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Influence of Weak Non-Thermic HF EMF on the Membrane Potential of Nerve Cells

Influence of Weak Non-Thermic HF EMF on the Membrane Potential of Nerve Cells

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The damage to normal physiological mechanisms of membrane potentials in both human beings and minor species constitutes an objective proof of a causal relationship of injury between cell phone telephony and electromagnetic fields. This has already been reported in Man. As such, it obeys the causal criteria as described by the Koch-Henle epidemiological postulates, the Susser criteria and the Bradford Hill criteria.
The damage to normal physiological mechanisms of membrane potentials in both human beings and minor species constitutes an objective proof of a causal relationship of injury between cell phone telephony and electromagnetic fields. This has already been reported in Man. As such, it obeys the causal criteria as described by the Koch-Henle epidemiological postulates, the Susser criteria and the Bradford Hill criteria.

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Bioelectrochemistry and Bioenergetics, 27 (1992) 2 9 3 - 3 0 4 A section o f J. Electroanal. Chem., and constituting Vol. 342 (1992)
Elsevier Sequoia S.A., L a u s a n n e

J E C BB 01485

Influence of weak non-thermic high-frequency e l e c t r o m a g n e t i c fields o n t h e m e m b r a n e p o t e n t i a l o f n e r v e cells *
U . Kullnick Zoologisches Institut, Physiologic, Technische Unioersitiit Braunschweig, D-3300 Braunschweig (Germany)
( R e c e i v e d 9 O c t o b e r 1991)

Abstract

N e r v e cells o f the snail Helix pomatia w e r e subjected to high-frequency (159 M H z , 8.3 H z m o d u l a t e d ) , n o n - t h e r m i e (maximum .qux density 124/~T) e l e c t r o m a g n e t i c fields. T h e e f f e c t o f the fields on the m e m b r a n e potential o f various nerve cells was investigated. It was o b s e r v e d that short and u n i q u e B e f e l d u n g e n t led to an alteration o f the m e m b r a n e potential o f the n e u r o n s examined. T h e alteration almost always expressed itself as a long-term h y p e r p o l a r i z a t i o n o f the resting potential. A clear c o n n e c t i o n b e t w e e n the negativity o f the m e m b r a n e p o t e n t i a l o f a nerve cell b e f o r e the B e f e l d u n g and the strength o f the hyperpolarization c a u s e d by the B e f e l d u n g was seen. As well as this effect, an a l t e r a t i o n in the threshold o f excitation o f b e f e l d e t cells could be m e a s u r e d .

INTRODUCTION

T h e influences o f so-called n o n - t h e r m i c h i g h - f r e q u e n c y ( H F ) e l e c t r o m a g n e t i c fields ( E M F ) on biological systems a r e b e c o m i n g a g r e a t e r p a r t o f public interest. T h e fields are b e i n g m a d e use o f m o r e a n d m o r e extensively for c o m m e r c i a l t h e r a p y a p p a r a t u s . This explains t h e discussion a b o u t t h e i r m e d i c a l i m p o r t a n c e a n d t h e i r possible effects on t h e e n v i r o n m e n t . It was against this b a c k g r o u n d t h a t t h e effects of w e a k ac m a g n e t i c fields b e g a n to b e c o m e the object o f n u m e r o u s r e s e a r c h projects [1]. In the m e a n t i m e , a n u m b e r o f clinical findings seem to
* P a p e r p r e s e n t e d at the syrn~osium " H i g h - F r e q u e n c y E l e c t r o m a g n e t i c ac Fields a n d their• Effects on Biological Systems", B r a u n s c h w e i g , 9.-.1.0 . .July. . 1991. . . .... .... .1 Definition: B e f e l d u n g is t r e a t m e n t with a w e a k (non-ionizing, n o n - t h e r m i c ) e l e c t r o m a g n e t i c field.
0302-4598/92/$05.00 © 1992 - Elsevier Sequoia S.A. All rights reserved

