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AN EVALUATION OF ORNITHINE DECARBOXYLASE ACTIVITY IN BRAIN REGIONS OF FETAL RATS EXPOSED TO RADIOFREQUENCY FIELDS. L.E. Anderson, J.E. Morris, L.B. Sasser, J.A. Creim, A.B Desta*, L. Cress* and R. Owen*. Battelle, Richland, Washington 99352, and *FDA, Rockville, MD 20850, USA .

OBJECTIVE: Ornithine decarboxylase activity (ODC) has been shown to be a sensitive marker in the brain tissue of the developing organism in terms of both growth and stress. The overall objective of this study is to conduct an experiment designed to measure the effect of exposure to 1.62 GHz radiofrequency electromagnetic fields on the activity level of ODC in the cerebellum and cerebrum of fetal Fischer 344 rats.

METHOD: Timed-pregnant Fischer-344 rats were assigned (randomized by weight) to five treatment groups as follows; 0.16 W/kg; 1.6 W/kg; 5.0 W/kg, sham (tube restraint) controls, and cage controls. Each treatment group was designated by a color code to facilitate correct assignment and tracking of the groups, while still preserving the blinded nature (for exposure and sham-exposure groups) of the study. Exposures began on day 18 of gestation and continued to day 21 at two hours per day. Sham-exposed control animals were placed in the exposure system with no field present during the two-hour exposure period. The fifth group remained as cage controls without exposure or sham exposure treatment. On day 21 the dams were sacrificed, the fetuses removed and brain sections were taken for analysis. ODC activity was measured by a modified method of Seely and Pegg in which the brain sections were placed in 0.5 ml of ice-cold ODC buffer (50 mM Tris-HCl, pH7.2, 2.5 mM dithiothreitol, 1.25 mM EDTA, 0.1 mM pyridoxyl 5'-phosphate, and 0.1 mM PMSF protease inhibitor). Cerebral and cerebellar tissues were homogenized and then centrifuged at 30,000 x g for 30 minutes. Two 100 µl aliquots of the supernatant per sample were assayed for ODC activity by measuring the amount of ¹⁴CO₂ released from L-1-¹⁴C-ornithine. Samples were incubated for one hour at 37 C in the Tris assay buffer containing both unlabeled (0.4mM) and labeled (2.77x10⁵ dpm) L-ornithine. Enzymatically released ¹⁴CO₂ was trapped in 100 µl of 1.0 N NaOH. Following the incubation, the reaction was terminated by addition of 300 µl of 20% trichloroacetic acid directly into the reaction mixture. After an additional 2-hour incubation at room temperature, the NaOH in center-well was removed and radioactivity measured in 10 ml of nonaqueous scintillation fluid. One enzyme unit of ODC is defined as the amount of enzyme required to catalyze the release of 1 picomole of CO₂ from L-ornithine per milligram of protein in 30 minutes.

RESULTS AND DISCUSSION: In the analysis, 80 litters were evaluated. From each litter, samples were prepared for analysis that included brain tissue from 2 fetuses per sample. Up to 4 samples per litter thus produced 250 fetal samples, representing 500 fetuses. ODC assays were performed blind at the FDA. Preliminary analysis of the data indicate no significant differences between ODC activity levels in animals exposed to 0.16, 1.6 or 5.0 W/kg when compared with sham exposed controls.

Updated 10/30/01