International Journal of Engineering Research and Development e-ISSN: 2278-067X, p-ISSN : 2278-800X, www.ijerd.

com Volume 5, Issue 9 (January 2013), PP. 22-29

Study the Window Effect of Rectangular Electrical Pulse in Membrane Potential of Dielectric Model of Osteoblast Cell under Different Microelectrodes
Swarup Sarkar1, Rajat Mahapatra2, Soumen Das3, Akash Ku Bhoi4
1,4

Applied Electronics and Instrumentation Department, Sikkim Manipal Institute of Technology Majitar, Rangpo, Sikkim-737136, India 2 Electronics and Communication Engineering Department, National Institute of Technology Durgapur, West Bengal, India. 3 School of Medical Science & Technology, Indian Institute of Technology Kharagpur, West Bengal, India

Abstract:- With refarances of bilayer dielectric model, for the spherical osteoblast cell subjected to expose on time domain electric field such as rectangular pulse, an equivalent circuit model is presented. This paper presents the window effect of different linear and non linear electric field on osteoblast cell in different microelectrode. It is found that because of different charging time constants & relaxation time, different durations and different geometry of microelectrode have selective effects on inner and outer membranes. We find that when duration of applied signal is reduced from microsecond to nanosecond, and to pecosecond, the generated inner and outer membrane potential are different & target induced is changed from the outer membrane to the inner membrane gradually for a specific type electrode geometry. In this paper we also explore the comparative analysis of window effect on cancer cell. The window effect gives preliminary explanation for various bioelectric effects such as electroporation, intracellular electromanipulation and nanopores, providing help to the applications of various cancer treatments. Keywords:- osteoblast cells, window effect, linear and non linear electric field, charging time constants, relaxation time and cancer treatment.

I.

INTRODUCTION

Osteoblast cells are the most abundant and fundamental elements in bone, and their concentration in serum is closely linked to bone metabolism and serves as a biological marker for the clinical assessment of bone disease. For clinical investigation the dielectric property of osteoblast cell plays an important role. Like other biological cell the dielectric properties of osteoblast cells are very remarkable. They typically display extremely high dielectric constants at low frequencies, falling off in more or less distinct steps as the excitation frequency is increased. Their frequency dependence permits identification and investigation of a number of completely different underlying mechanisms, and hence, dielectric studies of osteomaterials have long been important in electrophysiology and biomedical application. In drug delivery system the potential of the membrane plays an important role to control the radius and density of generated pores which allows the drugs enter into the cell. This concept is also applicable for clinical application in bone cell specially in osteoblast cell. Now a days the rectangular ,triangular & sawtooth pulsed electric field, because of their unique biomedical effects, have opened a new exposer to tumor treatment and become a research focus in the area of bioelectromagnetics. Many experimental researches show that rectangular pulse with different parameters can cause different bioelectric effects. Weaver et al found that in response to microsecond PEF (typical parameters: 1 kV/cm, 100 s) [1-2], many reversible aqueous channels, which are often called pores (radius 20-110 nm), appear at the cell membrane (outer membrane), while there is no obvious effect on the intramembranous organelles. This physical procedure is termed electroporation, which can make cell membrane more permeable to drug molecule, has been successfully applied to tumor treatment. On the other hand, Schoenbach et al found that the nanosecond PEF with amplitude above 10 kV/cm can induce a series of cellular responses [3-9], which are different from electroporation effect. This ns PEF can lead to poration of organelle membranes (inner membrane), such as nucleolus, mitochondria, etc, without losing the integrity of outer membrane, and may induce apoptosis of tumor cell. This phenomenon is termed intracellular electromanipulation (IEM). The ns PEF might be a new and promising therapy for tumor treatment, but how to accurately decide its parameters is a key point for its clinical application. Most recently, molecular dynamics modeling results and B16 cell experiments

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Study the Window Effect of Rectangular Electrical Pulse in Membrane...

[10] show that in response to pico second pulses (800 ps, 150 kV/cm, 1 MV/cm), reversible nanopores (radius 0.4 nm) appear at the outer membrane of B16 cells. From the above experiments, it could be concluded that PEF with different duration and intensity can cause various biomedical effects on cell, which suggests a window effect between. external PEF and biological cell. In this paper, based on an equivalent circuit model of spherical osteoblast cell, we studied the window effect in between rectangular pulse and transmembrane potential of osteoblast cell in different microelectrode through time domain analysis .We also compair the window effect in osteoblast cell with cancer cell.

II.

THE EQUIVALENT DIELECTRIC MODEL OF OSTEOBLAST CELL

A. Time domain analysis To study the window effect of osteoblast cell we consider the bilayer structure of the cell [15] [16] qnd verify the time domain analysis of outer & inner membrane potential describe by C .Yao et al.

