scholarly journals The Cyanotoxin 2,4-DAB Reduces Viability and Causes Behavioral and Molecular Dysfunctions Associated with Neurodegeneration in Larval Zebrafish

2022 ◽  
Author(s):  
Rubia M. Martin ◽  
Michael S. Bereman ◽  
Kurt C. Marsden
Keyword(s):  
Metabolic Pathways ◽  
Protein Profile ◽  
Acoustic Startle ◽  
Motor Dysfunction ◽  
Zebrafish Model ◽  
Larval Zebrafish ◽  
Neurotoxic Effects ◽  
And Behavior ◽  
Lateral Sclerosis

AbstractExposure to cyanotoxins has been linked to neurodegenerative diseases, including amyotrophic lateral sclerosis, Alzheimer’s, and Parkinson’s disease. While the cyanotoxin β-methylamino-L-alanine (BMAA) has received much attention, cyanobacteria produce many cyanotoxic compounds, several of which have been detected in nature alongside BMAA, including 2,4-diaminobutyric acid (2,4-DAB) and N-(2-aminoethyl)glycine (AEG). Thus, the question of whether 2,4-DAB and AEG also cause neurotoxic effects in vivo is of great interest, as is the question of whether they interact to enhance toxicity. Here, we evaluate the toxic and neurotoxic effects of these cyanotoxins alone or in combination by measuring zebrafish larval viability and behavior after exposure. 2,4-DAB was the most potent cyanotoxin as it decreased larval viability by approximately 50% at 6 days post fertilization, while BMAA and AEG decreased viability by just 16% and 8%, respectively. Although we only observed minor neurotoxic effects on spontaneous locomotion, BMAA and AEG enhanced acoustic startle sensitivity, and they interacted in an additive manner to exert their effects. 2,4-DAB; however, only modulated startle kinematics, an indication of motor dysfunction. To investigate the mechanisms of 2,4-DAB’s effects, we analyzed the protein profile of larval zebrafish exposed to 500 µM 2,4-DAB at two time points and identified molecular signatures consistent with neurodegeneration, including disruption of metabolic pathways and downregulation of the ALS-associated genes SOD1 and UBQLN4. Together, our data demonstrate that BMAA and its isomers AEG and 2,4-DAB cause neurotoxic effects in vivo, with 2,4-DAB as the most potent of the three in the zebrafish model.

scholarly journals The cyanotoxin 2,4-DAB enhances mortality and causes behavioral and molecular dysfunctions associated with neurodegeneration in larval zebrafish

2021 ◽  
Author(s):  
Rubia M. Martin ◽  
Michael S. Bereman ◽  
Kurt C. Marsden
Keyword(s):  
Metabolic Pathways ◽  
Protein Profile ◽  
Acoustic Startle ◽  
Motor Dysfunction ◽  
Zebrafish Model ◽  
Larval Zebrafish ◽  
Neurotoxic Effects ◽  
And Behavior ◽  
Lateral Sclerosis

Exposure to cyanotoxins has been linked to neurodegenerative diseases, including amyotrophic lateral sclerosis, Alzheimer's, and Parkinson's disease. While the cyanotoxin beta-methylamino-L-alanine (BMAA) has received much attention, cyanobacteria produce many cyanotoxic compounds, several of which have been detected in nature alongside BMAA including 2,4-diaminobutyric acid (2,4-DAB), and N-(2-aminoethyl)glycine (AEG). Thus, the question of whether DAB and AEG also cause neurotoxic effects in vivo is of great interest, as is the question of whether they interact to enhance toxicity. Here, we evaluate the toxic and neurotoxic effects of these cyanotoxins alone or in combination by measuring zebrafish larval viability and behavior after exposure. 2,4-DAB was the most potent cyanotoxin as it decreased larval viability by approximately 50% at 6 days post fertilization, while BMAA and AEG decreased viability by just 16% and 8%, respectively. Although we only observed minor neurotoxic effects on spontaneous locomotion, BMAA and AEG enhanced acoustic startle sensitivity, and they interacted in an additive manner to exert their effects. 2,4-DAB, however, only modulated the startle kinematics, an indication of motor dysfunction. To investigate the mechanisms of 2,4-DAB's effects, we analyzed the protein profile of larval zebrafish exposed to 500 uM 2,4-DAB at two time points and identified molecular signatures consistent with neurodegeneration, including disruption of metabolic pathways and downregulation of the ALS-associated genes SOD1 and UBQLN4. Together, our data demonstrate that BMAA and its isomers AEG and 2,4-DAB cause neurotoxic effects in vivo, with 2,4-DAB as the most potent of the three in the zebrafish model.

