Bioimpedance Spectroscopy in Subjects with Auricular Vagus Nerve

Stimulation
Y. Ülgen, B. Büyüksaraç, B. Tunç and H. Solmaz
Department of Biomedical Engineering, Faculty of Engineering and Life Sciences
Bahçeşehir University, Istanbul -Turkey.

Abstract: In this paper, single side body impedance spectroscopy is performed between wrist and ankle
in response to auricular transcutaneous electrical vagus stimulation, to obtain the Cole parameters Ro, R∞
and fc. Following a resting period of 5 minutes in supine position, left ear vagus nerve was stimulated for 3
minutes and BIS performed every 10 seconds, on 8 healthy subjects with age: 23,6+1,5 years, height:
175+8,3 cm; weight: 69,5+18,7 kg and BMI: 22,6+5,5 kg/cm2. Cole circles moved to the right (inreased Ro
and R∞ ) with stimulation while the centers shifted upwards. Intracellular and extracellular resistances and
the capacitance Cm all increased at the onset of vagus stimulation; resulting in a decrease of 3,4+1.33% in fc.
Changes in Ri mainly indicate water shift between cells and extracellular fluid; physiological changes
occurred on cell membrane. fc independent of the participants’ anthropomorphic factors could be used as
an indicator to vagus nerve stimulation, without further normalization. This preliminary study was realized
with a limited number of participants; however, to reach statistically meaningful results measurements will
be performed over a larger sample size.

Keywords: bioimpedance spectroscopy, extracellular and intracellular body fluids, transcutaneous vagus nerve
stimulation.

I. INTRODUCTION
Vagus nerve, is targeting almost every major organ and highly involved in the parasympathetic
nervous system, capable of regulating the autonomic nerve function for maintaining homeostasis [1, 2,
3, 4]. Current reasearch efforts indicated that the stimulation of vagus nerve can have therapeutic
effects over wide range of physiological conditions including migraine, tinnitus, trigeminal nevralgy,
gastrointestinal motility, stress, etc [5 - 10]. Devices and methods designed to stimulate the auricular
branch of vagus nerve require a controller that operates in the closed loop system to regulate stimulus
parameters such as amplitude, pulse width, pulse frequency and duration. Commonly used feedback
parameters are heart rate, blood pressure and respiratory rate.
In the present study, tragus and cavum conchae regions of the outer ear were electrically
stimulated at the perception threshold level and body impedance spectroscopy (BIS) from 3 kHz to
1MHz was performed, between wrist and ankle, to obtain the Cole parameters (Ro, R∞, fc and α) and
investigate if they could be potential feedback parameters in vagus stimulation [11, 12, 13, 14, 15, 16].

II. METHOD

Since the ear vagus nerve has no direct fibers connecting to the heart and no known physiological
side effects exist, both ears are be safe for applying (atVN) [17-18]. Electrical pulses were applied at
the left ear since the efferent vagus fibers to the heart are located on the right side and stimulus current
was continuously tracked [19]. The study was approved by the Ethics Committee of Bahcesehir
University and conducted in accordance with the Declaration of Helsinki. All participants signed an
informed consent form prior to experiments.
For transcutaneous auricular stimulation of the vagus nerve taVNS the tragus (ear canal) and
cymba concha areas were targeted. Tragus is innervated by the auricular branch of the vagus nerve.
Excessive activation of the vagus nerve typically leads to a reduction in heart rate, blood pressure, or
respiratory rate [20 - 24]. Parasympathetic overcompensation of the sympathetic nervous system
response can cause vasovagal syncope because of a sudden drop in blood pressure and heart rate.
Biphasic pulse waveform was applied to avoid the electrolytic and iontophoretic effects of the
unidirectional current. Biphasic pulses reduce the threshold voltage by the mechanism that the positive
phase does not synchronize as many cells since the negative phase removes the residual charges caused
by the first phase.
Different nerve groups respond to different pulse widths and usually 50 µs to 500 µs is a good
range of pulse widths that can be used while longer pulses generate a much stronger stimulation to
elicit the morphine like endorphin release.
 For the stimulation of the vagus nerve, the instrumentation system consisted of a (TENS) (Comfy
TENS, model EV-804, by iSTIM) current stimulator and an electrode assembly (VAGUSTIM  by  Health  
Technologies) (Figure 1). Few studies have systematically investigated the optimal settings suggesting
the frequency range of 1 to 30 Hz, pulse widths from 130 to 1000 µs and an average intensity of about
5 mA to obtain a therapeutical effect [25]. In this study, the stimulator was operated in the normal
continuous mode with asymmetrical biphasic rectangular pulses and PW and PR were set to 250 µs and
25 Hz respectively. The output current peak level was set to an average of 4 mA in the subjects.

Figure: 1 Vagus stimulation and BIS measurements.

Figure: 2 Electrode connection and stimulus current monitoring.

BIS was performed by using the Impedimed SFB7 (Impedimed Ltd. Queensland, Australia) a
single channel, tetrapolar bioimpedance device; calibrated with its own RRC calibrator. Subjects had to
remove shoes and socks, all metallic objects and lied in supine position. Measurements were taken
from the left side of the body and both heart rate and blood oxygenation were monitored continuously.
Electrodes were placed on hand and foot on the same side, 5 cm apart when measured from center to
center. BIS measurements were taken from 5 male and 3 female subjects (age 22-27 years; height 158-
187 cm and weight 46,5-107 Kg). The vagus stimulating bipolar electrode assembly was attached to the
left ear and the TENS unit output level was adjusted until participants reported a feeling of sensation
without pain, while observing the stimulus current on the storage oscilloscope (Fig. 2). Following 3
minutes of resting period, a total vagus stimulation of 3 minutes was applied and the IMPEDIMED
provided BIS measurements every 10 seconds. Post stimulus measurements lasted for 3 minutes.