Fig 1. Equivalent circuit model of cell subjected to PEF ( =0) From the above Figure, the complex-domain relationship between cb the cell voltage V and the external field E is () = () ----------( 1) The term f [19] is a compensation factor considering the effect of extracellular medium and cytoplasm, etc. Since the cell parameters are always under the conditions of , >> m and C c , >> , the term f can be approximated by a constant, f 1.5. Therefore, the transmembrane potential across the cell membrane is = . () ---------------(2) +1 Actually, the charging course of the outer membrane is affected by both the extracellular and intracellular mediums. According to the reference [14], the charging time constant of the outer membrane is given by Schwan =
1 20 1.5

----------------(3)

Where, notes the vacuum permittivity. Therefore, at any given , the ratio of transmembrane potential across the outer membrane to field intensity in complex-domain is 1.5 cos = . +1 On the other hand the transmembrane potential across the inner membrane is 1 = (4) 1 + 1 Therefore, at any given , the ratio of transmembrane potential to field intensity in complex-domain is 1.5 cos =
+1 + +1

The charging time constant of inner membrane is[14] =

1 2c

---------------- (5)

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Study the Window Effect of Rectangular Electrical Pulse in Membrane...

According to the dielectric cell parameters, it find out that > . Outer membrane: According to the transfer functions defined by equations (2) and (5), the time responses of inner and outer membranes to a given rectangular pulse electric field E(s) can be obtained () = 1 . () -------(6) & () = 1 . () -------(7) Put the value E(S)= ------- in equation no (6) & ( 7) we get the outer membrane potential ( ) s = 1.5 1 + . 1 cos -------( 8 ) At the end of the pulse, the transmembrane potential of outer membrane is = 1.5 1 cos --------------------------------------(9) From the equation no (9) it is come to know that if very low with respect to than outer membrane is poorly charged and when >= 4 than the outer membrane is fully charged. After simplification of equation (6) the membrane potential of inner membrane as And at the end of the pulse, the transmembrane potential of inner membrane is = ( ) cos -------( 10 ) From the equation no (10) it is come to know that if very low with respect to & than outer membrane is poorly charged and when >= 4 than the the inner membrane is fully charged, but its transmembrane potential of outer membrane is close to zero at the end of pulse. The charging of the inner membrane is affected by the time constants ( , ) of inner and outer membranes. As << , after the transmembrane potential across inner membrane reaching to its maximum, it starts to rapidly decline even if the field still persists.these shifting of generating membrane potential from outer to inner membrane shown in our study for osteoblast cell in rectangular electric pulse.
1.5

III.

USED PARAMETER
value 10 103 1.2 107 0.039s 10 101 0.08s 80 22 93 22 93 12 0.006m 6

A. Dielectric Value of Osteoblast Cell parameter Cell parameters conductivity Extracellular medium (S/m) Cell membrane Cell cytoplasm Nuclear membrane Nuclear cytoplasm Extracellular medium Cell membrane Cell cytoplasm Nuclear membrane Nuclear cytoplasm Cell radius Cell membrane thickness Nuclear radius

relative permittivity

Geometry parameter (m) B. Simulation Tool For fast and accurate simulation results, we chose the semiconductor, RF, and industry preferred NI Circuit Simulator and lab-view, product version(2.5.1).Lab view is commonly used for complex analogue and mixed-signal circuits .In an application, NI Circuit Simulator is used to study the circuit design .The analytical and mathematical simulation is done by the latest version MATLAB 10.5 and Math AutoCAD. Similarly, the electric field analysis of various electrode is done by COMsol multiphysics software. C. Used Electrode In our study we used different type of microelectrodes whose geometry and intra arrangement are shown in fig:4 & specification and generated electric field intensity of the used microelectrodes are explore in table I & II .

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Study the Window Effect of Rectangular Electrical Pulse in Membrane...

Table-I (Electrode specification) 1mm Length of electrode (l) 100 m Width of electrode (w) Inter-electrode distance 100 m 1m Thickness of electrode(d) 0 8.856 1012 F TABLE:II (ELECTRIC FIELD INTENSITY CALCULATING FROM COMSOLE MULTI PHYSICS S/W) Type of Rotation of Electric field electrode electrode intensity() Rectangular In line(c) ig:4 3.68 Shift (d) fig:4 3.75 Semicircular In line a)fig:4 4.22 Shift (b) fig:4 4.07 Saw tooth In line(e) ig:4 8.87 Shift (f) fig:4 3.61 Co-centric V/log(b/a)

Fig2. Configurations of electrode for FEA analysis. IV. THE WINDOW EFFECT

A. SIMULATION OF THE WINDOW EFFECT IN BETWEEN OF RECTANGULAR PULSE AND TRANSMEMBRANE POTENTIAL: a. Rectangular Electrode
For Ta=1e-012 0.5

For Ta=1e-010 0.5

-0.5 -20 10 0.5

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For Ta=1e-012

For Ta=1e-010

TMP in volts--------->

TMP in volts--------->
-15 -10 -5 0

-0.5 -20 10

10

10 time in sec ------>

10

10

-0.5 -20 10

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-15

10 time in sec ------>

-10

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Study the Window Effect of Rectangular Electrical Pulse in Membrane...

