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| EVIDENCE FOR A BIOPHYSICAL COMMUNICATION SYSTEM BETWEEN MAMMALIAN BRAINS AND EUKARYOTIC CELLS | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Tamara Galonja and Roger Coghill INTRODUCTION Biochemical mechanisms of inter- and intra-cellular signal transduction have become increasingly well characterised, stimulated by studies of weak bioeffects from electromagnetic (EM) fields. Interest in possible adverse effects of weak EM fields and radiation from mobile telephones has also given new impetus to EM field effects on cerebral rhythmic emissions. This paper proposes the existence of a mechanism of interaction between mammalian brains and individual cells by direct physical information transfer, and offers supporting experimental evidence. The hypothesis that endogenous electric fields from the brain can influence cellular processes directly without chemical action is tested by examining their influence on lymphocyte viability as shown by trypan blue exclusion. METHOD & MATERIALS Human peripheral blood lymphocytes were isolated from 30ml. of whole blood drawn from the v. cubitale by differential centrifugation at 800g (Baird & Tatlock Auto bench Centrifuge Mk. 4) in Ficoll-Triosil and micropipetting and diluted (1:1 v/v) in buffered saline solution (BSS) with added glucose. They were then cultured for c. 4 hours at 37° C (+/- 0.1° C) in a medium consisting of RPMI-1640, antibiotics and antimycotics (Sigma-Aldrich, Gillingham, Dorset, UK), and heat inactivated human serum from the same donor, in an incubator (Stuart Scientific S.I. 60, Redhill, Surrey) whose EM fields were shielded (<50nT rms) by a double skinned 0.8mm mu-metal casing (Magnetic Shields Ltd., Staplehurst, Kent). Endogenous electric fields emanating from this human donor (mean c.30mV, 100mV peak to peak at the skin surface) were led by means of a 60cm gold wire to a sterile glass screw-capped container (12 x 38mm O.D.) containing c. 1ml of the lymphocyte culture. The container was enclosed in a mu-metal box (30 x 30 x 45mm O.D.) together with similar volumes of the culture in control and sham-exposed (i.e. with gold wire but unattached to subject) containers (see Diag. 1). Temperature in the box after attachment varied between 32-33° C during the exposure period, due to small nocturnal variations in donor body temperature. Many studies show that lymphocytes in culture can survive for long periods when supplied with a 5% CO2 rich air atmosphere. In this study however the cells were deliberately not cultured in a 5% CO2 atmosphere, in order to lower mean viability levels. This mu-metal box exposure system, containing also the sham and control samples, was taped with masking tape to the donor’s forearm for 7.5 hours (2330hrs to 0800 hrs.) on the day of collection. At the end of this period (some 30 hours since original drawing) an aliquot from each container was tested for viability by trypan blue exclusion, and the ratio of viable/non viable cells ascertained for each. Counting by means of light microscopy was blinded, so that the counter was unaware of which sample was being counted. Two samples (a and b) were counted from each container. RESULTS Six separate exposure runs using the same donor were made over a 2 week period. The results of these are given below (Table 1). Table 1: Numbers of viable /non viable lymphocytes (human peripheral blood) Exposed Sham Control
These results showed a significant difference between the exposed and control samples (chi squared and Student’s t-test, p =<0.001). Testing for effects of gold wire To test for the possibility that the effect observed was solely due to the 60cm. gold wire feeding into the box or to temperature effects, a box containing a 60cm. gold wire and the lymphocyte culture was enclosed in a separate mu-metal box of similar dimensions together with a container containing only the lymphocyte culture (control) and maintained at 37° C (+/- 0.1° C). The results of these "exposures" for the same overnight period of about 7.5 hours, also commencing on the day of collection, are given in Table 2. Table 2: Viability of lymphocytes maintained for 7.5 hours (2330 hrs. to 0800 hrs, no EM fields, at 37° C): With gold wire Control (no wire)
Testing for effects of 50Hz electric fields It might also be argued that exposure to any electric field would produce an improvement in lymphocyte viability, not just the endogenous electric fields from the body. To test this a signal generator set to generate 50 Hz. with a square wave but with the circuit uncompleted was substituted for the endogenous fields. (By leaving the circuit uncompleted, though there was a net zero voltage the electrons were nevertheless moving in the wire, thereby creating a 50Hz. electric field in the wire but no magnetic field). The potential was set at about the same level as the endogenous field (c. 30-40 mV). The reason for using a square wave was because it has a very fast rise-time, and therefore a higher electric field. These two containers (50Hz-exposed and control) were also enclosed in a mu-metal box (30 x 30 x 45mm O.D.) to keep out stray EM fields, and maintained in the incubator at 37° C for the same 7.5 hours overnight period. The results are given in Table 3. Table 3: Viability of lymphocytes exposed to 50Hz. electric fields for 7.5 hours (2330hrs. to 0800hrs.) Exposed Control
