Zebrafish as a tool in Alzheimer's disease research

Alzheimer's disease is the most prevalent form of neurodegenerative disease. Despite many years of intensive research our understanding of the molecular events leading to this pathology is far from complete. No effective treatments have been defined and questions surround the validity and utility of existing animal models. The zebrafish (and, in particular, its embryos) is a malleable and accessible model possessing a vertebrate neural structure and genome. Zebrafish genes orthologous to those mutated in human familial Alzheimer's disease have been defined. Work in zebrafish has permitted discovery of unique characteristics of these genes that would have been difficult to observe with other models. In this brief review we give an overview of Alzheimer's disease and transgenic animal models before examining the current contribution of zebrafish to this research area. This article is part of a Special Issue entitled Zebrafish Models of Neurological Diseases.     

Zebrafish as a genomics model for human neurological and polygenic disorders

Whole exome sequencing and, to a lesser extent, genome-wide association studies, have provided unprecedented advances in identifying genes and candidate genomic regions involved in the development of human disease. Further progress will come from sequencing the entire genome of multiple patients and normal controls to evaluate overall mutational burden and disease risk. A major challenge will be the interpretation of the resulting data and distinguishing true pathogenic mutations from rare benign variants.While in model organisms such as the zebrafish,mutants are sought that disrupt the function of individual genes, human mutations that cause, or are associated with, the development of disease, are often not acting in a Mendelian fashion, are frequently of small effect size, are late onset, and may reside in noncoding parts of the genome. The zebrafish model is uniquely poised for understanding human coding- and noncoding variants because of its sequenced genome, a large body of knowledge on gene expression and function, rapid generation time, and easy access to embryos. A critical advantage is the ease of zebrafish transgenesis, both for the testing of human regulatory DNA driving expression of fluorescent reporter proteins, and the expression of mutated disease-associated human proteins in specific neurons to rapidly model aspects of neurological disorders. The zebrafish affords progress both through its model genome and it is rapidly developing transparent model vertebrate embryo.     

Proteostasis and neurodegeneration: The roles of proteasomal degradation and autophagy

All proteins in a cell continuously turn over, each at its own rate, contributing to a cell's development, differentiation, or aging. Of course, unnecessary protein(s), or those synthesized in excess, that hamper cellular homeostasis should be discarded rapidly. Furthermore, cells that have been subjected to various environmental stresses, e.g., reactive oxygen species (ROS) and UV irradiation, may incur various types of protein damage, which vitiate normal and homeostatic functions in the cell. Thereby, the prompt elimination of impaired proteins is essential for cell viability. This housekeeping is accomplished by two major catabolic routes—proteasomal digestion and autophagy. Strict maintenance of proteostasis is particularly important in non-proliferative cells, especially neurons, and it is plausible that its failure leads to a number of the neurodegenerative diseases becoming prominent in the growing elderly population. This article is part of a Special Issue entitled: Ubiquitin–Proteasome System. Guest Editors: Thomas Sommer and Dieter H. Wolf.     

Chronic expression of RCAN1-1L protein induces mitochondrial autophagy and metabolic shift from oxidative phosphorylation to glycolysis in neuronal cells

Expression of the RCAN1 gene can be induced by multiple stresses. RCAN1 proteins (RCAN1s) have both protective and harmful effects and are implicated in common human pathologies. The mechanisms by which RCAN1s function, however, remain poorly understood. We identify RCAN1s as regulators of mitochondrial autophagy (mitophagy) and demonstrate that induction of RCAN1-1L can cause dramatic degradation of mitochondria. The mechanisms of such degradation involve the adenine nucleotide translocator and mitochondrial permeability transition pore opening. We also demonstrate that RCAN1-1L induction can shift cellular bioenergetics from aerobic respiration to glycolysis, yet RCAN1-1L has very little effect on cell division, whereas it has a cumulative negative effect on cell survival. These results shed the light on mechanisms by which RCAN1s can protect or harm cells and by which they may operate in human pathologies. They also suggest that RCAN1s are important players in autophagy and such elusive phenomena as the mitochondrial permeability transition pore.     

