Fig. 1. The photosynthetic unicellular flagellate Euglena gracilis.
While the light factor can be manipulated easily in the laboratory, the effects of gravity cannot be studied with the same ease because the terrestrial gravitational field cannot be switched off. This may be one of the reasons why our knowledge concerning the gravireceptor of flagellates is still rather limited. According to an earlier hypothesis, the orientation of the cells is due to a passive physical orientation in the water column, assuming that the rear end is heavier than the front end.¹ The alternative hypothesis is that the cell possesses an active physiological gravireceptor that determines the vector of the gravitational field.² In addition to gravity, flagellates use light as a cue for their orientation in the water column,³ as indicated above. Experiments using light and gravity in different vectorial combinations under terrestrial conditions have indicated that the cells integrate over the two stimuli and generate an integrated response.4 A similar problem exists for fungi and higher plants in which phototropism is superimposed by gravitropism under terrestrial conditions.5
The aims of the TEXUS experiments were, first, to demonstrate that gravity is indeed the factor that orients the movements of E. gracilis; second, to characterise the orientation under µg conditions; and, third, to study both positive and negative phototaxis during weightlessness.
The unicellular photosynthetic freshwater flagellate, Euglena gracilis Klebs, strain Z, was used for all experiments (Fig. 1). The cells were grown as described earlier in a mineral medium.6 The cells were harvested after two weeks of growth and transferred into a circular cuvette (0.2 mm depth and 55 mm diameter) made from stainless steel; it was kept at 22.5±1°C by four Peltier elements. The window facing the microscope light source was made from an infrared transmitting filter (RG 715, Schott & Gen., Mainz, Germany). The cuvette was oriented vertically in the experiment module (TEM 06- 19, developed and manufactured by MBB-ERNO in Bremen) while the rocket stood vertically before launch. The cuvette could be rotated around its short axis by remote control to redistribute the cells. The cuvette was constantly rotated (35 rpm) during launch in order to minimise the effects of acceleration. The actinic light beam to elicit phototaxis impinged at an angle of 15° with respect to the surface of the cuvette.
Fig. 1. The photosynthetic unicellular flagellate Euglena
gracilis.
The image of the moving cells was recorded by a CCD camera and transmitted to the ground for recording. During playback the video signal was digitised in real time at a spatial resolution of 512 x 512 pixels with 256 possible grey levels. The tracking software had been written in the computer language C with assembly language modules for the time-critical input and output routines as well as the mathematical analysis of the data.7, 8 The precision of orientation was determined using the Rayleigh test.
The first experiment on TEXUS 23 showed that the Earth's gravitational field is indeed responsible for the orientation of E. gracilis in the absence of other orienting factors, such as light. Under µg conditions the cells moved randomly, so orientation with respect to the magnetic field lines or thermal gradients could be ruled out (Figs. 2a-2b).9 Similar results were found on the fast rotating horizontal clinostat.10 Reorientation of the cells occurs within less than a minute. Under terrestrial conditions the cells are subjected to sedimentation as they have a specific weight of 1.04, higher than that of the surrounding water. In accordance with the Rayleigh law, this results in a faster downward swimming rate and a slower upward swimming rate compared to the velocity in a horizontal direction. Consequently, the cells move at a higher velocity under µg conditions. In contrast, the ciliate Paramecium partially compensates the slower upward movement by gravikinesis.11, 12
Fig. 2. Distribution of the swimming directions of E. gracilis
at 1 g before the TEXUS flight (a) and 0 g during the first
minute on TEXUS 23 (b).
On the TEXUS 28-30 flights, the phototactic orientation was investigated. The cells showed both positive and negative phototaxis under µg conditions.13 The fluence rates for these responses are comparable to those at 1 g. However, phototactic orientation was reached faster than at 1 g. Also, the precision of orientation was higher in space than in the control measured simultaneously under terrestrial conditions. Measurements of the velocity distributions confirmed earlier results during gravitaxis experiments in space. Measurements after the TEXUS flights showed a similar behaviour both for phototaxis and gravitaxis as before the flight, indicating that the cells did not adapt to the conditions at 0 g. Recent experiments during the Space Shuttle's International Microgravity Laboratory-2 mission using the slow rotating centrifuge microscope (NIZEMI) indicated that the threshold for gravitactic orientation in E. gracilis is at about 0.16 g.14 Also, there was no adaptation to the altered gravity under the prolonged conditions at 0 g.
This work was supported by financial support from the Deutsche Agentur für Luftund Raumfahrtangelegenheiten (DARA).
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