Zebrafish Muscle Research Articles

CHCHD10P80L knock-in zebrafish display a mild ALS-like phenotype

Mutations in the nuclear-encoded mitochondrial gene CHCHD10 have been observed in patients with a spectrum of diseases that include amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). To investigate the pathogenic nature of disease-associated variants of CHCHD10 we generated a zebrafish knock-in (KI) model expressing the orthologous ALS-associated CHCHD10P80L variant (zebrafish: Chchd10P83L). Larval chchd10P83L/P83L fish displayed reduced Chchd10 protein expression levels, motor impairment, reduced survival and abnormal neuromuscular junctions (NMJ). These deficits were not accompanied by changes in transcripts involved in the integrated stress response (ISR), phenocopying previous findings in our knockout (chchd10−/−). Adult, 11-month old chchd10P83L/P83L zebrafish, displayed smaller slow- and fast-twitch muscle cell cross-sectional areas compared to wild type zebrafish muscle cells. Motoneurons in the spinal cord of chchd10P83L/P83L zebrafish displayed similar cross-sectional areas to that of wild type motor neurons and significantly fewer motor neurons were observed when compared to chchd2−/− adult spinal cords. Bulk RNA sequencing using whole spinal cords of 7-month old fish revealed transcriptional changes associated with neuroinflammation, apoptosis, amino acid metabolism and mt-DNA inflammatory response in our chchd10P83L/P83L model. The findings presented here, suggest that the CHCHD10P80L variant confers an ALS-like phenotype when expressed in zebrafish.

Triptolide, a Cancer Cell Proliferation Inhibitor, Causes Zebrafish Muscle Defects by Regulating Notch and STAT3 Signaling Pathways.

Triptolide is a natural compound in herbal remedies with anti-inflammatory and anti-proliferative properties. We studied its effects on critical signaling processes within the cell, including Notch1 and STAT3 signaling. Our research showed that triptolide reduces cancer cell proliferation by decreasing the expression of downstream targets of these signals. The levels of each signal-related protein and mRNA were analyzed using Western blot and qPCR methods. Interestingly, inhibiting one signal with a single inhibitor alone did not significantly reduce cancer cell proliferation. Instead, MTT assays showed that the simultaneous inhibition of Notch1 and STAT3 signaling reduced cell proliferation. The effect of triptolide was similar to a combination treatment with inhibitors for both signals. When we conducted a study on the impact of triptolide on zebrafish larvae, we found that it inhibited muscle development and interfered with muscle cell proliferation, as evidenced by differences in the staining of myosin heavy chain and F-actin proteins in confocal fluorescence microscopy. Additionally, we noticed that inhibiting a single type of signaling did not lead to any significant muscle defects. This implies that triptolide obstructs multiple signals simultaneously, including Notch1 and STAT3, during muscle development. Chemotherapy is commonly used to treat cancer, but it may cause muscle loss due to drug-related adverse reactions or other complex mechanisms. Our study suggests that anticancer agents like triptolide, inhibiting essential signaling pathways including Notch1 and STAT3 signaling, may cause muscle atrophy through anti-proliferative activity.

Aflatoxin B1 inhibited the development of primary myoblasts of grass carp (Ctenopharyngodon idella) by degrading extracellular matrix

