CHARACTERIZATION OF THE ENCAPSULATION

PROCESS OF DEEP BRAIN STIMULATION ELECTRODES

USING IMPEDANCE SPECTROSCOPY IN A RODENT MODEL

K. Badstübner

, T. Kröger

, E. Mix

, U. Gimsa

, R. Benecke

and J. Gimsa

2**

Department of Neurology, University of Rostock, Gehlsheimer Str. 20, 18147 Rostock, Germany

Chair of Biophysics, Institute of Biology, University of Rostock, Gertrudenstr. 11A, 18157 Rostock, Germany

Research Unit Behavioural Physiology, Leibniz Institute for Farm Animal Biology,

Wilhelm-Stahl-Allee 2,18196 Dummerstorf, Germany

* These authors contributed equally to the work.

** Corresponding author.

Keywords: EIS, Intracerebral electrodes, Basal ganglia, Subthalamic nucleus, Rat brain, Chronic instrumentation,

6-OHDA, Parkinson’s disease.

Abstract: Deep brain stimulation (DBS) is effective for the treatment of patients with Parkinson’s disease (PD),

especially in advanced stages which are refractory to conventional therapy. Despite of the regular use in

clinical therapy, rodent models for basic research into DBS are not routinely available. The main reason is

the geometry difference from rodents to humans, imposing larger problems in the transfer of the stimulation

conditions than from primates to humans. For rodents, the development of miniaturized mobile stimulators

and stimulation parameters, as well as improved electrode materials and geometry are desirable. The

impedance of custom made, cylindrical (contact diameter 200 µm, length 100 µm), platinum/iridium

electrodes has been measured in vivo for two weeks to characterize the influence of electrochemical

processes and of the adherent cell growth at the electrode surface. During the encapsulation process, the real

part of the electrode impedance at 10 kHz doubled with respect to its initial value after a characteristic

decrease by approximately one third at the second day. An outlook is given on further investigations with

different electrode designs for long-term DBS.

1 INTRODUCTION

Parkinson’s disease (PD) is a widespread

degenerative disorder of the central nervous system

that affects motor function, speech, cognition and

vegetative functions. The cardinal symptoms such as

tremor, rigidity, bradykinesia and postural instability

result mainly from the death of dopaminergic cells

in the substantia nigra pars compacta and the

subsequent lack of dopaminergic inputs into the

striatum. This causes an alteration of the activity

pattern in the basal ganglia (Braak and Braak, 2000).

Deep brain stimulation (DBS) is a novel therapeutic

option for PD as well as an increasing number of

neuropsychiatric disorders. Before DBS became a

therapeutic intervention, electric stimulation of basal

ganglia had been used to guide neurosurgeons to the

precise position for a surgical lesion, the ultimate

therapy of a late-stage PD. The main advantage of

DBS over surgical lesions is the reversibility and

possibility to modulate stimulation parameters

(Benabid et al., 1987). The small volume of the

target region for DBS in the human brain requires a

highly specific adaption of the electrodes which

need to be thoroughly tested in animal models,

including different materials and geometries. So far,

DBS data of animal models of PD are scarce. During

in vivo stimulation, the properties of the DBS

electrodes are changing as a function of time caused

by electrochemical processes at the surface of the

implant and the subsequent tissue response (Gimsa

et al., 2005). The tissue response is a foreign

substance reaction. Its intensity depends on the

material (Grill and Mortimer, 1994) and is correlated

with the thickness of the adventitia finally

125

Badstübner K., Kröger T., Mix E., Gimsa U., Benecke R. and Gimsa J..

CHARACTERIZATION OF THE ENCAPSULATION PROCESS OF DEEP BRAIN STIMULATION ELECTRODES USING IMPEDANCE SPECTROS-

COPY IN A RODENT MODEL.

DOI: 10.5220/0003790301250130

In Proceedings of the International Conference on Bio-inspired Systems and Signal Processing (BIOSIGNALS-2012), pages 125-130

ISBN: 978-989-8425-89-8

 2012 SCITEPRESS (Science and Technology Publications, Lda.)

encapsulating the implant (Wintermantel et al.,

2002). Adventitia formation causes a steady change

in the impedance of the electrodes leading to

changes in the attenuation of the stimulating signal.

