LOCALIZATION, QUANTITATION, AND CHARACTERIZATION OF NEUROPEPTIDE-F-IMMUNOREACTIVE AND FMRFAMIDE-IMMUNOREACTIVE PEPTIDES IN TURBELLARIANS AND A MONOGENEAN - A COMPARATIVE-STUDY

Over the past decade it has become clear that the nervous systems of platyhelminths are both complex and highly developed, particularly in peptidergic elements. The central position of an ancestral flatworm in the evolution of the Bilateria has placed a greater importance on the study of modern flatworms. Using antisera generated to the C-terminal region of platyhelminth neuropeptide F and the molluscan neuropeptide, FMRFamide, in immunocytochemistry at both Light and ultrastructural levels, immunoreactivities have been localised within the nervous systems of three species of triclad turbellarians, Dugesia lugubris, Dendrocoelum lacteum, and Polycelis nigra, and one species of monogenean trematode, Diclidophora merlangi. Extensive immunostaining was obtained with both antisera throughout the central and peripheral nervous systems of all species studied, but intensity and abundance was significantly greater in the turbellarians. Indirect electron-immunogold labeling demonstrated that immunoreactivity to both neuropeptides was often colocalised in neurosecretory vesicles, although discrete populations of vesicles were also observed. Radioimmunoassay of extracts of all species confirmed that neuropeptide F immunoreactivity was consistently more abundant than FMRFamide immunoreactivity, and that the levels of both in the three turbellarians were several orders of magnitude greater than those found in the monogenean. Chromatographic analyses of turbellarian extracts revealed that neuropeptide F and FMRFamide immunoreactivities were attributable to different peptides. These data imply that the neuropeptidergic systems of turbellarians are considerably more extensive than those of monogeneans, and would suggest that a regression has occurred in the latter as a consequence of the adoption of a more sedentary parasitic lifestyle. (C) 1995 Wiley-Liss, Inc.

ELECTRON IMMUNOGOLD LABELING OF REGULATORY PEPTIDE IMMUNOREACTIVITY IN THE NERVOUS-SYSTEM OF MONIEZIA-EXPANSA (CESTODA, CYCLOPHYLLIDEA)

An electron immunogold-labeling technique was used in conjunction with a post-embedding procedure to demonstrate for the first time the ultrastructural distribution of the parasitic platyhelminth neuropeptide, neuropeptide F (NPF), in the nervous system of the cestode Moniezia expansa. Two axon types, distinguished by their populations of different-sized electron-dense vesicles, were identified. Immunogold labeling demonstrated an apparent homogeneity of PP, FMRFamide and NPF (M. expansa) antigenic sites throughout the larger dense-cored vesicles within the central nervous system. Triple labeling clearly demonstrated the co-localisation of immunoreactivities (IR) for NPF, PP and FMRFamide within the same dense-cored vesicles. The presence of NPF-IR within the vesicles occupying the perikaryon of the neuronal cell body indicated that the peptides had undergone post-translational C-terminal amidation prior to entering the axon. Antigen pre-absorption experiments using NPF prevented labeling with either PP or FMRFamide antisera, and the failure of these antisera to block NPF-IR supports the view that some, if not all, of the PP/FMRFamide-IR is due to NPF-like peptides.

NEUROPEPTIDE F-IMMUNOREACTIVITY IN THE MONOGENEAN PARASITE DICLIDOPHORA-MERLANGI

The localisation and distribution of neuropeptide F (NPF)-immunoreactivity (IR) in the monogenean fish-gill parasite, Diclidophora merlangi, have been investigated by whole-mount immunocytochemistry interfaced with confocal scanning laser microscopy and, at the ultrastructural level, by indirect immunogold labeling. Using antisera directed to intact synthetic NPF (Moniezia expansa, residues 1-39) or to the C-terminal decapeptide (residues 30-39) of synthetic NPF (M. expansa), immunostaining was found throughout the central (CNS) and peripheral nervous systems (PNS), including the innervation of the reproductive system. Immunoreactivity was found to be more intense using the antiserum to the C-terminal decapeptide fragment of NPF. At the subcellular level, gold labeling of NPF-IR was found exclusively over the contents of dense-cored vesicles that occupied nerve axons of both the CNS and the PNS. The distribution pattern of immunostaining for NPF mirrored exactly that previously documented for the vertebrate pancreatic polypeptide (PP) family of peptides and for FMRFamide. This finding and the results of preabsorption experiments strongly suggest that NPF is the predominant native neuropeptide in D. merlangi and that it accounts for most of the immunostaining previously obtained with PP and FMRFamide antisera.

