Bisphenols disrupt differentiation of the pigmented cells during larval brain formation in the ascidian.

The endocrine disruptor Bisphenol A (BPA), a widely employed molecule in plastics, has been shown to affect several biological processes in vertebrates, mostly via binding to nuclear receptors. Neurodevelopmental effects of BPA have been documented in vertebrates and linked to neurodevelopmental disorders, probably because some nuclear receptors are present in the vertebrate brain. Similarly, endocrine disruptors have been shown to affect neurodevelopment in marine invertebrates such as ascidians, mollusks or echinoderms, but whether invertebrate nuclear receptors are involved in the mode-of-action is largely unknown. In this study, we assessed the effect of BPA on larval brain development of the ascidian Phallusia mammillata. We found that BPA is toxic to P. mammillata embryos in a dose-dependent manner (EC50: 11.8µM; LC50: 21µM). Furthermore, micromolar doses of BPA impaired differentiation of the ascidian pigmented cells, by inhibiting otolith movement within the sensory vesicle. We further show that this phenotype is specific to other two bisphenols (BPE and BPF) over a bisphenyl (2,2 DPP). Because in vertebrates the estrogen-related receptor gamma (ERRγ) can bind bisphenols with high affinity but not bisphenyls, we tested whether the ascidian ERR participates in the neurodevelopmental phenotype induced by BPA. Interestingly, P. mammillata ERR is expressed in the larval brain, adjacent to the differentiating otolith. Furthermore, antagonists of vertebrate ERRs also inhibited the otolith movement but not pigmentation. Together our observations suggest that BPA may affect ascidian otolith differentiation by altering Pm-ERR activity whereas otolith pigmentation defects might be due to the known inhibitory effect of bisphenols on tyrosinase enzymatic activity.

Exposure to cadmium causes inhibition of otolith development and behavioral impairment in zebrafish larvae.

Otolith consisting largely of calcium carbonate, fibrous and proteins, is vital for maintaining body balance and/or hearing of fish. The formation of otolith involves Ca2+ transport and deposition. In the present study, we investigated the effects of Cd2+ on otoliths development by using zebrafish embryos as model. The results showed that exposure to Cd2+ inhibited the utricular and saccular otoliths growth, indicated by reduced lateral areas. Swimming speeds were reduced and a losing balance control was observed in Cd2+ exposed larvae. The genes related to Ca2+ transport (e.g. plasma membrane Ca2+-ATPase isoform 2, pmca2; Ca2+-ATPase isoform 2, atp2b1a) and regulation (e.g. parathyroid hormone ligand type-1, pth1; stanniocalcin isoform 1, stc1) were significantly downregulated. However, the adverse effects of Cd2+ on otoliths growth and swimming activity can be protected by supplementation of Ca2+ in exposure medium. Body burden of Cd2+ in larvae was reduced upon the supplement with Ca2+. The overall results suggest that exposure to Cd2+ can inhibit influx of Ca2+, leading to less deposition of CaCO3 for otolith growth, and finally result in impaired balance control and swimming activity in zebrafish larvae.

Regulatory mechanisms of miR-96 and miR-184 abnormal expressions on otic vesicle development of zebrafish following exposure to ß-diketone antibiotics.

Chronic ototoxicity of ß-diketone antibiotics (DKAs) to zebrafish (Danio rerio) was explored in detail by following abnormal expressions of two hearing-related miRNAs. Dose-dependent down-regulation of miR-96 and miR-184 was observed in otoliths during embryonic-larval development. Continuous DKA exposure to 120-hpf larva decreased sensitivity to acoustic stimulation. Development of otolith was delayed in treatment groups, showing unclear boundaries and vacuolization at 72-hpf, and utricular enlargement as well as decreased saccular volume in 96-hpf or latter larval otoliths. If one miRNA was knocked-down and another over-expressed, only a slight influence on morphological development of the otic vesicle occurred, but knocked-down or over-expressed miRNA both significantly affected zebrafish normal development. Injection of miR-96, miR-184 or both micRNA mimics to yolk sac resulted in marked improvement of otic vesicle phenotype. However, hair cell staining showed that only the injected miR-96 mimic restored hair cell numbers after DKA exposure, demonstrating that miR-96 played an important role in otic vesicle development and formation of hearing, while miR-184 was only involved in otic vesicle construction during embryonic development. These observations advance our understanding of hearing loss owing to acute antibiotic exposure and provide theoretical guidance for early intervention and gene therapy for drug-induced diseases.