294

confirm t h a t t h e r e are effects which can be u s e d therapeutically. N o t only t h e u n k n o w n effect mechanisms, but also t h e lack of knowledge about t h e best t h e r a p e u t i c p a r a m e t e r s of t h e fields used (intensity, frequency, m o d u l a t i o n ) a r e against sensible use. Also with regard to work a n d e n v i r o n m e n t a l risks, g r e a t e r k n o w l e d g e of the interactions b e t w e e n E M F a n d biological systems is n e c e s s a r y at all costs. This is of special i m p o r t a n c e b e c a u s e s t a n d a r d s for the p r o t e c t i o n of m a n k i n d in the future could be f o r m u l a t e d not only from an e n e r g y p o i n t of view [2]. It can be seen t h a t the o p i n i o n of m a n y r e s e a r c h e r s ( f r e q u e n t l y expressed up to now) t h a t the effect of E M F can only be s u b s t a n t i a t e d e n e r g e t i c a l l y must now be r e c o n s i d e r e d in the wake of more m o d e r n knowledge. F o r investigations of t h e effect of w e a k e l e c t r o m a g n e t i c fields in this study, the central nervous system (CNS) of the vineyard snail is used. T h e a d v a n t a g e s of using a snail as the object of investigation can be found in the c o m p a r a t i v e l y small n u m b e r o f n e u r o n s (approximately 50 000), their easy accessibility, their specific size (up to 150 /xm) a n d t h e i r robustness against i n t e r v e n t i o n s in t h e nervous system (e.g. p r e p a r a t i o n ) . F u r t h e r , it is a well-known fact t h a t t h e f u n d a m e n t a l functions of nerve ceils are practically identical in t h e various groups of animals, b o t h f l o m a m e t a b o l i c a n d an electrophysical point of view (signal g e n e r a t i o n , c o n d u c t i o n a n d transmission). A great deal of c o r r e s p o n d e n c e can be f o u n d especially in molluscs a n d t h e n e u r o n s of v e r t e b r a t e s ( H o d g k i n , Huxley 1952). A p r i n c i p a l transferability of t h e effects of e l e c t r o m a g n e t i c fields observed in the n e u r o n s of snails to h u m a n nerve cells can t h u s be p r e s u p p o s e d . T h e n e u r o n s of molluscs m a k e it possible to recognize not only short-term, brief effects, but also l o n g - t e r m or irreversible ones, as t h e y can be m e a s u r e d intracellularly for a n u m b e r of hours a n d even days, as o p p o s e d to the nerve cells of vertebrates. Helix pomatia is thus a suitable m o d e l o r g a n i s m for t h e investigation of n e u r o n a l questions. In addition, e x p e r i m e n t s with v e r t e b r a t e cell cultures a n d p r e p a r a t i o n s of brain slices will be carried out in our l a b o r a t o r y in t h e n e a r future. T h e first investigations of t h e nerve ceils of Helix pomatia a n d t h e i r bioelectricity u n d e r t h e effects of w e a k H F m a g n e t i c fields (150 M H z , 8.3 Hz, L F pulsed) gave i n t e r e s t i n g results [3]. T h e p a r a m e t e r which was p r e d o m i n a n t l y investigated, t h e m e m b r a n e potential, showed clear h y p e r p o l a r i z a t i o n of the nerve cells. T h e s e results w e r e confirmed, c o m p l e t e d a n d e x t e n d e d in f u r t h e r examinations. P e r h a p s t h e y can form t h e basis for t h e solution to t h e p r o b l e m of cell-physiological m e c h a n i s m s of t h e a t h e r m i c effect of H F - E M fields.
METHODS

F o r t h e experiments, adult v i n e y a r d snails ( 3 - 4 years old) of t h e species Helix pomatia (t3astropoda, P u l m o n a t a ) w e r e used. T h e C N S of t h e animals was removed w i t h o u t t h e use of medicines. T h e C N S consists of an o e s o p h a g e a I g a n g l i o n w i t h cerebral a n d s u b o e s o p h a g e a l ganglia. F o r intracellular m e a s u r e m e n t s , only giant nerve cells of the s u b o e s o p h a g e a l g a n g l i o n w e r e used (the a r e a m a r k e d by a

295

Fig. 1. CNS of Helix pomatia with intact connective tissue. CG: cerebral ganglion; CPLK: cerebro-pleural connective; CPK: cerebro-pedal connective; P1-P10: pedal nerves; P A R G : parietal ganglion; PD: pedal ganglion; PLG: pleural ganglion; ST: statoeyst; STN: stato nerve; VISC: visceral ganglion. T h e neurons measured are those in the circle.

