Gravitaxis in the flagellate Euglena gracilis--results from NiZeMi, clinostat and sounding rocket flights

Many motile microorganisms including flagellates such as the green Euglena gracilis move up and down within the water column and use a number of external clues for their orientation, the most important of which may be light and gravity. The cells use positive phototaxis and negative gravitaxis to move closer to the surface of the water column which for energetic reasons is vital for their survival. However, most phytoplankton organisms cannot tolerate the bright irradiance of unfiltered solar radiation at the surface which also bleaches the photosynthetic pigments, disables the photosynthetic apparatus and impairs phototaxis, gravitaxis and motility in Euglena. Thus, it is not surprising that at higher irradiances negative phototaxis operates antagonistically to the responses described above to guide the cells into deeper water where they are protected from excessive radiation. Phototaxis and gravitaxis are not independent from one another: in a vertically positioned cuvette negative gravitaxis can be "titrated" by light impinging from above and is compensated at about 30 W m-2. While the photoreceptor for phototaxis has been identified in Euglena gracilis biochemically and spectroscopically, the gravireceptor is not yet known. Young cultures of Euglena gracilis show a positive gravitaxis, the ecological signficance of which is not yet understood while older cultures show negative gravitaxis. One hypothesis concerning the nature of graviperception is based on a passive physical process such as an asymmetric distribution of the mass within the cell. However, the observation that short term UV irradiation decreases the precision of negative gravitaxis rather indicates the involvement of an active physiological gravireceptor. Furthermore, some heavy metal ions have been found to change the direction of movement from positive to negative gravitaxis in young cells.     

Rhodopsin Receptors of Phototaxis in Green Flagellate Algae

Green flagellate algae are capable of the active adjustment of their swimming path according to the light direction (phototaxis). This direction is detected by a special photoreceptor apparatus consisting of the photoreceptor membrane and eyespot. Receptor photoexcitation in green flagellates triggers a cascade of rapid electrical events in the cell membrane which plays a crucial role in the signal transduction chain of phototaxis and the photophobic response. The photoreceptor current is the earliest so far detectable process in this cascade. Measurement of the photoreceptor current is at present the most suitable approach to investigation of the photoreceptor pigment in green flagellate algae, since a low receptor concentration in the cell makes application of optical and biochemical methods so far impossible. A set of physiological evidences shows that the phototaxis receptor in green flagellate algae is a unique rhodopsin-type protein. It shares common chromophore properties with retinal proteins from archaea. However, the involvement of photoelectric processes in the signal transduction chain relates it to animal visual rhodopsins. The presence of some enzymatic components of the animal visual cascade in isolated eyespot preparations might also point to this relation. A retinal-binding protein has been identified in such preparations, the amino acid sequence of which shows a certain homology to sequences of animal visual rhodopsins. However, potential function of this protein as the phototaxis receptor has been questioned in recent time.     

Behavior of red king crab larvae: Phototaxis,geotaxis and rheotaxis

Zoeae of Paralithodes camtschatica were positively phototactic to white light intensities above 1 × 10¹³ q cm^?2 s^?1. Negative phototaxis occurred at low (1 × 10¹² q cm^?2 s^?1), but not high intensities (2.2 × 10¹⁶q cm^?2 s^?1). Phototactic response was directly related to light intensity. Zoeae also responded to red, green and blue light. Zoeae were negatively geotactic, but geotaxis was dominated by phototaxis. Horizontal swimming speed of stage 1 zoeae <4 d old was 2.4 ± 0.1 (SE) cms^?1 and decreased to 1.7 ± 0.1 cm s^?1 in older zoeae (P <0.01). Horizontal swimming speed of stage 2 zoeae was not significantly different from ≥4 d old stage 1 zoeae. Vertical swimming speed, 1.6 ± 0.1 cm s^?1, and sinking rate, 0.7 ± 0.1 cm s^?1, did not change with ontogeny. King crab zoeae were positively rheotactic and maintained position in horizontal currents less than 1.4 cm s^?1. Starvation reduced swimming and sinking rates and phototactic response.     

