Inner Ear Otolith Asymmetry in Late-Larval Cichlid Fish (Oreochromis mossambicus, Perciformes) Showing Kinetotic Behaviour Under Diminished Gravity.

The inner ears of all vertebrates are designed to perceive auditory and vestibular inputs. Although a tremendous diversity in the inner ear can be found even among bony fishes, the morphologies of the utricle and of the semicircular canals are rather conservative among vertebrates. Fish show kinetoses under reduced gravity (spinning movements and looping responses) and are regarded model organisms concerning the performance of the otolithic organs. Otoliths can be analysed easily because they are compact, in contrast to the otoconial masses of other vertebrates. Here, late-larval Oreochromis mossambicus were subjected to 0.0001 × g and 0.04 × g aboard a sounding rocket, their behaviour was observed and morphometrical analyses on otoliths were carried out. Fish swimming kinetotically at 0.0001 × g had a higher asymmetry of utricular otoliths (gravity perception) but not of saccular otoliths (hearing process) than specimens behaving normally at this gravity level (p = 0.0055). Also, asymmetries of lapilli in animals swimming normally at 0.0001 × g were lower than asymmetries in specimens swimming normally at 0.04 × g (p = 0.06). This supports the "otolith asymmetry hypothesis", an explanation for the susceptibility to kinetosis, particularly concerning the utricular otoliths. It would be interesting to identify processes generating asymmetric otoliths, also in regard to human motion sickness.

Formation of the inner ear during embryonic and larval development of the cichlid fish (Oreochromis mossambicus).

BACKGROUND: The vertebrate inner ear comprises mineralized elements, namely the otoliths (fishes) or the otoconia (mammals). These elements serve vestibular and auditory functions. The formation of otoconia and otoliths is described as a stepwise process, and in fish, it is generally divided into an aggregation of the otolith primordia from precursor particles and then a growth process that continues throughout life. RESULTS: This study was undertaken to investigate the complex transition between these two steps. Therefore, we investigated the developmental profiles of several inner ear structural and calcium-binding proteins during the complete embryonic and larval development of the cichlid fish Oreochromis mossambicus in parallel with the morphology of inner ear and especially otoliths. We show that the formation of otoliths is a highly regulated temporal and spatial process which takes place throughout embryonic and larval development. CONCLUSIONS: Based on our data we defined eight phases of otolith differentiation from the primordia to the mature otolith.

Not All Inner Ears are the Same: Otolith Matrix Proteins in the Inner Ear of Sub-Adult Cichlid Fish, Oreochromis Mossambicus, Reveal Insights Into the Biomineralization Process.

The fish ear stones (otoliths) consist mainly of calcium carbonate and have lower amounts of a proteinous matrix. This matrix consists of macromolecules, which directly control the biomineralization process. We analyzed the composition of this proteinous matrix by mass spectrometry in a shotgun approach. For this purpose, an enhanced protein purification technique was developed that excludes any potential contamination of proteins from body fluids. Using this method we identified eight proteins in the inner ear of Oreochromis mossambicus. These include the common otolith matrix proteins (OMP-1, otolin-1, neuroserpin, SPARC and otoconin), and three proteins (alpha tectorin, otogelin and transferrin) not previously localized to the otoliths. Moreover, we were able to exclude the occurrence of two matrix proteins (starmaker and pre-cerebellin-like protein) known from other fish species. In further analyses, we show that the absence of the OMP starmaker corresponds to calcitic otoliths and that pre-cerebellin-like protein is not present at any stage during the development of the otoliths of the inner ear. This study shows O. mossambicus does not have all of the known otolith proteins indicating that the matrix proteins in the inner ear of fish are not the same across species. Further functional studies of the novel proteins we identified during otolith development are required.

Spatial Expression of Otolith Matrix Protein-1 and Otolin-1 in Normally and Kinetotically Swimming Fish.