For Ta=1e-011 0.5
0.5 For Ta=1e-009

-0.5 -20 10 0.5

10

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-10

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-0.5 -20 10 0.5

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For Ta=1e-011

For Ta=1e-009

TMP in volts--------->

TMP in volts--------->

-0.5 -20 10

10

-15

10 time in sec ------>

-10

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-5

10

-0.5 -20 10

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-15

10 time in sec ------>

-10

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For Ta=1e-008 0.5

-0.5 -20 10 0.5

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-15

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-10

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-5

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For Ta=1e-008

TMP in volts--------->

-0.5 -20 10

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-15

10 time in sec ------>

-10

10

-5

10

Fig: 3.Transmembrane potential of outer and inner membrane in different pulse width (Ta) & duration of rectangular pulse in rectangular electrode. We Plot the relationship curves between transmembrane potentials (at the end of the pulse) and duration ( for rectangular electrode where E=3.68104 ),at different pulse width (Ta) of rectangular pulse for outer and inner membrane of osteoblast cell , as shown in above Figure. It could be concluded that rectangular pulse with different duration has obvious selective effect on the inner and outer membranes, i.e. window effect ,which are as follows. (1) At Ta=10-9 s(nano second): As compared to time constants, the duration is so long that both the inner and outer membranes can be fully charged. But the transmembrane potential across inner membrane is close to zero at the end of the pulse. This field contains abundant signal components at low frequency, mainly acting on the outer membrane.The field about 3.68104 V/cm can make the transmembrane potential across outer membrane reach the critical value for poration. As a result, the outer membrane is porated, but the inner membrane is not prorated. (2) At = 1010 1011 ( ) i.e. = 4 nuc<4 cell: At the end of the pulse, the transmembrane potential of the inner membrane is 0.25V, which is close to membrane potential of the outer membrane, with almost similar effective action time. As external field with enough intensity may have similar action on inner and outer membranes, so both of them might be expected to porate. (3) At = 1012 ( )i.e. 0.05 nuc<<4 nucFor the extremely fast charging speed, the inner membrane fully charged and much greater transmembrane potential than the outer membrane. This field contains abundant signal components at middle-high frequency, mainly acting on the inner membrane. The high intensity may cause the inner membrane porated, but the outer membrane not porated.. (4)At > 1012 s ( above picosecond), i.e. <0.05 nuc:The inner and outer membranes are extremely poorly charged by the rectangular pulse which exhibit their filtering action on the extremely high frequency components. But when responding to the strong electric field (several hundred kV/cm), the transmembrane potentials can still reach the critical value, so the porations of the inner and outer membranes can also be expected. Considering the extremely short action time, the pores should be different from those induced by electroporation. In summary, it is come to know from above figure that the very short pulse duration causes to low transmembrane potential across outer membrane, while both the very short and very long pulse durations decrease the transmembrane potential across inner membrane for osteoblast cell . This thoroughly exhibits the band-pass and low-pass filter characteristics of inner and outer membranes, respectively which is also explore through our frequency domain analysis.

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Study the Window Effect of Rectangular Electrical Pulse in Membrane...

