Impact of Osteopathic Techniques on Autonomic Regulation: A Study of Heart Rate Variability in Healthy Adults

Introduction

AIM OF THE STUDY:

The aim of this study was to evaluate whether specific osteopathic techniques, including compression of the fourth ventricle (CV4) and rib raising (RR), influence autonomic nervous system (ANS) function, as assessed by heart rate variability (HRV). Additionally, the study aimed to compare the effectiveness of these techniques individually and in combination, specifically investigating whether adding the RR technique to CV4 enhances the therapeutic effect. A further objective was to assess the sustainability of the observed effects 1 month after completing the treatment protocol.

Therefore, this study aimed to evaluate the effects of the fourth ventricle compression technique and the rib raising osteopathic technique on the activity of the autonomic nervous system measured by heart rate variability in 100 healthy adults.

Material and Methods

ETHICS STATEMENT:

The study was conducted in accordance with ethical guidelines and approved by the Senate Ethics Committee for Scientific Research at Olsztyn Higher School (opinion no. 1.2023, dated January 12, 2023). All participants were fully informed about the purpose, procedures, and potential risks of the study. Written informed consent was obtained from each participant prior to their inclusion in the research.

REGISTRATION:

This study is registered at ClinicalTrials.gov with the identifier NCT06554834, unique protocol ID: SOMATIC2024-001.

POPULATION:

The research was carried out at the SomaticMed Clinic in Wołomin, Poland, specializing in physiotherapeutic and osteopathic therapy, between June and October 2024.

INCLUSION AND EXCLUSION CRITERIA:

The study included 100 healthy volunteers who met the following inclusion and exclusion criteria and completed all stages of the study, and only their data were included in the statistics and final results. The subjects were divided into 3 groups: CV4 – 34 (34%) participants, CV4+RR – 33 (33%) participants, and control – 33 (33%) participants.

Inclusion criteria for the study were:

Exclusion criteria for the study were:

After meeting the above criteria, the study group was randomly divided into an experimental group and a control group.

RANDOMIZATION:

From a specially designated container, the 100 participants selected cards marked with either “E1” for experimental group 1, “E2” for experimental group 2, or “P” for the control group. After drawing a card, each participant shared their selection with the study coordinator, who logged the information and confirmed their group assignment. All participants provided written consent and received comprehensive materials outlining the study’s purpose, as well as its indications and contraindications. Participants remained unaware of their group assignment throughout the study. The individual responsible for post-treatment assessments was also blinded to the group allocations of all participants. In the end, the experimental group 1 (CV4+RR) comprised 33 subjects, the experimental group 2 (CV4) consisted of 34 subjects, and the control group consisted of 33 subjects. Characteristics of the participants are presented in Tables 1–3.

The study included 100 patients, 68 (68%) women and 32 (32%) men. The average age of the participants was 44.5 years, with a standard deviation of 9.2 years. The oldest participant was 63 years old, and the youngest was 30 years old. The subjects were divided into 3 groups: CV4–34 (34%) participants, CV4+RR – 33 (33%) participants, and control – 33 (33%) participants.

INTERVENTION IN THE EXPERIMENTAL GROUP 1 (CV4+RR):

The procedure applied in the experimental group 1 (CV4+RR) was conducted using 2 techniques: 1) the CV4 technique and 2) the RR technique, which are aimed at normalizing the autonomic system by reducing heart contraction rate, potentially affecting the autonomic system function [14,16]. Each participant received three 30-minute sessions, 1 per week. Measurements were taken before the procedure, immediately after the third session, and 1 month after the third session.

INTERVENTION IN THE EXPERIMENTAL GROUP 2 (CV4):

The procedure applied in the experimental group 2 (CV4) was conducted using a single technique – the CV4 technique – which is aimed at normalizing the autonomic system by reducing heart contraction rate, potentially affecting the autonomic system function [14,16]. Each participant received three 30-minute sessions, 1 per week. Measurements were taken before the procedure, immediately after the third session, and 1 month after the third session.

INTERVENTION IN THE CONTROL GROUP:

The intervention was conducted with the participant lying face down on the therapy table. The examiner guided the ultrasound transducer along the thoracic spine region, positioning it approximately 10 cm from the spinous processes on both sides and around the suboccipital area. Before starting, the participant was informed that the therapist would use the ultrasound device to locate areas of increased tissue tension, which would be shown as “confirmation” in red on the ultrasound screen, while a change to green would signify simulated tissue relaxation. The session lasted 30 minutes [17]. Each participant underwent three 30-minute sessions, spaced 1 week apart. Measurements were taken before the first session, immediately after the third session, and 1 month following the last session. The procedures were conducted by an osteopath with dual qualifications in osteopathy and physiotherapy, with 15 years of clinical experience.

