Autonomic Responses to Ultrasound-Guided Percutaneous Needle Electrolysis: Effect of Needle Puncture or Electrical Current?

Abstract

Objective:

The aim of this study was to establish if the changes in sympathetic and parasympathetic activity (analyzed through heart-rate variability [HRV]) during ultrasound (US)-guided percutaneous needle electrolysis (PNE) is due to the effect of needle puncture only or of the PNE technique per se where the puncture and galvanic current are combined.

Methods:

This was an experimental, case-control study that took place at the University of Seville. Subjects were 36 male footballers who were randomly allocated to three groups: a control group (CG; 12 players), for whom HRV was recorded for 10 min, both at rest and during an exhaustive US examination of the patellar tendon and adjacent structures; a first experimental group (PNE group; 12 players), for whom HRV was recorded for 10 min, both at rest and during application of US-guided PNE in the patellar tendon; and a second experimental group (needle group; 12 players), for whom HRV was recorded for 10 min, both at rest and during application of US-guided PNE without electrical current in the patellar tendon. The outcome measures were the diameters of the Poincaré plot (SD1, SD2), stress score, and sympathetic/parasympathetic ratio.

Results:

There were no differences between groups in any baseline measurements, nor were there any significant differences between CG measurements (baseline vs. intervention). The PNE group exhibited statistically significant increases in SD1 (p = 0.01) and SD2 (p = 0.004) and statistically significant decreases in SS and S/PS ratio (p = 0.03), indicating increased parasympathetic and decreased sympathetic activity, respectively. The needle group exhibited statistically significant increases in SD2 (p = 0.02) and statistically significant decreases in SS (p = 0.02), indicating decreased sympathetic activity.

Conclusions:

The application of the US-guided PNE technique caused a measurable increase in parasympathetic activity (detected by HRV), which was due to the combination of needle puncture and electric current.

Introduction

Ultrasound (US)-guided percutaneous needle electrolysis (PNE) is a novel and minimally invasive approach that involves the application of a galvanic current through an acupuncture needle.^1
–3 This technique has mainly been applied to treat musculoskeletal pathologies.^1,4

–7 Much like acupuncture,^8

–11 this technique is believed to influence mechanotransduction in the tendons.^1,2

Techniques involving puncture of the skin are common in physiotherapy. These procedures may use a mechanical stimulus, as in acupuncture or dry needling,^12,13 or apply electrical current, as in electroacupuncture,¹⁴ stimulation of myofascial trigger points,¹⁵ or electrostimulation using galvanic current.^1,2

In these procedures, in which needling is applied, alterations of the autonomous behavior have been observed during the technique application. Heart-rate variability (HRV) has been validated as an accurate tool to assess the status of the autonomic nervous system (both sympathetic and parasympathetic components) under various conditions.^16
–18 In fact, an autonomic imbalance, analyzed through HRV, has been shown in physiotherapy techniques involving puncture. Kitagawa et al.¹⁹ suggested that acupuncture stimulation of trigger points of the tibialis anterior muscle transiently increases parasympathetic nerve activity. Others^20
–22 have shown increased sympathetic activity during acupuncture needling, which is generally followed by increased parasympathetic tone afterwards. De la Cruz et al.²³ showed US-guided PNE in men was associated with an autonomic imbalance characterized by greater parasympathetic activity. Occasionally, vasovagal reactions may occur. These clinical phenomena are mediated by the vagus nerve and characterized by lightheadedness and malaise. Exceptionally, vasovagal reactions may result in brief and reversible loss of consciousness (vasovagal syncope). The relevance of this finding to clinical practice lies in the avoidance of clinically significant vagal reactions (including syncope). Therefore, it is important to advise physiotherapists that this invasive technique should be performed with due care, and one must be fully prepared to attend to any adverse reaction.

For this reason, the aim of this study was to establish whether the changes in sympathetic and parasympathetic activity (analyzed through HRV) during US-guided PNE are due to the effect of needle puncture only or of the PNE technique per se where the needle puncture and galvanic current were combined. In other words, does galvanic current play any role in the autonomous changes observed?

