Altered calcium dynamics mediates P19-derived neuron-like cell responses to millimeter-wave radiation.

Intracellular calcium oscillations have long been recognized as a principal mediator of many vital cellular activities. Furthermore, Ca(2+) dynamics can be modulated by external physical cues, including electromagnetic fields. While cellular responses to low-frequency electric fields have been established, the possible non-thermal effects of millimeter-wave (MMW) radiation are still a subject of discussion and debate. We used mouse embryonic stem cell-derived neuronal cells and a custom-built 94 GHz applicator to examine in real time the altered Ca(2+) oscillations associated with MMW stimulation. MMW irradiation at 18.6 kW/m(2) nominal power density significantly increased the Ca(2+) spiking frequency in the cells exhibiting Ca(2+) activity. The N-type calcium channels, phospholipase C enzyme, and actin cytoskeleton appear to be involved in mediating increased Ca(2+) spiking. Reorganization of the actin microfilaments by a 94 GHz field seems to play a crucial role in modulating not only Ca(2+) activity but also cell biomechanics. Many but not all observed cellular responses to MMW were similar to thermally induced effects. For example, cell exposure to a 94 GHz field induced nitric oxide production in some morphologically distinct neuronal cells that could not be reproduced by thermal heating of the cells up to 42 degrees C. The highest observed average temperature rise in the MMW exposure chamber was approximately 8 degrees C above the room temperature, with possible complex non-uniform microscopic distribution of heating rates at the cell level. Our findings may be useful to establish quantitative molecular benchmarks for elucidation of nociception mechanisms and evaluation of potential adverse bioeffects associated with MMW exposure. Moreover, control of Ca(2+) dynamics by MMW stimulation may offer new tools for regulation of Ca(2+)-dependent cellular and molecular activities, for example, in tissue engineering applications.

Nonthermal effects of radiofrequency-field exposure on calcium dynamics in stem cell-derived neuronal cells: elucidation of calcium pathways.

Intracellular Ca(2+) spikes trigger cell proliferation, differentiation and cytoskeletal reorganization. In addition to Ca(2+) spiking that can be initiated by a ligand binding to its receptor, exposure to electromagnetic stimuli has also been shown to alter Ca(2+) dynamics. Using neuronal cells differentiated from a mouse embryonic stem cell line and a custom-built, frequency-tunable applicator, we examined in real time the altered Ca(2+) dynamics and observed increases in the cytosolic Ca(2+) in response to nonthermal radiofrequency (RF)-radiation exposure of cells from 700 to 1100 MHz. While about 60% of control cells (not exposed to RF radiation) were observed to exhibit about five spontaneous Ca(2+) spikes per cell in 60 min, exposure of cells to an 800 MHz, 0.5 W/kg RF radiation, for example, significantly increased the number of Ca(2+) spikes to 15.7+/-0.8 (P<0.05). The increase in the Ca(2+) spiking activities was dependent on the frequency but not on the SAR between 0.5 to 5 W/kg. Using pharmacological agents, it was found that both the N-type Ca(2+) channels and phospholipase C enzymes appear to be involved in mediating increased Ca(2+) spiking. Interestingly, microfilament disruption also prevented the Ca(2+) spikes. Regulation of Ca(2+) dynamics by external physical stimulation such as RF radiation may provide a noninvasive and useful tool for modulating the Ca(2+)-dependent cellular and molecular activities of cells seeded in a 3D environment for which only a few techniques are currently available to influence the cells.

Tailed forisomes of Canavalia gladiata: a new model to study Ca2+-driven protein contractility.