Growth and otolith morphology vary with alternative reproductive tactics and contaminant exposure in the round goby Neogobius melanostomus.

Round goby Neogobius melanostomus sagittal (saccular) otolith morphology was compared between males of the two alternative reproductive tactics (termed guarder and sneaker males) and between males captured from sites of high or low contamination. Otolith size increased with fish size and also displayed an ontogenetic shift in shape, becoming relatively taller as otoliths grew in size. Despite a considerable overlap in age between males adopting the two reproductive tactics, size-at-age measurements revealed that guarder males are significantly larger than sneakers at any given age and that they invest more into somatic growth than sneaker males. Controlling for body size, sneaker males possessed heavier sagittal otoliths than guarder males. Subtle otolith shape differences were also found between the two male tactics and between sites of high and low contaminant exposure. Sneaker males had relatively shorter otoliths with more pronounced notching than guarder males. Fish captured at sites of high contamination had otoliths showing slower growth rates in relation to body size and their shapes had more pronounced caudal points and ventral protrusions when compared with fish captured at sites of low contamination. The results are discussed in relation to life-history tradeoffs between the male tactics in terms of reproductive and somatic investment as well as the putative metabolic costs of exposure to contaminants. Overall, this study reveals that male alternative reproductive tactics and environmental contaminants can have small, yet measurable, effects on otolith morphology and these factors should be accounted for in future research when possible.

Ocean acidification promotes otolith growth and calcite deposition in gilthead sea bream (Sparus aurata) larvae.

The effects of ocean acidification on otolith crystallization and growth rates were investigated in gilthead sea bream (Sparus aurata) larvae. Larvae were exposed to three different pH levels: pH8.2, pH7.7 and pH7.3 for a period of 18 days post-fertilization. For the first time, we demonstrate that pH has a significant impact on the carbonate polymorph composition, showing calcite in a significant percentage of individuals at low pH. Around 21% of the larvae exposed to pH7.3 showed irregular calcitic otoliths rather than commonly found round aragonitic otoliths. Calcitic otoliths showed a moderate level of heritability suggesting an important role of genetic factors. We also observed significantly larger otoliths in larvae reared at pH7.7 and pH7.3 compared to pH8.2 in both sagittae and lapilli. Our results demonstrate that otolith growth rates in gilthead sea bream larvae increase at low pH while a significant proportion of larvae are prone to the formation of calcitic otoliths at pH7.3.

Mechanism Underlying the Effects of Estrogen Deficiency on Otoconia.

Otoconia-related vertigo and balance deficits, particularly benign paroxysmal positional vertigo (BPPV), are common. Our recent studies in humans show that, while BPPV prevalence greatly increases with age in both genders, peri-menopausal women are especially susceptible. In the present study, we show that bilateral ovariectomized (OVX) mice have significant balance behavioral deficits, and that estrogen deficiency compromises otoconia maintenance and anchoring by reducing the expression of otoconial component and anchoring proteins. There is ectopic debris formation in the ampulla under estrogen deficiency due to aberrant matrix protein expression. Furthermore, phytoestrogen is effective in rescuing the otoconia abnormalities. By comparing the expression levels of known estrogen receptor (Esr) subtypes, and by examining the otoconia phenotypes of null mice for selected receptors, we postulate that Esr2 may be critical in mediating the effects of estrogen in otoconia maintenance.