According to the International Agency for Research on Cancer (IARC), aflatoxin B1 (AFB1) has been recognized as a major contaminant in food and animal feed and which is a common mycotoxin with high toxicity. Previous research has found that AFB1 inhibited zebrafish muscle development. However, the potential mechanism of AFB1 on fish muscle development is unknown, so it is necessary to conduct further investigation. In the present research, the primary myoblast of grass carp was used as a model, we treated myoblasts with AFB1 for 24 h. Our results found that 5 μM AFB1 significantly inhibited cell proliferation and migration (P < 0.05), and 10 μM AFB1 promoted lactate dehydrogenase (LDH) release (P < 0.05). Reactive oxygen species (ROS), protein carbonyl (PC) and malondialdehyde (MDA) levels were increased in 15, 5 and 10 μM AFB1 (P < 0.05), respectively. Catalase (CAT), glutathione peroxidase (GPx) and total superoxide dismutase (T-SOD) activities were decreased in 10, 10 and 15 μM AFB1 (P < 0.05), respectively. Furthermore, 15 μM AFB1 induced oxidative damage by Nrf2 pathway, also induced apoptosis in primary myoblast of grass carp. Meanwhile, 15 μM AFB1 decreased MyoD gene and protein expression (P < 0.05). Importantly, 15 μM AFB1 decreased the protein expression of collagen Ⅰ and fibronectin (P < 0.05), and increased the protein levels of urokinase plasminogen activator (uPA), matrix metalloproteinase 9 (MMP-9), matrix metalloproteinase 2 (MMP-2), and p38 mitogen-activated protein kinase (p38MAPK) (P < 0.05). As a result, our findings suggested that AFB1 damaged the cell morphology, induced oxidative damage and apoptosis, degraded ECM components, in turn inhibiting myoblast development by activating the p38MAPK/urokinase-type plasminogen activator (uPA)/matrix metalloproteinase (MMPs)/extracellular matrix (ECM) signaling pathway.

Systematic Identification of Long Noncoding RNAs during Three Key Organogenesis Stages in Zebrafish.

Thousands of lncRNAs have been found in zebrafish embryogenesis and adult tissues, but their identification and organogenesis-related functions have not yet been elucidated. In this study, high-throughput sequencing was performed at three different organogenesis stages of zebrafish embryos that are important for zebrafish muscle development. The three stages were 10 hpf (hours post fertilization) (T1), 24 hpf (T2), and 36 hpf (T3). LncRNA gas5, associated with muscle development, was screened out as the next research target by high-throughput sequencing and qPCR validation. The spatiotemporal expression of lncRNA gas5 in zebrafish embryonic muscle development was studied through qPCR and in situ hybridization, and functional analysis was conducted using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/Cas9, CRISPR/Cas9). The results were as follows: (1) A total of 1486 differentially expressed lncRNAs were identified between T2 and T1, among which 843 lncRNAs were upregulated and 643 were downregulated. The comparison with T3 and T2 resulted in 844 differentially expressed lncRNAs, among which 482 lncRNAs were upregulated and 362 lncRNAs were downregulated. A total of 2137 differentially expressed lncRNAs were found between T3 and T1, among which 1148 lncRNAs were upregulated and 989 lncRNAs were downregulated, including lncRNA gas5, which was selected as the target gene. (2) The results of spatiotemporal expression analysis showed that lncRNA gas5 was expressed in almost all detected embryos of different developmental stages (0, 2, 6, 10, 16, 24, 36, 48, 72, 96 hpf) and detected tissues of adult zebrafish. (3) After lncRNA gas5 knockout using CRISPR/Cas9 technology, the expression levels of detected genes related to muscle development and adjacent to lncRNA gas5 were more highly affected in the knockout group compared with the control group, suggesting that lncRNA gas5 may play a role in embryonic muscle development in zebrafish. (4) The results of the expression of the skeletal myogenesis marker myod showed that the expression of myod in myotomes was abnormal, suggesting that skeletal myogenesis was affected after lncRNA gas5 knockout. The results of this study provide an experimental basis for further studies on the role of lncRNA gas5 in the embryonic skeletal muscle development of zebrafish.

Animal Models for Understanding the Mechanisms of Beta Cell Death during Type 2 Diabetes Pathogenesis.

Type 2 diabetes (T2D) has become a worldwide epidemic, primarily driven by obesity from overnutrition and sedentariness. Recent results reveal there is heterogeneity in both pathology and treatment responses in T2D patients. Therefore, a variety of T2D animal models are necessary to obtain a mechanistic understanding of distinct disease processes. T2D results from insufficient insulin, either due to beta cell loss or inborn deficiency. Although decreases in beta cell mass can occur through loss of identity or cell death, in this review, we will highlight the T2D animal models that display beta cell death, including the Zucker Diabetic Fatty Rat, sand rat, db/db mouse, and a novel diabetic zebrafish model, the Zebrafish Muscle Insulin-Resistant (zMIR) fish. Procuring a mechanistic understanding of different T2D progression trajectories under a variety of contexts is paramount for developing and testing more individualized treatments.