circle in Fig. 1). T h e cell d i a m e t e r was a p p r o x i m a t e l y 100 tzm. Selection of t h e m e a s u r e d cells d e p e n d e d o n t h e i r position relative to the electrode. M e a s u r e m e n t s of t h e m e m b r a n e p o t e n t i a l ( M P ) o f living cells, which w e r e not obviously stimulated, by use o f suitable m e a s u r e m e n t systems always show characteristic voltage values, called resting p o t e n t i a l s (RP), which can r e m a i n c o n s t a n t for a long time as a rule. In all resting potentials, t h e m e m b r a n e o f t h e nerve cell on t h e inside has a negative load c o m p a r e d with t h e outside. In this study, this r e s t i n g p o t e n t i a l was t h e p r e d o m i n a n t l y investigated p a r a m e t e r , in o r d e r to establish t h e effects of h i g h - f r e q u e n c y e l e c t r o m a g n e t i c fields on nerve cells. A l t e r a t i o n s of the m e m b r a n e p o t e n t i a l occur in physiological excitations or artificial electrical stimulation of t h e cell. A n increase in the m e m b r a n e potential, i.e. i n c r e a s e d n e g a t i v i z a t i o n o f t h e inside, is called h y p e r p o l a r i z a t i o n , a r e d u c t i o n in t h e p o t e n t i a l b e i n g d e p o l a r i z a t i o n . T h e m e m b r a n e p o t e n t i a l of nerve cells m a i n l y arises t h r o u g h : (1) a s e m i - p e r m e a b l e cell m e m b r a n e ; or ( 2 ) an u n e v e n d i s t r i b u t i o n o f ions b e t w e e n t h e i n t r a c e l l u l a r a n d the extracellular fluid, which is p r o d u c e d , i n t e r alia, by e n e r g y - c o n s u m i n g t r a n s p o r t processes.

296 •

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Fig. 2. Experimental set-up for intracellular measurements. (1) Signal connection; (2) signal-ground connection; (3) intercellular preamplifier; (4) oscilloscope; (5) D A T recorder; (6) Mega-Wave 150/1; (7) E M F transmitter; (8) off-line computer.

• Resting p o t e n t i a l and o t h e r bioelectrical signals (AP, EPSP, IPSP) are r e c o r d e d by m e a s u r e m e n t probes (electrodes). For intracellular m e a s u r e m e n t s , these are fine glass capillaries with tip o p e n i n g s of a r o u n d 1 /xm, filled with a 3 M KC1 solution. W i t h the help o f such m i c r o - e l e c t r o d e s and their c o n d u c t i v e c o n n e c t i o n ( A g - A g C I wire) and a m e t a l s i g n a l - g r o u n d c o n n e c t i o n in the R i n g e r solution in which the objects to be m e a s u r e d can be found, a n d also m e a s u r i n g i n s t r u m e n t s with a high electrical resistance, the m e m b r a n e polarity o f a nerve cell can be m e a s u r e d and portrayed. F o r reasons to be f o u n d in the physics o f h i g h - f r e q u e n c y e l e c t r o m a g n e t i c fields, t h e r e should be no m e t a l parts in the a r e a o f the effects o f the e l e c t r o m a g n e t i c fields used. In the a r e a of the m a g n e t i c field, we t h e r e f o r e m a n a g e d w i t h o u t metal conductors, especially in the m e a s u r e m e n t area, in o r d e r to avoid physical interactions which could falsify the m e a s u r e m e n t or possibly even m a k e it useless. T h e s e d e m a n d s were fulfilled by the d e v e l o p m e n t o f a new, m o d i f i e d intracellular m e a s u r i n g m e t h o d . M e a s u r e m e n t s o f the m e m b r a n e p o t e n t i a l in the m a g n e t i c field were m a d e possible by replacing the c h l o r i n a t e d silver wire (sig,ial a n d s i g n a l - g r o u n d connection) inside the field with a 3 M H C l - a g a r - a g a r bridge in thin plastic hoses. T h e agar bridges only pass into the silver wires, which c o n d u c t the m e a s u r e d signals to t h e amplifier (Fig. 2), at a distance which basically rules out an i n f l u e n c e of the a p p l i e d m a g n e t i c field. This m e t h o d m a k e s intracellular m e a s u r e m e n t o f nerve Cells in high-frequency e l e c t r o m a g n e t i c fields possible. In o r d e r to avoid f u r t h e r possible m e a s u r e m e n t artefacts caused by an u n f a v o u r a b l e m e a s u r e m e n t geometry, t h e signal a n d s i g n a l - g r o u n d c o n n e c t i o n s were a r r a n g e d m o r e or less in the d i r e c t i o n o f the spread of the e l e c t r o m a g n e t i c field (Fig. 3).