b. Semicircular Electrode
For Ta=1e-012 1 0.5 0 -0.5 -1 -20 10

For Ta=1e-010 1 0.5 0 -0.5 -1 -20 10

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For Ta=1e-012

For Ta=1e-010

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0.2 0.1 0 -0.1 -0.2 10

-20

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0.2 0.1 0 -0.1 -0.2 10

-20

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10 time in sec ------>

For Ta=1e-009

-10

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For Ta=1e-008

-10

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1 0.5 0 -0.5 -1 -20 10

1 0.5 0 -0.5 -1 -20 10

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For Ta=1e-009

For Ta=1e-008

TMP in volts--------->

0.1 0 -0.1 -0.2 10

-20

TMP in volts--------->
-15 -10 -5 0

0.2

0.2 0.1 0 -0.1 -0.2 10

-20

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10 time in sec ------>

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Fig: 4 transmembrane potential of outer and inner membrane in different pulse width & duration of rectangular pulse in semicircular electrode. We Plot the relationship curves between transmembrane potentials (at the end of the pulse) and duration ( for semicircular electrode where E=4.22104 ),at different pulse width (Ta) of rectangular pulse for outer and inner membrane of osteoblast cell, as shown in above Figure. It could be concluded that rectangular pulse with different duration has obvious selective effect on the inner and outer membranes, i.e. window effect. All the plots are as same as rectangular electrode only the value of membrane potential for both outer & inner membrane are different but when Ta=1010 a negative voltage is obtain in inner membrane due to the presences of rest potential. Moreover, the negative transmembrane potential shown when Ta=1010 is a peculiar property of the inner membrane; in other words, the voltage across the inner membrane is bipolar.Positive voltage produced due to the charging of the membrane. While the charges stored in the outer membrane discharges at the negative edge of rectangular pulse, which is the forms of a discharge current. The discharge current will charge the inner membrane again. As the discharge current is in the direction opposite to the charged one, as a result, the transmembrane potential induced by discharge current is negative. c. Sawtooth Electrode
For Ta=1e-011 2
2 For Ta=1e-010

1
1

0
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For Ta=1e-011
For Ta=1e-010

TMP in volts--------->

0.5 0 -0.5 -1 -20 10

TMP in volts--------->
-15 -10 -5 0

0.5 0 -0.5 -1 -20 10

10

10 time in sec ------>

For Ta=1e-013

10

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-1 -20 10 1

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For Ta=1e-013

For Ta=1e-012

TMP in volts--------->

0.5 0 -0.5 -1 -20 10

TMP in volts--------->
-15 -10 -5 0

0.5 0 -0.5 -1 -20 10

10

10 time in sec ------>

10

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10 time in sec ------>

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Study the Window Effect of Rectangular Electrical Pulse in Membrane...

For Ta=1e-009 2

For Ta=1e-008 2

-1 -20 10 1

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For Ta=1e-009

For Ta=1e-008

TMP in volts--------->

TMP in volts--------->

0.5 0 -0.5 -1 -20 10

0.5 0 -0.5 -1 -20 10

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10 time in sec ------>

-10

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Fig: 5.Transmembrane potential of outer and inner membrane in different pulse width & duration of rectangular pulse in trangular electrode. We Plot the relationship curves between transmembrane potentials (at the end of the pulse) and duration ( for trangular electrode where E=8.87104 ),at different pulse width (Ta) of rectangular pulse for outer and inner membrane of osteoblast cell, as shown in above figure. It could be concluded that rectangular pulse with different duration has obvious selective effect on the inner and outer membranes, i.e. window effect. All the plots are as same semicircular electrode only the value of membrane potential for both outer & inner membrane are different.

V.

DISCUSSIONS

Though the window effect is based on the spherical osteoblast cells in suspension, it could be expected to extend to practical applications. The pulse duration and intensity jointly determine the selective effect on the various parts of the cell, to induce different bioelectric and therapeutic effects. When the duration is reduced from microsecond to nano- second, and to peco-second, the element induction is changed from the outer membrane to inner membrane gradually. At the same time, the amplitude of field intensity should be increased to reach the threshold potential across the membrane. Window effect may give preliminary explanation for various bioelectric effects such as electroporation, and nanopores, and can be helpful in choosing suitable duration intensity combination for inducing corresponding poration effects of the inner and outer membranes of osteoblast cell.It is also observed that the window effect does not depends on the shape of electrode although the peak value of membrane potential is changed with the geometry of microelectrode .Because when the transmembrane potential is increased the threshold voltage is also increased in same ratio.The rectangular pulse may also have selective effect on different cancer cells, for they have different cell parameters, i.e. different time constants (cell and nuc). For the healthy and cancer cells of a same type, it is also possible to determine a reasonable rectangular pulse effect , with its main action on cancer cells.

VI.

COMPARITIVE STUDY

In our study we also compair the window effect of rectangular pulse in Cancer cell with osteoblast cell and we find that in cancer cell when the pulse duration is bellow micro level no voltage will generate at outer & inner membrane but in microlevel outer membrane is fully charged whereas inner membrane has no response.This condition remain constant upto peco range .Above this the membrane voltage is shifted from outer membrane to inner membrane.

VII.

CONCLUSION

From the frequency domain analysis it is come to know that the inner and outer membranes act as a band-pass and low-pass filter characteristics respectively, so they has different filter activity in different frequency range with the influences of the external electric field .Depondeng on different field duration decides different transmembrane potentials across inner and outer membranes, so the field has selective action or window effect on them. In summary, the rectangular electric pulse has a window effect on the osteoblast cells. When the duration is reduced from microsecond to nanosecond and to pecosecond, the pulse is induced from the outer membrane to inner membrane gradually. At the same time, we also find that the induced membrane potential is varied in different microelectrode to generate poors in the membrane.. Furthermore, rectangular pulse may also have selective effect on different parts of a osteoblast or different type of cancer cells. The study of window effect in between membrane and rectangular pulse should be useful for the drug delivery system & experimental treatment for bone cancer.

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