Porcine Circovirus Type 2 Morphogenesis in a Clone Derived from the L35 Lymphoblastoid Cell Line

Porcine circovirus type 2 (PCV2) is the essential infectious agent of post-weaning multisystemic wasting syndrome (PMWS), one of the most important diseases of swine. Although several studies have described different biological properties of the virus, some aspects of its replication cycle, including ultrastructural alterations, remain unknown. The aim of the present study was to describe for the first time a complete morphogenesis study of PCV2 in a clone of the lymphoblastoid L35 cell line at the ultrastructural level using electron microscopy techniques. Cells were infected with PCV2 at a multiplicity of infection of 10 and examined at 0, 6, 12, 24, 48, 60 and 72 h post-infection. PCV2 was internalized by endocytosis, after which the virus aggregated in intracytoplasmic inclusion bodies (ICIs). Subsequently, PCV2 was closely associated with mitochondria, completing a first cytoplasmic phase. The virus entered the nucleus for replication and virus assembly and encapsidation occurred with the participation of the nuclear membrane. Immature virions left the nucleus and formed ICIs in a second cytoplasmic phase. The results suggest that at the end of the replication cycle (between 24 and 48 h), PCV2 was released either by budding of mature virion clusters or by lysis of apoptotic or dead cells. In conclusion, the L35-derived clone represents a suitable in-vitro model for PCV2 morphogenesis studies and characterization of the PCV2 replication cycle. (C) 2010 Elsevier Ltd. All rights reserved.

Development of an in vitro model for study of the efficacy of ischemic preconditioning in human skeletal muscle against ischemia-reperfusion injury

Ischemia-reperfusion (I/R) injury causes skeletal muscle infarction and ischemic preconditioning (IPC) augments ischemic tolerance in animal models. To date, this has not been demonstrated in human skeletal muscle. This study aimed to develop an in vitro model to investigate the efficacy of simulated IPC in human skeletal muscle. Human skeletal muscle strips were equilibrated in oxygenated Krebs-Henseleit-HEPES buffer (37 degrees C). Aerobic and reperfusion phases were simulated by normoxic incubation and reoxygenation, respectively. Ischemia was simulated by hypoxic incubation. Energy store, cell viability, and cellular injury were assessed using ATP, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), and lactate dehydrogenase (LDH) assays, respectively. Morphological integrity was assessed using electron microscopy. Studies were designed to test stability of the preparation (n = 5-11) under normoxic incubation over 24 h; the effect of 1, 2, 3, 4, or 6 h hypoxia followed by 2 h of reoxygenation; and the protective effect of hypoxic preconditioning (HPC; 5 min of hypoxia/5 min of reoxygenation) before 3 h of hypoxia/2 h of reoxygenation. Over 24 h of normoxic incubation, muscle strips remained physiologically intact as assessed by MTT, ATP, and LDH assays. After 3 h of hypoxia/2 h of reoxygenation, MTT reduction levels declined to 50.1 +/- 5.5% (P <0.05). MTT reduction levels in HPC (82.3 +/- 10.8%) and normoxic control (81.3 +/- 10.2%) groups were similar and higher (P <0.05) than the 3 h of hypoxia/2 h of reoxygenation group (45.2 +/- 5.8%). Ultrastructural morphology was preserved in normoxic and HPC groups but not in the hypoxia/reoxygenation group. This is the first study to characterize a stable in vitro model of human skeletal muscle and to demonstrate a protective effect of HPC in human skeletal muscle against hypoxia/reoxygenation-induced injury.