Materials and Methods

Participants

A total of 36 members of an amateur male football club were studied, split randomly into three groups, each comprising 12 players: a control group (CG) and two experimental groups. Exclusion criteria were: (1) pathology in the patellar tendon; (2) a Personal Psychological Apprehension Scale (PPAS) score >37.5²⁴; (3) commonly accepted contraindications to PNE, including chronic joint disease, surgery, prosthesis, or osteosynthesis in the area of intervention, cardiac disease, neoplasia, coagulopathy, use of certain drugs (fluoroquinoles, anticoagulants, corticosteroids, or non-steroidal anti-inflammatories)²⁵; (4) any contraindications to needling per se, including unsurmountable fear of needles, history of adverse reaction to needling, immunocompromise, difficulty expressing feelings appropriately, and/or allergy to metals²⁵; and (5) epilepsy.

The local ethics committee approved the study, which complied with all the principles set out in the Declaration of Helsinki. All subjects signed informed written consent to participate in this study.

Procedures

First, all subjects filled in the PPAS, the general aim of which was to measure the psychological apprehension of the subjects during electrophysiotherapeutic treatment. This scale aims to evaluate, by means of the opinion and self-positioning of the subjects, an individual's degree of apprehension to the application of an electrical current.²⁴ Subsequently, HRV was recorded in the early morning and after fasting overnight. HRV was recorded for the CG for 10 min, both at rest and during an exhaustive US examination of the patellar tendon and adjacent structures using a high-resolution grayscale US machine (Logiq; GE Healthcare, Madison, WI) with a linear probe (12 MHz), according to European Society of Musculoskeletal Radiology guidelines.²⁶ HRV was recorded for two experimental groups: a first experimental group (PNE-G) for 10 min, both at rest and during application of US-guided PNE in the patellar tendon (Fig. 1); and a second experimental group (Needle-G) for 10 min, both at rest and during application of US-guided PNE in the patellar tendon without application of electric current.

FIG. 1.

Heart-rate variability recorded during application of ultrasound (US)-guided percutaneous needle electrolysis (PNE) in the patellar tendon. Subjects sat on the bed, leaning back on the recliner at an angle of 75° with a pillow below the knee.

The patellar tendon was selected because patellar tendinopathy is one of the most commonly accepted indications for this technique,^6,7 although all study participants had healthy patellar tendons and did not suffer from any patellar pathology. The US-guided PNE technique was applied using a specifically developed medically certified (Directive 93/42/EEC) device (EPI Advanced Medicine, Barcelona, Spain), which produces galvanic electricity through the negative electrode cathodic flow. This is applied using a modified electrosurgical scalpel that incorporates acupuncture needles (0.3 mm in diameter) of different lengths. The intensity can be adjusted by changing either the duration of stimulation or the output current (mA) of the device. Conversely, the machine's polarity is fixed (i.e., only cathodic flow is usable). During each procedure, all of which were performed by the same experienced operator, volunteers sat on the bed, leaning back on the recliner at an angle of 75° with a pillow below their knees (Fig. 1) in order to minimize the risk of any potential vagal reaction (Fig. 1). Finally, three US-guided precise applications at a fixed intensity of 3 mA for 3 s were performed, according to Valera-Garrido and Minaya-Muñoz's protocol.²⁷ The applications were in the tendon.

Measurement of HRV

A Firstbeat Bodyguard (Firstbeat Technologies, Jyväskylä, Finland) heart rate (HR) monitor was used to record HR data for 10 min in every session (at rest and during US or US-guided PNE). Data were downloaded from the devices to a computer using the Firstbeat Uploader software (Firstbeat Technologies), and all the RRI series were imported into the Kubios software package (University of Eastern Finland, Kuopio, Finland), which was used to calculate all HRV parameters. To calculate the autonomic balance, the HRV method commonly used for this purpose is based on the Poincaré plot.²⁸ This method reflects HRV fluctuations.²⁹ Physiologically, the transverse axis (SD1) is a measurement of the short-term changes in RRIs and is considered an indicator of parasympathetic activity.³⁰ The physiological meaning of the longitudinal axis (SD2) is not as clear, but it is thought that it reflects the long-term changes in RRIs, and it is considered an inverse indicator of sympathetic activity.³¹ Naranjo et al.³² recently defined two new indexes to facilitate the physiological interpretation of the Poincaré plot: the stress score (SS) and the sympathetic–parasympathetic ratio (S/PS), respectively. The SS is expressed as the inverse of the diameter SD2 multiplied by 1000, and it is considered to be directly proportional to sympathetic activity at the sinus node. The S/PS ratio is expressed as the quotient of SS and SD1, and it is considered to reflect autonomic balance, that is, the relationship between sympathetic and parasympathetic activity.

