Suitability of Novel Algal Biomass as Fish Feed: Accumulation and Distribution of Omega-3 Long-Chain Polyunsaturated Fatty Acid in Zebrafish.

A 28-day feeding experiment with formulated feed using docosahexaenoic acid (DHA)-rich whole cells of freeze-dried marine microalgae Schizochytrium sp. to understand the distribution of fatty acids in a laboratory model zebrafish was conducted. Three feeds, commercial feed, 50:50 feed (50% commercial and 50% algae), and pure algae, were investigated. All feeds were consumed by zebrafish and showed optimal growth and weight gain with a survival rate of 100%. Lipids were extracted from four different tissues, brain, liver, muscle, and blood, to understand the distribution of fatty acids with respect to the feed. Maximum lipid was observed in zebrafish fed with 50:50 feed in all tissue samples. An increasing concentration of fatty acids was observed upon increasing the experimental time. Algae feed supported the DHA accumulation in all tissue samples compared to other feeds and resulted in an overall increment of polyunsaturated fatty acid content. To understand the role of fatty acids during zebrafish embryogenesis, eggs were collected at the end of the experiment and fatty acid content was analyzed. However, no significant difference was observed in fatty acid composition of embryos fed with algae. This provides a base for the understanding of fatty acid distribution in zebrafish with commercial and algae feeds and support the utilization of Schizochytrium biomass as a potential replacement for fishmeal.

Rapid quantification of neutral lipids and triglycerides during zebrafish embryogenesis.

The zebrafish is a useful vertebrate model to study lipid metabolism. Oil Red-O (ORO) staining of zebrafish embryos, though sufficient for visualizing the localization of triglycerides, was previously inadequate to quantify neutral lipid abundance. For metabolic studies, it is crucial to be able to quantify lipids during embryogenesis. Currently no cost effective, rapid and reliable method exists to quantify the deposition of neutral lipids and triglycerides. Thin layer chromatography (TLC), gas chromatography and mass spectrometry can be used to accurately measure lipid levels, but are time consuming and costly in their use. Hence, we developed a rapid and reliable method to quantify neutral lipids and triglycerides. Zebrafish embryos were exposed to Rimonabant (Rimo) or WIN 55,212-2 mesylate (WIN), compounds previously shown to modify lipid content during zebrafish embryogenesis. Following this, ORO stain was extracted out of both the zebrafish body and yolk sac and optical density was measured to give an indication of neutral lipid and triglyceride accumulation. Embryos treated with 0.3 microM WIN resulted in increased lipid accumulation, whereas 3 microM Rimo caused a decrease in lipid accumulation during embryogenesis. TLC was performed on zebrafish bodies to validate the developed method. In addition, BODIPY free fatty acids were injected into zebrafish embryos to confirm quantification of changes in lipid content in the embryo. Previously, ORO was limited to qualitative assessment; now ORO can be used as a quantitative tool to directly determine changes in the levels of neutral lipids and triglycerides.

The Use of the Zebrafish Model to Aid in Drug Discovery and Target Validation.

The zebrafish is fast becoming a leading and prominent model organism used by researchers for developmental biology, and research in modeling human diseases in zebrafish is being undertaken at a fast pace. Many therapeutic areas, including oncology and cardiovascular diseases to name a few all have zebrafish models based on known disease mechanisms that are translatable to modes of action in humans. Many novel assays have been and are continuing to be developed to study human disease in zebrafish. Prominent methods to identify novel drug targets within the organism include, chemical mutagenesis, insertional mutagenesis and high throughput small molecule screens. Methods to validate potential drug targets include reverse and forward genetics, transgenesis and gene knockout. This review summarizes the important contributions made using the zebrafish model in recent years to aid in drug discovery and target validations in the highly important medical field of cancer medicine, cardiovascular disease and the emerging role of the zebrafish model as a platform for toxicity screening.