BACKGROUND AND AIMS: Forisomes are Ca(2+)-dependent contractile protein bodies that form reversible plugs in sieve tubes of faboid legumes. Previous work employed Vicia faba forisomes, a not entirely unproblematic experimental system. The aim of this study was to seek to establish a superior model to study these intriguing actuators. METHODS: Existing isolation procedures were modified to study the exceptionally large, tailed forisomes of Canavalia gladiata by differential interference contrast microscopy in vitro. To analyse contraction/expansion kinetics quantitatively, a geometric model was devised which enabled the computation of time-courses of derived parameters such as forisome volume from simple parameters readily determined on micrographs. KEY RESULTS: Advantages of C. gladiata over previously utilized species include the enormous size of its forisomes (up to 55 microm long), the presence of tails which facilitate micromanipulation of individual forisomes, and the possibility of collecting material repeatedly from these fast-growing vines without sacrificing the plants. The main bodies of isolated Canavalia forisomes were box-shaped with square cross-sections and basically retained this shape in all stages of contraction. Ca(2+)-induced a 6-fold volume increase within about 10-15 s; the reverse reaction following Ca(2+)-depletion proceeded in a fraction of that time. CONCLUSIONS: The sword bean C. gladiata provides a superior experimental system which will prove indispensable in physiological, biophysical, ultrastructural and molecular studies on the unique ATP-independent contractility of forisomes.

Measurement of DNA damage after acute exposure to pulsed-wave 2450 MHz microwaves in rat brain cells by two alkaline comet assay methods.

PURPOSE: To investigate the effect of 2450 MHz pulsed-wave microwaves on the induction of DNA damage in brain cells of exposed rats and to discover whether proteinase K is needed to detect DNA damage in the brain cells of rats exposed to 2450 MHz microwaves. MATERIALS AND METHODS: Sprague-Dawley rats were exposed to 2450 MHz pulsed-wave microwaves and sacrificed 4 h after a 2-h exposure. Rats irradiated whole-body with 1 Gy (137)Cs were included as positive controls. DNA damage was assayed by two variants of the alkaline comet assay on separate aliquots of the same cell preparation. RESULTS: Significant DNA damage was observed in the rat brain cells of rats exposed to gamma-rays using both versions of the alkaline comet assay independent of the presence or absence of proteinase K. However, neither version of the assay could detect any difference in comet length and/or normalized comet moment between sham- and 2450 MHz pulsed-wave microwave-exposed rats, regardless of the inclusion or omission of proteinase K in the comet assay. CONCLUSIONS: No DNA damage in brain cells was detected following exposure of rats to 2450 MHz microwaves pulsed-wave at a specific absorption rate of 1.2 W kg(-1) regardless of whether or not proteinase K was included in the assay. Thus, the results support the conclusion that low-level 2450 MHz pulsed-wave microwave exposures do not induce DNA damage detectable by the alkaline comet assay.

Micronuclei in the peripheral blood and bone marrow cells of rats exposed to 2450 MHz radiofrequency radiation.

PURPOSE: To determine the incidence of micronuclei in peripheral blood and bone marrow cells of rats exposed continuously for 24h to 2450 MHz continuous wave radiofrequency radiation (RFR) at an average whole-body specific absorption rate (SAR) of 12W/kg. MATERIALS AND METHODS: Eight adult male Sprague-Dawley rats were exposed to 2450 MHz RFR in circularly polarized waveguides. Eight sham-exposed rats were kept in similar waveguides without the transmission of RFR. Four rats were treated with mitomycin-C (MMC) and used as positive controls. All rats were necropsied 24h after the end of RFR and sham exposures, and after the 24h treatment with MMC. Peripheral blood and bone marrow smears were examined to determine the frequency of micronuclei (MN) in polychromatic erythrocytes (PCE). RESULTS: The results indicated that the incidence of MN/2000 PCE were not significantly different between RFR- and sham-exposed rats. The group mean frequencies of MN in the peripheral blood were 2.3+/-0.7 in RFR-exposed rats and 2.1+/-0.6 in sham-exposed rats. In bone marrow cells, the average MN incidence was 3.8+/-1.0 in RFR-exposed rats and 3.4+/-0.7 in sham-exposed rats. The corresponding values in positive control rats treated with MMC were 23.5+/-4.7 in the peripheral blood and 33.8+/-7.4 in bone marrow cells. CONCLUSION: There was no evidence for the induction of MN in peripheral blood and bone marrow cells of rats exposed for 24h to 2450 MHz continuous wave RFR at a whole body average SAR of 12 W/kg.