297
15 c m

I
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Fig, 3. Modified intracellular measurement. (1) Glass signal connection (3 M KCI); (2) KCl-agar bridges (3 M KCI); (3) connection of agar bridges and Ag-AgCI wife; (4) EMF transmitter; (5) ganglion.

Cells with high s p o n t a n e o u s activity w e r e not used in the e x p e r i m e n t . C e l l s whose m e m b r a n e p o t e n t i a l showed many E P S P (excitatory postsynaptical p o t e n tials) a n d IPSP (inhibitory postsynaptical p o t e n t i a l s ) were also n o t u s e d f o r t h e evaluation. Basically, only nerve cells with a resting p o t e n t i a l which r e m a i n e d u n c h a n g e d (2 rnV) for at least 40 min ( p r e l i m i n a r y stage) w e r e used in the evaluation. Following the experiment, no i d e n t i f i c a t i o n of the p e n e t r a t e d "cell was carried o u t by n e u r o - a n a t o m i c staining or f u n c t i o n a l checks. T h e m e a s u r e m e n t w a s always carried out in t h e following sequence: 40 rain p r e l i m i n a r y phase, 40 rain run-up, 10 min Befeldung, 40 min run-down, 40 rain s u b s e q u e n t phase. T h e fields used h a d t h e following characteristics: c a r r i e r frequency: 150 MHz; m o d u l a t i o n frequency; 8.3 Hz; form o f m o d u l a t i o n : n e e d l e impulse; m a g n e t i c flux density; 124 tzT 50% (2 cm from the t r a n s m i t t e r , I n s t i t u t e for E x p e r i m e n t a l Physics, F r e e University o f Berlin); time of application: 10 min+ T h e d a t a r e c o r d e d by a D A T r e c o r d e r (48 kHz scan frequency) were r e a d i n t o an off-line c o m p u t e r . T h e c o m p u t e r e v a l u a t i o n was c a r r i e d o u t o n a C o m m o d o r e PC by m e a n s of an analysis p r o g r a m d e v e l o p e d in o u r d e p a r t m e n t . This p r o g r a m works with a scan f r e q u e n c y o f 32 kHz. T h e m e a n v a l u e s a r e t a k e n from t e n m e a s u r e d values. This averaging is carried o u t until a m e a n value for 1 min of m e a s u r i n g is p r o d u c e d . T h e p r o g r a m recognizes technical i n t e r f e r e n c e , action p o t e n t i a l s a n d s u d d e n slight a l t e r a t i o n s o f t h e m e m b r a n e potential. T h e s e are f a d e d o u t with slight r u n - u p or r u n - d o w n times (1 s) a n d are n o t used in t h e c o m p u t a t i o n of the m e a n value. T h e times f a d e d out are displayed a n d should n o t be longer t h a n 1 rain in an e x p e r i m e n t a l p e r i o d of 9 0 - 1 4 0 min.

Measurement o f the threshold o f excitation
T h e n e r v e cells were excited intracellulary just above t h e t h r e s h o l d by signal c o n n e c t i o n s w i t h r e c t a n g u l a r impulses in a c c o r d a n c e with the s t a n d a r d m e t h o d . E a c h cell was excited ten times at intervals of 10 s. This t o o k p l a c e 10 rain b e f o r e the Befeldung, during t h e Befeldung, 10 rain after this and, for a final time, 1 h after t h e end o f t h e Befeldung.