Zebrafish as a model for infectious disease and immune function

The zebrafish, Danio rerio, has come to the forefront of biomedical research as a powerful model for the study of development, neurobiology, and genetics of humans. In recent years, use of the zebrafish system has extended into studies in behaviour, immunology and toxicology, retaining the concept that it will serve as a model for human disease. As one of the most thoroughly studied teleosts, with a wealth of genetic and genomic information available, the zebrafish is now being considered as a model for pathogen studies in finfishes. Its genome is currently being sequenced and annotated, and gene microarrays and insertional mutants are commercially available. The use of gene-specific knockdown of translation through morpholino oligonucleotides is widespread. As a result, several laboratories have developed bacterial and viral disease models with the zebrafish to study immune responses to infection. Although many of the zebrafish pathogen models were developed to address human infectious disease, the results of these studies should provide important clues for the development of effective vaccines and prophylactic measures against bacterial and viral pathogens in economically important fishes. In this review, the capabilities and potential of the zebrafish model system will be discussed and an overview of information on zebrafish infectious disease models will be presented.     

The role of autophagy in doxorubicin-induced cardiotoxicity

Doxorubicin (Dox) is an effective chemotherapeutic agent, however, its use is limited by cardiotoxicity. The mechanisms causing cardiotoxicity have not been clearly elucidated, but known to involve, at least in part, oxidative stress, mitochondrial dysfunction and apoptosis. More recently, it has been suggested that dysregulation of autophagy may also play an important role in Dox-induced cardiotoxicity. Autophagy has dual functions. Under physiological conditions, autophagy is essential for optimal cellular function and survival by ridding the cell of damaged or unwanted proteins and organelles. Under pathological conditions, autophagy may be stimulated in order to protect the cell from stress stimuli or, alternatively, to contribute to cell death. Thus, appropriate regulation of autophagy can be a matter of life or death. The role of autophagy in Dox-induced cardiotoxicity has recently been explored, however, conflicting reports on the effects of Dox on autophagy and its role in cardiotoxicity exist. Most, but not all, of the studies conclude that Dox upregulates cardiac autophagy and contributes to the pathogenesis of Dox-induced toxicity. Dox may induce autophagy by suppressing the expression of GATA4 and/or S6K1, which may directly or indirectly regulate expression of essential autophagy genes such as Atg12, Atg5, Beclin1 and Bcl-2. Interestingly, the Dox-induced autophagic response may be species specific as Dox treatment has been shown to stimulate autophagy in rat models, but suppress autophagy in mouse models. Additional studies will elucidate this possibility.     

Zebrafish as a Model to Assess the Teratogenic Potential of Nitrite

High nitrate levels in the environment may result in congenital defects or miscarriages in humans. Presumably, this is due to the conversion of nitrate to nitrite by gut and salivary bacteria. However, in other mammalian studies, high nitrite levels do not cause birth defects, although they can lead to poor reproductive outcomes. Thus, the teratogenic potential of nitrite is not clear. It would be useful to have a vertebrate model system to easily assess teratogenic effects of nitrite or any other chemical of interest. Here, we demonstrate the utility of zebrafish (Danio rerio) to screen compounds for toxicity and embryonic defects. Zebrafish embryos are fertilized externally and have rapid development, making them a good model for teratogenic studies. We show that increasing the time of exposure to nitrite negatively affects survival. Increasing the concentration of nitrite also adversely affects survival, whereas nitrate does not. For embryos that survive nitrite exposure, various defects can occur, including pericardial and yolk sac edema, swim bladder noninflation, and craniofacial malformation. Our results indicate that the zebrafish is a convenient system for studying the teratogenic potential of nitrite. This approach can easily be adapted to test other chemicals for their effects on early vertebrate development.     