Chromosome damage and micronucleus formation in human blood lymphocytes exposed in vitro to radiofrequency radiation at a cellular telephone frequency (847.74 MHz, CDMA).

Peripheral blood samples collected from four healthy nonsmoking human volunteers were diluted with tissue culture medium and exposed in vitro for 24 h to 847.74 MHz radiofrequency (RF) radiation (continuous wave), a frequency employed for cellular telephone communications. A code division multiple access (CDMA) technology was used with a nominal net forward power of 75 W and a nominal power density of 950 W/m(2) (95 mW/cm(2)). The mean specific absorption rate (SAR) was 4.9 or 5.5 W/kg. Blood aliquots that were sham-exposed or exposed in vitro to an acute dose of 1.5 Gy of gamma radiation were included in the study as controls. The temperatures of the medium during RF-radiation and sham exposures in the Radial Transmission Line facility were controlled at 37 +/- 0.3 degrees C. Immediately after the exposures, lymphocytes were cultured at 37 +/- 1 degrees C for 48 or 72 h. The extent of genetic damage was assessed from the incidence of chromosome aberrations and micronuclei. The kinetics of cell proliferation was determined from the mitotic indices in 48-h cultures and from the incidence of binucleate cells in 72-h cultures. The data indicated no significant differences between RF-radiation-exposed and sham-exposed lymphocytes with respect to mitotic indices, frequencies of exchange aberrations, excess fragments, binucleate cells, and micronuclei. The response of gamma-irradiated lymphocytes was significantly different from that of both RF-radiation-exposed and sham-exposed cells for all of these indices. Thus there was no evidence for induction of chromosome aberrations and micronuclei in human blood lymphocytes exposed in vitro for 24 h to 847.74 MHz RF radiation (CDMA) at SARs of 4.9 or 5.5 W/kg.

Energy deposition processes in biological tissue: nonthermal biohazards seem unlikely in the ultra-high frequency range.

The prospects of ultra high frequency (UHF, 300--3000 MHz) irradiation producing a nonthermal bioeffect are considered theoretically and found to be small. First, a general formula is derived within the framework of macroscopic electrodynamics for the specific absorption rate of microwaves in a biological tissue; this involves the complex Poynting vector, the mass density of the medium, the angular frequency of the electromagnetic field, and the three complex electromagnetic constitutive parameters of the medium. In the frequency ranges used for cellular telephony and personal communication systems, this model predicts that the chief physical loss mechanism will be ionic conduction, with increasingly important contributions from dielectric relaxation as the frequency rises. However, even in a magnetite unit cell within a magnetosome the deposition rate should not exceed 1/10 k(B)T per second. This supports previous arguments for the improbability of biological effects at UHF frequencies unless a mechanism can be found for accumulating energy over time and space and focussing it. Second, three possible nonthermal accumulation mechanisms are then considered and shown to be unlikely: (i) multiphoton absorption processes; (ii) direct electric field effects on ions; (iii) cooperative effects and/or coherent excitations. Finally, it is concluded that the rate of energy deposition from a typical field and within a typical tissue is so small as to make unlikely any significant nonthermal biological effect.

Cytogenetic studies in human blood lymphocytes exposed in vitro to radiofrequency radiation at a cellular telephone frequency (835.62 MHz, FDMA).