298 RESULTS

Measurement of the resting potential of the so-called quiet cell
T h e single Befeldung of a nerve cell almost always resulted in the resting p o t e n t i a l being more or less strongly h y p e r p o l a r i z e d . T h e h y p e r p o l a r i z a t i o n sometimes started shortly after the start of the Befeldung. However, h y p e r p o l a r i z a t i o n n o r m a l l y occurred only in the last m i n u t e of t h e B e f e l d u n g or, quite frequently, only 1 - 1 0 rain after the e n d o f the B e f e l d u n g (Fig. 4). T h e nerve cell s o m a t a m e a s u r e d had m e m b r a n e p o t e n t i a l s b e t w e e n - 2 3 and - 6 8 mV. T h e h y p e r p o l a r izatmn in these cases was - 2 and - 1 3 mV (cf. T a b l e 1). T h e strength o f the evoked h y p e r p o l a r i z a t i o n s is closely c o n n e c t e d with the m e m b r a n e p o t e n t i a l s o f the cells, Nerve cells with highly negative R P (e.g. --68 mV) were only slightly h y p e r p o l a r i z e d by the B e f e l d u n g (approximately - 2 mV). On the o t h e r hand, cells with a weak R P (e.g. - 2 3 mV) were strongly h y p e r p o l a r ized (e.g. 13 mV) (Fig. 5). Thus, t h e r e is a c o n n e c t i o n b e t w e e n the value o f a resting p o t e n t i a l set by the cell and its h y p e r p o l a r i z a t i o n caused by the field. T h e h y p e r p o l a r i z a t i o n also lasted for a n u m b e r of hours. As the p r e p a r a t i o n , as described in the M e t h o d s section, was not in a n u t r i t i o n solution b u t in a b l o o d r e p l a c e m e n t solution, it was not possible to establish u n d e r these exper.lmental c o n d i t i o n s w h e t h e r the d e p o l a r i z a t i o n of t h e m e m b r a n e p o t e n t i a l , which sometimes only occurred after several hours, was d u e to a d r o p in the effect of the B e f e l d u n g or to physiological reactions of the cell to a lack of supply (Fig. 6).

Measurement of the hyperpolarized resting potential after a second Befeldung
W h e n the h y p e r p o l a r i z a t i o n of a b e f e l d e t ceil had c o m e to a standstill, i.e. the m e m b r a n e p o t e n t i a l s h a d t a k e n on a stable value, a s e c o n d B e f e l d u n g was carried
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Fig. 4. Single Befeldung after one Befeldung. T h e resting potential o f the measured hyperpolarizations was approximately 7 mV.

299 TABLE 1 Dependence of the hyperpolarization values on the membrane potential of some nerve cells. Neurons with highly negative membrane potentials hyperpolarize slightly after one Befeldungl Neurons with weakly negative membrane potentials hyperpolarize strongly after one Befeldung Resting potential ~tart v a l u e / m V
- 68 - 67

Resting potential after 1st B e f e l d u n g / m V
--70 --68

Hyperpolarization/mV
2 I

--67
-- 66 -- 63 - 62 -58 - 57

--70
-- 69 -- 68 --65 -62 -61

3
3 5 3 4 4

-56 -46 --40 --39 --39 --38 --28
-- 27 -- 25 -- 24

-62 -55 -47 --45 --47 -47 -38
-37 -36 -37

5 9 7 6 8 9 10
10 11 13

out on a n u m b e r of occasions. T h e newly set m e m b r a n e potential did not hyperpolarize further. A third B e f e l d u n g also h a d no m e a s u r a b l e effect on the d e v e l o p m e n t of potential (Fig. 7).

Befeldung of non-quiet cells
(a) A l o n g s i d e quiet cells, o t h e r nerve cells were also included in the experiment. N e u r o n s w e r e befeldet, the resting potential of which was not stable but h a d a depolarizing tendency. This t e n d e n c y was s t o p p e d by t h e B e f e l d u n g and, in some cases, even reversed (Fig. 8). (b) S o m e ceils, which w e r e possibly excessively d a m a g e d in the p e n e t r a t i o n of the signal connection, showed strong depolarization. T h e Befeldung did not, as in case (a), achieve a d r o p in the depolarization, b u t the exact opposite (Fig. 9). T h e d e p o l a r i z a t i o n was clearly increased a n d practically led to potential c o m p e n s a t i o n after a short time (approximately 1 h).

Measurement o f the threshold of excitation
N a t u r a l l y , i t w a s interesting to see how the t h r e s h o l d of excitation of a nerve cell would b e h a v e if its m e m b r a n e potential was hyperpolarized by Befeldung.