The Ascidian Embryo Teratogenicity assay in Ciona intestinalis as a new teratological screening to test the mixture effect of the co-exposure to ethanol and fluconazole.

The aim of this work was to evaluate the Ascidian Embryo Teratogenicity assay (AET) as new alternative invertebrate model to test the developmental effects of the co-exposure to ethanol and fluconazole. Ciona intestinalis embryos were exposed to the azolic fungicide fluconazole, (FLUCO, 7.8-250µM), to ethanol (Eth, 0.01-0.5%) and to their mixture (0.01% Eth+FLUCO 7.8-250µM) from neurula to larval stage. At the end of the exposure period, larvae were morphologically evaluated and benchmark analysis performed by using the PROAST modelling software. Both compounds were teratogenic in a concentration-related manner, particularly affecting the pigmented organs. The co-exposure to Eth enhanced the effects of FLUCO, the additive hypothesis was not rejected by the modelling. The results demonstrated that AET could be considered a good vertebrate-free alternative model for toxicological investigation in embryos.

Effect of betel nut chewing on the otolithic reflex system.

OBJECTIVE: This study investigated the effect of betel nut chewing on the otolithic reflex system. METHODS: Seventeen healthy volunteers without any experience of chewing betel nut (fresh chewers) and 17 habitual chewers underwent vital sign measurements, ocular vestibular-evoked myogenic potential (oVEMP), and cervical VEMP (cVEMP) tests prior to the study. Each subject then chewed two pieces of betel nut for 2min (dosing). The same paradigm was repeated immediately, 10min, and 20min after chewing. On a different day, 10 fresh chewers masticated chewing gum as control. RESULTS: Fresh chewers exhibited significantly decreased response rates of oVEMP (53%) and cVEMP (71%) after dosing compared with those from the predosing period. These abnormal VEMPs returned to normal 20min after dosing. In contrast, 100% response rates of oVEMP and cVEMP were observed before and after masticating chewing gum. In habitual chewers, the response rates of oVEMP and cVEMP were 32% and 29%, respectively, 20min after dosing. CONCLUSION: Chewing betel nuts induced a transient loss of the otolithic reflexes in fresh chewers but may cause permanent loss in habitual chewers. SIGNIFICANCE: Chewing betel nuts can cause a loss of otholitic reflex function. This creates a risk for disturbed balance and malfunction, for instance, during driving.

Is Exposure to Macondo Oil Reflected in the Otolith Chemistry of Marsh-Resident Fish?

Genomic and physiological responses in Gulf killifish (Fundulus grandis) in the northern Gulf of Mexico have confirmed oil exposure of resident marsh fish following the Macondo blowout in 2010. Using these same fish, we evaluated otolith microchemistry as a method for assessing oil exposure history. Laser-ablation inductively-coupled-plasma mass spectrometry was used to analyze the chemical composition of sagittal otoliths to assess whether a trace metal signature could be detected in the otoliths of F. grandis collected from a Macondo-oil impacted site in 2010, post-spill relative to pre-spill, as well as versus fish from areas not impacted by the spill. We found no evidence of increased concentrations of two elements associated with oil contamination (nickel and vanadium) in F. grandis otoliths regardless of Macondo oil exposure history. One potential explanation for this is that Macondo oil is relatively depleted of those metals compared to other crude oils globally. During and after the spill, however, elevated levels of barium, lead, and to a lesser degree, copper were detected in killifish otoliths at the oil-impacted collection site in coastal Louisiana. This may reflect oil contact or other environmental perturbations that occurred concomitant with oiling. For example, increases in barium in otoliths from oil-exposed fish followed (temporally) freshwater diversions in Louisiana in 2010. This implicates (but does not conclusively demonstrate) freshwater diversions from the Mississippi River (with previously recorded higher concentrations of lead and copper), designed to halt the ingress of oil, as a mechanism for elevated elemental uptake in otoliths of Louisiana marsh fishes. These results highlight the potentially complex and indirect effects of the Macondo oil spill and human responses to it on Gulf of Mexico ecosystems, and emphasize the need to consider the multiple stressors acting simultaneously on inshore fish communities.