scholarly journals BMAA and MCLR interact to modulate behavior and exacerbate molecular changes related to neurodegeneration in larval zebrafish

2020 ◽  
Author(s):  
Rubia M Martin ◽  
Michael S Bereman ◽  
Kurt C Marsden
Keyword(s):  
Acoustic Startle ◽  
Fold Increase ◽  
Health Concern ◽  
P Value ◽  
Dependent Manner ◽  
Zebrafish Model ◽  
Molecular Changes ◽  
Larval Zebrafish ◽  
Morphological Defects

Abstract Exposure to toxins produced by cyanobacteria (i.e., cyanotoxins) is an emerging health concern due to their increasing prevalence and previous associations with neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). The objective of this study was to evaluate the neurotoxic effects of a mixture of two co-occurring cyanotoxins, β-methylamino-L-alanine (BMAA) and microcystin leucine and arginine (MCLR), using the larval zebrafish model. We combined high-throughput behavior-based toxicity assays with discovery proteomic techniques to identify behavioral and molecular changes following 6 days of exposure. While neither toxin caused mortality, morphological defects, or altered general locomotor behavior in zebrafish larvae, both toxins increased acoustic startle sensitivity in a dose-dependent manner by at least 40% (p < 0.0001). Furthermore, startle sensitivity was enhanced by an additional 40% in larvae exposed to the BMAA/MCLR mixture relative to those exposed to the individual toxins. Supporting these behavioral results, our proteomic analysis revealed a 4-fold increase in the number of differentially expressed proteins (DEPs) in the mixture-exposed group. Additionally, prediction analysis reveals activation and/or inhibition of 8 enriched canonical pathways (enrichment p-value < 0.01; z-score ≥|2|), including ILK, Rho Family GTPase, RhoGDI, and calcium signaling pathways, which have been implicated in neurodegeneration. We also found that expression of TDP-43, of which cytoplasmic aggregates are a hallmark of ALS pathology, was significantly upregulated by 5.7-fold following BMAA/MCLR mixture exposure. Together, our results emphasize the importance of including mixtures of cyanotoxins when investigating the link between environmental cyanotoxins and neurodegeneration as we reveal that BMAA and MCLR interact in vivo to enhance neurotoxicity.

scholarly journals Exposure to a mixture of BMAA and MCLR synergistically modulates behavior in larval zebrafish while exacerbating molecular changes related to neurodegeneration

2020 ◽  
Author(s):  
Rubia M. Martin ◽  
Michael S. Bereman ◽  
Kurt C. Marsden
Keyword(s):  
Acoustic Startle ◽  
Fold Increase ◽  
Health Concern ◽  
P Value ◽  
Dependent Manner ◽  
Zebrafish Model ◽  
Molecular Changes ◽  
Larval Zebrafish ◽  
Morphological Defects

AbstractExposure to toxins produced by cyanobacteria (i.e., cyanotoxins) is an emerging health concern due to their increased occurrence and previous associations with neurodegenerative disease including amyotrophic lateral sclerosis (ALS). The objective of this study was to evaluate the neurotoxic effects of a mixture of two co-occurring cyanotoxins, β-methylamino-L-alanine (BMAA) and microcystin leucine and arginine (MCLR), using the larval zebrafish model. We combined high-throughput behavior based toxicity assays with discovery proteomic techniques to identify behavioral and molecular changes following 6 days of exposure. While neither toxin caused mortality, morphological defects, or altered general locomotor behavior in zebrafish larvae, both toxins increased acoustic startle sensitivity in a dose-dependent manner by at least 40% (p<0.0001). Furthermore, startle sensitivity was enhanced by an additional 40% in larvae exposed to the BMAA/MCLR mixture relative to those exposed to the individual toxins. Supporting these behavioral results, our proteomic analysis revealed a 4-fold increase in the number of differentially expressed proteins (DEPs) in the mixture exposed group. Additionally, prediction analysis reveals activation and/or inhibition of 8 enriched canonical pathways (enrichment p-value<0.01; z-score≥|2|), including ILK, Rho Family GTPase, RhoGDI, and calcium signaling pathways, which have been implicated in neurodegeneration. We also found that expression of TDP-43, of which cytoplasmic aggregates are a hallmark of ALS pathology, was significantly upregulated by 5.7-fold following BMAA/MCLR mixture exposure. Together, our results emphasize the importance of including mixtures of cyanotoxins when investigating the link between environmental cyanotoxins and neurodegeneration as we reveal that BMAA and MCLR interact in vivo to enhance neurotoxicity.