300
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Fig. 5. H y p e r p o l a r i z a t i o n s after one B e f e l d u n g as a function o f the initial m e m b r a n e potential. U p p e r line: m e m b r a n e potentials of various cells b e f o r e the Befeldung; lower line: hyperpolarizations o f these cells after c n e Befeldung.

U n f o r t u n a t e l y , t h e r e are few r e s u l t s c o n c e r n i n g this q u e s t i o n . Y e t t h e s e a r e v e r y i n t e r e s t i n g , for w h i c h r e a s o n t h e y a r e p o r t r a y e d h e r e ( T a b l e 2). U n f o r t u n a t e l y , it h a s only b e e n possible to c a r r y o u t t h e e x p e r i m e n t t h r e e t i m e s u p to now, t h e r e s u l t b e i n g t h e s a m e e a c h time. T h e e l e c t r i c a l i n t r a c e l l u l a r s t i m u l a t i o n s , to w h i c h a r e a c t i o n c a m e w i t h a c t i o n p o t e n t i a l s ( A P ) in a b o u t 8 3 % o f t h e c a s e s in t h e r u n - u p , w h i c h b r o u g h t a b o u t a r e a c t i o n of 3 6 . 6 % d u r i n g t h e B e f e l d u n g . T e n

TABLE 2 M e a s u r e m e n t of the threshold o f excitation before, during and after a Befeldung (three different nerve cells). Evoked action potentials ( c A P ) c a u s e d by electrical r e c t a n g u l a r stimuli before, during, 10 min and 1 h after a Befeldung RP/ mV
--

Before Befeldung cAP 28 27 28 83%

During Befeldung cAP 12 10 11 36.6%

10 min after Befeldung eAP 17 15 16 53.3%

1 h after Befeldung cAP 22 20 22 71%

Hyperpolarization mV
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Fig. 6. A l t e r a t i o n s o f the m e m b r a n e p o t e n t i a l o f a nerve cell a f t e r o n e h y p e r p o l a r i z a t i o n . A p p r o x i m a t e l y 90 rain a f t e r the e n d of the B e f e l d u n g , t h e h y p e r p o l a r i z a t i o n o f t h e m e m b r a n e p o t e n t i a l due to the field has finished in this ease. A f t e r a p l a t e a u phase, slow d e p o l a r i z a t i o n starts a b o u t 3 h a f t e r the e n d o f the B e f e l d u n g .

m i n u t e s after the e n d of the B e f e l d u n g , this figure rose to 53.3% again, e v e n rising to 71% after 1 h. H o w e v e r , it m u s t be m e n t i o n e d that t h e cells u s e d in t h e s e e x p e r i m e n t s had a relatively strong negative resting potential, w h i c h m e a n s that the h y p e r p o l a r i z a t i o n w a s not very strong.

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Fig. 7. A l t e r a t i o n o f the m e m b r a n e p o t e n t i a l o f a nerve cell with t h r e e B e f e l d u n g e n . O n l y the first B e f e l d u n g leads to h y p e r p o l a r i z a t i o n o f the m e m b r a n e potential. B e f e l d u n g 2 a n d 3 have n o effect.

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Fig, 8. A l t e r a t i o n o f the m e m b r a n e p o t e n t i a l o f a n o n - q u i e t cell a f t e r a single B e f e l d u n g . T h e low d e p o l a r i z a t i o n of the m e m b r a n e p o t e n t i a l o f a nerve cell is s t o p p e d a f t e r o n e B e f e l d u n g a n d t h e n even b r o u g h t to hyperpolarization.

r e c o r d i n g electrode. T h e r e are some physical p h e n o m e n a ( m a g n e t i c induction, r a d i o aerial effect, influention, skin effect, etc.) which have p r o p e r t i e s m a s k i n g t h e ability to influence the m e a s u r e m e n t s . In o r d e r to escape from m a g n e t i c voltage i n d u c t i o n we i n t e r r u p t e d the m e a s u r i n g loop d u r i n g the e l e c t r o m a g n e t i c Befeldung. T h e d i s a d v a n t a g e of this type of m e a s u r e m e n t , however, is t h a t t h e n e u r o n s ( R P ) c a n n o t actually be t e s t e d d u r i n g E M F stimulation. T h e electrical i n f l u e n t i o n was tested e x p e r i m e n t a l l y a n d we could not see any artefact. O t h e r effects w e r e calculated (Institute of E x p e r i m e n t a l Physics, F U , Berlin) but t h e r e was no indication of any electrical or o t h e r influence on the m e a s u r i n g system.