High magnetic field induced otolith fusion in the zebrafish larvae.

Magnetoreception in animals illustrates the interaction of biological systems with the geomagnetic field (geoMF). However, there are few studies that identified the impact of high magnetic field (MF) exposure from Magnetic Resonance Imaging (MRI) scanners (>100,000 times of geoMF) on specific biological targets. Here, we investigated the effects of a 14 Tesla MRI scanner on zebrafish larvae. All zebrafish larvae aligned parallel to the B0 field, i.e. the static MF, in the MRI scanner. The two otoliths (ear stones) in the otic vesicles of zebrafish larvae older than 24 hours post fertilization (hpf) fused together after the high MF exposure as short as 2 hours, yielding a single-otolith phenotype with aberrant swimming behavior. The otolith fusion was blocked in zebrafish larvae under anesthesia or embedded in agarose. Hair cells may play an important role on the MF-induced otolith fusion. This work provided direct evidence to show that high MF interacts with the otic vesicle of zebrafish larvae and causes otolith fusion in an "all-or-none" manner. The MF-induced otolith fusion may facilitate the searching for MF sensors using genetically amenable vertebrate animal models, such as zebrafish.

The role of GABAB receptors in the vestibular oculomotor system in mice.

Systemic administration of a gamma-amino butyric acid type B (GABAB) receptor agonist, baclofen, affects various physiological and psychological processes. To date, the effects on oculomotor system have been well characterized in primates, however those in mice have not been explored. In this study, we investigated the effects of baclofen focusing on vestibular-related eye movements. Two rotational paradigms, i.e. sinusoidal rotation and counter rotation were employed to stimulate semicircular canals and otolith organs in the inner ear. Experimental conditions (dosage, routes and onset of recording) were determined based on the prior studies exploring the behavioral effects of baclofen in mice. With an increase in dosage, both canal and otolith induced ocular responses were gradually affected. There was a clear distinction in the drug sensitivity showing that eye movements derived from direct vestibulo-ocular reflex pathways were relatively unaltered, while the responses through higher-order neural networks in the vestibular system were substantially decreased. These findings were consistent with those observed in primates suggesting a well-conserved role of GABAB receptors in the oculomotor system across frontal-eyed and lateral-eyed animals. We showed here a previously unrecognized effect of baclofen on the vestibular oculomotor function in mice. When interpreting general animal performance under the drug, the potential contribution of altered balance system should be taken into consideration.

Assessment of D-methionine protecting cisplatin-induced otolith toxicity by vestibular-evoked myogenic potential tests, ATPase activities and oxidative state in guinea pigs.