Super-resolution imaging reveals the nanoscale distributions of dystroglycan and integrin ITG7A in zebrafish muscle fibers

Super-resolution imaging reveals the nanoscale distributions of dystroglycan and integrin ITG7A in zebrafish muscle fibers

Expression profile analysis of muscle regulation genes under growth and water flow stress in zebrafish

Muscle is one of the important quality of fish. However, there is still a lack of sufficient understanding of the mechanism of muscle growth regulation in fish. In this study, the expression patterns of 10 reported muscle development related genes are analyzed in zebrafish during growth or under water flow stress. Myogenic regulatory factors (MyoD, MyoG, MRF4), myostatin regulatory genes (Mstna, Mstnb) and myoblast development-related genes (Pax7, Prmt5, Desmin, MYHC) are significantly increased expression at somite stage and muscle effect stage embryos in zebrafish. The mRNA levels of the muscle development related genes are up-regulated in 7-day fries and 30-day juvenile of zebrafish. And there are differences in the expression levels of seven muscle development related genes (MyoD, Myf5, MyoG, Mstna, Mstnb, Desmin, MYHC) between the large and small individuals of 20-day-old zebrafish. Results indicated that short-term water flow stressing can promote zebrafish muscle growth, while prolonged water flow stress may lead to motor fatigue and affect growth in zebrafish. This research provides a foundation for further exploration of the growth regulation mechanism in fish, and is helpful on fish farming.

Spatial Evaluation of the Heavy Metal Iron in Soil, Pond Water and its Mobility into the Muscles of Zebrafish using ICP-OES

Spatial Evaluation of the Heavy Metal Iron in Soil, Pond Water and its Mobility into the Muscles of Zebrafish using ICP-OES

Tissue-specific toxic effects of nano-copper on zebrafish

Understanding the behavior and potential toxicity of copper nanoparticles (nano-Cu) in the aquatic environment is a primary way to assess their environmental risks. In this study, RNA-seq was performed on three different tissues (gills, intestines, and muscles) of zebrafish exposed to nano-Cu, to explore the potential toxic mechanism of nano-Cu on zebrafish. The results indicated that the toxic mechanism of nano-Cu on zebrafish was tissue-specific. Nano-Cu enables the CB1 receptor of the presynaptic membrane of gill cells to affect short-term synaptic plasticity or long-term synaptic changes (ECB-LTD) through DSI and DSE, causing dysfunction of intercellular signal transmission. Imbalance of de novo synthesis of UMP in intestinal cells and its transformation to UDP, UTP, uridine, and uracil, resulted in many functions involved in the pyrimidine metabolic pathway being blocked. Meanwhile, the toxicity of nano-Cu caused abnormal expression of RAD51 gene in muscle cells, which affects the repair of damaged DNA through Fanconi anemia and homologous recombination pathway, thus causing cell cycle disorder. These results provide insights for us to better understand the differences in toxicity of nano-Cu on zebrafish tissues and are helpful for a comprehensive assessment of nano-Cu's effects on aquatic organisms.

Red Fluorescent Protein Variant with a Dual-Peak Emission of Fluorescence.