As a result, the efficiency of the surrounding tissue

stimulation is changing (Lempka et al., 2009 and

2011; Grill and Mortimer, 1994). One opportunity to

minimize this problem is to choose an appropriate

electrode material. Previous investigations of our

group (Gimsa et al. 2006; Henning et al., 2007) have

shown that the use of stainless steel electrodes is not

appropriate because of the corrosion and erosion

processes intensified by electrolytic electrode

processes. Electrochemically induced alterations are

negligible for inert platinum electrodes, even though

electrode processes may still influence the

surrounding tissue (Gimsa et al., 2005). For an

optimal adjustment of the DBS signal, the kinetics of

the electrode impedance alterations caused by the

adventitia formation must be taken into account

(Lempka et al., 2009 and 2011).

2 MATERIALS AND METHODS

2.1 Animal Treatment

A number of 30, adult, male Wistar Han rats (240-

260 g) were obtained from Charles River

Laboratory, Sulzfeld, Germany) and housed under

temperature-controlled conditions in a 12 h light-

dark cycle with conventional rodent chow and water

provided ad libitum. The rats were subject to the

following treatments:

 anesthesia

 6-OHDA-lesioning (28 rats)

 electrode implantation (6 rats)

 impedance measurement (2 rats)

The study was carried out in accordance with

European Community Council directive 86/609/EEC

for the care of laboratory animals and was approved

by Rostock’s Animal Care Committee (LALLF M-

V/TSD/7221.3-1.2-043/06).

2.1.1 Anesthesia

The rats were anesthetized by ketamine-

hydrochloride (10 mg per 100 g body weight, i.p.,

Ketanest S

, Pfizer, Karlsruhe, Germany) and

xylazine (0,5 mg per 100 g body weight, i.p.,

Rompun

, Pfizer). Before surgery, the eyes of the

rats were medicated with Vidisic (Bausch and

Lomb, Berlin Germany). After surgery, the wound

was sutured and the rats received 0.1 ml

novaminsulfone (Ratiopharm, Ulm, Germany) and

4 ml saline subcutaneously. Rats were exposed to

red light (Petra, Burgau, Germany) until

normalization of vital functions.

2.1.2 6-Hydroxydopamine (6-OHDA)

Lesioning

The 6-OHDA (Sigma, Deisenhofen, Germany)

lesioning was performed by stereotactic surgery in

adaption to Kirik et al. (1998). The lesions of the

right medial forebrain bundle of rats were made by

injection of 26 µg 6-OHDA in 4 µl saline with 1 g/l

ascorbic acid delivered over 4 min via a 5 µl

hamilton microsyringe (Postnova Analytics,

Landsberg/Lech, Germany). The coordinates relative

to bregma were: anterior-posterior

(AP) = -2.3 mm, medial-lateral (ML) = 1.5 mm and

dorsal-ventral (DV) = -8.5 mm (Paxinos and

Watson, 1998).

2.1.3 Electrode Implantation

Electrodes were implanted into the subthalamic

nucleus (STN), which is the most common target

region for treatment of PD patients. The surgical

procedure was performed using a stereotactic frame

(Stoelting, Wood Dale, IL, USA) modified

according to Harnack et al. (2004). To support the

surgical procedure, a cold light source (KL 1500

LCD, Schott, Mainz, Germany) was used in

combination with a stereo-microscope (Leica,

Wetzlar, Germany). The skull was opened by a

dental rose-head bur (Kaniedenta, Germany). The

coordinates relative to bregma were: anterior-

posterior (AP) = -3.5 mm, medial-lateral

(ML) = 2.4 mm and dorsal-ventral (DV) = -7.6 mm

(Paxinos and Watson, 1998). A dental drill was used

to bore an additional hole in the skull for an anchor

screw. The electrode was fixed to the skull by an

adhesive-glue bridge (Technovit 5071, Heraeus,

Germany) to the anchor screw.

Counter-electrode wires were implanted into the

neck of the rats.

2.2 Stimulation- and Counter-

electrodes

For impedance measurements, we designed unipolar

platinum-iridium (Pt/Ir) microelectrodes which were

covered with polyesterimide insulation and custom-

made by Polyfil (Zug, Switzerland; Figure 1).

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126

Figure 1: Photograph (a) and scheme (b) of the unipolar

Pt/Ir electrode. The electrode pole was a round wire made

from Pt90Ir10 with a diameter of 200 µm. The length of

the non-insulated tip of the electrode pole was 100 µm.

The insulation consisted of polyesterimide 180 with a

thickness of 25 µm (Nowak et al., 2011).

Dental wires made of biocompatible, nickelfree steel

alloy (18% Cr, 18% Mn, 2% Mo, 1% N, remander

iron) of 1 mm diameter and 10 mm length were used

as counter electrodes (see: Figure 4) in combination

with the unipolar DBS electrodes (Figure 1).

