Breathwork and Stress: Investigating the Mechanisms of Action and Effectiveness of Breathing Interventions in Modulating the Psychophysiological Response to Acute Stress Test

Breathwork and Stress: Investigating the Mechanisms of Action and Effectiveness of Breathing Interventions in Modulating the Psychophysiological Response to Acute Stress Test

This study investigates whether the psychophysiological benefits of breathing exercises are driven by a specific physiological rhythm (6 breaths per minute) or by the general psychological experience of performing a structured, mindful activity. Researchers aim to determine if "coherent breathing", which is hypothesized to synchronize heart and respiratory rhythms, offers unique physiological protection against stress compared to breathing at a natural pace or simple resting. The main questions it aims to answer are:

• Does slow, steady breathing at 6 breaths per minute lower physical stress markers (like heart rate variability and cortisol) better than faster, but structured breathing or just sitting still?
• Is the calming effect caused by the specific breathing rhythm or simply by performing a structured, relaxing activity?

Researchers will compare three groups to see if the specific rhythm of "coherent breathing" offers unique benefits:

1. Group (Interventional): Coherent Breathing: Slow breathing at 6 breaths per minute.
2. Group (Sham Breathing): Regular breathing at 15 breaths per minute (matching a natural pace).
3. Spontaneous Breathing (Control Group): Natural, unguided breathing.

Participants will:

• Complete a one-day preparation phase to become familiar with the breathing technique.
• Visit the research center for one experimental session.
• Perform their assigned breathing method before and after a stress test.
• Take the Maastricht Acute Stress Test (MAST), which involves putting a hand in cold water and doing mental math.
• Provide saliva samples and have their heart rate variability, and mood measured multiple times.

This three-arm randomized controlled trial is designed to investigate whether the therapeutic benefits of slow-paced breathing stem from specific physiological mechanisms (cardiorespiratory resonance) or non-specific psychological factors, such as attention, expectancy, and structured relaxation.

The study directly compares resonant-frequency breathing against a structurally identical sham intervention and a passive control group.

Experimental Protocol:

The study utilizes the Maastricht Acute Stress Test (MAST) as a highly structured and validated laboratory stressor to capture the dynamic psychophysiological response to acute distress.

The protocol consists of the following phases:

Baseline Phase (BASE): A 5-minute measurement of baseline physiological (HRV - Heart Rate Variability) and psychological (STAI-S - State-Trait Anxiety Inventory-State, VAMS - Visual Analogue Mood Scales) parameters.

First Intervention (10 min): Participants perform their assigned breathing technique or sit quietly to assess the preventive "buffering" effect of the intervention.

Acute Stress Induction (MAST): A 15-minute structured stressor combining periods of hand immersion in cold water and mental arithmetic under social-evaluative pressure. The water temperature is strictly maintained within the range of 4.5 - 5.5 °C using a TECO chiller to ensure consistent stress induction across all participants.

Post-Stress Recovery (POST): Immediate 5-minute measurement following the conclusion of the MAST protocol.

Second Intervention (10 min): A repeat of the breathing/control protocol to evaluate its impact on psychophysiological recovery.

Final Assessments (END & FOLLOW): Measurements are taken immediately after the second intervention and following a 15-minute recovery phase to capture the peak glucocorticoid (cortisol) response.

Data Collection and Analysis:

• Heart Rate Variability: RR interval (time duration between successive heartbeats, specifically the time intervals between each R-wave peak) data is collected using a Polar H10 heart rate monitor. To ensure data integrity and redundancy, the signal is recorded simultaneously through two mobile applications: Elite HRV and Kubios HRV. All subsequent advanced HRV analyses, including time-domain, frequency-domain, and non-linear indices (e.g., PNS/SNS Index [Parasympathetic Nervous System Index and Sympathetic Nervous System Index], RMSSD [Root Mean Square of Successive Differences]), are performed using Kubios HRV Scientific software.
• Biochemical Markers: Salivary cortisol levels are measured to assess the hypothalamic-pituitary-adrenal (HPA) axis response using ELISA kits.
• Psychological Scales: Subjective stress and anxiety are tracked using the STAI-VAMS.

Adherence Check: Participant adherence to the paced breathing protocols is verified using the Kubios RESP (respiratory rate estimation) algorithm. This software-based solution extracts respiratory frequency from the RR interval data to confirm that participants maintained the target rates of 6 bpm (breaths per minute) or 15 bpm.

Research Arms:

1. Coherent Breathing (6 bpm): Guided nasal diaphragmatic breathing (5.5 s inhale / 5.5 s exhale) focused on inducing cardiorespiratory resonance.
2. Sham Breathing (15 bpm): Guided breathing at a natural pace (2 s inhale / 2 s exhale) to control for the effects of a structured intervention.
3. Spontaneous Breathing (Control): Passive resting with eyes closed, serving as a neutral baseline for the natural course of the stress response.

Inclusion Criteria:

• Healthy adults
• Aged 18 to 60 years
• Willingness to participate in all study phases, including preparation and laboratory session.
• Professionally active individuals or university students.

Exclusion Criteria:

• Severe chronic diseases, including metabolic disorders (e.g., diabetes) and mental disorders.
• Cardiac arrhythmia, history of heart attacks, strokes, or heart surgery.
• Regular use of medications such as anxiolytics or beta-blockers (excluding hormonal contraception)
• Pregnancy.
• Current participation in other scientific experiments.
• Significant previous experience with breathing techniques or current independent breathwork/meditation practice (defined as regular practice for more than 7 days in total within the last 12 months).
• Professional sports practice.
• Raynaud's disease
• Inability to abstain from alcohol, caffeine, and nicotine for the required periods before the experiment