The marine environment is a rich reservoir of diverse biological entities, many of which possess unique properties that are of immense value to biotechnological applications. One such example is the red fluorescent protein derived from the coral Discosoma sp. This protein, encoded by the DsRed gene, has been the subject of extensive research due to its potential applications in various fields. In the study, a variant of the red fluorescent protein was generated through random mutagenesis using the DsRed2 gene as a template. The process employed error-prone PCR (epPCR) to introduce random mutations, leading to the isolation of twelve gene variants. Among these, one variant stood out due to its unique spectral properties, exhibiting dual fluorescence emission at both 480nm (green) and 550nm (red). This novel variant was expressed in both Escherichia coli and zebrafish (Danio rerio) muscle, confirming the dual fluorescence emission in both model systems. One of the immediate applications of this novel protein variant is in ornamental aquaculture. The dual fluorescence can serve as a unique marker or trait, enhancing the aesthetic appeal of aquatic species in ornamental settings.

Allosteric inhibition of muscle-type nicotinic acetylcholine receptors by a neuromuscular blocking agent pancuronium.

Muscle relaxants are indispensable for surgical anesthesia. Early studies suggested that a classical non-depolarizing muscle relaxant pancuronium competitively binds to the ligand binding site to block nicotinic acetylcholine receptors (nAChR). Our group recently showed that nAChR which has two distinct subunit combinations are expressed in zebrafish muscles, αβδε and αβδ, for which potencies of pancuronium are different. Taking advantage of the distinct potencies, we generated chimeras between two types of nAChRs and found that the extracellular ACh binding site is not associated with the pancuronium sensitivity. Furthermore, application of either 2 μM or 100 μM ACh in native αβδε or αβδ subunits yielded similar IC50 of pancuronium. These data suggest that pancuronium allosterically inhibits the activity of zebrafish nAChRs.

Human Mutated MYOT and CRYAB Genes Cause a Myopathic Phenotype in Zebrafish.

Myofibrillar myopathies (MFMs) are a group of hereditary neuromuscular disorders sharing common histological features, such as myofibrillar derangement, Z-disk disintegration, and the accumulation of degradation products into protein aggregates. They are caused by mutations in several genes that encode either structural proteins or molecular chaperones. Nevertheless, the mechanisms by which mutated genes result in protein aggregation are still unknown. To unveil the role of myotilin and αB-crystallin in the pathogenesis of MFM, we injected zebrafish fertilized eggs at the one-cell stage with expression plasmids harboring cDNA sequences of human wildtype or mutated MYOT (p.Ser95Ile) and human wildtype or mutated CRYAB (p.Gly154Ser). We evaluated the effects on fish survival, motor behavior, muscle structure and development. We found that transgenic zebrafish showed morphological defects that were more severe in those overexpressing mutant genes. which developed a myopathic phenotype consistent with that of human myofibrillar myopathy, including the formation of protein aggregates. Results indicate that pathogenic mutations in myotilin and αB-crystallin genes associated with MFM cause a structural and functional impairment of the skeletal muscle in zebrafish, thereby making this non-mammalian organism a powerful model to dissect disease pathogenesis and find possible druggable targets.

Super-Resolution Imaging Reveals the Nanoscale Distributions of Dystroglycan and Integrin Itga7 in Zebrafish Muscle Fibers.

Cell signaling is determined partially by the localization and abundance of proteins. Dystroglycan and integrin are both transmembrane receptors that connect the cytoskeleton inside muscle cells to the extracellular matrix outside muscle cells, maintaining proper adhesion and function of muscle. The position and abundance of Dystroglycan relative to integrins is thought to be important for muscle adhesion and function. The subcellular localization and quantification of these receptor proteins can be determined at the nanometer scale by FPALM super-resolution microscopy. We used FPALM to determine localizations of Dystroglycan and integrin proteins in muscle fibers of intact zebrafish (Danio rerio). Results were consistent with confocal imaging data, but illuminate further details at the nanoscale and show the feasibility of using FPALM to quantify interactions of two proteins in a whole organism.

Selenium deficiency-induced high concentration of reactive oxygen species restricts hypertrophic growth of skeletal muscle in juvenile zebrafish by suppressing TORC1-mediated protein synthesis.

