Hypothesis

Baseline heart-rate variability (HRV) predicts the magnitude of anxiety reduction following supplementation with Lactobacillus plantarum JYLP-326, reflecting individual differences in vagal afferent readiness to transmit gut-derived signals.

Mechanistic Rationale

Recent work shows that Lactobacillus plantarum JYLP-326 restores beneficial taxa such as Bacteroides, Bifidobacterium, Prevotella and Roseburia, shifts tryptophan metabolism toward serotonin, and lowers proinflammatory cytokines [https://pmc.ncbi.nlm.nih.gov/articles/PMC12386460/][https://pmc.ncbi.nlm.nih.gov/articles/PMC10077425/]. These changes increase luminal serotonin and indole-derived metabolites that activate enterochromaffin cells and vagal afferents via 5-HT3 receptors and transient receptor potential channels. The strength of this gut-to-brain signal depends on the excitability of vagal fibers, which is modulated by their baseline tone. High baseline HRV indicates greater parasympathetic reserve and heightened sensitivity of vagal afferents, allowing microbiota‑induced neurochemical changes to be efficiently transmitted to the nucleus tractus solitarius and subsequently to limbic circuits that regulate anxiety. Conversely, low baseline HRV reflects reduced vagal excitability, limiting signal propagation despite equivalent microbiota shifts.

Testable Predictions

1. Participants with baseline HRV above the cohort median will show a larger decrease in HAMA/HDRS scores after 3 weeks of Lactobacillus plantarum JYLP-326 (≥1.5×10¹⁰ CFU/day) compared with those below the median.
2. The same high‑HRV subgroup will exhibit a greater increase in HRV (time‑domain RMSSD) from baseline to week 3, indicating enhanced vagal tone as a downstream effect of successful gut‑brain signaling.
3. Microbiota alterations (increase in Bacteroides/Bifidobacterium, decrease in IL-6) will occur irrespective of baseline HRV, confirming that HRV modulates signal transmission rather than microbial engraftment.

Experimental Design

• Population: 120 university students with baseline HAMA ≥15 (moderate anxiety). Stratify randomization by baseline HRV (high vs low) to ensure balanced groups.
• Intervention: Lactobacillus plantarum JYLP-326 1.5×10¹⁰ CFU/day or matched placebo for 3 weeks.
• Measurements:
  • Baseline: 5‑min resting ECG for HRV (RMSSD, HF power), HAMA, HDRS, fecal microbiota (16S rRNA), plasma tryptophan/kynurenine ratio, IL-6, TNF-α.
  • Week 3: repeat all measures.
• Analysis: Mixed‑effects model with fixed effects for treatment, HRV stratum, and their interaction; random intercept for participant. Primary outcome: change in HAMA score. Interaction term tests prediction 1. Secondary outcomes: change in HRV, microbiota shifts, cytokine levels.

Potential Outcomes and Falsifiability

• Support: Significant treatment × HRV interaction showing greater anxiety reduction in high‑HRV probiotic group, accompanied by HRV increase only in this subgroup, while microbiota changes are similar across strata.
• Refutation: No interaction; anxiety improvement independent of baseline HRV, or HRV does not change despite anxiety reduction. This would falsify the claim that baseline vagal tone gates psychobiotic efficacy, prompting alternative explanations such as central serotonergic mechanisms.