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Carbon-ion Microbeam Induces Behavioral Changes in
the Salt Chemotaxis Learning of C. elegans
T. Sakashita a) , M. Suzuki a) , Y. Mutou a) , Y. Yokota a) , T. Funayama a) , N. Hamada b) ,
K. Fukamoto c) and Y. Kobayashi a)
a) Radiation-Applied Biology Division, QuBS, JAEA,
b) Radiation Safety Research Center, Central Research Institute of Electric Power Industry,
c) Faculty of Textile Science and Technology, Shinshu University
Introduction
An increasing body of data indicates that ionizing
radiation affects the nervous system and alters its function 1) .
Chemotaxis of C. elegans during the salt chemotaxis
2)
learning was modulated by gamma irradiation . Our
preliminary results showed the similar response of the salt
chemotaxis learning to whole-body carbon-ion irradiation.
However, we have no direct evidence for the interaction of
ionizing radiation with the central neuronal tissue (nerve
ring) in C. elegans. Microbeam irradiation is useful to
analyze direct radiation effects at a cellular or tissue level.
Thus, we investigate the effects of energetic carbon ions
( 12 C 5+ , 18.3 MeV/u, LET = 119 keV/µm) on the salt
chemotaxis learning of C. elegans using microbeam
irradiation to its nerve ring and also combined effects with
anesthesia that inhibits nerve function.
Materials and Methods
Well-fed adults of C. elegans grown at 20 ˚C on the plate
spread with E. coli OP50 were used in all experiments.
The area of nerve ring, “Head”, of non-anesthetized C.
elegans with S-basal (100 mM NaCl, 50 mM KPO4 pH 6.0,
and 5 mg/L cholesterol) in a ditch of the micro total analysis
systems (µTAS) 3) was irradiated with 12,000 carbon ions
corresponding to 500 Gy at 20 µmφ micro-aperture area
(Fig. 1). Immediately after irradiation, chemotaxis to NaCl
during learning was measured. The TaKaRa Slide Seal
(Fig. 2; TKR 9066, TaKaRa Bio Inc., Shiga, Japan) was
4)
used for the microbeam irradiation of anesthetized animals .
Irradiation effects during learning could not be evaluated in
anesthetized animals, so that the ability of the salt
chemotaxis learning was tested. Using the assay plate with
a gradient of NaCl concentration, we evaluated chemotaxis
based on the chemotaxis index (CI) that was calculated as
{(number of animals at the high-concentration spot) –
(number of animals at the control spot)}/ (total number of
animals in the assay plate) 5) . The results of CI were
evaluated as mean ± 95% confidential interval.
Results and Discussion
To investigate direct radiation effects, non-anesthetized
animals was irradiated with carbon-ion microbeam, and
showed the modulation of the chemotaxis to NaCl during
learning (CI: 0.26 ± 0.12 in non-irradiated animals, -0.13 ±
0.09 in “Head” locally irradiated animals). It demonstrates
the direct evidence by the hit of carbon ions at the nerve ring.
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Also, to test the effects of carbon ion-induced damage in
Head area, we evaluated defects in the salt chemotaxis
learning of anesthetized animals. The ability of the salt
chemotaxis learning was not affected by carbon ions (CI:
0.33 ± 0.07 in non-irradiated animals, 0.32 ± 0.10 in “Head”
locally irradiated animals). Our findings indicate that the
modulation of the salt chemotaxis learning is induced by the
direct hit of ionizing radiation at the nerve ring of C. elegans,
and the salt chemotaxis learning in anesthetized C. elegans
is not affected by microbeam irradiation.
References
1) T. Sakashita et al., J. Radiat. Res. 51 (2010) 107.
2) T. Sakashita et al., FASEB J. 22 (2008) 713.
3) Lockery et al., J. Neurophysiol. 99 (2008) 3136.
4) Sugimoto et al., Int. J. Radiat. Biol. 82 (2006) 31.
5) Saeki et al., J. Exp. Biol., 204 (2001) 1757.
Head
60 µm
200 µm
Fig. 1 Non-anesthetized C. elegans in a ditch of µTAS.
Circle shows the area of nerve ring, “Head”.
1 cm
Fig. 2 Image of TaKaRa Slide Seal which is a gray frame,
and 25 µL buffer with anesthetized C. elegans is
instilled into this area.