Se deficiency causes impaired growth of fish skeletal muscle due to the retarded hypertrophy of muscle fibres. However, the inner mechanisms remain unclear. According to our previous researches, we infer this phenomenon is associated with Se deficiency-induced high concentration of reactive oxygen species (ROS), which could suppress the target of rapamycin complex 1 (TORC1) pathway-mediated protein synthesis by inhibiting protein kinase B (Akt), an upstream protein of TORC1. To test this hypothesis, juvenile zebrafish (45 d post-fertilisation) were fed a basal Se-adequate diet or a basal Se-deficient diet or them supplemented with an antioxidant (DL-α-tocopherol acetate, designed as VE) or a TOR activator (MHY1485) for 30 d. Zebrafish fed Se-deficient diets exhibited a clear Se-deficient status in skeletal muscle, which was not influenced by dietary VE and MHY1485. Se deficiency significantly elevated ROS concentrations, inhibited Akt activity and TORC1 pathway, suppressed protein synthesis in skeletal muscle, and impaired hypertrophy of skeletal muscle fibres. However, these negative effects of Se deficiency were partly (except that on ROS concentration) alleviated by dietary MHY1485 and completely alleviated by dietary VE. These data strongly support our speculation that Se deficiency-induced high concentration of ROS exerts a clear inhibiting effect on TORC1 pathway-mediated protein synthesis by regulating Akt activity, thereby restricting the hypertrophy of skeletal muscle fibres in fish. Our findings provide a mechanistic explanation for Se deficiency-caused retardation of fish skeletal muscle growth, contributing to a better understanding of the nutritional necessity and regulatory mechanisms of Se in fish muscle physiology.

Transcriptome Profiling Reveals Enhanced Mitochondrial Activity as a Cold Adaptive Strategy to Hypothermia in Zebrafish Muscle.

The utilisation of synthetic torpor for interplanetary travel once seemed farfetched. However, mounting evidence points to torpor-induced protective benefits from the main hazards of space travel, namely, exposure to radiation and microgravity. To determine the radio-protective effects of an induced torpor-like state we exploited the ectothermic nature of the Danio rerio (zebrafish) in reducing their body temperatures to replicate the hypothermic states seen during natural torpor. We also administered melatonin as a sedative to reduce physical activity. Zebrafish were then exposed to low-dose radiation (0.3 Gy) to simulate radiation exposure on long-term space missions. Transcriptomic analysis found that radiation exposure led to an upregulation of inflammatory and immune signatures and a differentiation and regeneration phenotype driven by STAT3 and MYOD1 transcription factors. In addition, DNA repair processes were downregulated in the muscle two days' post-irradiation. The effects of hypothermia led to an increase in mitochondrial translation including genes involved in oxidative phosphorylation and a downregulation of extracellular matrix and developmental genes. Upon radiation exposure, increases in endoplasmic reticulum stress genes were observed in a torpor+radiation group with downregulation of immune-related and ECM genes. Exposing hypothermic zebrafish to radiation also resulted in a downregulation of ECM and developmental genes however, immune/inflammatory related pathways were downregulated in contrast to that observed in the radiation only group. A cross-species comparison was performed with the muscle of hibernating Ursus arctos horribilis (brown bear) to define shared mechanisms of cold tolerance. Shared responses show an upregulation of protein translation and metabolism of amino acids, as well as a hypoxia response with the shared downregulation of glycolysis, ECM, and developmental genes.

Electrical impedance myography detects age-related skeletal muscle atrophy in adult zebrafish