Statistical analysis

All variables were expressed as the mean and standard deviation, and the normality of the data distribution was evaluated using the Shapiro–Wilk test. Data were first analyzed using a two-factor repeated-measures analysis of variance with three between-group factors (CG vs. PNE-G, CG vs. Needle-G, and PNE-G vs. Needle-G) and one within-group factor (baseline vs. intervention). The Games–Howell post hoc test was used for multiple comparisons. Data were analyzed with IBM SPSS Statistics for Windows v.21 (IBM Corp., Armonk, NY), and statistical significance was set at p < 0.05.

Results

There were no significant baseline differences between the CG, PNE-G, and Needle-G in any demographic variables, including age (22.92 ± 4.32 vs. 24.25 ± 4.05 vs. 25.92 ± 5.35 years), weight (74.33 ± 12.17 vs. 76.08 ± 6.42 vs. 77.33 ± 8.02 kg), height (176.50 ± 6.63 vs. 181.83 ± 3.21 vs. 176.17 ± 5.20 cm), and body mass index (23.77 ± 2.77 vs. 23.29 ± 2.13 vs. 24.92 ± 2.46 kg/m²). On the PPAS scale, the control and two experimental groups scored 23.17 ± 4.51, 22.50 ± 4.87, and 22.17 ± 4.39 (p > 0.05).

Table 1 shows the mean and standard deviation (SD) of the study variables (mean NN interval, mean HR, SD1, SD2, SS, and S/PS ratio) at rest (baseline measurement) and during US examination or application of the US-guided PNE technique or application of the US-guided PNE technique without electric current in the control, PNE, and needle groups, respectively. Table 1 also shows the difference (Δ, in %) between before and after interventions in each group.

Table 1.

Heart-Rate Variability Parameters Before and During US Examination, US-Guided PNE, or US-Guided PNE Without Electric Current

	Control group			PNE group			Needle group
	Baseline	Intervention	%Δ	Baseline	Intervention	%Δ	Baseline	Intervention	%Δ
NN interval (ms)	907.37 ± 165.55	931.88 ± 107.62	2.70	926.96 ± 126.53	952.41 ± 159.20	2.75	1008.58 ± 114.45	976.56 ± 91.24	−3.17
HR (bpm)	70.07 ± 11.69	66.50 ± 7.49	−5.09	69.05 ± 8.93	66.18 ± 10.69	−4.16	61.84 ± 12.38	66.53 ± 13.80	7.58
SD1	46.90 ± 25.11	48.85 ± 19.98	4.14	42.22 ± 15.22	72.92 ± 31.17^*	72.71	51.50 ± 31.80	66.60 ± 45.87	29.31
SD2	129.34 ± 44.35	139.05 ± 53.02	7.51	120.08 ± 47.74	189.15 ± 55.99^{*, †}	57.53	103.14 ± 28.53	142.06 ± 44.74^*	37.74
SS	8.58 ± 2.87	7.98 ± 2.39	−7.02	9.73 ± 4.16	5.72 ± 1.68^{*, †}	−41.21	10.48 ± 3.26	7.71 ± 2.51^*	−26.40
S/PS ratio	0.27 ± 0.24	0.20 ± 0.12	−25.84	0.31 ± 0.28	0.09 ± 0.06^{*, †}	−69.49	0.34 ± 0.41	0.21 ± 0.25	−39.39

Heart-rate variability (HRV) parameters before and during intervention in each group. The mean ± SD for each HRV parameter and group, and the difference (Δ, in %) between before and during interventions in the control, PNE group, and needle group consisted of US examination, US-guided PNE, and US-guided PNE without electric current, respectively.

p < 0.05 baseline versus intervention scores for PNE group and needle group.