Evolution of phototaxis

Phototaxis in the broadest sense means positive or negative displacement along a light gradient or vector. Prokaryotes most often use a biased random walk strategy, employing type I sensory rhodopsin photoreceptors and two-component signalling to regulate flagellar reversal. This strategy only allows phototaxis along steep light gradients, as found in microbial mats or sediments. Some filamentous cyanobacteria evolved the ability to steer towards a light vector. Even these cyanobacteria, however, can only navigate in two dimensions, gliding on a surface. In contrast, eukaryotes evolved the capacity to follow a light vector in three dimensions in open water. This strategy requires a polarized organism with a stable form, helical swimming with cilia and a shading or focusing body adjacent to a light sensor to allow for discrimination of light direction. Such arrangement and the ability of three-dimensional phototactic navigation evolved at least eight times independently in eukaryotes. The origin of three-dimensional phototaxis often followed a transition from a benthic to a pelagic lifestyle and the acquisition of chloroplasts either via primary or secondary endosymbiosis. Based on our understanding of the mechanism of phototaxis in single-celled eukaryotes and animal larvae, it is possible to define a series of elementary evolutionary steps, each of potential selective advantage, which can lead to pelagic phototactic navigation. We can conclude that it is relatively easy to evolve phototaxis once cell polarity, ciliary swimming and a stable cell shape are present.     

Action spectra for phototaxis in zoospores of the brown algaPseudochorda gracilis

Summary Action spectra for phototaxis in zoospores of brown alga,Pseudochorda gracilis (Laminariales), were examined in the wavelength range between 300 and 600 nm using the Okazaki Large Spectrograph and a video tracking system. The direction of swimming (both in percent cells swimming in parallel with the stimulating light, and in mean angle of cell movement) was dependent on the wavelength. The action spectra had two peaks at 420 and 460 nm, while light above 500 nm was not effective in changing the swimming direction of the cells.Abbreviations TCMA tracker-cell movement analyzer system - CMA cell movement analyzer program     

Automated Recording of Vertical Negative Phototactic Behaviour in Daphnia magna Straus (Crustacea)

Diurnal vertical migration is a well-known phenomenon in the circadian activity rhythms of zooplankton. Our goal was to test whether negative phototaxis in Daphnia magna clone BEAK (provoked by artificially induced light stress, alternating light and dark phases in 2 h intervals), and its interference with the endogenous rhythm of diurnal vertical migration, can be automatically registered with a biomonitor. For the first time the vertical swimming behaviour of D. magna was recorded quantitatively based on non-optical data recording in a fully automated biotest system, the Multispecies Freshwater Biomonitor in a new experimental setup consisting of a column of three recording units (3-level chambers). Circadian vertical migration was clearly recorded in the 3-level chambers and the rhythm was more clear with 5 than with 1 organism per chamber. The organisms clearly responded to induced light stress with negative phototaxis, however best in larger chambers. The artificially induced rhythm was influenced by the endogenous rhythm. This approach may facilitate long-term observations of vertical swimming activity of zooplankton in the future.     

New evidence for the mechanism of phototactic orientation ofEuglena gracilis

When rotated horizontally in a cuvette in a strong lateral light beam, the flagellateEuglena gracilis effectively corrects its course and shows negative phototaxis, provided the angular velocity does not exceed 20⁰s^–1. Faster rotations cannot be corrected effeciently. In two strong light beams of equal illuminance perpendicular to each other, the cells move along the resultant away from the light beams. Decreasing the illuminance of one beam causes increasing numbers of the organisms to orient with respect to the stronger light source. In two perpendicular low illuminance beams (>200lx), the population splits into two components moving towards either light source. The percentage of cells in each component depends on the relative illuminances. The results can be explained by the shading hypothesis combined with a dichroic orientation of the photoreceptor molecules perpendicular to the long axis of the cells. Externally applied electric dc fields have no effect on positive or negative phototaxis; this supports the hypothesis that electrical potential changes are not involved in the sensory transduction chain of photoorientation inEuglena.     