scholarly journals A Larval Zebrafish Model for Assessing Hypoxic‐Induced In Vivo Cardiomyocyte Damage: Time Course for Induction and Cardiac Output Recovery

2020 ◽  
Vol 34 (S1) ◽  
pp. 1-1
Author(s):  
Warren Burggren ◽  
Regina Abramova ◽  
Naim Bautista ◽  
Regina Fritsche Danielson ◽  
Avi Gupta ◽  
...  
Keyword(s):  
Cardiac Output ◽  
Time Course ◽  
Zebrafish Model ◽  
Larval Zebrafish

Abstract 712: Screening of Thrombolytic Agents in vivo Using a Novel Zebrafish Model of Ischemic Stroke Induced by Photochemical Thrombosis

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Jia-Wen Hsu ◽  
I-Ju Lee ◽  
Wei-Tien Chang ◽  
Yung-Jen Chuang ◽  
Ian Liau
Keyword(s):  
Blood Flow ◽  
Ischemic Stroke ◽  
Blood Vessel ◽  
Acute Ischemic Stroke ◽  
Zebrafish Model ◽  
Thrombolytic Activity ◽  
Larval Zebrafish ◽  
Thrombolytic Agents ◽  
Partially Occluded

Background: Timely thrombolytic therapy for acute ischemic stroke is important for improving neurological prognosis. Screening of thrombolytic agents on contemporary animal models of focal ischemic stroke is generally difficult as they require sophisticated surgical procedures such as induction of cerebral artery occlusion through mechanical ligation. Herein we seek to develop a novel zebrafish (Danio rerio) model of acute ischemic stroke for screening of thrombolytic drugs. Methods: All experiments were performed on a modified confocal optical microscope, which allows the induction of thrombosis at a selected blood vessel of larval zebrafish and the imaging of thrombotic and thrombolytic processes in real time. To initiate photochemical thrombosis, a 532-nm laser was focused at a blood vessel of a larva (4 dpf) that had been injected with a photosensitizer (rose bengal). To test the thrombolytic activity of tPA, we injected tPA to the blood vessel of a larval with a thrombus that partially occluded the blood flow. Results: Through photochemical means, we induced endothelial injury at selected blood vessel which subsequently triggered thrombosis. We show that an occlusion at the 1st branch of central artery drastically diminished the hemodynamics and cardiac function of larval zebrafish and impaired their capability to maintain balance during swimming whereas that at the basilar artery resulted in a high death rate. Immunofluorescent imaging shows that the photochemically induced thrombus comprised fibrins and platelets. After the injection of tPA to a larval with a partially occluded blood vessel, the fibrin mesh on the thrombus appeared sparse and limp which eventually led to the restoration of blood flow. Conclusions: Our zebrafish model of ischemic stroke is convenient to adapt and is highly reproducible. This model can potentially become an effective platform that benefits the screening of thrombolytic agents and the optimization of their dose.

scholarly journals Kaempferol-3-O-Glucuronide Ameliorates Non-Alcoholic Steatohepatitis in High-Cholesterol-Diet-Induced Larval Zebrafish and HepG2 Cell Models via Regulating Oxidation Stress

Life ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 445
Author(s):  
Yang Deng ◽  
Ji Ma ◽  
Xin Weng ◽  
Yuqin Wang ◽  
Maoru Li ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Lipid Accumulation ◽  
Hepg2 Cells ◽  
High Cholesterol Diet ◽  
Cholesterol Diet ◽  
High Cholesterol ◽  
Zebrafish Model ◽  
Alcoholic Fatty Liver ◽  
Larval Zebrafish