†

p < 0.05 significant difference during intervention between PNE group and needle group.

US, ultrasound; PNE, percutaneous needle electrolysis; NN interval, normal RR interval (RRI); HR, heart rate; bpm, beats per minute; SD1, transverse axis of Poincaré plot; SD2, longitudinal axis of Poincaré plot; SS, stress score (inverse of diameter SD2 × 1000); S/PS ratio, quotient of SS and SD1.

There were no differences between the CG and two experimental groups in the baseline measurements of NN interval (p > 0.05).

There were no significant differences between the CG measurements taken at baseline and during the US examination (p > 0.05). By contrast, compared with baseline values, the PNE-G exhibited statistically significant increases in the following two parameters of HRV during intervention: SD1 (72.93 ± 31.17 vs. 42.22 ± 15.22; p = 0.01) and SD2 (189.15 ± 55.99 vs. 120.08 ± 47.74; p = 0.004). This group also exhibited statistically significant decreases in the following two parameters of HRV during intervention: SS (5.72 ± 1.68 vs. 9.73 ± 4.16; p = 0.03) and S/PS ratio (0.09 ± 0.06 vs. 0.31 ± 0.28; p = 0.03). Compared with baseline values, the Needle-G exhibited statistically significant increases in SD2 (142.06 ± 44.74 vs. 103.14 ± 28.53; p = 0.02) and statistically significant decreases in SS (7.71 ± 2.51 vs. 10.48 ± 3.26; p = 0.02) during intervention.

Figure 2 shows the comparison between the PNE-G and Needle-G during US-guided PNE technique or US-guided PNE technique without electric current, respectively. There were statistically significant differences in SD2 (189.15 ± 55.99 vs. 142.06 ± 44.74; p = 0.03), SS (5.72 ± 1.68 vs. 7.71 ± 2.51; p = 0.03), and ratio S/PS (0.09 ± 0.06 vs. 0.21 ± 0.25; p = 0.04).

FIG. 2.

Box-and-whiskers plot of SD2, SS, and ratio S/PS during US examination, US-guided PNE, and US-guided PNE without electric current, in control, PNE, and needle groups, respectively. The box represents the lower and upper quartiles, and the whiskers represent the total range. SD1, transverse axis of Poincaré plot; SD2, longitudinal axis of Poincaré plot; SS, stress score; S/PS, quotient of SS and SD1.

Figure 3 shows the comparison of SD1, SS, and S/PS ratio between the three groups.

FIG. 3.

Comparison of SD1, SS, and S/PS ratio between baseline and the intervention for each group. *p < 0.05.

Discussion

The main finding of this study was a significant autonomic imbalance (in keeping with a potential vasovagal reaction) during application of the US-guided PNE technique in healthy patellar tendons of male football players by means of simple HRV measurements. The relevance of this finding to clinical practice lies in the avoidance of clinically significant vasovagal reactions (including syncope). Therefore, it is important to advise physiotherapists that this invasive technique should be performed with due care, and one must be fully prepared to attend to any adverse reaction.

In the PNE-G, a statistically significant increases were observed in the marker of parasympathetic activity, SD1 (p = 0.01). Furthermore, a statistically significant increase was found in SD2 (p = 0.004), which is indicative of decreased sympathetic activity, as well as a statistically significant decrease in SS and S/PS ratio (p = 0.03). Overall, this suggests that the application of the US-guided PNE technique causes an autonomic imbalance with a predominance in parasympathetic activity (Fig. 3). However, in the Needle-G, a statistically significant increase in SD2 (p = 0.02) was demonstrated, which is indicative of decreased sympathetic activity, as well as a statistically significant decrease in SS (p = 0.02). Overall, this suggests that the application of the US-guided PNE technique without application of electrical current causes an autonomic imbalance but not large enough to be a predominance in parasympathetic activity. When the PNE-G and Needle-G were compared during the application of the US-guided PNE technique or application of the US-guided PNE technique without electric current, respectively, there were a statistically significant differences in SD2, SS, and ratio S/PS. Only the PNE-G during intervention showed a decrease in sympathetic activity enough to affect the balance between parasympathetic and sympathetic activity, reflected as ratio S/PS (Fig. 3). This result could be because the difference between before and during the interventions in the PNE-G was higher than in the Needle-G (Table 1). This means that the combination of needling and electric current was a cause of vagal response. However, these results have a positive effect because this technique causes a segmentary response (central nervous system activation) that starts the repair processes. This is known as the response of the organism to a threat (fight–flight response).^33,34