Electrochemical electrode effects were negligible

at the counter electrode due to the low current

density at its large surface.

2.3 Electric Impedance Spectroscopy

(EIS)

Electric impedance spectroscopy (EIS) is a common

measureing technique for determining the electrical

properties of tissues (Foster and Schwan, 1989). It is

used in a wide range of applications, such as breast

cancer detection (Kerner et al., 2002), the

monitoring of the lung volume (Adler et al., 1997)

and in material sciences. EIS is nondestructive and

therefore suitable for the characterization of the DBS

electrodes during the encapsulation process.

Equipment

The EIS measurements were conducted with an

impedance spectrometer Sciospec ISX3 (Sciospec

Scientific Instruments, Pausitz, Germany) and a test

fixture HP16047D (Hewlett-Packard, Japan)

connected to a personal computer with Sciospec-

measuring software. The two connectors of the test

fixture were connected to the DBS and the counter

electrodes.

Measurement

To characterize the electrode properties during

encapsulation, the impedance was recorded in the

frequency range from 100 Hz to 10 MHz over a

period of two weeks after implantation. Frequency

range, amplitude, number of points and the

averaging of the impedance spectrometer were

programmed by the measuring-software (Sciospec).

401 frequency points were logged which were

distributed equidistantly over a logarithmic

frequency scale. The measuring voltage (peak to

peak) was 12.5 mV

. The measuring-software

logged the measuring data of the impedance

spectrometer (real and imaginary parts of the

impedance vs. frequency), saving them to a file.

Before each measurement, the impedance

spectrometer was calibrated by open, short and load

measurements. Each measurement was repeated

three times to improve the statistical significance.

The measurements were repeated every day for one

week and every second day during the second week.

The stimulation pulse usually applied in DBS has

a frequency of 130 Hz and a pulse width of 60 µs.

Because of the steep slopes of the needle-shaped

pulse, the signal is rich in high harmonic frequencies

(Gimsa et al. 2005). For this reason, we measured

the impedance within the wide frequency range from

100 Hz to 10 MHz, which is beyond the range of up

to 10 kHz reported by Lempka et al. (2009).

Impedance theory

The electrical impedance describes the magnitude

ratio between the applied AC voltage and the

resulting current flowing with a certain phase shift.

Mathematically speaking, the impedance Z* is a

complex number with the unit [Ω], which is

composed of a real (Z´) and an orthogonal imaginary

part (Z´´) marked by the complex unit j =

√

1

Z* = Re(Z*) +

·Im(Z*) = Z´+

Z´´ (1)

For interpretation of the measuring data, an

equivalent circuit model is required to be fitted to

the measuring data. The aim was to model

electrochemical processes and adherent cell growths

by combinations of resistors, capacitors and constant

phase elements (Lempka et al., 2009).

Data analysis

The logged data were transferred to Matlab (The

MathWorks™, Version 7.9.0.529) to calculate

means and standard deviations. For their graphic

representation, they were finally copied to Sigma

Plot 11.0 (Systat Software, 11.0, Build 11.2.0.5).

2.4 Electron Microscopy Study of

Electrode Encapsulation by Tissue

Concentric bipolar microelectrodes with an inner

CHARACTERIZATION OF THE ENCAPSULATION PROCESS OF DEEP BRAIN STIMULATION ELECTRODES

USING IMPEDANCE SPECTROSCOPY IN A RODENT MODEL

127

pole diameter of 75 µm and an outer pole diameter

of 250 µm (CB CSG75; FHC, Bowdoinham, ME)

were placed into the STN of two anesthetized rats.

The rats were stimulated for 3 h with biphasic

constant-current pulses with a repetition frequency

of 130 Hz and pulse duration of 60 µs at 250 µA

with a stimulus generator (Multichannel Systems,

Reutlingen, Germany). The electrodes were removed

and one electrode was incubated in trypsin solution

(Trypsin-EDTA (1x) in HBSS W/O CA&MG

W/EDTA.4NA, Gibco, UK) at 37°C for 1 h. The

other electrode was postfixed overnight in 4%

glutaraldehyde (Merck, Germany) in PBS.

Electrodes were washed, fixed in 4%

glutaraldehyde in PBS, washed again, postfixed in

1% osmium tetroxide, dehydrated in acetone, and

subjected to critical-point drying (EMITECH K850,

Ashford, Kent, UK). The samples were sputtered

with colloidal gold using a Sputter Coater (BAL-

TEC SCD 004, Schalksmühle, Germany) before

examination in the SEM (DSM 960 A, Zeiss,

Oberkochen, Germany).

