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研究目的（著者による）
The effects of exposure of different strains of Drosophila melanogaster to static and different extremely low frequency magnetic fields and to visible light on the circadian and general locomotor activity should be examined. Additionally, the bioluminescence as a response to 50 Hz magnetic fields was examined in brain slices from the suprachiasmatic nucleus (brain region which is responsible for controlling circadian rhythms) of mice.
詳細情報
The mechanism of magnetoreception in animals cannot be explained by now. A hypothesis suggests that cryptochrome (a photoreceptor, activated by light and as a consequence becomes susceptible to magnetic fields) may act as a magnetoreceptor through a radical pair mechanism involving tryptophans and the flavin cofactor (FAD)). This hypothesis should be investigated by using wild type Drosophila melanogaster, different genetically modified cryptochrome mutants of Drosophila melanogaster, and mammalian cryptochrome both in transgenic Drosophila as well as in its "mammalian environment" (brain slices). While Drosophila melanogaster contains CRY as a photoreceptor, in non-drosophilid insects there can be type 1 CRY and type 2 CRY and in mammalian species there are two similar type 2 CRYs.
The wild type flies were exposed to different conditions: 1) static magnetic field (300 µT), 2) 50 Hz magnetic field (300 µT), 3) 3 Hz magnetic field (300 µT), 4) 3 Hz magnetic field (90 µT), and 5) 3 Hz magnetic field (1000 µT). The genetically modified strains were exposed to a 3 Hz magnetic field (300 µT). The suprachiasmatic nuclei slices were exposed to a 50 Hz magnetic field with different magnetic flux densities of 50 µT (number of slices =10), 150 µT (n=5), 300 µT (n=10), or 500 µT (n=10). The experimental design was as follows for the experiments with the flies: Two groups of flies of the same genotype were studied for seven days under constant dim blue light followed by eight days under the same illumination but exposed either to a magnetic field or a sham exposure. The tissue slices were magnetic field exposed for five days and afterwards 5 days sham exposed orvice versa. Different illumination conditions (wavelength) were used.
flies were exposed in a modified Schuderer apparatus (Schuderer et al., 2004); field was generated by two independent double-wrapped coils placed inside two µ-metal boxes within a commercial incubator; four quadratic Helmholtz coil systems produced a homogenous magnetic field (static or oscillating) with perpendicular orientation to the horizontal plane of the Trikinetics monitors, each coil was formed with a pair of wires with the current passing in the same direction through both wires for EMF exposure but in opposite directions to provide a sham exposure condition
slices were exposed through coils in a system based on the Schuderer apparatus (µ-metal shielded chamber, see Schuderer et al., 2004) within an incubator at 37°C
分子生合成: bioluminescence of brain slices (photon multiplier tube assembly); stability/protein expression of cryptochrome in Drosophila wild type and gentically modified flies (Western blot)
認知的／行動学的影響評価項目: circadian locomotor activity and general locomotor activity in allDrosophila strains (automated system "Drosophila Trikinetics Monitor")
In wild type flies (CRY), exposure to magnetic fields (field 1-5) significantly shortened the circadian periods and significantly increased locomotor activity levels compared to sham exposure. Furthermore, the Western blot analysis showed that the magnetic field exposure significantly increased the stability of cryptochrome compared to the sham exposure. In genetically modified flies which did not express cryptochrome, no response to the magnetic field was observed. Further experiments with cryptochrome mutant flies showed that the tryptophan hypothesized to be essential to the electron transfer in the radical pair mechanism is not necessary for magnetic field responses. Transformants bearing an hCRY1 were not able to detect the magnetic fields. The mammalian cryptochrome (hCRY2) responded to the magnetic field when cloned into flies but not in its "mammalian environment" i.e. in the brain slices of mice. The authors suggest that cryptochromes act as blue-light/magnetic field sensors depending on factors that are present in particular cellular environments.