CV4 TECHNIQUE:

The subject assumed a supine position on a therapy table. The therapist positioned themselves behind the subject’s head in a sitting posture, with their forearms resting against the therapeutic table. The therapist’s hands were positioned under the occipital bone, aligning the knuckles of the thumbs laterally to the external occipital protuberance and medially to the jugular foramen. Overlapping fingertips targeted the area around the Th1 spinous process. Subsequently, the therapist awaited the detection of the cranial rhythm.

During the “cranial exhalation” phase, the therapist applied gentle compression on the occipital bone in a medial direction, emphasizing this stage. Subsequently, during the “cranial inhalation” phase, pressure was momentarily released. The entire procedure lasted 30 minutes, and its methodology was chosen based on recommendations from the literature [15]. The technique is designed to activate the parasympathetic nervous system by targeting the vagus nerve and promoting the functioning of the brain’s glymphatic system [10].

RR TECHNIQUE: The RR technique was conducted with the participant lying in a supine position on a treatment table, while the therapist was seated on the participant’s left side. The therapist applied pressure near the rib angles, directing force towards the abdominal and lateral regions to achieve movement in the costovertebral joint. Each cycle consisted of 5 seconds of pressure followed by a 5-second pause. The procedure began with ribs I–IV, progressing to ribs V–VIII after completing 10 cycles, and then to ribs IX–XII for another 10 cycles. Once the left side was completed, the same process was repeated on the right side. The entire session for both sides lasted approximately 10 minutes. The methodology followed established recommendations from prior literature [15]. This technique is designed to mobilize the costovertebral joints, which in turn is believed to influence the local metabolism of autonomic sympathetic ganglia and reduce sympathetic nervous system activity [5].

HRV STUDY PROTOCOL:

To evaluate ANS functionality, we employed an HRV test [15,17–19]. The study made use of the HRV Infiniti 8 measurement system, in combination with the ProComp Infiniti encoder, as well as Biograph Infiniti and Physiology Suit software, provided by Biomed Neurotechnologie (www.biomed.org.pl, accessed on 01.12.2023).

The test was conducted with participants in a seated position. Before initiating the test, subjects received instructions to maintain calm and natural breathing throughout the 6-minute duration. They were also instructed to refrain from moving the finger connected to the measuring apparatus. Following the procedure, the test results were processed by a certified physiotherapist specializing in HRV testing to address any artifacts.

STATISTICAL ANALYSIS:

The statistical analysis was conducted to evaluate within-group changes and between-group differences using appropriate non-parametric tests. The Wilcoxon signed-rank test was applied for dependent samples to assess changes within groups, while the Kruskal-Wallis test was used for between-group comparisons. These non-parametric tests were selected due to the results of the Shapiro-Wilk test, which indicated a non-normal distribution for most parameters.

The analysis focused on HRV parameters (HRV1, HRV2, HRV3), VLF%, LF%, HF%, heart rate, and the LF/HF ratio. All tests were two-tailed, and a significance threshold of p<0.05 was adopted.

NORMALITY ASSESSMENT:

The Shapiro-Wilk test was performed to verify the normality of variable distributions. The following variables were found to exhibit a normal distribution:

(Table 4 provides detailed results of the normality assessment). For variables demonstrating normal distributions, additional parametric analyses were considered where appropriate.

Results

HRV Parameter

ANALYSIS OF DIFFERENCES BETWEEN HRV PARAMETER MEASUREMENTS TAKEN BEFORE THERAPY AND PATIENT GROUP MEMBERSHIP: To verify the differences between the HRV parameter measurements taken before therapy and patient group membership, a repeated measures analysis of variance in a mixed design was conducted. Here, the within-subject factors were the 3 HRV parameter measurements taken before the therapy, and the between-subject factor was patient group membership (Table 5).

As a result of the analysis, no statistically significant findings were observed:

ANALYSIS OF DIFFERENCES BETWEEN LF PARAMETER MEASUREMENTS TAKEN BEFORE THERAPY AND PATIENT GROUP MEMBERSHIP: To verify the differences between the LF parameter measurements taken before therapy and patient group membership, a repeated measures analysis of variance in a mixed design was conducted. Here, the within-subject factors were the 3 LF parameter measurements taken before the therapy, and the between-subject factor was patient group membership (Table 6).