To date, inadequate study has been devoted to the toxic vestibular effects caused by cisplatin. In addition, no electrophysiological examination has been conducted to assess cisplatin-induced otolith toxicity. The purposes of this study are thus two-fold: 1) to determine whether cervical vestibular-evoked myogenic potentials (VEMPs) and ocular VEMPs are practical electrophysiological methods of testing for cisplatin-induced otolith toxicity and 2) to examine if D-methionine (D-met) pre-injection would protect the otolith organs against cisplatin-induced changes in enzyme activities and/or oxidative status. Guinea pigs were intraperitoneally treated once daily with the following injections for seven consecutive days: sterile 0.9% saline control, cisplatin (5 mg/kg) only, D-met (300 mg/kg) only, or a combination of d-met (300 mg/kg) and cisplatin (5 mg/kg), respectively, with a 30 minute window in between. Each animal underwent the oVEMP and cVEMP tests before and after treatment. The changes in the biochemistry of the otolith organs, including membranous Na(+), K(+)-ATPase and Ca(2+)-ATPase, lipid peroxidation (LPO) levels and nitric oxide (NO) levels, were also evaluated. In the cisplatin-only treated guinea pigs, the mean amplitudes of the oVEMP tests were significantly (p<0.05) decreased when compared to the other three groups. In guinea pigs receiving both D-met and cisplatin, the amplitudes of their oVEMP tests were significantly larger (p<0.05) than those of the cisplatin-only group, but smaller (p<0.05) than those of the saline control or D-met-only group. However, no significant difference of the amplitudes of cVEMP tests was noted among the four groups. In comparison with the other three groups, the cisplatin-only group had the lowest (ps<0.05) mean Na(+), K(+)-ATPase and Ca(2+)-ATPase, and the highest (ps<0.05) LPO and NO levels. The oVEMP tests were feasible for the evaluation of cisplatin-related otolith dysfunction. D-Met attenuated the reduced ATPase activities and increased oxidative stress induced by cisplatin toxicity in the otolith organs.

Protective Effect of Ginkgo Biloba Extract on Gentamicin-Induced Structural Changes of Calcite-Gelatin Composite.

OBJECTIVE: To evaluate the protective effect of Ginkgo biloba extract (GBE) on gentamicin (GM)-induced morphological damage of an artificial otoconia. MATERIALS AND METHODS: An artificial otoconia powder was placed in a 12-well culture plate containing artificial endolymph. GM (500 µL; 40 mg/mL) was added to the first four wells. GM (500 µL; 40 mg/mL) with GBE (500 µL) was added to the next four wells. PBS (500 µL) was added to the remaining four wells as a control. The levels of nitric oxide (NO) synthesis were determined in the supernatants of each group. Alteration in surface morphology and calcium content were determined by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDX), respectively. RESULTS: NO concentration increased in the GM-treated group compared with that in the control group. When the calcite-gelatin composite was treated with GBE, GM-stimulated NO production significantly decreased. The surface of the calcite-gelatin composite exposed to GM showed erosive fissures and had a honeycomb-like appearance. GBE showed a protective effect against dissolution. EDX showed that the calcium content of the GM-treated group significantly decreased. However, the GBE-treated group demonstrated an insignificant change in the calcium concentration compared with the control. CONCLUSION: From these results, we can conclude that GBE may have a protective effect against GM-induced NO production and superficial structural changes.

The proteome of Atlantic herring (Clupea harengus L.) larvae is resistant to elevated pCO2.

Elevated anthropogenic pCO2 can delay growth and impair otolith structure and function in the larvae of some fishes. These effects may concurrently alter the larva's proteome expression pattern. To test this hypothesis, Atlantic herring larvae were exposed to ambient (370 µatm) and elevated (1800 µatm) pCO2 for one-month. The proteome structure of the larvae was examined using a 2-DE and mass spectrometry. The length of herring larvae was marginally less in the elevated pCO2 treatment compared to the control. The proteome structure was also different between the control and treatment, but only slightly: the expression of a small number of proteins was altered by a factor of less than 2-fold at elevated pCO2. This comparative proteome analysis suggests that the proteome of herring larvae is resilient to elevated pCO2. These observations suggest that herring larvae can cope with levels of CO2 projected for near future without significant proteome-wide changes.

Estrogen-related receptor γ is an in vivo receptor of bisphenol A.

