Brigitte Buchen

Evaluation of the three-dimensional clinostat as a simulator of weightlessness

Abstract. Concerns regarding the reliability of slow- and fast-rotating uni-axial clinostats in simulating weightlessness have induced the construction of devices considered to simulate weightlessness more adequately. A new three-dimensional (3-D) clinostat equipped with two rotation axes placed at right angles has been constructed. In the clinostat, the rotation achieved with two motors is computer-controlled and monitored with encoders attached to the motors. By rotating plants three-dimensionally at random rates on the clinostat, their dynamic stimulation by gravity in every direction can be eliminated. Some of the vegetative growth phases of plants dependent on the gravity vector, such as morphogenesis, are shown to be influenced by rotation on the 3-D clinostat. The validity of 3-D clinostatting has been evaluated by comparing structural parameters of cress roots and Chara rhizoids obtained under real microgravity with those obtained after 3-D clinostatting. The parameters analyzed up to now (organization of the root cap, integrity and polarity of statocytes, dislocation of statoliths, amount of starch and ER) demonstrate that the 3-D clinostat is a valuable device for simulating weightlessness.

Oriented movement of statoliths studied in a reduced gravitational field during parabolic flights of rockets.

During five rocket flights (TEXUS 18, 19, 21, 23 and 25), experiments were performed to investigate the behaviour of statoliths in rhizoids of the green alga Chara globularia Thuill. and in statocytes of cress (Lepidium sativum L.) roots, when the gravitational field changed to approx. 10−4 · g (i.e. microgravity) during the parabolic flight (lasting for 301–390 s) of the rockets. The position of statoliths was only slightly influenced by the conditions during launch, e.g. vibration, acceleration and rotation of the rocket. Within approx. 6 min of microgravity conditions the shape of the statolith complex in the rhizoids changed from a transversely oriented lens into a longitudinally oriented spindle. The center of the statolith complex moved approx. 14 μm and 3.6 μm in rhizoids and root statocytes, respectively, in the opposite direction to the originally acting gravity vector. The kinetics of statolith displacement in rhizoids demonstrate that the velocity was nearly constant under microgravity whereas it decreased remarkably after inversion of rhizoids on Earth. It can be concluded that on Earth the position of statoliths in both rhizoids and root statocytes depends on the balance of two forces, i.e. the gravitational force and the counteracting force mediated by microfilaments.

Statoliths pull on microfilaments: experiments under microgravity.

SummaryPrevious videomicroscopy ofChara rhizoids during parabolic flights of rockets showed that the weightless statoliths moved basipetally. A hypothesis was offered that the removal of gravity force disturbed the initial balance between this force and the basipetally acting forces generated in a dynamic interaction of statoliths with microfilaments (MFs). The prediction of this hypothesis that the statoliths would not be displaced basipetally during the microgravity phase (MG-phase) after disorganizing the MFs was tested by videomicroscopy of a rhizoid treated with cytochalasin D (CD) immediately before the flight. The prediction was fully supported by the flight experiment. Additionally, by chemical fixation of many rhizoids at the end of the MG-phase it was shown that all rhizoids treated with CD before the flight had statoliths at the same location, i.e., sedimented on the apical cell wall, while all untreated rhizoids had statoliths considerably displaced basipetally from their normal position. Thus, a dynamical interaction involving shearing forces between MFs and statoliths appears highly probable.

Sporogenesis and Pollen Grain Formation

Pollen grains and the spores of nonseed vascular plants1 develop from diploid somatic cells in a largely identical way. Ultimately, the cells are characterized by a highly ordered cell wall, the so-called sporoderm (Figs. 1 and 2). Both the chemical and the principal ultrastructural features of sporoderms show little Variation in different taxa, although the sculpture, number, and thickness of the individual wall layers display species-specificity. Fig. 3 shows a schematic cross section of a sporoderm and the surface view of a tectum, using in part the terminology proposed by Erdtman (1952).

Possible use of a 3-D clinostat to analyze plant growth processes under microgravity conditions.

A three-dimensional (3-D) clinostat equipped with two rotation axes placed at right angles was constructed, and various growth processes of higher plants grown on this clinostat were compared with ground controls, with plants grown on the conventional horizontal clinostat, and with those under real microgravity in space. On the 3-D clinostat, cress roots developed a normal root cap and the statocytes showed the typical polar organization except a random distribution of statoliths. The structural features of clinostatted statocytes were fundamentally similar to those observed under real microgravity. The graviresponse of cress roots grown on the 3-D clinostat was the same as the control roots. On the 3-D clinostat, shoots and roots exhibited a spontaneous curvature as well as an altered growth direction. Such an automorphogenesis was sometimes exaggerated when plants were subjected to the horizontal rotation, whereas the curvature was suppressed on the vertical rotation. These discrepancies in curvature between the 3-D clinostat and the conventional ones appear to be brought about by the centrifugal force produced. Thus, the 3-D clinostat was proven as a useful device to simulate microgravity.

Cytoplasmic streaming in Chara rhizoids: studies in a reduced gravitational field during parabolic flights of rockets.

SummaryIn-vivo videomicroscopy ofChara rhizoids under 10–4g demonstrated that gravity affected the velocities of cytoplasmic streaming. Both, the acropetal and basipetal streaming velocities increased on the change to microgravity. The endogenous difference in the velocities of the oppositely directed cytoplasmic streams was maintained under microgravity, yet the difference was diminished as the basipetal streaming velocity increased more than the acropetal streaming velocity. Direction and structure of microfilaments labeled by rhodamine-phalloidin had not changed after 6 min of microgravity.