As a result of the analysis, the following were observed:

Multiple comparisons for the main effect of group membership showed that the highest LF parameter values were observed in the CVR+RR group (M=57.3; SE=2.37), which obtained significantly higher results than the CV4 group (M=39.6; SE=2.33) and the control group (M=42.8; SE=2.37), as shown in Figure 1.

Multiple comparisons for the interaction effect between the LF parameter and group membership showed that:

ANALYSIS OF DIFFERENCES BETWEEN HF PARAMETER MEASUREMENTS TAKEN BEFORE THERAPY AND PATIENT GROUP MEMBERSHIP: To verify the differences between the HF parameter measurements taken before therapy and patient group membership, a repeated measures analysis of variance in a mixed design was conducted. Here, the within-subject factors were the 3 HF parameter measurements taken before the therapy, and the between-subject factor was patient group membership (Table 8).

As a result of the analysis, the following were observed:

However, a statistically significant interaction effect between group membership and HF parameter measurements was observed: F2=(4,194)=3.262; p=0.013; eta2=0.024, indicating the presence of statistically significant differences between patient groups and HF parameter measurements. To verify the significance of this effect, multiple comparisons were conducted using Tukey’s test.

Multiple comparisons for the interaction effect between the HF parameter and group membership showed that:

ANALYSIS OF DIFFERENCES BETWEEN LF/HF PARAMETER MEASUREMENTS TAKEN BEFORE THERAPY AND PATIENT GROUP MEMBERSHIP: To verify the differences between the LF/HF parameter measurements taken before therapy and patient group membership, a repeated measures analysis of variance in a mixed design was conducted. Here, the within-subject factors were the 3 LF/HF parameter measurements taken before the therapy, and the between-subject factor was patient group membership (Table 9).

As a result of the analysis, the following were observed:

To verify the significance of these effects, multiple comparisons were conducted.

Multiple comparisons for the main effect of the LF/HF parameter showed that the average LF/HF parameter value (M=2.31; SE=0.25) in the first measurement was significantly lower than in the second measurement (M=5.53; SE=0.51). The average LF/HF parameter value in the second measurement was significantly higher than in the third measurement (M=2.21; SE=0.22) (Table 10), as illustrated in Figure 4.

Multiple comparisons for the main effect of group membership showed that the average LF/HF parameter value in the CV4 group (M=1.84; SE=0.42) was significantly lower than in the CV4+RR group (4.26; SE=0.43). It was also observed that the average LF/HF parameter value in the CV4+RR group (4.26; SE=0.43) was significantly higher than in the control group (M=1.95; SE=0.43) (Table 11), as illustrated in Figure 5.

Multiple comparisons for the interaction effect between the LF/HF parameter and group membership showed that:

Discussion

The aim of the study was to evaluate the potential effects of osteopathic manipulative treatment (OMT) on autonomic nervous system (ANS) function, measured through heart rate variability (HRV). Thirty-minute treatments were conducted 3 times, at weekly intervals, to replicate clinical practice conditions commonly observed by physiotherapists and osteopaths. To assess the durability of the therapeutic effects, a third measurement was performed 1 month after the final treatment.

A significant change was observed in the LF parameter (p<0.001), which is associated with sympathetic nervous system activity. In the CV4+RR group, the LF parameter increased significantly following the 3 therapy sessions, as seen in the second measurement. However, by the third measurement, taken 1 month later, the value had decreased, though it remained higher than the baseline. This decrease, however, was not statistically significant, suggesting that additional treatments may be required to sustain the effect over a longer period. After the 3 treatments, the LF parameter was significantly higher in the CV4+RR group compared to the CV4 and control groups, highlighting the procedure’s efficacy in increasing sympathetic nervous system activity. One month after the therapy, a significant difference was observed only between the CV4+RR and CV4 groups, with no significant difference from the control group, indicating that more treatments may be needed for a longer-lasting effect.

The HF parameter, indicative of parasympathetic activity, showed only minor changes. A trend-level decrease (p=0.057) was observed in the CV4+RR group between the baseline and the second measurement. This suggests that with a larger sample size, a statistically significant result is likely. In the second measurement, the CV4+RR group exhibited a significant difference compared to the CV4 group, indicating opposing effects on ANS activity: CV4+RR reduced parasympathetic activity, while CV4 increased it. However, neither group showed a significant difference from the control group, suggesting that the effects might be insufficiently robust or require additional interventions.