Ultrastructure of head organs (anterior adhesive apparatus) and posterior secretory systems of Caballeria liewi Lim, 1995 (Monogenea, Ancyrocephalidae)

Caballeria liewi Lim, 1995, uses adhesive secretions from the head organs and posterior secretory systems to assist in locomotion and attachment. Ultrastructural investigations show that the head organs of C. liewi consist of three pairs of antero-lateral pit-like openings bearing microvilli and ducts leading from two types of uninucleated gland cells (located lateral to the pharynx), one type producing rod-like (S1) bodies with an electron-dense matrix containing less electron-dense vesicles and the second type producing oval (S2) bodies with a homogeneous electron-dense matrix. Interlinking band-like structures are observed between S1 bodies and between S2 bodies. S1 body is synthesised in the granular endoplasmic reticulum, transported to a Golgi complex to be packaged into vesicles and routed into ducts for exudation. The synthesis of the S2 body is unresolved. Haptoral secretions manifested externally as net-like structures are derived from dual electron-dense (DED) secretory body produced in the peduncular gland cells. The DED body consists of a less electron-dense oval core in a homogeneous electron-dense matrix. On exocytosis into the pyriform haptoral reservoir, DED bodies are transformed into a secretion with two types of inclusions (less electron-dense oval and electron-dense spherical inclusions) in an electron-dense matrix. The secretions are further transformed (as small, oval, electron-dense bodies) when transported to the superficial anchor grooves, and on exudation into the gill tissues, the secretions become an electron-dense matrix. Secretory bodies associated with uniciliated structures, anchor sleeves and marginal hooks are also observed.

Synapsin- and actin-dependent frequency enhancement in mouse hippocampal mossy fiber synapses

The synapsin proteins have different roles in excitatory and inhibitory synaptic terminals. We demonstrate a differential role between types of excitatory terminals. Structural and functional aspects of the hippocampal mossy fiber (MF) synapses were studied in wild-type (WT) mice and in synapsin double-knockout mice (DKO). A severe reduction in the number of synaptic vesicles situated more than 100 nm away from the presynaptic membrane active zone was found in the synapsin DKO animals. The ultrastructural level gave concomitant reduction in F-actin immunoreactivity observed at the periactive endocytic zone of the MF terminals. Frequency facilitation was normal in synapsin DKO mice at low firing rates (approximately 0.1 Hz) but was impaired at firing rates within the physiological range (approximately 2 Hz). Synapses made by associational/commissural fibers showed comparatively small frequency facilitation at the same frequencies. Synapsin-dependent facilitation in MF synapses of WT mice was attenuated by blocking F-actin polymerization with cytochalasin B in hippocampal slices. Synapsin III, selectively seen in MF synapses, is enriched specifically in the area adjacent to the synaptic cleft. This may underlie the ability of synapsin III to promote synaptic depression, contributing to the reduced frequency facilitation observed in the absence of synapsins I and II.