In this particular study, it would appear that this autonomic imbalance was elicited by the needling technique per se, regardless of the participants' attitudes, which were generally positive. In fact, subjects in both control and experimental groups scored an average of 23 points on the PPAS, which is much less than the threshold of 37.5 that is considered to indicate apprehension.²⁴ Thus, it appears unlikely that any study subject manifested any significant degree of apprehension to the needling or the electrical current that may otherwise have had an impact on the results. Consequently, the HRV changes were attributable to the study's physiotherapy technique, which it is considered important to acknowledge in order to avoid serious vasovagal reactions such as vasovagal syncope.

The analysis of HRV has been applied to several pathological conditions¹⁶ in addition to sports performance.^17,18 In this context, papers on HRV have been identified involving therapeutic massage,^35
–37 craniosacral therapy,³⁸ acupuncture,^39
–41 and PNE.²³ Some authors have reported an increase in HRV based on increased parasympathetic activity that has been translated into techniques designed to reduce physiological stress.^35,36,38 Other authors have reported increased sympathetic activity during acupuncture needling, which is generally followed by increased parasympathetic tone afterwards.^39
–41 In this study, comparing two experimental groups of homogeneous subjects, a trend toward increased parasympathetic activity and decreased sympathetic activity was observed in both groups, although the changes were only statistically significant in the PNE-G. In light of the fact that US-guided PNE is a needling technique, it is likely that subjects were experiencing an autonomic imbalance, and this should be borne in mind during clinical treatment to avoid potentially serious adverse reactions such as vasovagal syncope.

Limitations and recommendations

The PNE technique causes an autonomic imbalance. Therefore, the technique has a local and a segmental effect. It would be interesting for future studies to clarify the local and segmental therapeutic mechanisms of this technique.

However, HRV changes are different between females and males.^42
–44 Generally, women have a higher prevalence of parasympathetic activity than men do, and therefore women show greater HRV.⁴² Also, it would be interesting for future studies to determine the autonomic response to PNE in women.

Conclusion

In conclusion, application of the US-guided PNE technique caused a measurable increase in parasympathetic activity (detected by HRV), which was due to the combination of needle puncture and electric current.

Footnotes

Author Disclosure Statement

There are no financial interests to disclose.

References

Valera-Garrido

, Minaya-Muñoz

, Medina-Mirapeix

. Ultrasound-guided percutaneous needle electrolysis in chronic lateral epicondylitis: Short-term and long-term results. Acupunct Med, 2014; 32:446–454.

Abat

, Valles

, Gelber

, et al. An experimental study of muscular injury repair in a mouse model of notexin-induced lesion with EPI^® technique. BMC Sports Sci Med Rehabil, 2015; 17:7.

Abat

, Valles

, Gelber

, et al. Molecular repair mechanisms using the intratissue percutaneous electrolysis technique in patellar tendinosis. Rev Esp Cir Ortop Traumatol, 2014; 58:201–205.

Moreno

, Mattiussi

, Nuñez

. Therapeutic results after ultrasound-guided intratissue percutaneous electrolysis (EPI^®) in the treatment of rectus abdominis-related groin pain in professional footballers: A pilot study. J Sports Med Phys Fitness, 2016; 56:1171–1178.