Behavior of red king crab larvae: phototaxis, geotaxis and rheotaxis

Zoeae of Paralithodes camtschatica were positively phototactic to white light intensities above 1 x 10(13) q cm-2 s-1. Negative phototaxis occurred at low (1 x 10(12) q cm-2 s-1), but not high intensities (2.2 x 10(16) q cm-2 s-1). Phototactic response was directly related to light intensity. Zoeae also responded to red, green and blue light. Zoeae were negatively geotactic, but geotaxis was dominated by phototaxis. Horizontal swimming speed of stage 1 zoeae < 4 d old was 2.4 +/- 0.1 (SE) cm s-1 and decreased to 1.7 +/- 0.1 cm s-1 in older zoeae (P < 0.01). Horizontal swimming speed of stage 2 zoeae was not significantly different from > or = 4 d old stage 1 zoeae. Vertical swimming speed, 1.6 +/- 0.1 cm s-1, and sinking rate, 0.7 +/- 0.1 cm s-1, did not change with ontogeny. King crab zoeae were positively rheotactic and maintained position in horizontal currents less than 1.4 cm s-1. Starvation reduced swimming and sinking rates and phototactic response.     

The involvement of a protein kinase in phototaxis and gravitaxis of <Emphasis Type="Italic">Euglena gracilis</Emphasis>

The unicellular flagellate Euglena gracilis shows positive phototaxis at low-light intensities (<10 W/m²) and a negative one at higher irradiances (>10 W/m²). Phototaxis is based on blue light-activated adenylyl cyclases, which produce cAMP upon irradiation. In the absence of light the cells swim upward in the water column (negative gravitaxis). The results of sounding rocket campaigns and of a large number of ground experiments led to the following model of signal perception and transduction in gravitaxis of E. gracilis: The body of the cell is heavier than the surrounding medium, sediments and thereby exerts a force onto the lower membrane. Upon deviation from a vertical swimming path mechano-sensitive ion channels are activated. Calcium is gated inwards which leads to an increase in the intracellular calcium concentration and causes a change of the membrane potential. After influx, calcium activates one of several calmodulins found in Euglena, which in turn activates an adenylyl cyclase (different from the one involved in phototaxis) to produce cAMP from ATP. One further element in the sensory transduction chain of both phototaxis and gravitaxis is a specific protein kinase A. We found five different protein kinases A in E. gracilis. The blockage of only one of these (PK.4, accession No. EU935859) by means of RNAi inhibited both phototaxis and gravitaxis, while inhibition of the other four affected neither phototaxis nor gravitaxis. It is assumed that cAMP directly activates this protein kinase A which may in turn phosphorylate a protein involved in the flagellar beating mechanism.     

Simulation of phototaxis in the flagellateEuglena gracilis

A three-dimensional model of the flagellateEuglena gracilis was developed to simulate phototaxis and movement in space. The simulation of the phototactic behavior was compared with thein vivo behavior in order to determine the mechanism of orientation with respect to light. Phototactic behavior with respect to one light source, can be explained by the shading hypothesis as well as by a dichroic orientation of the absorbing vectors of the photoreceptor pigments. In contrast, the behavior of the cells when exposed to two perpendicular light beams is not compatible with the shading hypothesis. Likewise, the phototactic orientation of stigmaless cells cannot be accounted for on the basis of the shading hypothesis. In contrast, simulations andin vivo observations of the behavior under polarized light strongly indicate the validity of the dichroic orientation of the photoreceptor pigments.     