These samples were some 20% less viable but not significantly different from the controls in Table 1, since the data was not sufficient to establish significance. To address the possibility that RF/MW frequency may be a factor, signals from three cellphones on standby were substituted for the endogenous and ELF electric fields, and the experiment was repeated. The results are given elsewhere (being submitted for peer review) . They show that overnight exposure to these radiations also have an important adverse effect on lymphocyte viability DISCUSSION Chemical communication between cells through gap junctions is routinely complemented by charge-related signal transduction mechanisms via sialic acid residues on glycoproteins with subsequent cytosolic cascades implicating [Ca2+]i as a second messenger. These systems facilitate processes such as contact inhibition, and without them cells proliferate out of regulatory growth control, leading, with failure of immune cell surveillance or apoptotic competence, to solid tumour formation (Rosenberg & Terry, 1977). Similarly electric fields from a special pacemaker region of the heart (the sino-atrial node) synchronise the orderly contraction of muscle cells in the heart, also Ca2+ dependent (Orchard, 1998). Electric fields are also rhythmically emitted by mammalian brains. All these fields can be detected at or near the skin surface (EEG, EKG). The glycocalyx of lymphocytes is exceptionally rich in glycoproteins capable of almost infinite structural variation (Sharon & Lis, 1993). Oligosaccharide side-chains of glycoproteins and glycolipids, though usually containing fewer than 15 sugar residues, are normally branched, and are enormously diverse in their arrangement of sugars: "even three sugar residues can be put together to form hundreds of different trisaccharides" (Alberts, Bray et al., 1994, p 502). This variety permits exquisite signal recognition both chemically and electrically. Uckun, Kurosaki et al., (1995) showed that the protein tyrosine kinases (PTK) which play pivotal roles in the initiation of signal cascades affecting the proliferation and survival of human B-lineage lymphoid cells are stimulated by a 1 Gauss 60 Hz field, implying a receptor role. In seeking a corresponding natural "transmitter" source similar to that of the heart, detectable rhythmic ultra low frequency (ULF) rhythms from the brain have been characterised, but not functionally explained (Berger, 1929). During sleep these rhythms include regular periods of high amplitude (paradoxical or REM sleep), punctuated by periods of comparatively low electrical activity (Dement, Greenberg et al., 1965). The structures responsible for these emissions are also capable of infinite variety, since they emanate from trans-hemispheral potential differences of c. 70mV along the 30,000 or so intercerebral connections between pyramidal cells (Hodgkins & Huxley, 1952), individually under thalamic control (Andersen & Andersson, 1968). Consistent changes in EEG records in pigtailed macaque monkeys in response to 7Hz (but not 10Hz) electric fields of 2.8V p-p were reported (Gavalas, Walter et al., 1970), together with altered reaction times, and epileptiform activity in rat hippocampal slices can be suppressed by electric fields (Gluckman, Neel et al., 1996), indicating that, as well as cells, the mammalian brain is also sensitive to such fields. Organic saline solutions include 0.9% Na+Cl-, and though these ions are in balance, they are separated by hydration, assisting thereby in transmission processes through the medium. Connected by this bathing solution the brain and cells may thus be prepared by evolution to send/receive electric fields via the physiological saline solutions of the body almost losslessly, and to convey frequency- and amplitude-modulated information. In a normal adult human some 500 million cells are renewed each night, mainly during REM sleep, suggesting that the brain fulfils its role of morphological control by instructing for mitosis during REM sleep, rather than relying on local or neuro-chemical events. (No mechanism for initiating mitosis at a local level has yet been identified). It has been assumed that intercellular signals and neural signals from the brain via the CNS are solely electrochemical in nature. The results of this study show