Age-related deficits in skeletal muscle function, termed sarcopenia, are due to loss of muscle mass and changes in the intrinsic mechanisms underlying contraction. Sarcopenia is associated with falls, functional decline, and mortality. Electrical impedance myography (EIM)—a minimally invasive, rapid electrophysiological tool—can be applied to animals and humans to monitor muscle health, thereby serving as a biomarker in both preclinical and clinical studies. EIM has been successfully employed in several species; however, the application of EIM to the assessment of zebrafish—a model organism amenable to high-throughput experimentation—has not been reported. Here, we demonstrated differences in EIM measures between the skeletal muscles of young (6 months of age) and aged (33 months of age) zebrafish. For example, EIM phase angle and reactance at 2 kHz showed significantly decreased phase angle (5.3 ± 2.1 versus 10.7 ± 1.5°; p = 0.001) and reactance (89.0 ± 3.9 versus 172.2 ± 54.8 ohms; p = 0.007) in aged versus young animals. Total muscle area, in addition to other morphometric features, was also strongly correlated to EIM 2 kHz phase angle across both groups (r = 0.7133, p = 0.01). Moreover, there was a strong correlation between 2 kHz phase angle and established metrics of zebrafish swimming performance, including turn angle, angular velocity, and lateral motion (r = 0.7253, r = 0.7308, r = 0.7857, respectively, p < 0.01 for all). In addition, the technique was shown to have high reproducibility between repeated measurements with a mean percentage difference of 5.34 ± 1.17% for phase angle. These relationships were also confirmed in a separate replication cohort. Together, these findings establish EIM as a fast, sensitive method for quantifying zebrafish muscle function and quality. Moreover, identifying the abnormalities in the bioelectrical properties of sarcopenic zebrafish provides new opportunities to evaluate potential therapeutics for age-related neuromuscular disorders and to interrogate the disease mechanisms of muscle degeneration.

Di-butyl phthalate disrupts muscle, motor and sensory neuron development in embryonic zebrafish

Phthalates are added to plastics to enhance its flexibility, durability and transparency. Phthalates are not covalently bound to plastics and leach into the environment. Phthalates are now pervasive and ubiquitously present in the atmosphere, soil and sediment, surface and wastewater. Phthalates are known endocrine disruptors and their effects on male and female reproduction are well noted. However, studies on the developmental effects of di-butyl phthalate (DBP) are limited. Here we investigated the developmental toxicity of DBP on motor and sensory neuron populations and the muscle structures motor neurons innervate using the zebrafish vertebrate model system. We investigated these effects during the time window of development spanning the period where embryonic patterning determines adult structures. We found that treatment with 2.5 μM DBP from 6 h post-fertilization (hpf) until 72hpf induces loss and disorganization of primary motor neuron innervation of the somatic tissue with concomitant disruptions to muscle fiber organization. Furthermore, we found disruptions to sensory motor neuron development including defects in dorsal root ganglion and their peripherally extending axons. Rohon-Beard sensory neurons were also disrupted showing loss of the bilateral soma positioning along the length of the spinal cord and their afferent axonal projections to the epithelium. Thus, we concluded that DBP is toxic to developing motor and sensory neurons during embryonic development.

Probing microstructural changes in muscles of leptin-deficient zebrafish by non-invasive ex-vivo magnetic resonance microimaging.

Leptin is a hormone that plays a key role in controlling food intake and energy homeostasis. Skeletal muscle is an important target for leptin and recent studies have shown that leptin deficiency may lead to muscular atrophy. However, leptin deficiency-induced structural changes in muscles are poorly understood. The zebrafish has emerged as an excellent model organism for studies of vertebrate diseases and hormone response mechanisms. In this study, we explored ex-vivo magnetic resonance microimaging (μMRI) methods to non-invasively assess muscle wasting in leptin-deficient (lepb-/-) zebrafish model. The fat mapping performed by using chemical shift selective imaging shows significant fat infiltration in muscles of lepb-/- zebrafish compared to control zebrafish. T2 relaxation measurements show considerably longer T2 values in the muscle of lepb-/- zebrafish. Multiexponential T2 analysis detected a significantly higher value and magnitude of long T2 component in the muscles of lepb-/- as compared to control zebrafish. For further zooming into the microstructural changes, we applied diffusion-weighted MRI. The results show a significant decrease in the apparent diffusion coefficient indicating increased constraints of molecular movements within the muscle regions of lepb-/- zebrafish. The use of the phasor transformation for the separation of diffusion-weighted decay signals showed a bi-component diffusion system which allows us to estimate each fraction on a voxel-wise basis. A substantial difference was found between the ratio of two components in lepb-/- and control zebrafish muscles, indicating alterations in diffusion behavior associated with the tissue microstructural changes in muscles of lepb-/- zebrafish as compared to control zebrafish. Taken together, our results demonstrate that the muscles of lepb-/- zebrafish undergo significant fat infiltration and microstructural changes leading to muscle wasting. This study also demonstrates that μMRI provides excellent means to non-invasively study the microstructural changes in the muscles of the zebrafish model.