The effect of temperature on phototaxis and geotaxis by larvae of the crab Rhithropanopeus harrisi (Gould)

Investigations of the effect of sudden temperature change on the phototaxis of Stage I and IV zoeae upon stimulation from horizontal and vertical directions with 500-nm light indicate a temperature-induced geotactic response in larvae of the crab Rhithropanopeus harrisi (Gould). For the horizontal tests both zoea stages were reared at 20 °C. Stage I showed positive phototaxis at temperatures between 15 ° and 35 °C, while Stage IV responded over the range of 10–30 °C. For the vertical tests, larvae, reared at 25 °C, were stimulated with overhead lights. Stage I zoeae ascended at 15 °, 20 ° and 25 °C and descended at 5 °, 10 °, 30 ° and 35 °C. Stage IV zoeae ascended at 20 ° and 25 °C and descended at 5 °, 10 °, 15 °, 30 ° and 35 °C. Although the descent at high temperatures could result from a negative phototaxis, a reversal in phototactic sign at high temperatures was not found in the horizontal experiments and the same vertical movement pattern is observed in total darkness. Upon exposure to high temperatures near the water surface, larvae would descend by means of a positive geotaxis rather than a negative phototaxis. This response involves active swimming by Stage IV larvae and passive sinking by Stage I.     

Photoresponses of Chaoborus larvae

The diel vertical migration of Chaoborus larvae is a well known phenomenon. In order to quantify the ability of larvae to utilize underwater light cues in their migration, we measured photoresponses of fourth-instar Chaoborus punctipennis larvae in the laboratory. The action spectrum for these larvae was characterized by a maximum in sensitivity at 400 nm, a plateau at a lower sensitivity from 480 to 560 nm, and a region of much lower sensitivity at wavelengths longer than 620 nm. Dark-adapted larvae exhibited a positive phototaxis at low light intensity which shifted to a negative phototaxix as light intensity increased. At 540 nm the threshold intensity was 1.5 × 10^?9 W/m² for positive phototaxis and about 10^?6 W/m² for negative phototaxis. Light adaptation decreased sensitivity and altered the phototactic pattern. Larvae have a clear circadian rhythm in negative phototaxis, in which greatest responsiveness occurs early in the day. We suggest that the rhythm in photoresponsiveness primarily controls the timing of the downward migration at dawn.     

Effects of gravity on positive phototaxis in fruit fly Drosophila melanogaster

Geotaxis and phototaxis are movements in response to gravity and light, respectively, and are commonly observed in nature. The interactions between these two types of movement have been shown to confer ecological advantages to many taxa. Although several studies have been conducted on phototaxis and geotaxis in various organisms, reports on the interactions between positive phototaxis and negative geotaxis are lacking. In the fruit fly, Drosophila melanogaster, any direct interactions that exist between positive phototaxis and negative geotaxis are yet to be determined and the ecological significance of such interactions remains unclear. In the present study, the effects of gravity on positive phototaxis in a Y‐maze were investigated using the Canton‐S wild type and gravity‐sensing‐deficient pyx³ mutant fruit flies. Gravity sensing was not necessary for horizontal positive phototaxis, but was required for vertical positive phototaxis. These results suggest that gravitoreception may selectively modulate positive phototaxis depending on the vertical and horizontal movements of the fruit flies.     