Ultrastructure and phylogeny of peridinioid dinoflagellates

Os dinoflagelados são um grupo muito diverso de protistas que possuem um conjunto de características pouco comuns. Os peridinióides são dinoflagelados com teca que é formada por seis séries latitudinais de placas, incluindo a série cingular e um anel incompleto de placas intercalares anteriores, embora as últimas estejam ausentes em algumas espécies de Peridiniopsis. São dinoflagelados com simetria bilateral em relação ao plano apical que contem o eixo dorso-ventral. Na série sulcal há apenas uma placa posterior que contacta com o limite ventral de duas grandes placas antapicais. Entre os peridinióides, a presença ou ausência de um poro apical e o número de placas no cíngulo são geralmente consideradas marcas filogenéticas importantes ao nível de género ou família. Actualmente, a definição de Peridinium Ehrenberg, o dinoflagelado mais comum de água doce, inclui organismos com combinações diferentes destas duas características. Trabalhos anteriores sobre a ultrastrutura e afinidade filogenética das espécies tipo de Peridinium, P. cinctum, e Peridiniopsis Lemmermann, P. borgei também sugerem a necessidade de reexaminar as relações taxonómicas dos peridinióides. Esta tese combina o estudo ultrastrutural de uma selecção de espécies com hipóteses filogenéticas baseadas nas sequências de LSU rDNA, para aumentar o nosso conhecimento das diferenças e afinidades dentro dos peridinióides. Tem como objectivo aumentar o nosso conhecimento das características individuais das células que possam levar a reconhecer sinapomorfias que possam ser usadas como marcadores dos peridinióides como um todo e dos seus subgrupos. As espécies escolhidas para exame pormenorizado foram: Peridinium palatinum Lauterborn, de um grupo com duas placas intercalares anteriores, seis placas cingulares e sem poro apical; Peridinium lomnickii Wo!oszy"ska, de um grupo com poro apical, três placas intercalares e seis cingulares; Peridiniopsis berolinensis (Lemmermann) Bourrelly, uma espécie heterotrófica com poro apical, sem placas intercalares e com seis placas cingulares; e Sphaerodinium cracoviense Wo!oszy"ska, um membro de um género de formas com teca com um tipo de tabulação marginalmente peridinióide, com um suposto poro apical e quatro placas intercalares anteriores. Peridinium palatinum difere de Peridinium e Peridiniopsis típicos, quer em características da teca, quer internas. As diferenças estimadas entre as sequências parciais de LSU rDNA de P. palatinum e a espécie próxima P. pseudolaeve, relativamente a P. cinctum são comparativamente grandes e, juntamente com a topologia da árvore filogenética, apoiam a separação de P. palatinum e formas próximas ao nível de género. Palatinus nov. gen. foi, então, descrito com as novas combinações Palatinus apiculatus nov. comb. (espécie tipo; sin. Peridinium palatinum), P. apiculatus var. laevis nov. comb. e P. pseudolaevis nov. comb.. As características distintivas de Palatinus incluem uma superfície das placas lisa ou um tanto granulosa, mas não areolada, um grande pirenóide central penetrado por canais citoplasmáticos e de onde radiam lobos plastidiais, e a presença de uma fiada microtubular homóloga à de um pedúnculo. As células de Palatinus saem da teca pela zona antapicalpos- cingular. Peridinium lomnickii apresenta tabulação semelhante às formas marinhas, produtoras de quistos calcários, do género Scrippsiella A.R. Loeblich. Para comparação, adicionámos novas observações ultrastruturais de S. trochoidea. Peridinium lomnickii tem uma combinação de características diferente de Peridinium, Peridiniopsis e Scrippsiella. As hipóteses filogenéticas baseadas em DNA colocam P. lomnickii no mesmo ramo que Pfiesteria Steidinger et Burkholder, Tyrannodinium e outras Pfiesteriaceae, com as quais partilha um "microtubular basket" e uma ligação peculiar entre duas placas do sulco. As características distintivas do novo género proposto Chimonodinium gen. ined. incluem, além da tabulação, a ausência de pirenóides, a presença de um "microtubular basket" com quatro ou cinco fiadas sobrepostas de microtúbulos associados a um pequeno pedúnculo, um sistema pusular com tubos pusulares bem definidos ligados aos canais flagelares, e a produção de quistos não calcários. Peridiniopsis berolinensis partilha várias características significativas com Pfiesteria e afins, como um "microtubular basket" com a capacidade de suportar um tubo de alimentação, quimiossensibilidade para encontrar presas apropriadas, o modo de natação junto às presas e a organização geral da célula. Hipóteses filogenéticas com base em LSU rDNA confirmam a afinidade entre P. berolinensis e Pfiesteria bem como a relação mais remota com a espécie tipo de Peridiniopsis, P. borgei. Estas razões justificam a proposta de Tyrannodinium gen. nov., uma nova Pfiesteriaceae que difere de outros membros do grupo por viver em água doce e nos pormenores da tabulação. Sphaerodinium cracoviense revelou a tabulação típica do género Sphaerodinium, que apresenta um número de placas intercalares superiores e pos-cingulares maior que o que é típico em peridinióides: 4 e 6, respectivamente. Observações em SEM mostraram uma estrutura apical diferente da dos peridinióides, e um sulco apical numa das placas fazendo lembrar a área apical de alguns woloszynskióides. Os pormenores do aparelho flagelar e do sistema pusular ligam o Sphaerodinium aos woloszynskióides em geral e ao género Baldinia em particular, mas não aos peridinióides. O volumoso estigma de S. cracoviense revelou ser extraplastidial e de um modelo único, composto por elementos que se encontram em woloszynskióides, mas nunca encontrados anteriormente juntos. A análise filogenética baseada nas sequências parciais de LSU rDNA também sugerem uma maior proximidade de S. cracoviense com os woloszynskióides do que com os peridinióides. Futuras análises pormenorizadas de dinoflagelados peridinióides, em especial entre os do numeroso grupo de espécies com poro apical, serão necessárias para clarificar as suas relações taxonómicas; e a produção de descrições melhoradas das características finas particulares das células serão um requisito para perceber a evolução dos caracteres dos peridinióides por forma a podermos identificar marcadores filogenéticos.