Freshly collected peripheral blood samples from four healthy human volunteers were diluted with RPMI 1640 tissue culture medium and exposed in sterile T-75 tissue culture flasks in vitro for 24 h to 835.62 MHz radiofrequency (RF) radiation, a frequency employed for customer-to-base station transmission of cellular telephone communications. An analog signal was used, and the access technology was frequency division multiple access (FDMA, continuous wave). A nominal net forward power of 68 W was used, and the nominal power density at the center of the exposure flask was 860 W/m(2). The mean specific absorption rate in the exposure flask was 4.4 or 5.0 W/kg. Aliquots of diluted blood that were sham-exposed or exposed in vitro to an acute dose of 1.50 Gy of gamma radiation were used as negative or positive controls. Immediately after the exposures, the lymphocytes were stimulated with a mitogen, phytohemagglutinin, and cultured for 48 or 72 h to determine the extent of genetic damage, as assessed from the frequencies of chromosomal aberrations and micronuclei. The extent of alteration in the kinetics of cell proliferation was determined from the mitotic indices in 48-h cultures and from the incidence of binucleate cells in 72-h cultures. The data indicated no significant differences between RF-radiation- and sham-exposed lymphocytes with respect to mitotic indices, incidence of exchange aberrations, excess fragments, binucleate cells, and micronuclei. In contrast, the response of the lymphocytes exposed to gamma radiation was significantly different from both RF-radiation- and sham-exposed cells for all of these indices. Thus, under the experimental conditions tested, there is no evidence for the induction of chromosomal aberrations and micronuclei in human blood lymphocytes exposed in vitro for 24 h to 835.62 MHz RF radiation at SARs of 4.4 or 5.0 W/kg.

Neoplastic transformation in C3H 10T(1/2) cells after exposure to 835.62 MHz FDMA and 847.74 MHz CDMA radiations.

The effect of radiofrequency (RF) radiation in the cellular phone communication range (835.62 MHz frequency division multiple access, FDMA; 847.74 MHz code division multiple access, CDMA) on neoplastic transformation frequency was measured using the in vitro C3H 10T(1/2) cell transformation assay system. To determine if 835.62 MHz FDMA or 847.74 MHz CDMA radiations have any genotoxic effects that induce neoplastic transformation, C3H 10T(1/2) cells were exposed at 37 degrees C to either of the above radiations [each at a specific absorption rate (SAR) of 0.6 W/kg] or sham-exposed at the same time for 7 days. After the culture medium was changed, the cultures were transferred to incubators and refed with fresh growth medium every 7 days. After 42 days, the cells were fixed and stained with Giemsa, and transformed foci were scored. To determine if exposure to 835.62 MHz FDMA or 847.74 MHz CDMA radiation has any epigenetic effects that can promote neoplastic transformation, cells were first exposed to 4.5 Gy of X rays to induce the transformation process and then exposed to the above radiations (SAR = 0.6 W/kg) in temperature-controlled irradiators with weekly refeeding for 42 days. After both the 7-day RF exposure and the 42-day RF exposure after X irradiation, no statistically significant differences in the transformation frequencies were observed between incubator controls, the sham-exposed (maintained in irradiators without power to the antenna), and the 835.62 MHz FDMA or 847.74 MHz CDMA-exposed groups.

How might spatial nonuniformity of dose in a homogeneous biological system affect its total response?

The total response of a homogeneous biological system to a fixed total dose of a biological agent is modeled by dividing the system into N cubical voxels, each of which can be associated with an individual dose D(n) and an individual response R(n) =F(D(n)). Among the results shown are the following: A. (Voxel Theorem). Let the average dose D(avg) be held fixed as the dose distribution is shifted from uniform u to arbitrary a. Then, if F' > or = 0 over [D(min), D(max)] and R = summation operator (n = 1)(N) R(n), a sufficient condition that NF(D(avg)) = R(u) < or = R(a) is that F be a concave-upwards function of dose; that is, F" > or = 0 over [D(min), D(max)]. B. If F' is constant over [D(min), D(max)], then R(a) = R(u). That is, the total response is a function of D(avg) only. The applications of these (and other) results are illustrated by examples from bioelectromagnetics.

Experimental and numerical determination of SAR distributions within culture flasks in a dielectric loaded radial transmission line.

The effect of dielectric loading on the cell layer specific absorption rate (SAR) within a T-75 culture flask being irradiated within a transverse electromagnetic (TEM) cell was studied both experimentally and numerically. Direct thermal measurements of a T-75 containing 40 mL of culture medium and resting upon a 3-mm-thick slab of alumina ceramic (epsilon r = 9.6) revealed that, compared to the same flask resting upon a foam slab (epsilon r = 1.0) of the same thickness, the average SAR at the cell layer was increased roughly fourfold. This fourfold increase is significant experimentally because it allows biologists to perform experiments over a larger range of SAR values needed to determine possible dose-response curves without the costs and difficulties of a fourfold increase in amplifier power. Finite-difference time-domain (FDTD) simulations of the SAR distribution were in good quantitative agreement with the experimental measurements. It is concluded that FDTD modeling can be a cost effective and scientifically acceptable means of obviating the thermal measurement of SAR.