Amino-terminal arginylation as a degradation signal for selective autophagy

The ubiquitin-proteasome system and the autophagy lysosome system are the two major protein degradation machineries in eukaryotic cells. These two systems coordinate the removal of unwanted intracellular materials, but the mechanism by which they achieve this synchronization is largely unknown. The ubiquitination of substrates serves as a universal degradation signal for both systems. Our study revealed that the amino-terminal Arg, a canonical N-degron in the ubiquitin-proteasome system, also acts as a degradation signal in autophagy. We showed that many ER residents, such as BiP, contain evolutionally conserved arginylation permissive pro-N-degrons, and that certain inducers like dsDNA or proteasome inhibitors cause their translocation into the cytoplasm where they bind misfolded proteins and undergo amino-terminal arginylation by arginyl transferase 1 (ATE1). The amino-terminal Arg of BiP binds p62, which triggers p62 oligomerization and enhances p62-LC3 interaction, thereby stimulating autophagic delivery and degradation of misfolded proteins, promoting cell survival. This study reveals a novel ubiquitin-independent mechanism for the selective autophagy pathway, and provides an insight into how these two major protein degradation pathways communicate in cells to dispose the unwanted proteins. [BMB Reports 2015; 48(9): 487-488]     

Zebrafish Jak2a plays a crucial role in definitive hematopoiesis and blood vessel formation

We examined the role of zebrafish (Danio rerio) Jak2a, a homolog of mammalian Jak2, in the developing embryo by injecting in vitro synthesized Jak2a shRNA into zebrafish zygotes. Blood circulation was suppressed in Jak2a shRNA-injected embryos from 24 hours post fertilization (hpf) and all embryos died with enlarged pericardium, shortened body lengths, and defects in some vasculature within 8 days post fertilization. O-dianisidine staining of red blood cells revealed normal blood island formation with no circulating red blood cells. As in Jak2^−/− transgenic mice, expression of definitive Ba1 globin was significantly reduced in Jak2a knockdown embryos at 36 hpf, whereas expression of other hematopoietic markers, primitive be1 globin, gata-1, and scl, were unaffected. More importantly, blood vessel formation was disturbed in Jak2a knockdown embryos as revealed by alkaline phosphatase staining at 72 hpf. Thus, our data indicate that zebrafish Jak2a is important in both definitive hematopoiesis and blood vessel formation.     

Enterovirus 71-induced autophagy increases viral replication and pathogenesis in a suckling mouse model

Background

We previously reported that Enterovirus 71 (EV71) infection activates autophagy, which promotes viral replication both in vitro and in vivo. In the present study we further investigated whether EV71 infection of neuronal SK-N-SH cells induces an autophagic flux. Furthermore, the effects of autophagy on EV71-related pathogenesis and viral load were evaluated after intracranial inoculation of mouse-adapted EV71 (MP4 strain) into 6-day-old ICR suckling mice.

Results

We demonstrated that in EV71-infected SK-N-SH cells, EV71 structural protein VP1 and nonstructural protein 2C co-localized with LC3 and mannose-6-phosphate receptor (MPR, endosome marker) proteins by immunofluorescence staining, indicating amphisome formation. Together with amphisome formation, EV71 induced an autophagic flux, which could be blocked by NH₄Cl (inhibitor of acidification) and vinblastine (inhibitor of fusion), as demonstrated by Western blotting. Suckling mice intracranially inoculated with EV71 showed EV71 VP1 protein expression (representing EV71 infection) in the cerebellum, medulla, and pons by immunohistochemical staining. Accompanied with these infected brain tissues, increased expression of LC3-II protein as well as formation of LC3 aggregates, autophagosomes and amphisomes were detected. Amphisome formation, which was confirmed by colocalization of EV71-VP1 protein or LC3 puncta and the endosome marker protein MPR. Thus, EV71-infected suckling mice (similar to EV71-infected SK-N-SH cells) also show an autophagic flux. The physiopathological parameters of EV71-MP4 infected mice, including body weight loss, disease symptoms, and mortality were increased compared to those of the uninfected mice. We further blocked EV71-induced autophagy with the inhibitor 3-methyladenine (3-MA), which attenuated the disease symptoms and decreased the viral load in the brain tissues of the infected mice.

Conclusions

In this study, we reveal that EV71 infection of suckling mice induces an amphisome formation accompanied with the autophagic flux in the brain tissues. Autophagy induced by EV71 promotes viral replication and EV71-related pathogenesis.     

