Espécies do gênero Valeriana são tradicionalmente utilizadas para tratar ansiedade, irritabilidade e desordens de sono. A Organização Mundial da Saúde indica o uso de preparações farmacêuticas como alternativa aos benzodiazepínicos para tratamento da ansiedade e insônia. Entre as substâncias ativas presentes no gênero Valeriana destacam-se os óleos voláteis, valepotriatos, flavonóides e lignanas. Entretanto, estudos farmacológicos com produtos isolados são escassos. Espécies nativas de Valeriana, que ocorrem no Rio Grande do Sul, têm sido estudadas em relação a sua composição química, sendo a espécie Valeriana glechomifolia a que possui maior teor de valepotriatos. Neste estudo, diferentes métodos de extração de valepotriatos foram comparados e extratos enriquecidos em valepotriatos foram testados em modelos animais de sedação, ansiedade e depressão. Valepotriatos isolados foram submetidos a ensaios de ligação a receptores benzodiazepínico e serotonérgico (5HT1A) e ensaios de atividade da enzima Na+K+ATPase. A extração por fluido supercrítico com dióxido de carbono (SCCO2) foi realizada em temperatura constante de 40 oC e pressões variáveis (90, 120, 150 e 200 bar) e demonstrou maior teor de valepotriatos que os métodos de extração por maceração em ultrassom e maceração Entre as diferentes pressões de extração utilizadas no método de SCCO2, o maior teor de valepotriatos foi apresentado pela fração obtida na pressão de 90 bar. Todos os extratos mostraram o mesmo perfil qualitativo de valepotriatos. Flavonóides também foram obtidos através de maceração por ultrassom em metanol. A dose letal mediana dos valepotriatos obtidos por maceração por ultrassom foi de 42±3 mg/kg, i.p. Esta mesma solução extrativa, na dose de 10 mg/kg, v.o. (gavage), foi inefetiva no labirinto em cruz elevado e tempo de sono barbitúrico, sugerindo a ausência de propriedades ansiolíticas e hipnótico-sedativas. Resultados similares foram obtidos com a solução extrativa enriquecida em flavonóides. Valtrato, acevaltrato, 1-b-acevaltrato, diavaltrato e o flavonóide codificado como B6 não apresentaram ligação ao sítio benzodiazepínico do complexo receptor GABAA, nem ao receptor serotonérgico (5HT1A) viii na faixa de concentração de 1-100 μM. Os valepotriatos inibiram a atividade da enzima Na+K+-ATPase na faixa de concentração micromolar. A fração de valepotriatos obtida por SCCO2 (10 mg/kg, gavage) foi efetiva no teste da natação forçada, sem interferir na atividade locomotora espontânea, sugerindo uma atividade do tipo antidepressiva. Em conclusão, a extração por fluido supercrítico com dióxido de carbono mostrou-se eficiente para a obtenção de frações enriquecidas em valepotriatos e estas substâncias representam uma nova classe química com atividade inibitória não seletiva da enzima Na+K+ATPase, e com atividade do tipo antidepressiva. / Species of the genus Valeriana are traditionally used to treat anxiety, irritability and sleep disorders. The World Health Organization indicates pharmaceutical preparations of V. officinalis as an alternative to benzodiazepine drugs for treating anxiety and insomnia. Volatile oil, valepotriates, flavonoids and lignan have been suggested as active substances of the Valeriana genus. However pharmacological studies on isolated compounds are scarce. Species of Valeriana native to Rio Grande do Sul have been studied regarding their chemical composition being Valeriana glechomifolia the species with highest valepotriate’ contents. In this study different methods of extraction of valepotriates from aerial parts of V. glechomifolia were compared, and valepotriate’ enriched extracts were tested in mice models of sedation, anxiety and depression. Isolated valepotriates were submitted to binding to benzodiazepine and 5HT1A receptors, and assayed for the Na+K+ATPase inhibitory activity. The extraction by supercritical carbon dioxide (SCCO2) was carried out at 40 oC under 90, 120, 150 or 200 bar affording higher valepotriates contents than maceration with dichloromethane and ultrasound. The highest valepotriates yielding was obtained with SCCO2 (40 oC , 90 bar). All extracts presented the same qualitative valepotriate’ profile. Flavonoids were also obtained from methanol extracts. The median lethal dose of valepotriates obtained by maceration was determined as 42±3 mg/kg, i.p. This valepotriate’ enriched extract (10 mg/kg, p.o., gavage) was ineffective in the elevated plus maze and barbiturate sleeping time tests suggesting