Radiofrequency electromagnetic fields have no effect on the in vivo proliferation of the 9L brain tumor.

The intracranial 9L tumor model was used to determine if exposure to a radiofrequency (RF) electromagnetic field similar to those used in cellular telephone has any effects on the growth of a central nervous system tumor. Fischer 344 rats implanted with different numbers of 9L gliosarcoma cells were exposed to 835.62 MHz frequency-modulated continuous wave (FMCW) or 847.74 MHz code division multiple access (CDMA) RF field with nominal slot-average specific absorption rates in the brain of 0.75 +/- 0.25 W/kg. The animals were exposed to the RF field for 4 h a day, 5 days a week starting 4 weeks prior to and up to 150 days after the implantation of tumor cells. Among sham-exposed animals injected with 2 to 10 viable cells (group 1), the median survival was 70 days, with 27% of the animals surviving at 150 days. The median survival length and final survival fraction for animals injected with 11 to 36 viable cells (group 2) were 52 days and 14%, respectively, while the values for those injected with 37 to 100 cells (group 3) were 45 days and 0%. The animals exposed to CDMA or FMCW had similar survival parameters, and the statistical comparison of the survival curves for each of the groups 1, 2 and 3 showed no significant differences compared to sham-exposed controls.

On the assumption of negligible heat diffusion during the thermal measurement of a nonuniform specific absorption rate.

The problem of measuring a spatially varying specific absorption rate (SAR) by thermal techniques is treated both analytically and numerically. It is shown that, unless the measurement is attempted at an inflection point of the SAR distribution, it will be confounded by thermal diffusion within the medium. Rules of thumb are provided to enable the experimenter to gauge the thermal conduction contributions (i.e. error) to the thermally determined SAR near a spatial extremum. The simplest of these is that the width t(m) [s] of the time window, over which temperature variation associated with SAR is measured, should satisfy the inequality t(m)

Cell phones and cancer: what is the evidence for a connection?

There have been allegations in the media and in the courts that cell phones and other types of hand-held transceivers are a cause of cancer. There have also been numerous public objections to the siting of TV, radio and cell phone transmission facilities because of a fear of cancer induction. A recent publication in Radiation Research by Repacholi et al. (147, 631-640, 1997) which suggests that exposure to radiofrequency (RF) radiation may increase lymphoma incidence in mice has contributed to this controversy. The goal of this review is to provide biomedical researchers a brief overview of the existing RF radiation-cancer studies. This article begins with a brief review of the physics and technology of cell phones. It then reviews the existing epidemiological studies of RF radiation, identifying gaps in our knowledge. Finally, the review discusses the cytogenetics literature on RF radiation and the whole-animal RF-radiation carcinogenesis studies. The epidemiological evidence for an association between RF radiation and cancer is found to be weak and inconsistent, the laboratory studies generally do not suggest that cell phone RF radiation has genotoxic or epigenetic activity, and a cell phone RF radiation-cancer connection is found to be physically implausible. Overall, the existing evidence for a causal relationship between RF radiation from cell phones and cancer is found to be weak to nonexistent.

Simplified model and measurement of specific absorption rate distribution in a culture flask within a transverse electromagnetic mode exposure system.