Zebrafish disease models in hematology: Highlights on biological and translational impact

Zebrafish (Danio rerio) has proven to be a versatile and reliable in vivo experimental model to study human hematopoiesis and hematological malignancies. As vertebrates, zebrafish has significant anatomical and biological similarities to humans, including the hematopoietic system. The powerful genome editing and genome-wide forward genetic screening tools have generated models that recapitulate human malignant hematopoietic pathologies in zebrafish and unravel cellular mechanisms involved in these diseases. Moreover, the use of zebrafish models in large-scale chemical screens has allowed the identification of new molecular targets and the design of alternative therapies. In this review we summarize the recent achievements in hematological research that highlight the power of the zebrafish model for discovery of new therapeutic molecules. We believe that the model is ready to give an immediate translational impact into the clinic.     

Context-dependent role of ATG4B as target for autophagy inhibition in prostate cancer therapy

ATG4B belongs to the autophagin family of cysteine proteases required for autophagy, an emerging target of cancer therapy. Developing pharmacological ATG4B inhibitors is a very active area of research. However, detailed studies on the role of ATG4B during anticancer therapy are lacking. By analyzing PC-3 and C4-2 prostate cancer cells overexpressing dominant negative ATG4B^C74Ain vitro and in vivo, we show that the effects of ATG4B^C74A are cell type, treatment, and context-dependent. ATG4B^C74A expression can either amplify the effects of cytotoxic therapies or contribute to treatment resistance. Thus, the successful clinical application of ATG4B inhibitors will depend on finding predictive markers of response.     

Unfolding the role of autophagy in the cancer metabolism

Autophagy is considered an indispensable process that scavenges toxins, recycles complex macromolecules, and sustains the essential cellular functions. In addition to its housekeeping role, autophagy plays a substantial role in many pathophysiological processes such as cancer. Certainly, it adapts cancer cells to thrive in the stress conditions such as hypoxia and starvation. Cancer cells indeed have also evolved by exploiting the autophagy process to fulfill energy requirements through the production of metabolic fuel sources and fundamentally altered metabolic pathways. Occasionally autophagy as a foe impedes tumorigenesis and promotes cell death. The complex role of autophagy in cancer makes it a potent therapeutic target and has been actively tested in clinical trials. Moreover, the versatility of autophagy has opened new avenues of effective combinatorial therapeutic strategies. Thereby, it is imperative to comprehend the specificity of autophagy in cancer-metabolism. This review summarizes the recent research and conceptual framework on the regulation of autophagy by various metabolic pathways, enzymes, and their cross-talk in the cancer milieu, including the implementation of altered metabolism and autophagy in clinically approved and experimental therapeutics.     