The endogenous difference in the rates of acropetal and basipetal cytoplasmic streaming inChara rhizoids is enhanced by gravity

SummaryMeasurements of cytoplasmic streaming inChara rhizoids were made by a streak-photography method combined with dark-field illumination. The velocity of cytoplasmic streaming in the acropetal direction was faster than in the basipetal direction. The difference in the streaming velocities in both morphological directions was apparently due to endogenous forces. In addition to this, a small difference attributable to gravity was superimposed if the rhizoid was oriented parallel to the gravity vector. The difference in the endogenous forces underlying the oppositely directed streams may be as high as about 12-fold the force imposed by gravity but, on average, it is about 5-fold the gravity force. In the normal vertical position of the rhizoid, this endogenously generated difference is enhanced by the gravity effect. In contrast to the variability of streaming rate due to endogenous forces, the effect of the gravity force is relatively uniform. The difference between acropetal and basipetal streaming velocities is perpetuated over the whole range of lowered velocities after treatment with cytochalasin B. After prolonged incubation in cytochalasin B, the basipetal streaming stops earlier than the acropetal streaming. A difference in the length of filaments on both sides of the streaming machinery in rhizoids is proposed as the structural basis for the difference in velocities.

Über den Feinbau der wachsenden Megaspore vonSelaginella

Im Gegensatz zu den ErgebnissenFittings (1900) ist das Cytoplasma derSelaginella- Megaspore nicht vom wachsenden Sporoderm getrennt. DieSelaginella-Megaspore stellt demnach kein Beispiel fur Zellwandwachstum ohne Kontakt mit dem Plasmalemma dar. Die Elemente der Exine anastomosieren. Die Sexine hat eine schwammartige Struktur und eine warzige und kurzstachelige Skulptur. Abgesehen vom proximalen Pol, wo zwischen Sexine und Nexine eine solide Sporopolleninschicht liegt, entsteht im inneren Teil der Sexine in einem fruhen Entwicklungsstadium ein breiter, artifizieller Spalt. Die Nexine besteht auf 7–13 Lamellen. Ihre dunnsten Stellen haben eine Dimension von 13 nm. Vor allem zwischen den Lamellen der Nexine liegt eine fibrillar-netzformige Matrix mit verschieden grosen, dunkel kontrastierten Granula.SummaryIn contrary to the results ofFitting (1900) the cytoplasm of theSelaginella- megaspore is not separated from the growing sporoderm. Therefore theSelaginella- megaspore is not an example for cell wall growth without contact with the plasmamembrane. The elements of the exine anastomose. The sexine has a spongy structure and a verrucate and spinulous sculpture. Apart from the proximal pole, where a solid layer of sporopollenin lies between sexine and nexine, in the inner part of the sexine in an early developmental stage a broad artificial cleft originates. The nexine consists of 7–13 lamellae. Their thinnest parts show a dimension of 13 nm. Mainly between the lamellae of the nexine a fibrillar-netlike matrix with darkly contrasted granula of different size is situated.ZusammenfassungIm Gegensatz zu den ErgebnissenFittings (1900) ist das Cytoplasma derSelaginella- Megaspore nicht vom wachsenden Sporoderm getrennt. DieSelaginella-Megaspore stellt demnach kein Beispiel für Zellwandwachstum ohne Kontakt mit dem Plasmalemma dar. Die Elemente der Exine anastomosieren. Die Sexine hat eine schwammartige Struktur und eine warzige und kurzstachelige Skulptur. Abgesehen vom proximalen Pol, wo zwischen Sexine und Nexine eine solide Sporopolleninschicht liegt, entsteht im inneren Teil der Sexine in einem frühen Entwicklungsstadium ein breiter, artifizieller Spalt. Die Nexine besteht auf 7–13 Lamellen. Ihre dünnsten Stellen haben eine Dimension von 13 nm. Vor allem zwischen den Lamellen der Nexine liegt eine fibrillär-netzförmige Matrix mit verschieden großen, dunkel kontrastierten Granula.

Polarity in mechanoreceptor cells of trigger hairs of Dionaea muscipula Ellis

Both the apical and the basal cell poles of the sensory cells in trigger hairs of Dionaea muscipula are structured identically. A complex of concentrically arranged endoplasmic reticulum cisternae occupies each of the poles. One to four vacuoles are enclosed within the central cisterna and contain polyphenols (deposits of “tannin”). Structural polarity, whether symmetric or asymmetric, as well as the occurrence of abundant endoplasmic reticulum and numerous mitochondria are characteristics of the perception cells of most animals and plants.

Structure of cress root statocytes in microgravity (Bion-10 mission)

Experiments on primary roots of Lepidium sativum L. have been performed on board the Bion-10 satellite. The experimental set-up was extremely miniaturized and completely automatic. The results demonstrate the effectiveness of the instrumentation. The spatial orientation, growth, root cap differentiation and statocyte structure of roots grown under microgravity (MG) have been compared with control roots grown on the ground (GC) and in a 1G-reference centrifuge in space (RC). Root length and cap shape did not differ between MG and control samples. Under MG, the mean distance of the statoliths from the distal cell wall of the statocytes increased significantly, the mean distance of the mitochondria decreased and the nucleus did not change its position in comparison to both controls. The number and the shape of the amyloplasts (statoliths) were not influenced by the space flight factors, but their size as well as their relative area in the cell decreased. The number of starch grains per statolith as well as their size and shape changed under MG. In MG and RC samples the number of lipid bodies in the statocytes was higher and the relative area larger than in GC samples. The relative area occupied by vacuoles was greater in RC statocytes than in GC and MG statocytes. These results partly confirm and, in addition, extend the data from earlier experiments in space.