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Fig. 9. A l t e r a t i o n o f the m e m b r a n e p o t e n t i a l o f a nerve cell d e p o l a r i z i n g d u e to injury, a f t e r o n e Befeldung. T h e d e p o l a r i z a t i o n of t h e cells is strongly a c c e l e r a t e d a f t e r the E;efeldung.

303

N a t u r a l l y , B e f e l d u n g w a s also c a r r i e d o u t o n cells w h i c h h a d c o m p l e t e l y c o l l a p s e d p h y s i o l o g i c a l l y ( a l m o s t c o m p l e t e c o m p e n s a t i o n of p o t e n t i a l , n o E P S P , I P S P o r A P , a n d n o electrical excitability). I n t h e B e f e l d u n g of t h e s e ceils, t h e r e w a s n o a l t e r a t i o n of t h e p o t e n t i a l m e a s u r e d . I n t h e e x p e r i m e n t s w i t h q u i e t cells, a r e l a t i v e l y u n i f o r m t e n d e n c y t o w a r d s h y p e r p o l a r i z a t i o n w a s seen. T h e r e l a t i o n s h i p b e t w e e n t h e r e s t i n g p o t e n t i a l a n d t h e s t r e n g t h o f t h e h y p e r p o l a r i z a t i o n to b e a t t r i b u t e d to t h e B e f e l d u n g ( h i g h r e s t i n g p o t e n t i a l - - w e a k h y p e r p o l a r i z a t i o n ; low r e s t i n g p o t e n t i a l - - s t r o n g h y p e r p o l a r i z a t i o n ) c o r r e s p o n d s to t h e q u a n t i t a t i v e relat i o n s h i p b e t w e e n t h e c o n c e n t r a t i o n r a t i o a n d t h e e q u i l i b r i u m p o t e n t i a l on n e r v e cell m e m b r a n e s ( N e r n s t ' s e q u a t i o n ) . T h i s is also w h y r e p e a t e d B e f e l d u n g in t h e e x p e r i m e n t did n o t l e a d to a n y f u r t h e r h y p e r p o l a r i z a t i o n s .
DISCUSSION

T h e r e s u l t s o f t h e B e f e l d u n g o f n o n - q u i e t cells m a t c h t h e a b o v e - m e n t i o n e d a t t e m p t s at e x p l a n a t i o n . T h e e n d of d e p o l a r i z a t i o n t e n d e n c i e s c o r r e s p o n d s to t h a t of t h e h y p e r p o l a r i z a t i o n a n d could b e s u b j e c t to t h e s a m e e f f e c t m e c h a n i s m s . T h e m e a s u r e m e n t o f h i g h l y d a m a g e d n e r v e cells is d i f f e r e n t . T h e a c c e l e r a t i o n of t h e potential c o m p e n s a t i o n could be substantiated by the fact that the mechanisms w h i c h led to t h e h y p e r p o l a r i z a t i o n of q u i e t cells c a n n o l o n g e r w o r k effectively. N e v e r t h e l e s s , t h e y c o u l d possibly r e p r e s e n t a p r o c e s s w h i c h c o n s u m e s a l a r g e a m o u n t of e n e r g y . T h i s e n e r g y c o n s u m p t i o n c o u l d l e a d to a q u i c k c o l l a p s e o f the b i o e t e c t r i c i t y of t h e n e r v e cell. T h e m e a s u r e m e n t o f t h e t h r e s h o l d o f e x c i t a t i o n o f a n e r v e ceil s h o w e d q u i t e s p e c i a l results. A n i n t e r p r e t a t i o n c a n o n l y b e m a d e very r e t i c e n t l y here. Obviously, t h e s t r o n g e s t effect o f t h e e l e c t r o m a g n e t i c fields d u r i n g t h e B e f e l d u n g is t h a t it c a u s e s a d r o p in t h e e v o k e d p o t e n t i a l s to 3 6 % . T h e r e a c t i o n to electrical stimuli i n c r e a s e s a g a i n a f t e r t h e B e f e l d u n g . T h i s a p p e a r s to c o n t r a d i c t the results o b t a i n e d u p to now, as t h e excitability is at its lowest at t h e m o m e n t w h e n t h e h y p e r p o l a r i z a t i o n c a u s e d b y t h e field is also at its lowest, s o m e t i m e s h a r d l y m e a s u r a b l e . T h e excitability i n c r e a s e s a g a i n w i t h t h e a m o u n t o f h y p e r p o l a r i z a t i o n o r w i t h t h e d r o p in t h e B e f e l d u n g . All t h e f i n d i n g s m e a s u r e d i n d i c a t e t h a t v a r i o u s effect m e c h a n i s m s exist in t h e field e f f e c t o n n e r v e cells. M o r e p r e c i s e a n d m o r e n u m e r o u s m e a s u r e m e n t s in o u r i n s t i t u t e , i n t e r alia o n b r a i n sclices a n d n e r v e cell cultures, a n d e x a m i n a t i o n s o n ion c h a n n e l s will p r o d u c e f u r t h e r r e s u l t s in t h e n e a r f u t u r e a n d t h u s possibly c o n t r i b u t e to t h e f o r m u l a t i o n of a g e n e r a l t h e o r y of effects. A s t h e r e h a s b e e n f r e q u e n t s p e c u l a t i o n in t h e m e d i c a l l i t e r a t u r e in t h e r e c e n t p a s t c o n c e r n i n g t h e effects ~f w e a k e l e c t r o m a g n e t i c fields in t h e i m m u n e s y s t e m o f m a n , i n v e s t i g a t i o n s a r e c u r r e n t l y b e i n g c a r r i e d o u t in o u r l a b o r a t o r y o n n e u r o s e c r e t o r y n e r v e cells o n m o l l u s c s (Lymnaea stagnalis), t h e a i m b e i n g to e x a m i n e t h e field effects r e f e r r e d to in this paper. O w i n g to t h e c h a n g e in m e t h o d s in t h e i n t r a c e l l u l a r m e a s u r e m e n t of n e r v e cells, it w a s p o s s i b l e for t h e first t i m e to m e a s u r e t h e m e m b r a n e p o t e n t i a l f r e e o f