In vitro experiments in bioelectromagnetics frequently require the determination of specific absorption rate (SAR) within a layer of cells on the bottom of a culture flask when the SAR has rapid spatial variation both horizontally within the cell layer and vertically in the medium bathing the cells. This problem has only recently been treated in the literature; and it is here approached differently for another irradiation system. It is shown that a simple two-dimensional frequency-domain guided-wave treatment yields results qualitatively comparable to those of more computationally intensive three-dimensional time-domain free-field scattering treatments. The problem of inferring local SARs from temperature-vs.-time curves is shown to be seriously confounded by thermal diffusion; and specific analytic and numerical results are presented to aid in understanding this effect. A novel experimental technique is introduced for measuring millikelvin temperature offsets with subsecond resolution, and illustrative experimental data are presented. Finally, present experimental and theoretical uncertainties are considered; and it is pessimistically asserted that, in a culture flask where spatial SAR variation is rapid, point SAR measurements by thermal methods may be in error by as much as +/- 3 dB. More reliable thermal determinations will require extreme care, challenging technological innovations, or both.

Proto-oncogene mRNA levels and activities of multiple transcription factors in C3H 10T 1/2 murine embryonic fibroblasts exposed to 835.62 and 847.74 MHz cellular phone communication frequency radiation.

This study was designed to determine whether two differently modulated radiofrequencies of the type generally used in cellular phone communications could elicit a general stress response in a biological system. The two modulations and frequencies studied were a frequency-modulated continuous wave (FMCW) with a carrier frequency of 835.62 MHz and a code division multiple-access (CDMA) modulation centered on 847.74 MHz. Changes in proto-oncogene expression, determined by measuring Fos, Jun, and Myc mRNA levels as well as by the DNA-binding activity of the AP1, AP2 and NF-kappaB transcription factors, were used as indicators of a general stress response. The effect of radiofrequency exposure on proto-oncogene expression was assessed (1) in exponentially growing C3H 10T 1/2 mouse embryo fibroblasts during their transition to plateau phase and (2) during transition of serum-deprived cells to the proliferation cycle after serum stimulation. Exposure of serum-deprived cells to 835.62 MHz FMCW or 847.74 MHz CDMA microwaves (at an average specific absorption rate, SAR, of 0.6 W/kg) did not significantly change the kinetics of proto-oncogene expression after serum stimulation. Similarly, these exposures did not affect either the Jun and Myc mRNA levels or the DNA-binding activity of AP1, AP2 and NF-kappaB in exponential cells during transit to plateau-phase growth. Therefore, these results suggest that the radiofrequency exposure is unlikely to elicit a general stress response in cells of this cell line under these conditions. However, statistically significant increases (approximately 2-fold, P = 0.001) in Fos mRNA levels were detected in exponential cells in transit to the plateau phase and in plateau-phase cells exposed to 835.62 MHz FMCW microwaves. For 847.74 MHz CDMA exposure, the increase was 1.4-fold (P = 0.04). This increase in Fos expression suggests that expression of specific genes could be affected by radiofrequency exposure.

The radial transmission line as a broad-band shielded exposure system for microwave irradiation of large numbers of culture flasks.

The problem of simultaneously exposing large numbers of culture flasks at nominally equivalent incident power densities and with good thermal control is considered, and the radial transmission line (RTL) is proposed as a solution. The electromagnetic design of this structure is discussed, and an extensively bench-tested realization is described. Referred to 1 W of net forward power, the following specific absorption rate (SAR) data were obtained: at 835.62 MHz, 16.0+/-2.5 mW/kg (mean+/-SD) with range (11-22); at 2450 MHz, 245+/-50 mW/kg with range (130-323). Radio-frequency interference from an RTL driven at roughly 100 W is so low as to be compatible with a cellular base station only 500 m distant. To avoid potential confounding by temperature differences among as many as 144 T-75 flasks distributed over 9 RTLs (six irradiates and three shams), temperature within all flasks was controlled to 37.0+/-0.3 degrees C. Experience with over two years of trouble-free operation suggests that the RTL offers a robust, logistically friendly, and environmentally satisfactory solution to the problem of large-scale in vitro experiments in bioelectromagnetics.

Compact shielded exposure system for the simultaneous long-term UHF irradiation of forty small mammals. II. Dosimetry.