Cone photoreceptor mechanisms and the detection of polarized light in fish

Summary Although numerous studies have demonstrated the detection of polarized light in vertebrates, little is known of the photoreceptor mechanisms involved. Recent evidence, however, indicates that cyprinid fishes possess both ultraviolet (UV) and polarization sensitivity suggesting that some vertebrates, like many invertebrates, may employ UV-sensitive cone receptors in polarization sensitivity. In this report, we describe experiments that determine which spectral types of receptors participate in the detection of polarized light. We used a heart-rate conditioning technique to measure increment thresholds of immobilized goldfish for plane-polarized, narrow-band (10 nm half max.) spectral stimuli (380 nm, 460 nm, 540 nm, 660 nm). A typical experiment involved isolating the activity of a cone photoreceptor mechanism by chromatic adaptation and measuring increment thresholds for spectral stimuli at e-vector orientations of the polarizer between 0° to 180° in 30° steps. The UV-, green- and red-sensitive cone receptor mechanisms showed clear evidence of polarization sensitivity while the blue-sensitive cone receptor mechanism was polarizationally insensitive. The average amplitude (base to peak height on Fig. 4) of the polarization sensitivity curves (UV-, green- and red-curves) was 0.67 log unit (standard deviation of 0.12 log unit), with the UV-sensitive cone receptor mechanism most sensitive to the vertical e-vector axis and the green- and red-sensitive cone receptor mechanisms most sensitive to the horizontal e-vector axis. The observation that different cone photoreceptor mechanisms have orthogonal polarization sensitivity in fish suggests that the perception of polarized light may enhance the capacity for visual discrimination in lower vertebrates.     

Phototaxis, gravitaxis and vertical migrations in the marine dinoflagellate Prorocentrum micans

Abstract The phototactic orientation of the marine dinoflagellate Prorocentrum micans was studied at three different ages and at several light intensities. High irradiances caused the cells to show negative phototaxis and low irradiances caused positive phototaxis. The precision of negative phototaxis reached a maximum in the early afternoon, while the precision of positive phototaxis was found to peak in the morning and at night. The cells also showed a pronounced negative gravitactic orientation, which had a maximum in precision in the early afternoon. The degree of gravitaxis was found to be constant over time when the cells were confined to a closed cuvette for up to 9 h. As a consequence of the orientation strategies, populations of Prorocentrum micans showed daily vertical migrations in a 3-m Plexiglas column. They accumulated in the top layers in the afternoon and were almost randomly distributed during the rest of the day.     

Growth of dinoflagellates, Ceratium furca and Ceratium fusus in Sagami Bay, Japan: The role of vertical migration and cell division

To better understand the mechanism underlying the bloom outbreaks of dinoflagellates, Ceratium furca, and Ceratium fusus in the temperate coastal area of Sagami Bay, we investigated the diel changes of vertical migration, swimming speed, cell volume, and cell division. Our results from both the field and laboratory indicate that C. furca and C. fusus can migrate vertically between surface and sub-surface layers to avoid strong sunlight (>1000 μmol m⁻² s⁻¹). Diel vertical migration (DVM) of C. furca was observed in the laboratory, while that of C. fusus was not observed. C. furca demonstrated a constant DVM rhythm, i.e., their cells began to descend from the surface before the light was extinguished, and ascended into the surface before the light was turned on. The downward and upward migrations of the cells occurred at every 3 h before turning on and off the light, suggesting that the DVM pattern was independent of nutrient concentration. The swimming speeds of C. furca (avg. 250 μm s⁻¹) were always faster than those of C. fusus (avg. 75 μm s⁻¹). In addition, the speeds of C. furca during light periods were faster than those during dark periods, whereas the speeds of C. fusus remained relatively constant. A higher proportion of dividing cells was recorded near dawn (05:00–07:00 h). Cell volumes of C. furca and C. fusus did not markedly change between 12:00 and 21:00 h, but gradually increased until 03:00 h and then sharply decreased. Furthermore, the cell volume of the two Ceratium species was significantly shifted to the temporal pattern of cell division. Combined with the DVM manner of two Ceratium and cell division timing, only C. furca divided at the bottom, and then moved toward the surface shortly before the dark to light transition. Based on our observations, C. furca has an ecological advantage due to their DVM activity, since nutrients can be obtained well in the near bottom layers, while during the daytime, light present in nutrient-depleted surface water can be obtained using their high swimming speed. On the other hand, C. fusus stimulated by low salinity conditions, might be dependent on external environmental conditions such as additional nutrients following freshwater discharge by heavy rainfall because they may not perform active DVM due to a slow swimming ability. Our findings support that specific characteristics, including the DVM behavior in C. furca, yield a competitive advantage over C. fusus in Sagami Bay.     