ROS removal by DJ-1: Arabidopsis as a new model to understand Parkinson disease

Reactive oxygen species represent one of the principal factors that cause cell death and scavenging of reactive oxygen species by superoxide dismutase-related pathway is essential for cell survival. The Parkinson disease-related DJ-1 protein (also known as PARK7) has been implicated in resistance against oxidative stress in dopaminergic neurons however, its molecular mechanism has to date been unknown. We have used Arabidopsis thaliana as a model system to demonstrate that DJ-1, in both plant and mammalian cells, directly influence SOD activity in a highly conserved manner thereby preventing cell death. These data not only provides evidence for the molecular mechanisms associated with DJ-1-induced Parkinson disease but also highlight the unprecedented value of plants as a tool in understanding human disease mechanisms.Key words: DJ-1, stress, cell death, Parkinson disease, ArabidopsisReactive oxygen species (ROS) are involved in a myriad of fundamental biological processes including cell signaling and cellular defense pathways in plants and animals.^1–3 Despite its role as a signaling molecule, inappropriate and elevated levels of ROS have a major impact on the etiology of neurodegenerative diseases, such as for example Parkinson disease (PD), and in oxidative stress responses in plants. In general ROS can cause damage to DNA, lipids, proteins and various cofactors. During normal physiological conditions, when ROS are continuously generated, antioxidant defense systems are adequately equipped to prevent ROS-induced tissue dysfunction.^4,5 However upon elevated ROS generation the cellular antioxidant systems either recruit additional factors to minimize ROS-induced damage or cells suffer the consequences of cell death. Because of this dichotomy, where ROS plays a vital role during growth and development but can also have overwhelming damaging effects, it is clear that strict regulatory mechanisms need to be in place to effectively control ROS levels. In both plant and mammalian cells elevated ROS levels lead to cell death and in various human disease such as PD, Alzheimer disease, amyotrophic lateral sclerosis and Huntington disease proteins involved in stress-related pathways are often mutated.⁶In both plants and mammals mitochondria act as an important source of ROS however, plants also produce ROS in chloroplasts as part of photosynthetic activity. Combined with the fact that plants are sessile organisms it suggests that, although similar in nature, plants most probably have more complex antioxidant systems than other organisms.Strategies for removing excess ROS are similar in plants and humans. The principle ROS removal pathway involves superoxide dismutases (SOD) (or copper/zinc superoxide dismutase-CSD in plants), glutathione peroxidases (GPX) and catalases (CAT) localized in the cytosol, mitochondria and chloroplasts (Fig. 1). SOD converts superoxide anion to H₂O₂, which is then detoxified to H₂O by GPX and CAT. In Arabidopsis, besides SOD, GPX and CAT, there are five ascorbate peroxidases (APX) located in the cytosol and chloroplasts, involved in scavenging ROS generated during photosynthesis.^7,8 Therefore, SOD, GPX, CAT and APX, together with other auxiliary proteins, form the main line of defense against ROS.Open in a separate windowFigure 1Involvement of DJ-1-like proteins in ROS scavenging pathways. Produced O₂^- is converted to H₂O₂ by SOD in human or CSD (Arabidopsis SOD) in Arabidopsis. H₂O₂ is then converted to H₂O by GPX or CAT (catalase) in human or APX, GPX or CAT in Arabidopsis. DJ-1-like proteins interact with SOD or GPX in humans and CSD, GPX and APX in Arabidopsis. It is assumed here that DJ-1-like proteins may also interact with catalases (broken arrows).DJ-1 was originally identified as an oncogene and represents a ubiquitous redox-responsive cytoprotective protein with diverse functions where one of its main roles have been attributed to oxidative stress protection.⁹ Numerous studies have shown that several DJ-1 mutations in humans cause autosomal recessive, early onset PD however, its mode of action has been elusive in terms of having a direct influence on neuronal cell death.¹⁰ In an attempt to clarify the mechanism of DJ-1 we established Arabidopsis thaliana as a new and novel model system.¹¹ The Arabidopsis genome contains three DJ-1 homologs compared to the single DJ-1 locus found in humans and we showed that two of these (AtDJ-1b and AtDJ-1c) localizes to chloroplasts whilst one, AtDJ-1a, localizes to the cytosol and nucleus as observed for human DJ-1.