FISH

Author: MONROIG MARZÁ ÓSCAR.
Year: 2006.
University: VALENCIA [www.uv.es].
Place of defense: FACULTAD DE MATEMÁTICAS - UNIVERSITAT DE VALÈNCIA.
Place of preparation: INSTITUTO DE ACUICULTURA DE TORRE DE LA SAL (C.S.I.C.).

Summary: The liposomes are a versatile tool for the administration of nutrients into marine fish larvae due to its structure and wings nutritional properties of the components that make them up. So, liposomes can be made with water-soluble nutrients dissolved in the aqueous phase and soluble immersed in the environment lipofilico between chains 8cidos fatty of the phospholipids. The study presented here appreciates the use of these vesículas system as nutrient supply to marine fish larvae through two ways. The first of these possibilities posed by vehiculacion nutrient to fish larvae through the bioencapsulacion of liposomes rich in essential nutrients in Artemia nauplii, as happens with other commercial products, including emulsions lipídicas employed as a product enriching reference in this study. The second alternative is related to the administration directly by the larvae of these dives in suspensions liposomes rich in essential nutrients. This last strategy allows control of the composition of the diet and the transformation obvious that the dams alive Ilevan out about what components of liposomes when these are used as enriching. Using liposomes as eoriquecedores Artemia fortification dams are living in conditions that can limit the aggressive product performance enriching. The liposomes are no exception and, despite. Ias promising properties described his job as enriching requires preliminary studies designed to assess their real potential. Thus, Chapter 3 contains a study of three properties that characterize the liposomes and -adem8s, distinguish them from other products enriching. The properties selected were: retention capacity of water soluble substances in the aqueous phase of the internal vesícula, susceptibility to peroxidación lipid and distribution tamalios of vesículas. The liposomes, unlike the goticulas forming enriching products based emulsions, have a space aqueous allowing the inclusion of water-soluble compounds. Under experimental conditions that tried to emulate those inherent in enrichments of Artemia nauplii, liposomes liberated part of substances dissolved in its internal aqueous phase. This rate of liberation depends mostly on the composition of the membrane. Thus, the liposomes made with long-chain unsaturated phospholipids (excerpt from phospholipids krill) show a low capacity for retention of water soluble substances as a result of the fluidity of the membrane. This fluidity in liposomes highly unsaturated phospholipids as those formulated with krill (KPL), is reduced by the inclusion of cholesterol, registr8ndose an improvement in the retention of nutrients dissolved in the aqueous phase internally. The use of other sources fosfolipidicas more pure as soy lecithin (SPC), the egg (APC) and fosfolipido synthetic dipalmitoylphosphatidylcholine (DPPC) liposomes can be made better able to accommodate water-soluble molecules Among them, the preparation of liposomes unilamelares preparations the method solubilizacion by detergent (LUVdet) and made exclusively with soy phosphatidylcholine represents a formulation recommended a priori because it combines an ability to encapsulate such substances and a low cost economy in comparison to other phospholipids. As for the formation of peroxides, the enrichment process is carried out under conditions that promote lipid peroxidación of enriching. The study Ilevado out in conditions of enriching Artemia indicates that 105 liposomes composed of phospholipids with long chains of fatty acids and highly unsaturated are more susceptible to this process of degradation. However, the peroxidación experime 8 ntada in 1ff8 the emulsion trade exceeds two orders of magnitude to the extent liposomes for the same time of incubation. This can have serious implications on the larvae fed nauplii high in peroxides due to the toxicity exhibited by these compounds. Lastly, tamalio Particle is a feature of great importance in an enriching d prey alive, as it determines the effectiveness with which these incorporate the product. In terms of enrichment, 105 liposomes unilamelares not being altered their tamalio original when incubated in the absence of nauplii, while the presence of these involves the appearance of particles larger in some formulations of liposomes unilamelares prepared by extrusion (LUVext). In any case, increased tamalio of 105 liposomes should not mean a loss of enriching their skills, which is not always associated with some instability causing the evacuation of substances dissolved in the aqueous phase. Given the results found in the preliminary study of Chapter 3, using various types of liposomes to enrich Artemia nauplii in essential nutrients of various kinds (Chapters 4 and 5). The Capftulo 4 focuses attention on improving the content of essential fatty acids (EFAs) Artemia through the use of liposomes made with KPL rich in EFAs. To this end, there were five enrichment experiments aimed at determining what kind of vesícula more efficient and