Bisphenol A (BPA) is an endocrine disruptor that displays estrogenic activity. Several reports suggest that BPA may have estrogen receptor-independent effects. In zebrafish, 50 µM BPA exposure induces otic vesicle abnormalities, including otolith aggregation. The purpose of this study was to test if BPA action was mediated in vivo during zebrafish development by the orphan nuclear estrogen related receptor (ERR) γ. Combining pharmacological and functional approaches, we demonstrate that the zebrafish ERRγ mediates BPA-induced malformations in otoliths. Using different bisphenol derivatives, we show that different compounds can induce a similar otolith phenotype than BPA and that the binding affinity of these derivatives to the zebrafish ERRγ correlates with their ability to induce otolith malformations. Morpholino knockdown of ERRγ function suppresses the BPA effect on otoliths whereas overexpression of ERRγ led to a BPA-like otolith phenotype. Moreover, a subphenotypical dose of BPA (1 µM) combined with ERRγ overexpression led to a full-dose (50 µM) BPA otolith phenotype. We therefore conclude that ERRγ mediates the otic vesicle phenotype generated by BPA. Our results suggest that the range of pathways perturbed by this compound and its potential harmful effect are larger than expected.-Tohmé, M., Prud'homme, S. M., Boulahtouf, A., Samarut, E., Brunet, F., Bernard, L., Bourguet, W., Gibert, Y., Balaguer, P., Laudet, V. Estrogen-related receptor γ is an in vivo receptor of bisphenol A.

The effect of alcohol on cervical and ocular vestibular evoked myogenic potentials in healthy volunteers.

OBJECTIVE: We investigated the effect of alcohol on the cervical and ocular vestibular evoked myogenic potentials (cVEMPs and oVEMPs). As alcohol produces gaze-evoked nystagmus (GEN), we also tested the effect of nystagmus independent of alcohol by recording oVEMPs during optokinetic stimulation (OKS). METHODS: The effect of alcohol was tested in 14 subjects over multiple rounds of alcohol consumption up to a maximum breath alcohol concentration (BrAC) of 1.5‰ (mean 0.97‰). The effect of OKS was tested in 11 subjects at 5, 10 and 15deg/sec. RESULTS: oVEMP amplitude decreased from baseline to the highest BrAC level by 27% (range 5-50%, P<0.001), but there was no significant effect on oVEMP latency or cVEMP amplitude or latency. There was a significant negative effect of OKS on oVEMP amplitude (16%, P=0.006). CONCLUSIONS: We found a selective effect of alcohol on oVEMP amplitude, but no effect on the cVEMP. Vertical nystagmus elicited by OKS reduced oVEMP amplitude. SIGNIFICANCE: Alcohol selectively affects oVEMP amplitude. Despite the effects of alcohol and nystagmus, both reflexes were reliably recorded in all subjects and conditions. An absent response in a patient affected by alcohol or nystagmus indicates a vestibular deficit.

Gentamicin-induced structural damage of human and artificial (biomimetic) otoconia.

CONCLUSIONS: Gentamicin causes irreversible structural damage of human and artificial otoconia by progressive dissolution of calcite. The inner architecture of otoconia is strongly affected by degradation scenarios during gentamicin exposure. Artificial otoconia can be used as a model system mimicking the chemical attacks for detailed investigations. OBJECTIVES: To investigate the chemical interactions of gentamicin with natural calcite and human and artificial otoconia under in vivo conditions. METHODS: Pure calcite crystals and artificial and human otoconia were exposed to gentamicin injection solutions at various concentrations. Morphological changes were observed in time steps by the use of environmental scanning electron microscopy (ESEM). RESULTS: Dissolution of pure calcite crystals results in the formation of well oriented nanoshoots indicating an irreversible chemical reaction with gentamicin. Human and artificial otoconia reveal irreversible structural changes of their surface areas as well as of their inner structure, resulting in characteristic changes at different gentamicin concentrations. Minor changes are first observed by surface alterations and dissolution of calcite in the belly region. Major changes result in further reduction of the belly area reaching the center of symmetry. Finally, a complete dissolution of the branches takes place. Artificial otoconia provide detailed insight into surface alterations.

Baclofen affects the semicircular canals but not the otoliths in humans.