¹¹ As mutated DJ-1 in mammals leads to cell death we identified and characterized a DJ-1 loss-of-function mutant which showed increased cell death in aging plants. Using Bimolecular Fluorescence Complementation (BiFC) and isothermal titration calorimetry (ITC) assays we showed that AtDJ-1a interacts with CSD1, the cytosolic SOD in Arabidopsis, and with human SOD1 in plant cells. Further we demonstrated that the human DJ-1 protein interacts with SOD1 in mammalian CHO cells.¹¹ Similar approaches were also employed to show that AtDJ-1a and human DJ-1 had an interaction with GPX2 in plant and mammalian cells.¹¹Enzyme assays revealed that AtDJ-1a and DJ-1 stimulated SOD/CSD1 activity and that only the copper-loaded forms of AtDJ-1a and DJ-1 had this effect suggesting that AtDJ-1a/DJ-1 may provide copper for SOD/CSD1.¹¹ Although the observed SOD activation provides clues towards the role of DJ-1 in detoxification of ROS, SOD only converts superoxide anion to H₂O₂ which must further be detoxified to H₂O by GPX and CAT. Although we showed that AtDJ-1a and human DJ-1 can interact with AtGPX2 and GPX2, respectively, we observed no changes in GPX2 activity upon DJ-1 interaction. The reason for this may be several-fold. First, cellular GPX2 activity levels may be sufficient to convert SOD-generated H₂O₂ to H₂O. Second, DJ-1 may indeed have no effect on GPX2 activity but simply act as an anchor to dock GPX2 in the vicinity of SOD. To test whether the DJ-1/SOD/GPX2 complex recruits other auxiliary proteins we have also shown that AtDJ-1a interacts with the Arabidopsis cytosolic APX1 protein (Fig. 2, unpublished data). It is also highly possible that DJ-1 interacts with catalase or at least influences its activity (Fig. 1). Although we have no data to date indicating a functional significance of the DJ-1/APX1 interaction we speculate that DJ-1 indeed acts as a scaffold protein bringing together SOD, GPX and possibly APX1 to mediate and control ROS scavenging, ultimately preventing oxidative stress-induced cell death (Fig. 3).Open in a separate windowFigure 2Interaction of AtDJ-1a with APX1. AtDJ-1a tagged with the N-terminal region of GFP and APX1 tagged with the C-terminal region of GFP gene were co-transformed into tobacco cells. The observed GFP signal in (B) demonstrates an AtDJ-1a/APX1 interaction through reconstitution of functional GFP molecules. (A) Negative control.Open in a separate windowFigure 3Working model of AtDJ-1a and DJ-1 mode of action. AtDJ-1a and DJ-1 interacts with SOD and GPX2 leading to SOD activation in a copper-dependent fashion. It is proposed that AtDJ-1a and DJ-1 delivers copper to SOD enhancing its activity whilst GPX2 is anchored by AtDJ-1 and DJ-1 to the protein complex to ensure conversion of the SOD-generated H₂O₂ to H₂O.The fact that Arabidopsis has three DJ-1 homologs where two of these, AtDJ-1b and AtDJ-1c, are localized to chloroplasts¹¹ underlines the protective role of DJ-1-like proteins during oxidative stress in plants. From our localization studies it appears that AtDJ-1b is localized to the chloroplast stroma whilst AtDJ-1c is localized to both the stroma and the thylakoid membranes (unpublished data). Whether AtDJ-1b and AtDJ-1c act in isolation or in concert and how these two proteins are involved in photosynthesis-induced ROS regulation is unclear but represent exciting future challenges.The notion that plants can be used as tools to increase our understanding of human disease mechanisms is somewhat obscure to the general scientific community. The fact remains that many discoveries with direct relevance to human health and disease have been elaborated using Arabidopsis, and several processes important to human biology are more easily studied in this versatile model plant.¹² The use of Arabidopsis to understand human disease states has several advantages: (1) Arabidopsis represents a well established model organism with a fully annotated genome, (2) The Arabidopsis genome contains homologs of numerous genes involved in human disease, (3) The identification and generation of Arabidopsis mutants is simple and requires little effort, (4) Arabidopsis growth and maintenance requires little infrastructure and running costs and (5) Arabidopsis research has few ethical constraints.Despite the advantages of Arabidopsis as a model system for elucidating human disease mechanisms it is important to appreciate that Arabidopsis and plant research in general can only reach its full potential in the field of medical research if combined with complementary, and perhaps more conventional, model systems.     