optimal conditions of use (of incubation time and how aeration, nauplii density, concentration of enriching and mode dose). The first experiment assessed the effectiveness of three types of liposomes (vesículas multilamelares, LUVext and LUVdet) made exclusively with K, PL to incorporate EFAs to 10 over time. The results indicate that 105 liposomes LUVdet reached levels of EFAs higher than 105 two other types of liposomes (vesículas multilamelares and LUVext), and moreover, matched the performance shown by a commercial emulsion with a higher content of polyunsaturated fatty acids long chain. Other experiments, led to the establishment of a protocol enrichment using liposomes, focuses on the use of this type of vesícula (LUVdet) used in incubaciones period of 21 h as optimal. The study on the effect of aeration used in enriching Artemia evidenced by the poor performance that 105 liposomes show in extreme conditions of aeration, both in terms of minimum contribution as when the turbulence of the medium is very high. As for the type of dissemination system, the use of systems with high capacity Diffuser (porous stone) improves enrichment nauplii with liposomes krill, but, in turn, is associated with massive mortandades nauplii. This phenomenon is manifest in a variety of experiments and appears to be motivated by the fact tensoactivo of 105 liposomes caused on the one hand, the formation of foam removed nauplii were middle and on the other, an effect of "Iavado" over 105 components fosfolipidicos of 105 tissue of 105 animals. In any case, this negative effect of using liposomes can be minimized by controlling the aeration during enrichment, as showed in the rest of experiments included in Chapter 4. They, moreover, just defining the parameters optimal enrichment Artemia with liposomes LUVdet krill. So, the protocol of use is resulting in an approximately 21 h of incubation, an aeration of 1.0 Ipm applied through hollow rods, a density naupliar of 300 ind ml "1 and a concentration of material enrich 0.50 g lipido 1 "1 administered in a single dose at the start of incubation. As discussed above, liposomes have a structure that allows for the inclusion, along with the phospholipids, other essential nutrients such as vitamins and amino acids. This possibility is examined in Chapter 5, which includes enrichment Artemia in vitamin A, vitamin C and methionine. The results reveal the ability of liposomes made with KPL to increase the content of retinol (vitamin A) of Artemia nauplii from retinol palmitate included in the liposome. This result indicates that the system enzimático of nauplio acts on the way up esterified into retinol. As for the vehiculacion of water-soluble nutrients, the experiment carried out show that liposomes unilamelares have a limited effectiveness in enriching enrichment in vitamin C as in the case of methionine. In both cases, the turbulence of the medium enrichment might limit the performance of the process. However, the liposomes multilamelares podrlan be more protected from this effect and, so, retaining the nutrient achieved dissolved in aqueous compartments between bicapas nearest to the nucleus, explaining This levels of methionine free nauplii found in fortified with this kind of vesículas and were higher than during nauplii incubated with liposomes unilamelares O methionine dissolved. The study on the use of liposomes as enriching of anemia complete fulfillment of larval feeding experiments. These tests demonstrate the possibility of using liposomes as nauplii enriched feed larvae bass and golden without being observed differences in survival and growth obtained with regard to treatment with a commercial product performance contrasted, addition, the results show the importance of controlling the biological parameters and fisico-qulmicos involved in the enrichment of nauplii of anemia, for obtaining an optimal performance in terms of incorporacion nutrients. This control parameters can reduce the intrinsic variability of the enrichment process prey alive, and consequently control the quality of diets, An alternative way to manage nutrients to fish larvae using liposomes is lainmersion these suspensions in liposomes. This approach is studied in experiments on bass and golden larvae in various stages of development (Chapter 7), The results suggest the possibility of hatching larvae of marine fish with liposomes made with essential nutrients of diverse nature without jeopardizing the survival of animals. However, the usefulness of this route of administration of nutrients has not been demonstrated despite the many mechanisms of interaction larva-Iiposoma involved These include entry through the ingestion of half made to meet osmorregulatorias, ingestion active capture of liposomes by the larvae and other mechanisms related to the properties of biocompatibility of liposomes (transfer intermembrana, fusion, "contacto-Iiberacion", etc..). The use of techniques marking nutrients, as well as the determination of indices nutritional condition that they find the nutritional benefits arising from the incorporacion of nutrients liposomated they are seen as future lines of investigation that must complete studies aqul presented.