Morphological and ecological studies on the phylum Cycliophora

Cycliophora é um filo animal descrito recentemente que acomoda, apenas, duas espécies: Symbion pandora Funch e Kristensen, 1995 e S. americanus Obst, Funch e Kristensen, 2006. Este filo é caracterizado por um ciclo de vida assaz complexo, cuja posição filogenética tem sido debatida desde a sua descoberta. Esta dissertação visa aprofundar o conhecimento geral existente acerca destes enigmáticos e pouco explorados metazoários. Assim, vários aspectos da morfologia e ecologia de ciclióforos foram estudados através de observações in vivo, técnicas de microscopia e reconstrução tridimensional. A mioanatomia de várias fases do ciclo de vida é descrita para S. pandora e S. americanus. Os nossos resultados revelam uma similaridade contundente entre a musculatura das duas espécies. A mioanatomia geral de Symbion é, ainda, comparada à de outros metazoários. A expressão de algumas substâncias imunorreactivas, como são exemplo a serotonina e as sinapsinas, é investigada em várias formas do ciclo de vida. Quando comparados com outros representantes de Spiralia, conclui-se que a neuroanatomia geral dos ciclióforos se assemelha mais às formas larvares do que aos adultos. Apesar de possuírem um plano corporal sofisticado, com extensas áreas ciliadas e uma mioanatomia complexa, descobrimos que o macho de ambas as espécies Symbion é composto por apenas algumas dezenas de células. Baseando-nos nestas observações, inferimos que a complexidade dos metazoários não se relaciona com o tamanho corporal nem com o número de células de um organismo. Estudos sobre a ultra-estrutura da fêmea revelaram, entre outras estruturas, um putativo poro genital, extensões citoplasmáticas do oócito e glândulas posteriores. Morfologia e implicações funcionais destas estruturas são aqui discutidas. A anatomia do protonefrídeo da larva cordóide é descrita. A arquitectura deste órgão diverge daquela presente noutros representantes de Nephrozoa, particularmente ao nível da área de filtração da célula terminal. As nossas observações são discutidas em termos filogenéticos. A maturação sexual em ciclióforos é investigada. Os nossos resultados sugerem que a transição de reprodução assexual a sexual se relacione com a idade da forma séssil, a “feeding stage”. A presença da larva Prometeus assente no tronco desta também poderá influenciar o processo, embora mais estudos sejam desejáveis para o comprovar. Os nossos resultados são discutidos integrativa e comparativamente com o conhecimento prévio sobre Cycliophora. A cumulação deste conhecimento será essencial para a compreensão da evolução e filogenia deste enigmático filo.

Citotoxicidade do ácido peracético: avaliação metabólica, estrutural e de morte em fibroblastos L929

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Données nouvelles sur l’innervation à dopamine du striatum et son co-phénotype glutamatergique

