No description available.

Cytokines are key regulators of the immune system and play a central role in the resistance to all infectious diseases. In the present thesis, and with the aim to determine the cytokine expression in response to Photobacterium damsefae ssp. piscicid6. (Phdp) exposure, sea bass TNF-a and IL-12 molecules were isolated. The molecular characterization of sea bass TNF.a showed that the putative protein conserves the TNF-a family signature, as well as the two cysteines usually involved in the formation of a disulfide bond. The mouse TNF-a Thr-Leu cleavage sequence and a potential transmembrane domain were also found, suggesting that sbTNF-a exists as a membrane bound form and a soluble protein. The single copy sbTNF-a gene contains a four exon-three intron structure similar to other known TNF a genes. Homology modelling of sbTNF-a is compatible with the trimeric quaternary architecture of its mammalian counterparts. The two subunits, IL12 p40 and IL-12 p35, that constitute the biological active IL-12 were also cloned and sequenced in sea bass. Sea bass IL-12 p40 and p35 conserve most cysteines involved in the intra-chain disulfide bonds of human IL-12 subunits as well as the important structural residues for human IL-12 heterodimerization. The gene organization of sea bass IL-12 p40 is si~ilar to the human orthologue, whilst the sea bass IL12 p35 gene structure, as reported fur pufferfish, differs from the human one in containing an additional exon and lacking a second copy of a duplicated exon present in the mammalian genes. Three differentially regulated IL-12 p40 genes (IL-12 p40 a, b and c) had been previously reported in bony fishes. In this thesis, the analysis of pufferfish and zebrafish genomes shows that two IL12 p35 genes (IL-12 p35 a and b) are present in these species. Furthermore, the synteny and phylogenetic analysis of pufferfish and zebrafish putative IL12 p40 and p35 genes showed that these are duplicated genes likely generated by the WGO in the ray-finned fish lineage. Sea bass IL-1, TNF-a, IL-10 and IL.12 expression profiles were then investigated in non-stimulated conditions and in response to Phdp or Phdp extracellular products. These studies contributed to the understanding of cytokine expression in sea bass but also to the finding that Phdp is able to inhibit IL-12 p40 expression most probably by a secreted protein. This raises the question about how IL-12 might influence the resistance to pasteurellosis, a disease caused by Phdp, and that induces high mortalities in marine fish farms. The promoters· of sea bass IL-10 and IL12 subunits were also sequenced and the presence of potential cis-elements discussed. The final experimental. chapter of this thesis describes the production of the recombinant sea bass TNF-a and IL-12 proteins, providing goOd tools for further studies on the immunobiology of these cytokines.

Diets containing high levels of raw kidney beans (Phaseolus vulgaris) are toxic to several species of animals. The main toxic component is a glycoprotein lectin. In nitrogen balance studies the inclusion of raw beans in an otherwise nutritionally adequate diet for rats increased both faecal and urinary losses of nitrogen and resulted in a negative nitrogen balance for the animals. The increased faecal nitrogen loss was due to the binding of the lectin to the luminal surfaces of enterocytes causing a severe disruption of the brush border of the proximal small intestine and resulting in an interference with digestion and absorption of dietary nitrogen. In addition, local hypersensitivity reactions, possibly involving lectin-specific IgE, might cause a stimulation of mucus secretion and an increase in plasma protein flow into the lumen. This would also contribute to the increased endogenous faecal nitrogen loss. The increased output of urinary nitrogen, mainly as urea, however, indicates the existence of systemic catabolic effects. Kidney bean lectins are resistant to intestinal proteolysis and a relatively large proportion of the lectin (1-5%) was shown to be transported across the intestinal barrier, possibly through a specific uptake mediated by receptors on the surface of the enterocyte. Circulating antibodies to the lectin (IgG class) were detected in the serum of bean-fed rats by an enzyme-linked immunosorbant assay. It is suggested that this systemic uptake of lectin and the ensuing immune response could result in hypersensitivity reactions (Type III) and could also affect the functioning of various cells by disrupting their normal metabolism.

No description available.

The effect of xenobiotic compounds on drug metabolising enzymes was assessed in ruminants; the development of an <i>in vivo </i>approach using a drug cocktail to measure specific enzyme families of phase I and II metabolism has been developed. Predictive clearances based on <i>in vitro </i>data from tolbutamide, phenacetin, chlorzoxazone, midazolam and oxazepam incubations were correlated with clearance determined <i>in vivo </i>(R²=<i> </i>0.695). A drug cocktail approach was used to assess the effect of plant secondary compounds on phase I and II enzymatic activities. This approach showed that phase I enzymes, e.g. CYP3A, can be affected by plant compounds, such as α-pinene or pyrrolizidine alkaloids. Additionally, due to carry over from one treatment period to the next, this experiment showed that interaction between plant secondary compounds can alter the overall effect of a single compound. Subsequently, <i>in vitro </i>experiments with hepatocytes or rumen microorganisms from sheep or goats, showed that pyrrolizidine alkaloids were not completely degraded in rumen fluid after 48h. Dose- and time-dependent toxicity of pyrrolizidine alkaloids to hepatocytes were observed. The magnitude of toxicity measured in goat and sheep hepatocytes was compatible with <i>in vivo </i>observations. Interaction between plant compounds observed <i>in vivo </i>was confirmed by <i>in vitro </i>assays. This concept of interaction between plant compounds was used to decrease the toxicity of pyrrolizidine alkaloids to hepatocytes. Exposure to certain plant compounds prior to the potentially toxic compounds could decrease pyrrolizidine alkaloid toxicity by acting on phase I or II enzymes. The mechanism of protection appears to be different between sheep and goats.