The LF/HF ratio, a key parameter reflecting the balance between the sympathetic and parasympathetic nervous systems, showed a statistically significant main effect (p=0.004). In the CV4+RR group, the LF/HF ratio significantly increased between the baseline and the second measurement, followed by a significant decrease between the second and third measurements. There was no significant difference between the baseline and the third measurement, indicating a temporary shift toward sympathetic dominance. These findings may inform therapeutic decisions for patients with excessive parasympathetic dominance. Additionally, in the second measurement, the LF/HF ratio in the CV4+RR group was significantly higher than in both the CV4 and control groups.

The control group did not exhibit significant changes in any HRV parameters. Similarly, HRV did not show significant changes across any groups.

These findings suggest that combining the RR technique with CV4 significantly influenced the final outcomes by stimulating the sympathetic nervous system, as demonstrated by all 3 parameters. This effect is likely attributable to the mobilization of the costovertebral joints and their mechanical impact on the sympathetic ganglia located ventrally.

Henderson et al (2010) demonstrated the effect of the RR technique on reducing sympathetic activity by lowering salivary alpha-amylase (SAA) levels, which differs significantly from the authors’ results [20]. The alpha-amylase index is correlated with sympathetic activity [21]. It is possible that the opposite results are due to a different measurement method, as alpha-amylase testing may have a more localized rather than systemic effect. Arienti et al (2020) studied 32 healthy individuals divided into a CV4 group, an RR group, and a control group. They measured heart rate variability (HRV) and skin conductance parameters. The RR technique group showed a significant decrease in the LF/HF (low-frequency/high-frequency) ratio (p<0.001) and low frequency (LF) (p<0.001), and an increase in HF (p<0.001). The CV4 technique group showed a significant decrease in the LF/HF ratio (p<0.001), a significant decrease in LF (p<0.001), and a significant increase in HF. The researchers suggest that both techniques have a beneficial effect on the autonomic system by increasing parasympathetic dominance [22]. For the effect of the CV4 technique alone, the results align with the authors’ findings, but the results for the CV4 and RR groups are contradictory. However, it should be emphasized that Arienti et al. (2020) only performed single sessions [22]. Ruffini et al (2015) studied 66 healthy individuals who underwent OMT selected according to the therapist’s assessment rather than a standardized protocol. In the OMT-treated groups, an increase in the HF parameter and a decrease in LF were observed after the procedure. No significant changes were noted in the control group [23]. Besson et al (2023) studied the effect of a single 10-minute session of the occipito-mastoid structure normalization (OMSN) technique on HRV parameters. The study showed a significant increase in parasympathetic nervous system activity, expressed by an increase in the RMSSD (root mean square of successive differences) index after a 10-minute application of the OMSN technique compared to the control group receiving a placebo procedure. The intervention did not affect other HRV parameters [24].

In the study by Stępnik et al (2023), the application of the fourth ventricle compression (CV4) and rib raising osteopathic techniques resulted in a significant decrease in heart rate among healthy individuals, suggesting enhanced parasympathetic activity. However, other heart rate variability (HRV) parameters did not show significant changes [17]. The present study was conducted on a larger group, with 3 therapeutic procedures performed, which positively influenced changes in parameters other than just heart rate (HR). This suggests that a single treatment is insufficient to significantly affect parameters such as HRV, LF, HF, and LF/HF. Further research is needed to determine whether performing a greater number of treatments would lead to more significant improvements in these parameters [17].

The study of OMT’s impact requires further research, as there are ongoing uncertainties about the measurement results of different authors. It would also be worth extending the research to individuals with conditions affecting the ANS. It would also be valuable to conduct a study only on individuals with significantly elevated HF or LF parameters before the study begins to see if OMT affects balancing significantly elevated parameters. If the group is mixed (some individuals have elevated, and others have decreased parameters at the study’s start), if OMT has a balancing effect (reducing HF when elevated and reducing LF when elevated), the results will cancel each other out, and the study’s conclusions may be incorrect.

Conclusions

This study demonstrated that the combined use of the compression of the fourth ventricle (CV4) and rib raising (RR) techniques significantly impacts certain parameters of heart rate variability (HRV), particularly those associated with sympathetic nervous system activity. The CV4+RR group showed notable changes in LF%, HF%, and LF/HF ratio compared to the CV4-only and control groups. These results suggest that adding the RR technique to CV4 enhances its effectiveness in modulating autonomic nervous system (ANS) activity. However, the temporary nature of these effects highlights the potential need for additional or more frequent treatments to achieve sustained outcomes. Further research is needed to confirm these findings and explore their broader clinical implications.