Une sous-population des neurones à dopamine (DA) du mésencéphale ventral du rat et de la souris étant connue pour exprimer l'ARN messager du transporteur vésiculaire 2 du glutamate (VGLUT2), nous avons eu recours à l'immunocytochimie en microscopie électronique, après simple ou double marquage de l'enzyme de synthèse tyrosine hydroxylase (TH) et de VGLUT2, pour déterminer la présence de l'une et/ou l'autre protéine dans les terminaisons (varicosités) axonales de ces neurones et caractériser leur morphologie ultrastructurale dans diverses conditions expérimentales. Dans un premier temps, des rats jeunes (P15) ou adultes (P90), ainsi que des rats des deux âges soumis à l'administration intraventriculaire cérébrale de la cytotoxine 6-hydroxydopamine (6-OHDA) dans les jours suivant la naissance, ont été examinés, afin d'étayer l'hypothèse d'un rôle de VGLUT2 au sein des neurones DA, au cours du développement normal ou pathologique de ces neurones. Chez le jeune rat, ces études ont montré: i) la présence de VGLUT2 dans une fraction importante des varicosités axonales TH immunoréactives du coeur du noyau accumbens ainsi que du néostriatum; ii) une augmentation de la proportion de ces terminaisons doublement marquées dans le noyau accumbens par suite de la lésion 6-OHDA néonatale; iii) le double marquage fréquent des varicosités axonales appartenant à l'innervation DA aberrante (néoinnervation), qui se développe dans la substance noire, par suite de la lésion 6-OHDA néonatale. Des différences significatives ont aussi été notées quant à la dimension des terminaisons axonales marquées pour la TH seulement, VGLUT2 seulement ou TH et VGLUT2. Enfin, à cet âge (P15), toutes les terminaisons doublement marquées sont apparues dotées d'une spécialisation membranaire synaptique, contrairement aux terminaisons marquées pour la TH ou pour VGLUT2 seulement. Dans un deuxième temps, nous avons voulu déterminer le devenir du double phénotype chez le rat adulte (P90) soumis ou non à la lésion 6-OHDA néonatale. Contrairement aux observations recueillies chez le jeune rat, nous avons alors constaté: i) l'absence complète de terminaisons doublement marquées dans le coeur du noyau accumbens et le néostriatum d'animaux intacts, de même que dans les restes de la substance noire des animaux 6-OHDA lésés; ii) une très forte baisse de leurnombre dans le coeur du noyau accumbens des animaux 6-OHDA lésés. Ces observations, suggérant une régression du double phénotype TH/VGLUT2 avec l'âge, sont venues renforcer l'hypothèse d'un rôle particulier d'une co-libération de glutamate par les neurones mésencéphaliques DA au cours du développement. Dans ces conditions, il est apparu des plus intéressants d'examiner l'innervation DA méso-striatale chez deux lignées de souris dont le gène Vglut2 avait été sélectivement invalidé dans les neurones DA du cerveau, ainsi que leurs témoins et des souris sauvages. D'autant que malgré l'utilisation croissante de la souris en neurobiologie, cette innervation DA n'avait jamais fait l'objet d’une caractérisation systématique en microscopie électronique. En raison de possibles différences entre le coeur et la coque du noyau accumbens, l'étude a donc porté sur les deux parties de ce noyau ainsi que le néostriatum et des souris jeunes (P15) et adultes (P70-90) de chaque lignée, préparées pour l'immunocytochimie de la TH, mais aussi pour le double marquage TH et VGLUT2, selon le protocole précédemment utilisé chez le rat. Les résultats ont surpris. Aux deux âges et quel que soit le génotype, les terminaisons axonales TH immunoréactives des trois régions sont apparues comparables quant à leur taille, leur contenu vésiculaire, le pourcentage contenant une mitochondrie et une très faible incidence synaptique (5% des varicosités, en moyenne). Ainsi, chez la souris, la régression du double phénotype pourrait être encore plus précoce que chez le rat, à moins que les deux protéines ne soient très tôt ségréguées dans des varicosités axonales distinctes des mêmes neurones DA. Ces données renforcent aussi l’hypothèse d’une transmission diffuse (volumique) et d’un niveau ambiant de DA comme élément déterminant du fonctionnement du système mésostriatal DA chez la souris comme chez le rat.

Récepteur EphA7 : expression régionale dans le cerveau et localisation ultrastructurale dans l’hippocampe chez le rat et la souris adultes

Bourse de maîtrise du Groupe de recherche sur le système nerveux central GRSNC, (2009,2010) Bourse d’études supérieures du Canada Frederick Banting et Charles Best, IRSC Instituts de recherche en santé du Canada, (2011)

Neuropathological changes in the brain of pigs with acute liver failure.