Lipid imbalance in the neurological disorder, Niemann-Pick C disease

Niemann-Pick C (NPC) disease is a progressive neurological disorder in which cholesterol, gangliosides and bis-monoacylglycerol phosphate accumulate in late endosomes/lysosomes. This disease is caused by mutations in either the NPC1 or NPC2 gene. NPC1 and NPC2 are involved in egress of lipids, particularly cholesterol, from late endosomes/lysosomes but the precise functions of these proteins are not clear. An important question regarding the function of NPC proteins is: why do mutations in these ubiquitously expressed proteins have such dire consequences in the brain? This review summarizes the roles of NPC proteins in lipid homeostasis particularly in the central nervous system.     

Protective role of cell division cycle 48 (CDC48) protein against neurodegeneration via ubiquitin-proteasome system dysfunction during zebrafish development

Cell division cycle 48 (CDC48), a ubiquitin-dependent molecular chaperone, is thought to mediate a variety of degradative and regulatory processes and maintain cellular homoeostasis. To investigate the protective function of CDC48 against accumulated ubiquitinated proteins during neurodevelopment, we developed an in vivo bioassay technique that detects expression and accumulation of fluorescent proteins with a polyubiquitination signal at the N terminus. When we introduced CDC48 antisense morpholino oligonucleotides into zebrafish embryos, the morphant embryos were lethal and showed defects in neuronal outgrowth and neurodegeneration, and polyubiquitinated fluorescent proteins accumulated in the inner plexiform and ganglion cell layers, as well as the diencephalon and mesencephalon, indicating that the degradation of polyubiquitinated proteins by the ubiquitin-proteasome system was blocked. These abnormal phenotypes in the morphant were rescued by CDC48 or human valosin-containing protein overexpression. Therefore, the protective function of CDC48 is essential for neurodevelopment.     

Zebrafish as a model for hearing and deafness

The zebrafish is an especially attractive model for the study of the development and function of the vertebrate inner ear. It combines rapid and accessible embryogenesis with a host of genetic and genomic tools for systematic gene discovery and analysis. A large collection of mutations affecting development and function of the ear and a related sensory system, the lateral line, have been isolated; several of these have now been cloned, and at least five provide models for human deafness disorders. Disruption of multiple genes, using both forward and reverse genetic approaches, has established key players--both signaling molecules and autonomous factors--responsible for induction and specification of the otic placode. Vestibular and auditory defects have been detected in adult animals, making the zebrafish a useful system in which to tackle the genetic causes of late onset deafness and vestibular disease.     

The Optokinetic Response as a Quantitative Measure of Visual Acuity in Zebrafish

Zebrafish are a proven model for vision research, however many of the earlier methods generally focused on larval fish or demonstrated a simple response. More recently adult visual behavior in zebrafish has become of interest, but methods to measure specific responses are new coming. To address this gap, we set out to develop a methodology to repeatedly and accurately utilize the optokinetic response (OKR) to measure visual acuity in adult zebrafish. Here we show that the adult zebrafish''s visual acuity can be measured, including both binocular and monocular acuities. Because the fish is not harmed during the procedure, the visual acuity can be measured and compared over short or long periods of time. The visual acuity measurements described here can also be done quickly allowing for high throughput and for additional visual procedures if desired. This type of analysis is conducive to drug intervention studies or investigations of disease progression.     

Zebrafish as a tool in Alzheimer's disease research

Alzheimer's disease is the most prevalent form of neurodegenerative disease. Despite many years of intensive research our understanding of the molecular events leading to this pathology is far from complete. No effective treatments have been defined and questions surround the validity and utility of existing animal models. The zebrafish (and, in particular, its embryos) is a malleable and accessible model possessing a vertebrate neural structure and genome. Zebrafish genes orthologous to those mutated in human familial Alzheimer's disease have been defined. Work in zebrafish has permitted discovery of unique characteristics of these genes that would have been difficult to observe with other models. In this brief review we give an overview of Alzheimer's disease and transgenic animal models before examining the current contribution of zebrafish to this research area. This article is part of a Special Issue entitled Zebrafish Models of Neurological Diseases.     

Zebrafish as a model for systems biology

Zebrafish offer a unique vertebrate model for research areas such as drug development, disease modeling and other biological exploration. There is significant conservation of genetics and other cellular networks among zebrafish and other vertebrate models, including humans. Here we discuss the recent work and efforts made in different fields of biology to explore the potential of zebrafish. Along with this, we also reviewed the concept of systems biology. A biological system is made up of a large number of components that interact in a huge variety of combinations. To understand completely the behavior of a system, it is important to know its components and interactions, and this can be achieved through a systems biology approach. At the end of the paper we present a concept of integrating zebrafish into the systems biology approach.