Author: SANTOS ATIENZA EVA.
Year: 2006.
University: OVIEDO [www.uniovi.es].
Place of defense: FACULTAD DE BIOLOGIA.
Place of preparation: DEPARTAMENTO DE BIOLOGIA DE ORGANISMOS Y SISTEMAS.

Summary: The acute exposure to low pH can cause the death of fish because of their deleterious effects on respiration and ionic regulation. Subletalmente, acidification of the water causes changes in the distribution of energy, taking as a result a decrease in the growth and disruption of reproduction. The skin of fish is a metabolically active tissue that acts as a protective barrier between the environment and osmotic and internal fluids. The skin responds quickly to external changes such as cleaner or histopathological and ultrastructural changes in the concentration of ionic water, but the effects of acidification are not well known. The objective of this paper is to describe the structural changes in the epidermis and the upper stratum of the dermis of the tegument of rainbow trout (Oncorhynchus mykiss), as well as some changes enzyme, which is producencuando the fish is exposed to water ácida.La acidification water was reached gradually, in five hours by the addition of H2 S O4. A constant pH of 5.5 was maintained throughout the remainder of the experiment. Contact with the atmospheric CO2 has been avoided to eliminate its effect tamponador. The skin samples of fish were taken 0, 1, 2, 4, 6, 8, 10, 12 and 16 hours after reaching the pH constant (TPA: times postacidificación). Pieces of fabric were processed without scales of the dorsal region of the head of the fish for optical and electronic microscopy. The hydrolytic activity of peptidil-AMC was measured in excerpts from an essay by epidermal cells sensitive skin fluorescencia.La filed an initial hyperplastic response. Subsequently, the treated skin showed a significant loss of cohesion in the upper layers of the epidermis and appeared spaces acantolíticos. As a result there was an increase in cell detachment and scaling on the block. These changes resulted in a decrease in the thickness of the epidermis. There was an increase in the frequency of cell death processes in the cells filamentous surface. The number and size of the cells increased mucous first, then fell again and eventually increase. It was noted a decrease in the number of lymphocytes epidermal. Leukocytes infiltrators were found throughout the epidermis. Finally, the samples showed a partial recovery of the normal characteristics of the epidermis. The release of AMC in the epidermal extract was affected by the values of pH.Los results reveal sublethal effects of low pH in the skin and, subsequently, an increase in the susceptibility of fish to other agents. However, the conditions applied, the fish have shown the ability to regenerate its normal structure.

Author: Youssef Boshra Hani.
Year: 2006.
University: AUTÓNOMA DE BARCELONA [www.uab.es].
Place of defense: Facultad de Ciencias.
Place of preparation: Facultad de Ciencias.

Summary: Activation of the complement system results in the generation of anaphylatoxins C3a, C4a and C5a. In mammals, it has been shown that C3a and C5a elicit biological responses through their interactions with C3 and C5 receptors respectively. Although both receptors share a high degree of homology, until recently, very little was understood about the evolution of these receptors. In particular, it was not known whether or not these receptors were represented by a single common ancestral molecule in lower vertebrates. Here, we present the clonning and functional characterization of bona fise C3aR and C5aR in rainbow trout. While both receptors share a high degree of homology with their mammalian counterparts, the secondary structure of C3aR is unique in its truncated second extracellular loop. Antibodies generated against C3aR inhibited trout C3a-mediated intracellular calcium mobilization in trout leukocytes. While recombinantly expressed C5a was able to induce chemotaxis in trout peripheral leukocytes, antibodies generated against C5aR were found to inhibit this activity. Both C3aR and C5aR are expressed primary on B-cells, and to a lesser extent, on granulocytes. Our resluts indicate that, although highly homologous, the divergence of these two molecules into distinct receptors occurred prior to the emergence of teleost fish.