304

artefacts during a n d after the B e f e l d u n g with w e a k H F e l e c t r o m a g n e t i c fields. Thus, the effects which the fields used had on the bioelectricity of the nerve cells (clear h y p e r p o l a r i z a t i o n ) were proven. T h e s e results were e s p e c i a l l y i m p o r t a n t because it was possible to show clear influences o n the electrical propt~rties of nerve cells, despite the use of field strengths below the m e a n energy of m o l e c u l a r t h e r m a l motion. As t h e r m i c effects t h r o u g h t h e fields used are ruled out [4], o n e or m o r e different effects must cause the a l t e r a t i o n of the bioelectricity of the cells investigated. As the m e m b r a n e p o t e n t i a l is basically a biophysical p h e n o m e n o n , which can be explained by the specific p r o p e r t i e s of the cell m e m b r a n e , it would be possible to imagine effect m e c h a n i s m s which have an effect o n e l e m e n t s of the m e m b r a n e , e.g. various p r o t e i n structures, on the basic structure o f the m e m b r a n e (phospholipid d o u b l e layer) in its i n t e r p r e t a t i o n as a liquid crystal or m e m b r a n e c o n d e n s e r or on the energetics forming the basis of the active t r a n s p o r t m e c h a nisms. T h e a l t e r a t i o n s of the ion c h a n n e l s which result from this, a n d t h e r e f o r e also of the p e r m e a b i l i t y and the resistance of the cell m e m b r a n e , could explain the m e a s u r e d a l t e r a t i o n s of the bioelectricity o f ~he iielwe cell.
REFERENCES 1 R. Glaser, Bioelectrochem. Bioenerg., 27 (1992) 255. 2 "Nichtionisierende Strahlung", Klausurtagung der Strahlenschutzkommission 7.-9. Dez. 1988, VerSffentlichung der Strahlensehutzkommission, Bd. 16, Bundesminister t-fir Umwelt, Naturschutz u. Reaktorsicherheit, Fischer, 1990. 3 LI. Kullniek and H.G. Wolff, in N. Eisner and H. Penzlin (Eds.), Synapse-Transmission-Modulation, Proceedings of the 19th GSttingen Neurobiology Conference, Thieme, Stuttgart, NY, 1991. 4 K.D. Kramer, personal communication.

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