CCN6 influences transcription and controls mitochondrial mass and muscle organization.

Mutations in Cellular Communication Network Factor 6 (CCN6) are linked to the debilitating musculoskeletal disease Progressive Pseudo Rheumatoid Dysplasia (PPRD), which disrupts mobility. Yet, much remains unknown about CCN6 function at the molecular level. In this study, we revealed a new function of CCN6 in transcriptional regulation. We demonstrated that CCN6 localizes to chromatin and associates with RNA Polymerase II in human chondrocyte lines. Using zebrafish as a model organism we validated the nuclear presence of CCN6 and its association with RNA Polymerase II in different developmental stages from 10 hpf embryo to adult fish muscle. In concurrence with these findings, we confirmed the requirement of CCN6 in the transcription of several genes encoding mitochondrial electron transport complex proteins in the zebrafish, both in the embryonic stages and in the adult muscle. Reduction in the expression of these genes upon morpholino-mediated knockdown of CCN6 protein expression led to reduced mitochondrial mass, which correlated with defective myotome organization during zebrafish muscle development. Overall, this study suggests that the developmental musculoskeletal abnormalities linked with PPRD could be contributed at least partly by impaired expression of genes encoding mitochondrial electron transport complexes due to defects in CCN6 associated transcriptional regulation.

Ryanodine receptor-active non-dioxin-like polychlorinated biphenyls cause neurobehavioral deficits in larval zebrafish.

Although their production was banned in the United States in 1977, polychlorinated biphenyls (PCBs) continue to pose significant risks to the developing nervous system. Perinatal exposure to PCBs is associated with increased risk of neuropsychiatric disorders, perhaps due to altered patterns of dendritic arborization of central neurons. Non-dioxin-like (NDL) PCB congeners enhance dendritic arborization of developing mammalian neurons via sensitization of ryanodine receptors (RYR). Structure-activity relationships (SAR) of RYR sensitization by PCBs have been demonstrated using mammalian and rainbow trout (Oncorhynchus mykiss) tissue homogenates. The purpose of this study is to determine whether this SAR translates to developmental neurotoxicity (DNT) of PCBs in vivo, a question that has yet to be tested. To address this gap, we leveraged a zebrafish model to evaluate the developmental neurotoxicity potential of PCBs 28, 66, 84, 95, 138, and 153, congeners previously shown to have broadly different potencies towards sensitizing RYR. We first confirmed that these PCB congeners exhibited differing potency in sensitizing RYR in zebrafish muscle ranging from negligible (PCB 66) to moderate (PCB 153) to high (PCB 95) RYR activity. Next, enzymatically dechorionated embryos were statically exposed to varying concentrations (0.1–10 μM) of each PCB congener from 6 h post-fertilization to 5 days post-fertilization (dpf). Embryos were observed daily using stereomicroscopy to assess mortality and gross malformations and photomotor behavior was assessed in larval zebrafish at 3, 4, and 5 dpf. The body burden of each PCB was measured by gas chromatography. The key findings are: 1) None of these PCBs caused death or overt teratology at the concentrations tested; 2) A subset of these PCB congeners altered photomotor behavior in larval zebrafish and the SAR for PCB behavioral effects mirrored the SAR for RYR sensitization; and 3) Quantification of PCB levels in larval zebrafish ruled out the possibility that congener-specific effects on behavior were due to differential uptake of PCB congeners. Collectively, the findings from this study provide in vivo evidence in support of the hypothesis that RYR sensitization contributes to the DNT of PCBs.