A four-antenna collinear array in an electromagnetically shielded chamber was designed and constructed to preferentially irradiate the brains of a large number of small mammals using cellular telephony microwave signals. Ten animals in special restrainers were positioned symmetrically around a centrally located antenna. These restrainers are resting on a circular structure made of acrylic plastic called a "carousel." Four carousels are stacked vertically, forming the array, inside a microwave anechoic chamber called a "chamberette." (Details of the design of this irradiator and of a 12-chamberette irradiation facility are given in a previous article.) In this article, the dosimetry on rats is reported. Both thermometric and thermographic measurements were performed. The average specific absorption rate (SAR) in brain tissue measured thermometrically was 0.85+/-0.34 W/kg per watt of net input power into the radiating antenna. This range agrees with the SAR levels reported in the literature for cellular telephones. Thermographic evaluation using splittable phantoms showed that most of the energy absorbed by the rats is concentrated in and around the brain. Moreover, it was found that the SAR in brain tissue can vary considerably for rats of similar weights, depending on position of the rats' heads inside the restrainers, and that there exists a significant dependence of SAR on animal weight. These variations may be of importance in the interpretation of results of lifelong studies. The data presented clearly show that the chamberette is, dosimetrically, a suitable irradiation system for electromagnetic bioeffects studies in the cellular communication frequency range, especially when a large number of laboratory animals is required.

A compact shielded exposure system for the simultaneous long-term UHF irradiation of forty small mammals: I. Electromagnetic and environmental design.

To carry out in vivo studies of the possible health effects of radiation from cellular telephone handsets, it is necessary to expose large numbers of small mammals at realistic power densities, modulations, and frequencies. Because even microwatt leakage could compromise the local cellular system, extreme care in shielding is required. Experimental logistics dictate, however, that the irradiated animals be easily accessed and that it be possible to irradiate them in small groups, while other groups are being loaded into or unloaded from the irradiators. This problem has been resolved by exposing the animals in aluminum-sheathed rectangular parallelepipeds, lined with microwave absorber and having doors that can be opened readily. Inside each of these microwave anechoic "chamberettes" is a vertical, four-element collinear array of dipole antennas; and around each antenna, 10 animal restrainers can be arranged like spokes on a wheel. The system has worked efficiently in studies of up to 480 rats. There is negligible coupling between antennas, and back reflection at an antenna's feed line is down 7-9 dB. Received CDMA power at the local base station is below the receiver's noise floor. Interior illumination reinforces the rats' diurnal rhythms, and the rats sleep during irradiation. Experimental logistics are excellent. In this paper, the irradiator design is presented.

DNA damage in rat brain cells after in vivo exposure to 2450 MHz electromagnetic radiation and various methods of euthanasia.

The present study was done to confirm the reported observation that low-intensity acute exposure to 2450 MHz radiation causes DNA single-strand breaks (Lai and Singh, Bioelectromagnetics 16, 207-210, 1995). Male Sprague-Dawley rats weighing approximately 250 g were irradiated with 2450 MHz continuous-wave (CW) microwaves for 2 h at a specific absorption rate of 1.2 W/kg in a cylindrical waveguide system (Guy et al., Radio Sci. 14, 63-74, 1979). There was no associated rise in the core body temperature of the rats. After the irradiation or sham treatments, rats were euthanized by either CO2 asphyxia or decapitation by guillotine (eight pairs of animals per euthanasia group). After euthanasia the brains were removed and immediately immersed in cold Ames medium and the cells of the cerebral cortex and the hippocampus were dissociated separately and subjected to the alkaline comet assay. Irrespective of whether the rats were euthanized by CO2 asphyxia or decapitated by guillotine, no significant differences were observed between either the comet length or the normalized comet moment of cells from either the cerebral cortex or the hippocampus of sham-treated rats and those from the irradiated rats. However, the data for the rats asphyxiated with CO2 showed more intrinsic DNA damage and more experiment-to-experiment variation than did the data for rats euthanized by guillotine. Therefore, the guillotine method of euthanasia is the most appropriate in studies relating to DNA damage. Furthermore, we did not confirm the observation that DNA damage is produced in cells of the rat cerebral cortex or the hippocampus after a 2-h exposure to 2450 MHz CW microwaves or at 4 h after the exposure.