3 RESULTS

3.1 Stimulation- and Counter

Electrode Implantation

As a first test, the localization of the electrode tip in

the target region was verified by ink injection via a

5 µl hamilton microsyringe of approximately the

same size as the implanted stimulation electrode

(Figure 2).

Figure 2: Rat brain fixed in formalin, (a): top view of a rat

brain with a puncture resembling an electrode canal (1),

(b): sagittal section of one hemisphere with the puncture

(1), (c): hemisphere with tissue removed to the injection

canal, (d): enlarged photo of (c) with ink injection canal

(2), striatum (3), and basal ganglia (4).

Chronic instrumentation was established and

applied to freely moving PD -rats (Figure 3). For

this, a commercial rat jacket (Lomir Biomedical,

Quebec, Canada) with a backpack was used. All

electronic components (Figure 2) were located in the

backpack. This allowed for a stimulation of up to six

weeks when the batteries were changed every

second day.

For the EIS measurements, counter electrodes were

pierced through the intact scalp close behind the cut

for the implantation of the stimulation electrode. An

example of implantation of a counter electrode using

dental wire of biocompatible steel alloy is shown in

Figure 4.

Figure 3: Photograph of a rat that was anesthetized for

implantation of chronic instrumentation consisting of: (1):

Pt/Ir electrode, (2): counter-electrode, (3): plug-connector

electrodes, (4): battery, (5): DBS-stimulator and (6):

carrying bag.

Figure 4: Implanted dental wire of biocompatible steel

alloy used as counter electrode for the EIS measurement.

3.2 Impedance Measurement

Our data showed the general tendency of an

impedance increase during the encapsulation

process. The measuring results are shown in

Figure 6. After 12 days, the value of Z´ (Figure 6a)

has almost doubled compared to its initial value at

10 kHz. In parallel, also the absolute value of Z´´

(Figure 6b) increased. The results match with

findings of Lempka et al. (2009) for the monkey

brain. The main reason for the impedance increase is

the formation of the adventitia as a foreign substance

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128

reaction. Indeed, the adventitia covered the electrode

surface as shown by Lempka et al. (2009). Our own

electron-microcopy study showed that stimulation

electrodes get overgrown by cells within 3h post

implantation (Figure 5).

Figure 5: Scanning electron-microscopical images of elec-

trodes that had been implanted and used for 3 h of stimu-

lation. (a): Electrode cleaned by trypsination prior to

electron microscopy. (b): Electrode fixed with adhering tissue.

Figure 6: In vivo impedance measurements with a unipolar

Pt/Ir electrode in combination with a steel alloy counter-

electrode ((a) Z´; (b) Z´´).

Our data showed an impedance decrease at the

second day after implantation followed by a

significant increase from the third day onwards.

Interestingly, the same impedance behavior has

already been reported by Lempka et al. (2009, p. 6,

Figure 4). However, no explanation has been given

for this phenomenon.

4 CONCLUSIONS

AND PERSPECTIVES

Pilot experiments in the rat model have shown that

the impedance of a unipolar DBS electrode is

significantly increasing with time after implantation.

In future experiments, different biocompatible

electrode materials such as titanium, diverse

titanium alloys, platinum, gold, silver and stainless

steel will be tested. For these tests, an array of

various counter electrodes will be pierced into the

necks of a number of rats.

In addition, bipolar electrodes possessing modified

configurations such as shifted tips will be used to

test the effects of non-axial symmetric field

distributions.

The equivalent circuit model will be improved

for a better understanding of the measuring data in

order to extract encapsulation parameters. These

investigations aim at clarifying the phenomenon of

the impedance drop at the second day after

implantation.

Potential effects at the electrode-tissue-interface

will be analyzed by histological, immunochemical

and electron-microscopical methods.

ACKNOWLEDGEMENTS

K.B. is grateful for a stipend of the German

Research Foundation (DFG, Research Training

Group 1505/1 “welisa”). T.K. acknowledges

financing by a project of the Federal Ministery of

Economics and Technology (BMWi, V230-630-08-

TVMV-S-031). Part of the work was conducted

within a project financed by the Federal Ministry of

Education and Research (BMBF, FKZ 01EZ0911).

The authors are grateful to Dr. J. Henning for help

with the electron microscopy study and would like

to thank the staff of the electron microscopy center

at the University of Rostock's Medical Faculty for

outstanding technical support.