Kinetosis (motion sickness) has been repeatedly shown to affect some fish of a given clutch following the transition from 1g to microgravity or from hypergravity to 1g. This susceptibility to kinetosis may be correlated with irregular inner ear otolith growth. Otoliths are mainly composed of calcium carbonate and matrix proteins, which play an important role in the process of otolith mineralization. Here, we examine the morphology of otoliths and the expression pattern of the major otolith proteins OMP-1 and otolin-1 in a series of hypergravity experiments. In the utricle, OMP-1 is present in centripetal (medial) and centrifugal (lateral) regions of the meshwork area. In the saccule, OMP-1 was expressed within a dorsal and a ventral narrow band of the meshwork area opposite to the periphery of the sulcus acusticus. In normal animals, the spatial expression pattern of OMP-1 reaches more posteriorly in the centrifugal aspect and is considerably broader in the centripetal portion of the utricle compared to kinetotic animals. However, otolin-1 was not expressed in the utricule. In the saccule, no differences were observed for either gene when comparing normal and kinetotically behaving fish. The difference in the utricular OMP-1 expression pattern between normally and kinetotically swimming fish indicates a different otolith morphology and thus a different geometry of the otoliths resting on the corresponding sensory maculae. As the utricle is the endorgan responsible for sensing gravity, the aberrant morphology of the utricular otoliths, based on OMP-1 expression, likely leads to the observed kinetotic behavior.

Expression of SPARC and the osteopontin-like protein during skeletal development in the cichlid fish Oreochromis mossambicus.

BACKGROUND: Bones are mainly composed of calcium hydroxyapatite and a proteinous matrix. In this study, we focus on the bone matrix proteins, the fish osteopontin orthologous protein (osteopontin-like protein; OP-L) and SPARC, because the current knowledge regarding their expression is fragmentary or contradictory. RESULTS: We first provide a comprehensive and detailed description of skeletal development in the cichlid fish Oreochromis mossambicus. Following this, we analyzed the expression pattern of OP-L and SPARC in detail during development. OP-L expression was only found in tissues that undergo ossification (i.e., developing bones and teeth). Furthermore, we show that there is a fundamental difference in cartilage formation of the splanchnocranium and all other cartilages, concerning SPARC expression. Significantly, we show that the initial calcification of cranial bones occurs simultaneously with the expression of OP-L and SPARC in the osteoblast-like cells, which appear early in development. CONCLUSIONS: The difference in SPARC expression during chondrogenesis of the splanchnocranium is likely based on its different evolutionary history compared with the dermatocranium and chondrocranium. Moreover, our results suggest a co-occurrence of the initial calcium deposition and bone matrix protein expression during osteogenesis. Overall, this study enhances our understanding of fish skeletal development and evolution.

Sensitivity differences in fish offer near-infrared vision as an adaptable evolutionary trait.

Near-infrared (NIR) light constitutes an integrated part of solar radiation. The principal ability to sense NIR under laboratory conditions has previously been demonstrated in fish. The availability of NIR in aquatic habitats, and thus its potential use as a cue for distinct behaviors such as orientation and detection of prey, however, depends on physical and environmental parameters. In clear water, blue and green light represents the dominating part of the illumination. In turbid waters, in contrast, the relative content of red and NIR radiation is enhanced, due to increased scattering and absorption of short and middle range wavelengths by suspended particles and dissolved colored materials. We have studied NIR detection thresholds using a phototactic swimming assay in five fish species, which are exposed to different NIR conditions in their natural habitats. Nile and Mozambique tilapia, which inhabit waters with increased turbidity, displayed the highest spectral sensitivity, with thresholds at wavelengths above 930 nm. Zebrafish, guppy and green swordtail, which prefer clearer waters, revealed significantly lower thresholds of spectral sensitivity with 825-845 nm for green swordtail and 845-910 nm for zebrafish and guppy. The present study revealed a clear correlation between NIR sensation thresholds and availability of NIR in the natural habitats, suggesting that NIR vision, as an integral part of the whole spectrum of visual abilities, can serve as an evolutionarily adaptable trait in fish.

Ground-based facilities for simulation of microgravity: organism-specific recommendations for their use, and recommended terminology.

Research in microgravity is indispensable to disclose the impact of gravity on biological processes and organisms. However, research in the near-Earth orbit is severely constrained by the limited number of flight opportunities. Ground-based simulators of microgravity are valuable tools for preparing spaceflight experiments, but they also facilitate stand-alone studies and thus provide additional and cost-efficient platforms for gravitational research. The various microgravity simulators that are frequently used by gravitational biologists are based on different physical principles. This comparative study gives an overview of the most frequently used microgravity simulators and demonstrates their individual capacities and limitations. The range of applicability of the various ground-based microgravity simulators for biological specimens was carefully evaluated by using organisms that have been studied extensively under the conditions of real microgravity in space. In addition, current heterogeneous terminology is discussed critically, and recommendations are given for appropriate selection of adequate simulators and consistent use of nomenclature.

Near-infrared orientation of Mozambique tilapia Oreochromis mossambicus.

