A Xenopus neuromast bioassay for chemical ototoxicity

Background: Ototoxic chemicals can impair the senses of hearing and balance in mammals through irreversible damage to the mechanosensory bundles of inner ear hair cells. Fish and amphibians are useful models for investigating ototoxicity because their inner ear hair cells, like those of mammals, are susceptible to damage by ototoxins. Moreover, amphibian mechanosensation is augmented by a lateral line organ on the body surface that comprises external mechanosensory hair cells. The lateral line hair cells are arranged in clusters (neuromasts) and are structurally and functionally similar to inner ear hair cells, but are more accessible for experimental manipulation. Herein, we implemented neuromasts of the amphibian (Xenopus) lateral line as an organ system for evaluating the effects of ototoxic chemicals, such as antibiotics, on mechanosensory hair cell bundles. Methods: We examined the ultrastructure of larval Xenopus laevis neuromasts with scanning electron microscopy (SEM) after larvae were continuously exposed to ototoxic aminoglycoside antibiotics at sub-lethal concentrations (gentamicin; streptomycin; neomycin) for 72 hours. Results: SEM images demonstrated that 72 hours of exposure to antibiotic concentrations greater than 25 μM reduced the hair cell bundle number in lateral line neuromasts. Conclusion: Therapeutic drug studies will benefit from the incorporation of bioassay strategies that evaluate ototoxicity across multiple species including genera of amphibian origin such as Xenopus. Our outcomes support the use of the Xenopus lateral line for identification of potential ototoxic chemicals and suggest that Xenopus neuromast hair cell bundles can withstand antibiotic exposure. The Xenopus bioassay presented here can be incorporated into drug discovery methodology as a high-resolution phenotypic screen for ototoxic effects.

Macrophages break interneuromast cell quiescence by intervening the inhibition of Schwann cells in zebrafish lateral line

In the zebrafish lateral line system, interneuromast cells (INCs) between neuromasts are normally kept quiescent by underlying Schwann cells (SWCs). Upon severe injuries that cause the complete loss of an entire NM, INCs can occasionally differentiate into NMs but how they escape from the inhibition by SWCs is still unclear. Using a genetic/chemical method to specifically ablate a neuromast, we found a small portion of larvae can regenerate a new neuromast, but the regeneration was hindered by inhibiting macrophages. We also demonstrated that the inhibition of macrophage can reduce the percentage of tail fin-amputated larvae to regenerate a new NM. By in toto imaging, we further discovered heterogeneities in macrophage behavior and distribution along lateral line. We witnessed the crawling of macrophages in between injured lateral line and SWCs during regeneration and also in between the second primordium and the first mature lateral line during development. It implies that macrophages may physically separate and alleviate the inhibition from pLLn and SWCs to break the quiescence of INCs during regeneration and development in the zebrafish lateral line.

Behavioural responses of threespine stickleback with lateral line asymmetries to experimental mechanosensory stimuli

Behavioural asymmetry, typically referred to as laterality, is widespread among bilaterians and is often associated with asymmetry in brain structure. However, the influence of sensory receptor asymmetry on laterality has undergone limited investigation. Here we use threespine stickleback (Gasterosteus aculeatus) to investigate the influence of lateral line asymmetry on laterality during lab simulations of three mechanosensation-dependent behaviours: predator evasion, prey localization and rheotaxis. We recorded the response of stickleback to impacts at the water surface and water flow in photic conditions and low-frequency oscillations in the dark, across four repeat trials. We then compared individuals’ laterality to asymmetry in the number of neuromasts on either side of their body. Stickleback hovered with their right side against the arena wall 57% of the time (P<0.001) in illuminated surface impact trials and 56% of the time in (P=0.085) dark low-frequency stimulation trials. Light regime modulated the effect of neuromast count on laterality, as fish with more neuromasts were more likely to hover with the wall on their right during illumination (P=0.007) but were less likely to do so in darkness (P=0.025). Population level laterality diminished in later trials across multiple behaviours and individuals did not show a consistent side bias in any behaviours. Our results demonstrate a complex relationship between sensory structure asymmetry and laterality, suggesting that laterality is modulated multiple sensory modalities and temporally dynamic.

Diversity of lateral line patterns and neuromast numbers in the genus Oryzias

ABSTRACT A remarkable diversity of lateral line patterns exists in adult teleost fishes, the basis of which is largely unknown. By analysing the lateral line patterns and organ numbers in 29 Oryzias species and strains we report a rapid diversification of the lateral line system within this genus. We show a strong dependence of lateral line elaboration (number of neuromasts per cluster, number of parallel lateral lines) on adult species body size irrespective of phylogenetic relationships. In addition, we report that the degree of elaboration of the anterior lateral line, posterior lateral line and caudal neuromast clusters is tightly linked within species, arguing for a globally coordinated mechanism controlling lateral line organ numbers and patterns. We provide evidence for a polygenic control over neuromast numbers and positioning in the genus Oryzias. Our data also indicate that the diversity in lateral lines can arise as a result of differences in patterning both during embryonic development and post-embryonically, where simpler embryonic patterns generate less complex adult patterns and organ numbers, arguing for a linkage between the two processes.

