758 weeks, mean absolute error (MAE) of 0.608 weeks, and coefficient of determination (R²) of 0.943. After validation, a final control-trained model was applied to the high-salt groups to estimate model-derived deviations from the control-derived autonomic trajectory. The 4% NaCl group showed a sustained positive shift in ANS Age Index, indicating an older-than-expected model-predicted autonomic profile relative to the control-derived trajectory. In contrast, the 8% NaCl group showed persistently lower and flattened predicted autonomic age trajectories, indicating a younger-than-expected or altered model-derived autonomic trajectory. These findings suggest that high-salt intake may be associated with non-linear deviations in model-predicted autonomic trajectories. The proposed HRV-based ANS Age Index should be interpreted as an exploratory computational proxy for detecting diet-associated shifts in HRV-derived autonomic patterns, rather than as a validated physiological biomarker of autonomic aging.