Real

log( f ) / Hz

4567

Z´ / kOhm

Day 1

Day 2

Day 4

Day 5

Day 6

Day 8

Day 12

Imaginary

log ( f ) / Hz

4567

Z´´ / Ohm

-10

-8

-6

-4

-2

(a)

(b)

Day 1

Day 2

Day 4

Day 5

Day 6

Day 8

Day 12

CHARACTERIZATION OF THE ENCAPSULATION PROCESS OF DEEP BRAIN STIMULATION ELECTRODES

USING IMPEDANCE SPECTROSCOPY IN A RODENT MODEL

129

REFERENCES

Adler, A., Amyot, R., Guardo, R., Bates, J. H.,

Berthiaume, Y., 1997. Monitoring changes in lung air

and liquid volumes with electrical impedance

tomography. Journal of Applied Physiology. 83(5):

1762-7.

Braak, H., Braak, E., 2000. Pathoanatomy of Parkinson's

disease. Journal of Neurology. 247(2): II3-10.

Benabid, A. L., Pollak, P., Louveau, A., Henry, S., de

Rougemont, J., 1987. Combined (thalamotomy and

stimulation) stereotactic surgery of the VIM thalamic

nucleus for bilateral Parkinson disease. Applied

Neurophysiology. 50(1-6): 344-6.

Foster, K. R., Schwan, H. P., 1989. Dielectric properties of

tissues and biological materials: a critical review.

Critical reviews in biomedical engineering.

17(1): 25-104.

Gimsa, J., Habel, B., Schreiber, U., van Rienen, U.,

Strauss, U., Gimsa, U., 2005. Choosing electrodes for

deep brain stimulation experiments – electrochemical

considerations. Journal of Neuroscience Methods.

142(2): 251-65.

Gimsa, U., Schreiber, U., Habel, B., Flehr, J., van Rienen,

U., Gimsa, J., 2006. Matching geometry and

simulation parameters of electrodes for deep brain

stimulation experiments – numerical considerations.

Journal of Neuroscience Methods. 150(2): 212-27.

Grill, W. M., Mortimer, J.T., 1994. Electrical properties of

implant encapsulation tissue. Annals of Biomedical

Engineering. 22(1): 23-33.

Henning, J., 2007. Wirkungen der tiefen Hirnstimulation –

Analyse der Gen- und Proteinexpression in einem

optimierten Rattenmodell. Dissertation. University of

Rostock.

Harnack, D., Winter, C., Meissner, W., Reum, T., Kupsch,

A., Morgenstern, R., 2004. The effects of electrode

material, charge density and stimulation duration on

the safety of high-frequency stimulation of the

subthalamic nucleus in rats. Journal of Neuroscience

Methods. 138(1-2): 207-16.

Kerner, T. E., Paulson, K. D., Hartov, A., Soho, S. K.,

Poplack, S. P., 2002. Electrical impedance

spectroscopy of the breast: clinical imaging results in

26 subjects. IEEE Transactions on medical imaging.

21(6): 638-45.

Kirik, D., Rosenblad, C., Björklund, A., 1998.

Characterization of behavioral and neurodegenerative

changes following partial lesions of the nigrostriatal

dopamine system induced by intrastriatal

6-hydroxydopamine in the rat. Experimental

Neurology. 152(2): 259-77.

Lempka, S. F., Miocinovic, S., Johnson, M. D., Vitek, J.

L., McIntyre, C. C., 2009. In vivo impedance

spectroscopy of deep brain stimulation electrodes.

Journal of Neural Engineering. 6, 046001, 11pp.

Lempka, S. F., Johnson, M. D., Moffittt, M. A., Otto, K.

J., Kipke, D. R., McIntyre, C. C., 2011. Theoretical

analysis of intracortical microelectrode recordings.

Journal of Neural Engineering. 8(4):045006.

Macdonald, J. R., 1992. Impedance spectroscopy. Annals

of Biomedical Engineering. 20(3): 289-305.

Nowak, K., Mix, E., Gimsa, J., Strauss, U.,

Sriperumbudur, K. K., Benecke, R., Gimsa, U., 2011.

Optimizing a rodent model of Parkinson’s disease for

exploring the effects and mechanisms of deep brain

stimulation. Parkinson’s Disease. 2011: 414682.

Wintermantel, E., Ha, S-W., 2002. Medizintechnik mit

biokompatiblen Werkstoffen und Verfahren. Springer,

Berlin, 3

edition: 134-5.

Paxinos, G., Watson, C., The rat brain in stereotaxic

coordinates. Academic Press, San Diego, 1998.

BIOSIGNALS 2012 - International Conference on Bio-inspired Systems and Signal Processing

130