A new record of a rare labrid, Suezichthys notatus (Actinopterygii: Labridae), from Taiwan, with comparison to related species from Taiwan

Three specimens of a rare labrid, Suezichthys notatus (Kamohara, 1958) were recently collected from local markets, which were captured from deep-water off northern and southwestern Taiwan, and represent a new record for Taiwan. Suezichthys notatus can be distinguished from its congeners by a combination of characters: scale rows above lateral line 2½; low scaly sheath present at base of dorsal and anal fins; dorsal-fin element IX, 11; anal-fin elements III, 10; lateral line scales 25‒26, each with simple, unbranched laterosensory canal tube; cheek scale rows behind and below eye 2 and 2‒3 respectively; a group of prominent dark blotches extending from the interorbital region dorsoposteriorly; body depth at dorsal-fin origin 3.7‒3.9 in standard length; short pelvic fin without filamentous extension, 2.2‒2.5 in head length. Suezichthys resembles the labrid genus Pseudolabrus, comparison of Taiwanese species of Suezichthys with those of Pseudolabrus are given.

Tetraspanin Cd9b and Cxcl12a/Cxcr4b have a synergistic effect on the control of collective cell migration

Collective cell migration is essential for embryonic development and homeostatic processes. During zebrafish development, the posterior lateral line primordium (pLLP) navigates along the embryo flank by collective cell migration. The chemokine receptors, Cxcr4b and Cxcr7b, as well as their cognate ligand, Cxcl12a, are essential for this process. We corroborate that knockdown of the zebrafish cd9 tetraspanin orthologue, cd9b, results in mild pLL abnormalities. Through generation of CRISPR and TALEN mutants, we show that cd9a and cd9b function partially redundantly in pLLP migration, which is delayed in the cd9b single and cd9a; cd9b double mutants. This delay led to a transient reduction in neuromast numbers. Loss of both Cd9a and Cd9b sensitized embryos to reduced Cxcr4b and Cxcl12a levels. Together these results provide evidence that Cd9 modulates collective cell migration of the pLLP during zebrafish development. One interpretation of these observations is that Cd9 contributes to more effective chemokine signalling.

Deep learning model inspired by lateral line system for underwater object detection

Inspired by the lateral line systems of various aquatic organisms that are capable of hydrodynamic imaging using ambient flow information, this study develops a deep learning-based object localization model that can detect the location of objects using flow information measured from a moving sensor array. In numerical simulations with the assumption of a potential flow, a two-dimensional hydrofoil navigates around four stationary cylinders in a uniform flow and obtains two types of sensory data during a simulation, namely flow velocity and pressure, from an array of sensors located on the surface of the hydrofoil. Several neural network models are constructed using the flow velocity and pressure data, and these are used to detect the positions of the hydrofoil and surrounding objects. The model based on a long short-term memory network, which is capable of learning order dependence in sequence prediction problems, outperforms the other models. The number of sensors is then optimized using feature selection techniques. This sensor optimization leads to a new object localization model that achieves impressive accuracy in predicting the locations of the hydrofoil and objects with only 40$\%$ of the sensors used in the original model.

Using light-sheet microscopy to study spontaneous activity in the developing lateral-line system

Hair cells are the sensory receptors in the auditory and vestibular systems of all vertebrates, and in the lateral-line system of aquatic vertebrates. During development, spontaneous activity in hair cells shapes the formation of these sensory systems. In auditory hair cells of mice, coordinated waves of spontaneous activity can be triggered by concomitant activity in nearby supporting cells. But in mammals, developing auditory and vestibular hair cells can also autonomously generate spontaneous events independent of supporting cell activity. To date, significant progress has been made studying spontaneous activity in the auditory and vestibular systems of mammals, in isolated cultures. The purpose of this work is to explore the zebrafish lateral-line system as a model to study and understand spontaneous activity in vivo. Our work applies genetically encoded calcium indicators along with light-sheet fluorescence microscopy to visualize spontaneous calcium activity in the developing lateral-line system. Consistent with our previous work, we show that spontaneous calcium activity is present in developing lateral-line hair cells. We now show that supporting cells that surround hair cells, and cholinergic efferent terminals that directly contact hair cells are also spontaneously active. Using two-color functional imaging we demonstrate that spontaneous activity in hair cells does not correlate with activity in either supporting cells or cholinergic terminals. We find that during lateral-line development, hair cells autonomously generate spontaneous events. Using localized calcium indicators, we show that within hair cells, spontaneous calcium activity occurs in two distinct domains-the mechanosensory bundle and the presynapse. Further, spontaneous activity in the mechanosensory bundle ultimately drives spontaneous calcium influx at the presynapse. Comprehensively, our results indicate that in developing lateral-line hair cells, autonomously generated spontaneous activity originates with spontaneous mechanosensory events. Overall, with robust spontaneous activity three different cell types, the developing lateral line is a rich model to study these activities in an intact sensory organ. Future work studying this model may further our understanding of these activities and their role in sensory system formation, function and regeneration.