III. THEORY          

Body fluids include intracellular fluids ICF and extracellular fluids ECF composed of plasma and
interstitial fluids. The main ions in ECF are Na+ and Cl−; K+ and PO−4 in ICF [20]. The electrical
properties of the body is modeled as in Fig. 3, as a capacitive cell membrane with conductive
extracellular and intracellular fluids. ECF and ICF charge separation across this membrane causes a
capacitive effect. Cole analysis and the Hanai mixture theory were used: the conductive properties of
the body were modeled as conductive medium containing an intracellular volume of membrane-bound
cells in suspension [13]. In biological tissues, plot of the real versus the imaginary parts of the
measured impedance Z*= Re(Z*) + jIm(Z*) results in a semicircle with the center below the real axis.
This depressed semicircular locus is called the Cole plot and represented by the equation below:

Z* = R∞ + (Ro - R∞ )/(1 + (jωτ )(1-a)) (1)

 Ro (Ω) and R∞ (Ω) the zero frequency and infinite frequency resistances are determined by
extrapolation as the intersection points of the Cole circle with the real axis; τ =1/2π fc is the relaxation
time constant. At low frequencies, cell membranes are electrically insulating and the electrical currents
run in the ECF only. The intracellular space resistance Ri is then equal to Ro R∞ /(Ro-R∞). Capacitive
effects of cell membranes are expressed in a constant phase angle impedance ZCPA:

ZCPA = K (jω ) -1 α
(2)

where K is a constant, and 0 < α < 1. The characteristic frequency corresponding to the top of the
circle when Im(Z*) is maximal is given by:

fc = (2π (Ri + Re)Cm ) -1 (3)

Figure: 3 The electrical model of body cells and the Cole circle. Re and Ri are the ECF and ICF resistances.

The accuracy of multifrequency impedance analysis depends primarily on the number of

measurement frequencies (256 frequencies with Impedimed SFB7) [21, 22].

IV. RESULTS

BIS on the IMPEDIMED with the calibrator provided by the manufacturer produced the  
following   performance,   Ro   (Ω)=   604.6   +   0.03%   and   R∞   (Ω) =   402.0   +   0.04%, with a precision of
0.04%.   BIS measurements results (Ro, R∞, Cm, fc ) from 8 subjects are displayed in Figs 4, 5, 6, 7, 8 and
9, during resting, stimulation and post stimulation periods. The period of time from 200 seconds to 400
seconds corresponds to the stimulus phase. All graphs were produced using the MatLab software. Data
from subject F1 and F3 exhibited too much fluctuations and were not considered for further
evaluation.

Figure: 4 Extrapolated Ro from 8 subjects ordered from left to right as M1, F1, M2, F2, F3, M4, M5
and M3. Vagus stimulation is from 200 s to 400 s.
 Figure: 5 Extrapolated R∞ from 8 subjects.

Figure: 6 Membrane capacitance Cm from 8 subjects.

Figure: 7 Characteristic frequency fc from the same subjects.

V. DISCUSSION
 With vagus stimulation, the Cole circles moved to the right (inreased Ro and R∞ ) while the centers
shifted upwards with the parameter α reduced.

Figure: 7 Cole circles for subject M4. Blue, green and red refer to10 s, 350 s and 760 s under vagus stimulation.

Intracellular and extracellular resistances and the capacitance Cm all increased at the onset of
vagus stimulation. Accordingly, the characteristic frequency fc decreased. Changes in Ri mainly
indicate water shift between cells and extracellular fluid; physiological changes occurred on cell
membrane. From Eq. 3, fc is independent of the participants’ anthropomorphic factors and could be
used as an indicator to vagus nerve stimulation, without normalization.

Table 1 Relative change in Cole parameters Ro, R∞ and fc with vagus stimulation.

Height (cm) Weight (Kg) Δ Ro (%) Δ R∞ (%) Δ fc (%)

M1 180 56 0,75 -0,21 -2,68
M2 187 74 1,67 1,02 -2,98
F2 158 46,5 -0,40 0,76 -4,57
M4 177 107 0,76 0,47 -1,30
M5 175 82 1,51 0,90 -4,20
M3 178 69 0,44 2,06 -4,74
Mean 0,79 0,83 -3,41
SD 0,75 0,75 1,33

Dilation of peripheral vasculature would mobilize water from other fluid compartments to
compensate the decrease in plasma water and body dehydration [28]. The autonomic nervous system
regulates balance of water and electrolytes automatically without a person’s conscious effort. Because
changes in solute concentrations lead to immediate changes in water distribution, regulation of fluid
and electrolyte balances are tightly intertwined [28]. Dilation of peripheral vasculature would mobilize
water from other fluid compartments to compensate the decrease in plasma water and body
dehydration.
Stimulus level was the threshold of sensation to minimize the risks to the subjects. But, with
threshold level stimuli applied, in some of the subjects the vagus nerve did not respond. Parameters
such as waveform, magnitude, frequency, burst frequency, pulse duration and periodic or intermittent
operation could also be regulated by using whole body impedance, as a physiological feedback tool
complementary to heart rate and blood pressure.
 
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