NAFLD (non-alcoholic fatty liver disease) is one of the most prominent liver diseases in the world. As a metabolic-related disease, the development of NAFLD is closely associated with various degrees of lipid accumulation, oxidation, inflammation, and fibrosis. Ilex chinensis Sims is a form of traditional Chinese medicine which is used to treat bronchitis, burns, pneumonia, ulceration, and chilblains. Kaempferol-3-O-glucuronide (K3O) is a natural chemical present in Ilex chinensis Sims. This study was designed to investigate the antioxidative, fat metabolism-regulating, and anti-inflammatory potential of K3O. A high-cholesterol diet (HCD) was used to establish steatosis in larval zebrafish, whereby 1mM free fatty acid (FFA) was used to induce lipid accumulation in HepG2 cells, while H2O2 was used to induce oxidative stress in HepG2. The results of this experiment showed that K3O reduced lipid accumulation and the level of reactive oxygen species (ROS) both in vivo (K3O, 40μM) and in vitro (K3O, 20μM). Additionally, K3O (40μM) reduced neutrophil aggregation in vivo. K3O (20μM) also decreased the level of malondialdehyde (MDA) and significantly increased the level of glutathione peroxidase (GSH-px) in both the HCD-induced larval zebrafish model and H2O2-exposed HepG2 cells. In the mechanism study, keap1, nrf2, tnf-α, and il-6 mRNA were all significantly reversed by K3O (20μM) in zebrafish. Changes in Keap1 and Nrf2 mRNA expression were also detected in H2O2-exposed HepG2 cells after they were treated with K3O (20μM). In conclusion, K3O exhibited a reduction in oxidative stress and lipid peroxidation, and this may be related to the Nrf2/Keap1 pathway in the NAFLD larval zebrafish model.

Exposure of a cryptic Hsp70 binding site determines the cytotoxicity of the ALS-associated SOD1-mutant A4V

2019 ◽  
Vol 32 (10) ◽  
pp. 443-457 ◽  
Author(s):  
Filip Claes ◽  
Stanislav Rudyak ◽  
Angela S Laird ◽  
Nikolaos Louros ◽  
Jacinte Beerten ◽  
...  
Keyword(s):  
Binding Site ◽  
Motor Neurons ◽  
Zebrafish Model ◽  
Surveillance Network ◽  
Denatured State ◽  
Inclusion Body Formation ◽  
Sod1 Mutant ◽  
Lateral Sclerosis

Abstract The accumulation of toxic protein aggregates is thought to play a key role in a range of degenerative pathologies, but it remains unclear why aggregation of polypeptides into non-native assemblies is toxic and why cellular clearance pathways offer ineffective protection. We here study the A4V mutant of SOD1, which forms toxic aggregates in motor neurons of patients with familial amyotrophic lateral sclerosis (ALS). A comparison of the location of aggregation prone regions (APRs) and Hsp70 binding sites in the denatured state of SOD1 reveals that ALS-associated mutations promote exposure of the APRs more than the strongest Hsc/Hsp70 binding site that we could detect. Mutations designed to increase the exposure of this Hsp70 interaction site in the denatured state promote aggregation but also display an increased interaction with Hsp70 chaperones. Depending on the cell type, in vitro this resulted in cellular inclusion body formation or increased clearance, accompanied with a suppression of cytotoxicity. The latter was also observed in a zebrafish model in vivo. Our results suggest that the uncontrolled accumulation of toxic SOD1A4V aggregates results from insufficient detection by the cellular surveillance network.

scholarly journals Efficacy of Voriconazole against Aspergillus fumigatus Infection Depends on Host Immune Function

2019 ◽  
Vol 64 (2) ◽  
Author(s):  
Emily E. Rosowski ◽  
Jiaye He ◽  
Jan Huisken ◽  
Nancy P. Keller ◽  
Anna Huttenlocher
Keyword(s):  
Aspergillus Fumigatus ◽  
Fungal Growth ◽  
Fungal Spore ◽  
Antifungal Drugs ◽  
Zebrafish Model ◽  
Content Type ◽  
Larval Zebrafish ◽  
Number Of Patients

ABSTRACT Antifungal therapy can fail in a remarkable number of patients with invasive fungal disease, resulting in significant morbidity worldwide. A major contributor to this failure is that while these drugs have high potency in vitro, we do not fully understand how they work inside infected hosts. Here, we used a transparent larval zebrafish model of Aspergillus fumigatus infection amenable to real-time imaging of invasive disease as an in vivo intermediate vertebrate model to investigate the efficacy and mechanism of the antifungal drug voriconazole. We found that the ability of voriconazole to protect against A. fumigatus infection depends on host innate immune cells and, specifically, on the presence of macrophages. While voriconazole inhibits fungal spore germination and growth in vitro, it does not do so in larval zebrafish. Instead, live imaging of whole, intact larvae over a multiday course of infection revealed that macrophages slow down initial fungal growth, allowing voriconazole time to target and kill A. fumigatus hyphae postgermination. These findings shed light on how antifungal drugs such as voriconazole may synergize with the immune response in living hosts.

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