Arias-Buría

, Truyols-Domínguez

, Valero-Alcaide

, et al. Ultrasound-guided percutaneous electrolysis and eccentric exercises for subacromial pain syndrome: A randomized clinical trial. Evid Based Complement Alternat Med, 2015; 2015:315219.

Abat

, Diesel

, Gelber

, Polidori

, et al. Effectiveness of the intratissue percutaneous electrolysis (EPI^®) technique and isoinertial eccentric exercise in the treatment of patellar tendinopathy at two years follow-up. Muscles Ligaments Tendons J, 2014; 4:188–193.

Abat

, Gelber

, Polidori

, et al. Clinical results after ultrasound-guided intratissue percutaneous electrolysis (EPI) and eccentric exercise in the treatment of patellar tendinopathy. Knee Surg Sports Traumatol Arthrosc, 2015; 23:1046–1052.

de Almedia Mdos

, Guerra Fda

, de Oliveira

, et al. A hypothesis for the anti-inflammatory and mechanotransduction molecular mechanisms underlying acupuncture tendon healing. Acupunct Med, 2014; 32:178–182.

Speed

. Acupuncture's role in tendinopathy: New possibilities. Acupunct Med, 2015; 33:7–8.

10.

Inoue

, Nakajima

, Oi

, et al. The effect of electroacupuncture on tendon repair in a rat Achilles tendon rupture model. Acupunct Med, 2015; 33:58–64.

11.

de Almedia Mdos

, de Freitas

, Oliveira

, et al. Acupuncture increases the diameter and reorganization of collagen fibrils during rat tendon healing. Acupunct Med, 2015; 33:51–57.

12.

Chou

, Hsieh

, Kuan

, et al. Needling therapy for myofascial pain: Recommended technique with multiple rapid needle insertion. BioMedicine, 2014; 4:13.

13.

Liu

, Wang

, Xu

, et al. Effect of electroacupuncture versus pelvic floor muscle training plus solifenacin for moderate and severe mixed urinary incontinence in women: A study protocol. BMC Complement Altern Med, 2014; 14:301.

14.

Dunning

, Butts

, Mourad

, et al. Dry needling: A literature review with implications for clinical practice guidelines. Phys Ther Rev, 2014; 19:252–265.

15.

Dommerholt

, Mayoral-del Moral

, Gröbli

. Trigger point dry needling. J Man Manip Ther, 2006; 14:70E–87E.

16.

Task Force of ESC and NASPE: Heart rate variability, standards of measurement, physiological interpretation, and clinical use. Circulation, 1996; 93:1043–1065.

17.

De la Cruz

, López

, Naranjo

. Analysis of heart rate variability at rest and during aerobic exercise: A study in healthy people and cardiac patients. Br J Sports Med, 2008; 42:715–720.

18.

Naranjo

, De la Cruz

, Sarabia

, et al. Heart rate variability: A follow-up in elite soccer players throughout the season. Int J Sports Med, 2015; 36:881–886.

19.

Kitagawa

, Kimura

, Yoshida

. Spectral analysis of heart rate variability during trigger point acupuncture. Acupunct Med, 2014; 32:273–278.

20.

Sakai

, Hori

, Umeno

, et al. Specific acupuncture sensation correlates with EEGs and autonomic changes in human subjects. Auton Neurosci, 2007; 133:158–169.

21.

Hideaki

, Tatsuya

, Shogo

, et al. Effect of 100 Hz electroacupuncture on salivary immunoglobulin A and the autonomic nervous system. Acupunct Med, 2015; 33:451–456.

22.

Huang

, Zhong

, Chen

, et al. Effect of acupuncture at HT7 on heart rate variability: An exploratory study. Acupunct Med, 2015; 33:30–35.

23.

De la Cruz

, Albornoz

, García

, et al. Autonomic responses to ultrasound-guided percutaneous needle electrolysis of the patellar tendon in healthy male footballers. Acupunct Med, 2016; 34:275–279.

24.

Albornoz

, Rebollo

, García

. Escala de Aprensión Psicológica Personal (EAPP) en Fisioterapia. Rev Iberoam Fisioter Kinesol, 2005; 2:77–87.

25.