Otolith increments are used extensively to gain information on age and growth of fish. This is based on two fundamental assumptions. First, that the primary increments are produced at a known and constant periodically, and secondly, that the distance between the consecutive otolith increments is proportional to the growth of the fish. This study proposes to investigate the validity of the second assumption on a daily and weekly scale. There was a significant correlation between otolith size and fish size in Atlantic salmon (<I>Salmo solar</I> L.) and haddock (<I>Melanogrammus aeglefinus</I> L.). However, no correlation was found between somatic growth and otolith growth in individually measured yolk-sac haddock larvae or demersal juvenile haddock. This supports the suggestion that there is no functional relationship between otolith accretion rate and somatic growth rate. Different back-calculation models were employed to illustrate how this apparent 'uncoupling' of somatic growth and otolith growth can increase the error in size and age predictions. The possibility that there is a relationship between one or more components of metabolic rate and otolith accretion was investigated, in individual demersal juvenile haddock, Atlantic salmon, and pike (<I>Esox, lucius</I> L.). Variations in otolith daily increment width and somatic growth were compared with experimentally induced variations in resting metabolic rate, specific dynamic action (the increase in metabolism during digestion) and activity metabolism. Salmon and haddock were subjected to acute variations in temperature to vary resting metabolic rate. Pike were fed a single meal to elicit specific dynamic action. Activity levels in salmon were manipulated by exploiting the territoriality of salmon in a fluvial aquarium. An experimental change in resting metabolic rate resulted in a change in increment width that was uncoupled from somatic growth. Metabolic rate and increment width increased simultaneously after feeding in pike. In both experiments there was a less than isometric relationship between the two parameters. No correlation was found between increment width and the activity level of salmon.

In <i>Eledone cirrhosa</i>, egg development proceeds along typical caphalopod lines, but the egg stalks are surrounded by a ciliated columnar epithelium and the eggs go through a phase where the follicular layer forms a tri-lobed structure. Niether of these phenomena have been reported in other species and these specializations may faciliate the exchange of substances between egg and gonadial fluid. Potential fecundity (total number of eggs) is not correlated with either body size or gonadial maturity, but larger body size is associated with an increase in the proportion of eggs which have become vitellogenic (actual fecundity). Egg development is asynchronous and the ovaries of mature animals contain large numbers of immature eggs. Mean egg dry weight is strongly correlated with gonadosomatic index and ovarian maturity can be estimated from the mean dry weight of small biopsied egg samples. The usefulness of ovarian biopsy in <i>Eledone cirrhosa</i> is limited by the relatively short post-operative longevity of these animals. Nonetheless the technique has provided some interesting results and is likely to be even more effective when used with a hardier species. Food intake and gross growth efficiency are greatly reduced in mature females and this seems to be coupled with a decrease in the endocrine activity of the posterior salivary glands so that 5-hydroxytryptamine levels in the posterior salivary glands and blood fall dramatically during gonadial maturation. The decrease in glandular and circulatory levels of 5-hydroxytryptamine can be mimicked by enforced starvation. There is some evidence to suggest that both posterior salivary gland removal and starvation may cause precocious gonadial maturation. The genitoperiocardial ducts drain the ovary but do not resorb glucose or amino acid. Ovarian venous blood contains lower levels of lipid, glucose and amino acid than ovarian arterial blood and these differences between arterial and venous blood are more marked in maturing females.

A combination of techniques were used to sub-divide the auditory cortex of the ferret. The primary auditory cortex (AI) could be identified as an area of dense myelin, acetylcholinesterase and succinate dehydrogenase staining with high 2-deoxyglucose activity which could be reduced by ablation of the inferior colliculus and which contained three callosally labelled bands which ran in a dorsoventral direction orthogonal to the isofrequency contours. In addition, up to five secondary auditory fields were also identified: (1) A ventroposterior area (VP) which had dense myelin, acetylcholinesterase and succinate dehydrogenase staining, high 2-deoxyglucose activity which could be reduced by ablation of the inferior colliculus and was reciprocally connected to AI. (2) A ventroanterior area (VA) which had dense myelin staining, high 2-deoxyglucose activity which could be reduced by ablation of the inferior colliculus and was reciprocally connected to AI. (3) An area around the suprasylvian gyrus/suprasylvian sulcus (A) which had dense myelin and succinate dehydrogenase staining, high 2-deoxyglucose activity which may be reduced by ablation of the inferior colliculus and was reciprocally connected to AI. (4) An area on the gyrus dorsal to the suprasylvian gyrus (M) which had dense myelin staining and was reciprocally connected to AI. (5) An area ventral to VP (V) which was reciprocally connected to AI. The density of certain neurochemicals, thought to be involved in plasticity, was also studied in ferret AI during development.

No description available.

No description available.