Characterization of Halobacterium halobium mutants defective in taxis.

Mutant derivatives of Halobacterium halobium previously isolated by using a procedure that selected for defective phototactic response to white light were examined for an array of phenotypic characteristics related to phototaxis and chemotaxis. The properties tested were unstimulated swimming behavior, behaviorial responses to temporal gradients of light and spatial gradients of chemoattractants, content of photoreceptor pigments, methylation of methyl-accepting taxis proteins, and transient increases in rate of release of volatile methyl groups induced by tactic stimulation. Several distinct phenotypes were identified, corresponding to a mutant missing photoreceptors, a mutant defective in the methyltransferase, a mutant altered in control of the methylesterase, and mutants apparently defective in intracellular signaling. All except the photoreceptor mutant were defective in both chemotaxis and phototaxis.     

Negative Phototaxis in Blepharisma japonicum

The protozoan Blepharisma japonicum showed negative phototaxis caused by transient reversal of the direction of ciliary beat and changes of swimming velocity induced with varying intensities of light. The ciliary reversal occurred at 1–2 sec after a sudden increase in light intensity. When light intensity was decreased, no response was observed. Moreover, the ciliates swam fast in light areas but slowly in dark areas; the mean velocity of swimming was 80 μ m/sec at 5 × 10² lux but reached about 400 μMm/sec at 5 × 10³ lux. In addition, the cell body elongated in response to light application; the mean length of the body was 308 μm at 5 × 10² lux, which increased to 397 μ m at 10⁴ lux. Such body elongation seems to contribute to rapid swimming. Negative phototaxis may be an important behavior in B. japonicum because the organisms are killed by exposure to strong light.     

Phototaxis in the flagellates,Euglena gracilis and Ochromonas danica

Oriented movement with respect to laterally impinging white light of the flagellates Euglena gracilis and Ochromonas danica has been analyzed in an individual cell study with a microvideographic technique. Using the deviation of track segments (in given time intervals of 1 s) from the light direction as raw data allowed a computer based analysis of the direction distribution. A number of statistical methods employed to test the significance of the obtained results demonstrated an obvious phototactic orientation in Ochromonas which was positive (toward the light source) in low illuminance (1.25 lx=5.3×10^-3 Wm^-2) and negative in higher illuminance (>12.5 lx=5.3×10^-2 Wm^-2). Since in this flagellate the threshold for negative phototaxis is much lower than that for the step-up photophobic response, the hypothesis that negative phototaxis may be brought about by repetitive step-up phobic responses can be rejected for at least this organism. In Euglena positive phototaxis was observed in 50 lx (=0.21 Wm^-2), while an illuminance of 500 lx (=2.1 Wm^-2) caused a negative phototaxis.The experiments were carried out in this laboratory     

Negative Phototaxis from Blue Light and the Role of Third Rhodopsinlike Pigment in Halobacterium Cutirubrum

Wild-type cells of Halobacterium cutirubrum show phototaxis. In negative phototaxis the cells are repelled by blue-near ultraviolet light, and in positive phototaxis the cells are attracted to green-red light. The extent of the responses are measured by monitoring the changes in the reversal frequency of the swimming direction of cells using a computer-linked automated method as described previously (Takahashi, T., and Y. Kobatake, 1982, Cell. Struct. Funct., 7:183-192). When the intensity of the background light (illumination for the observation) was dramatically reduced, the sensitivity of the cells to the repellent light decreased markedly. This result has been previously explained by Bogomolni and Spudich (1982, Proc. Natl. Acad. Sci. USA, 79:6250-6254), who proposed that the photoreceptor for negative phototaxis is the long-lifetime intermediate in the photocycle of slow-rhodospin. The behavioral response in the negative phototaxis is dependent upon the intensity of the actinic light and the background light. This agrees quantitatively with our model based on the aforementioned hypothesis.