however that lymphocyte viability is improved by exposure to endogenous electric fields from the same donor, in the absence of any chemical or magnetic field influences. Studies reporting negative EM effects have principally deployed magnetic rather than electric field exposure systems. For example, whilst Bawin and Adey (1976) found modification of Ca2+ efflux from freshly isolated chick and cat cerebral tissue exposed to ELF electric fields of less than 56 V/m, Coulton & Barker (1993) reported an absence of effect on calcium uptake by murine lymphocytes at the same (ion cyclotron resonance "ICR") frequencies using alternating magnetic fields of 46 microTesla. Similarly Prasad (1991) found no 45Ca2+ response to magnetic fields at ICR conditions in human peripheral blood lymphocytes. This suggests an electric rather than magnetic parameter of interaction and a communications role for endogenous fields. Endogenous fields emanating from the body originate largely in the brain and the heart. The fields used for exposure in this study almost certainly emanate from the brain rather than elsewhere. Penfield & Boldrey (1937) observed that sensory/motor cortical neurones were arrayed so that those body organs nearest the brain were furthest apart on the cortical surface and vice versa. This is consistent with the explanation that cortical neurones nearest the brain need to carry the lowest potential differences for transmission purposes. Indeed, cortical areas reserved for important drives (procreation, preservation) may have been displaced into the nearest and hence most heavily charged regions for that reason (see Diag. 3). Moreover evidence from a number of studies points towards a link between central neural activity and altered immune responses. For example lesions of the anterior/preoptic regions of the hypothalamus resulted in diminished lymphocyte competence (Felten, Cohen et al., 1991) and decreases in the helper/suppressor T-cell ratio (Katayama, Kobayashi et al., 1987). Lesions of the left cerebral cortex in mice resulted in decreased T-cell numbers and responses (Renoux , Biziere et al., 1987). It is equally consistent that REM sleep occurs only when signal-to-noise conditions are optimal (i.e. when corporeal electrical activity is minimal), and is inhibited or discontinued in the presence of competing exogenous alternating (AC) magnetic fields (Graham, Cook et al., 1997). Mitosis occurs largely during the periods of REM sleep (Palmer, 1976), and is associated with altered electric potential at the membrane surface (Cone, 1971). Similarly in cell studies it has been shown that exogenous ELF/ULF EM fields have important proliferative or inhibiting effects on growth rates (Blackman et al., 1985 etc.) as well as on the competence of cytotoxic lymphocytes (Lyle Shechter et al., 1983; Lyle, Ayotte et al., 1988). Given known speeds of nervous conduction via synapses (typically 20m/s) it is not possible to instruct individually for mitosis of (e.g) 500 million cells within one normal sleep period, ruling out chemico-physical control of morphology at a central level. In order to explain the results of this study we propose that during evolution this problem has been solved for multicellular organisms by the use of endogenous currents giving rise to electric fields alternating in frequency and amplitude as a means of intracorporeal information transfer. Ambient ELF electric fields and RF/MW radiations from artificial EM sources appear from our study to have adverse effects on lymphocyte viability. This would confirm the many studies recently reporting adverse bioeffects at a cellular and whole organism level from EM fields too low to induce thermal effects, and invokes renewed consideration of EM compatibility between modern electronic technologies and organic life. The system proposed here does not exclude the possibility of electrical cell-to-brain communication, possibly also via the reticular activating system and its connections to the hypothalamus, which has also been reported (Saphier, Abramsky et al., 1987; Besedovski, Sorkin et al., 1977). It would be interesting to see if endogenous electric fields from other subjects, or frequencies other than 50Hz., have similar effects on lymphocyte viability. CONCLUSIONS Exposure to the endogenous electric fields of their donor appears to improve the viability of human peripheral blood lymphocytes, whereas exposure to to artificial 50Hz. or RF/MW electric fields depresses viability. This finding may have implications for the avoidance and management of immune-related health disorders. The existence of a purposive biophysical communications system between brain and immune system cells cannot be ruled out. Note : References used in this text may be found at our references section on the website. |
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