Abstract Objective. Cerebral edema is a serious complication of acute liver failure (ALF), which may lead to intracranial hypertension and death. An accepted tenet has been that the blood-brain barrier is intact and that brain edema is primarily caused by a cytotoxic etiology due to hyperammonemia. However, the neuropathological changes in ALF have been poorly studied. Using a well characterized porcine model we aimed to investigate ultrastructural changes in the brain from pigs suffering from ALF. Materials and methods. Sixteen female Norwegian Landrace pigs weighing 27-35 kg were randomised into two groups: ALF (n = 8) and sham operated controls (n = 8). ALF was induced with an end-to-side portacaval shunt followed by ligation of the hepatic arteries. Biopsies were harvested from three different areas of the brain (frontal lobe, cerebellum, and brain stem) following eight hours of ALF and analyzed using electron microscopy. Results. Profound perivascular and interstitial edema were found in all three areas. Disruption of pericytic and astrocytic processes were seen, reflecting breakdown/lesion of the blood-brain barrier in animals suffering from ALF. Furthermore, neurons and axons were edematous and surrounded by vesicles. Severe damage to Purkinje neuron (necrosis) and damaged myelin were seen in the cerebellum and brain stem, respectively. Biopsies from sham operated animals were normal. Conclusions. Our data support the concept that vasogenic brain edema plays an important role in the development of intracranial hypertension in pigs with ALF.

AFLATOXICOSIS AND ITS AMELIORATION IN BLACK TIGER SHRIMP,PENAEUS MONODON FABRICIUS

The present study was undertaken to elucidate the nutritional and pathological changes associated with aflatoxin B1 toxicity in Penaeus monodon and to determine the efficacy of vitamins E and K, and Amrita Bindu, herbal mixture in ameliorating the toxicity of AFB1. The main objectives the study is to document the pathological and immunological changes in P.monodon fed with AFB1 incorporated diets and to delineate the histological and ultrastructural changes and determine the presence of AFB1 residue in the shrimp body, to evaluate the growth performance of feed efficiency in P. monodon post larvae fed AFB1 added diets, to assess the interactive effect of heavy metals like copper and cadmium at sub-lethal levels in P. monodon postlarve fed AFB1 added diets, to decipher the ameliorative action of Vitamins E & K and a spicy herbal mixture, Amrita Bindu on AFB1 in P.monodon sub-adults. The study has revealed that Aflatoxin B1 significantly affects protein, lipid and carbohydrate metabolism in the shrimp penaeus monodon. The remarkable effect was observed in the immune system, as AFB1 has elevatod the immune response during initial days of exposure and prolonged exposure to the toxin leads to weakening of the animal’s immunity. Aflatoxin B1 level above 50 ppb severely affected the growth and feed utilization which in turn reflects the damage caused to the hepatopancreas as evident from the histological and ultrastructural observations.

Some studies on the reproductive endocrinology of tiger prawn Penaeus monodon Fabricius

The growing demand for quality prawn seed from the farmers‘ and entrepreneurs, coupled with uncertainity of their availability from nature at the appropriate time in required quantities has prompted‘ research on problems connected with prawn seed production. Endocrine control of reproduction in the penaeid shrimp _P_. monodon has been investigated in detail by adopting a comprehensive approach to the problem. The major aspects of the study included in depth investigations of the cytological details of the reproductive and neuroendocrine organs in correlation with the process of gonadal maturation. Based on the conclusions drawn from such ultrastructural studies various endocrine manipulations were carried out to see their effects on gonadal maturation. Besides that studies on the reproductive quality of male P_. monodon and the cryopreservation of spermatophores form a part of the present investigation. The shrimp 3; Inonodon used in the present study were collected from the offshore waters of Cochin, Madras and Mandapam and from the culture ponds of Vypeen Island near Cochin (Kerala) . The entire investigation on the cytological aspects were carried out using standard histological and electron-microscopic methods. Endocrine manipulations and cryopreservation studies is also carried out using the standard methods.