CONCLUSION: This study showed that GABAB agonist baclofen (10 mg) affects the semicircular canals (SCCs), both centrally and peripherally, but does not influence the otolithic function. OBJECTIVES: The aim of the study was to identify the effects of baclofen on the complete vestibular system, i.e. semicircular canals, saccules and utricles. METHODS: The study had a double-blind, placebo-controlled, repeated measures design and was conducted on healthy male volunteers. With electronystagmography (ENG), the SCC function was evaluated, whereas utricular function was determined by means of unilateral centrifugation (UC). Cervical vestibular evoked myogenic potentials (cVEMPs) tested saccular integrity. RESULTS: Baclofen caused a significant increase of the vestibulo-ocular reflex (VOR) phase and a significant decrease of the total caloric response (TCR), both measured during ENG. The drug also decreased the maximal contribution of the SCCs to ocular counter-rolling (OCR) evaluated during UC. No effects on saccules and utricules were observed.

Differences in organochlorine accumulation accompanying life history in the catadromous eel Anguilla japonica and the marine eel Conger myriaster.

In order to examine the ecological risk for organic pollutants in diadromous fish migrating between sea and freshwater, organochlorine compounds (OCs) were determined in the catadromous eel Anguilla japonica having marine, estuarine and freshwater residence life histories. The eels were collected in Japanese coastal areas. We also compared the OCs accumulation with the marine eel Conger myriaster, which has a similar life history as A. japonica in the marine environment. The ontogenic changes in the otolith strontium (Sr) and calcium (Ca) concentrations were examined along the life history transect to discriminate the migration type. There were generally three different patterns, which were categorized as 'marine residence' (spent most of their life in the sea and did not enter freshwater), 'estuarine residence' (inhabited estuaries or switched between different habitats), and 'freshwater residence' (entered and remained in freshwater river habitats after arrival in the estuary) according to the otolith Sr:Ca ratio. There were generally no correlations between OCs such as Dichlorodiphenyltrichloroethanes (DDTs), Hexachlorobenzene (HCB), Hexachlorocyclohexanes (HCHs) and Chlordanes (CHLs) accumulation and each biological characteristic such as TL, BW and age in A. japonica. A positive correlation between the lipid content and concentrations of OCs were found. Additionally, the concentrations of HCB, ∑HCHs, ∑CHLs and ∑DDTs in A. japonica were significantly higher than those of C. myriaster, associating with the higher lipid contents (14% on average) in the former than the latter eels (9% on average). A negative linear relationship was found between the otolith Sr:Ca ratios and concentration of each OCs in A. japonica. The ecological risk of OCs increase as the freshwater residence period in the eel becomes longer. It is clear that migratory histories and lipid contents directly affected OCs accumulation in the anguillid eels.

Role of Chd7 in zebrafish: a model for CHARGE syndrome.

CHARGE syndrome is caused by mutations in the CHD7 gene. Several organ systems including the retina, cranial nerves, inner ear and heart are affected in CHARGE syndrome. However, the mechanistic link between mutations in CHD7 and many of the organ systems dysfunction remains elusive. Here, we show that Chd7 is required for the organization of the neural retina in zebrafish. We observe an abnormal expression or a complete absence of molecular markers for the retinal ganglion cells and photoreceptors, indicating that Chd7 regulates the differentiation of retinal cells and plays an essential role in retinal cell development. In addition, zebrafish with reduced Chd7 display an abnormal organization and clustering of cranial motor neurons. We also note a pronounced reduction in the facial branchiomotor neurons and the vagal motor neurons display aberrant positioning. Further, these fish exhibit a severe loss of the facial nerves. Knock-down of Chd7 results in a curvature of the long body axis and these fish develop irregular shaped vertebrae and have a reduction in bone mineralization. Chd7 knockdown also results in a loss of proper segment polarity illustrated by flawed efnb2a and ttna expression, which is associated with later vascular segmentation defects. These critical roles for Chd7 in retinal and vertebral development were previously unrecognized and our results provide new insights into the role of Chd7 during development and in CHARGE syndrome pathogenesis.