Light plays a pivotal role in animal orientation. Aquatic animals face the problem that penetration of light in water is restricted through high attenuation which limits the use of visual cues. In pure water, blue and green light penetrates considerably deeper than red and infrared spectral components. Submicroscopic particles and coloured dissolved organic matter, however, may cause increased scattering and absorption of short-wave components of the solar spectrum, resulting in a relative increase of red and infrared illumination. Here we investigated the potential of near-infrared (NIR) light as a cue for swimming orientation of the African cichlid fish (Cichlidae) Oreochromis mossambicus. A high-throughput semi-automated video tracking assay was used to analyse innate behavioural NIR-sensitivity. Fish revealed a strong preference to swim in the direction of NIR light of a spectral range of 850-950nm at an irradiance similar to values typical of natural surface waters. Our study demonstrates the ability of teleost fish to sense NIR and use it for phototactic swimming orientation.

Histology of the utricle in kinetotically swimming fish: a parabolic aircraft flight study.

OBJECTIVES: Humans taking part in parabolic aircraft flights (PAFs) may suffer from space motion sickness, which is a form of kinetosis. As it has been repeatedly shown that some fish in a given batch also reveal kinetotic behaviour (especially so-called spinning movements and looping responses) during PAFs, and as a result of the homology of the vestibular apparatus of all vertebrates, fish can be used as model systems to investigate the origin of susceptibility to motion sickness. Therefore. we were prompted to examine the utricular maculae, which are responsible for the internalization of gravity in teleosteans of fish swimming kinetotically in microgravity (microg) in comparison with those of animals from the same batch who swam normally. MATERIAL AND METHODS: Larval cichlid fish (Oreochromis mossambicus) were subjected to PAFs. Post-flight, animals which had behaved normally or kinetotically during the microg phases were examined histologically The sizes of the inner ear utricular maculae as well as the numbers of sensory and supporting cells were determined. RESULTS: The total numbers of both sensory and supporting cells of the utricular maculae did not differ between kinetotic and normally swimming fish. Cell density (number of sensory and supporting cells/100 microm2) was, however, reduced in kinetotic animals (p < 0.0001), which seemed to be due to the presence of malformed epithelial cells of increased size in the kinetotic specimens. CONCLUSION: These results indicate that susceptibility to kinetosis may originate from genetically predisposed malformed sensory epithelia.

On the origin of susceptibility to kinetotic swimming behaviour in fish: a parabolic aircraft flight study.

Humans taking part in parabolic aircraft flights (PAFs) may suffer from motion sickness (SMS, a kinetosis; it comprises a dynamic and a static component). It has been argued that the so-called static variety of SMS during PAFs might be based on asymmetric statoliths (i.e., differently weighed statoliths on the right and the left side of the head), with asymmetric inputs to the brain being disclosed in microgravity. Since it has been repeatedly shown earlier that some fish of a given batch reveal a kinetotic behaviour during PAFs (especially so-called spinning movements and looping responses), we investigated whether fish swimming kinetotically in microgravity have a pronounced inner ear otolith asymmetry. Therefore, the swimming behaviour of larval cichlid fish was video-recorded during PAFs and subsequently, size and asymmetry (size difference between the left and the right side) of inner ear otoliths were determined. The asymmetry of utricular otoliths of kinetotic samples was found to be significantly higher than that of normally behaving experimental specimens. Regarding the asymmetry of saccular otoliths of the two groups, statistically different results were not obtained. The findings strongly support the earlier theoretical concept, according to which otolith asymmetry causes (static) SMS.

Developmental profiles of gangliosides in mouse and rat cerebral cortex.

Developmental profiles of 11 gangliosides, concentration of lipid- and glycoprotein-bound sialic acid, and activity of AChE of the rat and mouse cerebral cortex were followed from the 7th day of gestation to the 21st postnatal day.There are three main changes in ganglioside concentration, which are similar in both species. The first occurs from gestation day 10 until birth: parallel to decreased proliferation, cell migration, and neuroblast differentiation, GM3 and GD3 in mouse cortex and GD3 in the rat's decreases in favor of GQ1b, GT1b, and GD1a.The second occurs from birth until the first postnatal week: Parallel to increased growth and arborization of dendrites and axons as well as synaptogenesis in rats and mice, there is a two-fold rise of GD1a, whereas GQ1b and GT1b remain on a nearly constant level. Concomitantly, GM3 and GD3 decreases. The third period of ganglioside changes starts in the second postnatal week, parallel to onset of myelination, and is characterized by an increase of GM1 in parallel with a decrease of the polysialogangliosides GT1b and GQ1b.