Valera-Garrido

, Minaya-Muñoz

, eds. Fisioterapia Invasiva. 1st ed. Barcelona: Elsevier España, S.L., 2013.

26.

Beggs

, Bianchi

, Bueno

, Cohen

, et al. Musculoskeletal ultrasound technical guidelines. Online document at: www.essr.org/html/img/pool/ knee.pdf, accessed September 1, 2016 .

27.

Kamen

, Tonkin

. Application of the poincaré plot of heart rate variability: A new measure of functional status in heart failure. Aust N Z J Med, 1995; 25:18–26.

28.

Valera-Garrido

, Minaya-Muñoz

, eds. Fisioterapia Invasiva. 2nd ed. Barcelona: Elsevier España, S.L., 2016.

29.

Brennan

, Palaniswami

, Kamen

. Do existing measures of Poincaré plot geometry reflect nonlinear features of heart rate variability?. IEEE Trans on Biomed Eng, 2001; 48:1342–1347.

30.

Mourot

, Bouhaddi

, Perrey

, et al. Quantitative Poincaré plot analysis of heart rate variability: Effect of endurance training. Eur J Appl Physiol, 2004; 91:79–87.

31.

Hoshi

, Pastre

, Vanderlei

, et al. Poincaré plot indexes of heart rate variability: Relationships with other nonlinear variables. Auton Neurosci, 2013; 177:271–274.

32.

Naranjo

, De la Cruz

, Sarabia

, et al. Two new indexes for the assessment of autonomic balance in elite soccer players. Int J Sports Physiol Perform, 2015; 10:452–457.

33.

MaYun-Tao

, Zang

, eds. Biomedical Acupuncture for Pain Management: An Integrative Approach. St. Louis, MO: Elsevier, 2005.

34.

Carvajal

, Álvarez

, Medina

, et al. Efectos de la electrolisis percutánea en el tendón rotuliano sobre la temperatura local y contralateral medida con termografía infrarroja. Rev Fisioter Invasiva, 2016; 1:18–25.

35.

Wälchli

, Saltzwedel

, Krüerke

, et al. Physiologic effects of rhythmical massage: A prospective exploratory cohort study. J Altern Complement Med, 2014; 20:507–515.

36.

Chuang

C-Y

, Tsai

C-N

, Kao

M-T

, et al. Effects of massage therapy intervention on autonomic nervous system promotion in integrated circuit design company employees. IFMBE Proc, 2014; 43:562–564.

37.

Tbuttagat

, Eungpinichpong

, Chatchawan

, et al. The immediate effects of traditional Thai massage on heart rate variability and stress-related parameters in patients with back pain associated with myofascial trigger points. J Bodyw Mov Ther, 2011; 15:15.

38.

Girsberger

, Bänziger

, Lingg

, et al. Heart rate variability and the influence of cranosacral therapy on autonomous nervous system regulation in persons with subjective discomforts: A pilot study. J Integr Med, 2014; 12:156–161.

39.

Sakai

, Hori

, Umeno

, et al. Specific acupuncture sensation correlates with EEGs and autonomic changes in human subjects. Auton Neurosci, 2007; 30; 133:158–169.

40.

Hideaki

, Tatsuya

, Shogo

, et al. Effect of 100 Hz electroacupuncture on salivary immunoglobulin A and the autonomic nervous system. Acupunct Med, 2015; 33:451–456.

41.

Huang

, Zhong

, Huang

, et al. Effect of acupuncture at HT7 on heart rate variability: An exploratory study. Acupunct Med, 2015; 33:30–35.

42.

Koening

, Thayer

. Sex differences in healthy human heart rate variability: A meta-analysis. Neurosci Biobehav Rev, 2016; 64:288–310.

43.

Moodithaya

, Avadhany

. Gender differences in age-related changes in cardiac autonomic nervous function. J Aging Res, 2012; 2012:679345.

44.

Reulecke

, Charleston-Villalobos

, Voss

, et al. Men and women should be separately investigated in studies of orthostatic challenge due to different gender-related dynamics of autonomic response. Physiol Meas, 2016; 37:314–332.