that it does not present anxiolitic or hypnotic-sedative properties. Similar results were obtained with flavonoids’ enriched extract. Valtrate, acevaltrate, 1-b-acevaltrate, diavaltrate and the flavonoid named B6 did not bind to benzodiazepine site of receptor GABAA complex neither to serotonergic receptor (5HT1A) at 1-100 μM. The valepotriates inhibited the Na+K+ATPase activity at micromolar concentration range. The valepotriates fraction obtained by SCCO2 (10 mg/kg, gavage) was effective in the forced swimming test without interfering with the spontaneous locomotor activity suggesting that it presents an antidepressant-like activity. In conclusion the extraction by supercritical carbon dioxide is valuable to obtain valepotriates enriched fractions; and valepotriates seem to represent new chemical entities with no selective inhibitory activity of Na+K+ATPase, as well as with antidepressant-like activity.

All living cells contain the electrogenic enzyme Na/K ATPase, whose function is to pump ions against the electrochemical gradient, in order to provide potential energy which is later used for cellular processes such as action potentials, muscle contraction and facilitated transport. Using a technique developed in our lab, exploiting the molecule's voltage dependence, it is possible to increase this pump function by many folds. Optical measurement of the membrane potential of living cells was made using a potentiommetric dye, with successful manipulation of the ionic concentration and membrane potential reported. Additional supporting results are presented, along with extension of this field to the study of cardiac Myocytes, representing a progression to Mammalian cells, with advantages to future clinical research evident. Successful manipulation of membrane potential was also achieved using cells in a two dimensional tissue matrix, which more closely approximates the living system, and hence is closer to an eventual clinical application. Also, expedited recovery from electrical injury was recorded, demonstrating a possible therapeutic application of the technique.

<p>During the smoltification the Atlantic salmon (Salmo salar) develop different adaptations to survive in oceanic environment. One of the most important adaptations is the ability to excrete the surplus of salt through the gills. The excretion is controlled by an enzyme called Na+/K+-ATPase which is produced in an α-subunit by two gene isoforms called α1a and α1b. Enzyme activity is increasing during the smoltification process and is a strong indicator for that the process is taking place. The aim of this study was to investigate a landlocked strain of Atlantic salmon and see how the enzyme activity is developing in comparison with the gene expression of the mRNA that is coded for the enzyme. The study was carried out between March and May in the hatchery in Brattfors, Värmland. Fish were sampled at four occasions. The enzyme activity was compared between two groups of salmon where one group had full ration of food, 100% and the other group had a 15% food ration. The enzyme activity for the 100% group was then compared with the gene expression from the same group. The hypothesis was that food availability should effect smoltification and that the 15% group would have a faster increase in activity compared with the 100% group. There should also be some correlation between enzyme activity and gene expression. Na+/K+-ATPase enzyme activity showed no major differences between the groups except for a significant difference at the last sampling. Both groups had a large increase in activity from the second to the third sampling with a peak on 3.16 µmol ADP/mg/h at most. This was followed by a drop in activity at the last sampling date. The gene expression showed a fast increase of the α1b gene over the study with drop in the last sampling and the α1a gene had a constant increase from the first to the last sampling. The comparison with enzyme activity and gene expression showed a weak correlation. Compared with studies done on anadromous salmon and the land locked salmon in this study had a different development in gene expression. This could be explained that the different life strategies play an important role how the genes are expressed.</p> / <p>Under smoltifieringen utvecklar atlantlaxen (Salmo salar) olika anpassningar för att överleva i havsmiljö. En av de vikigaste anpassningarna är att utsöndra överskott av salt via gällarna. Exkretionen är kontrollerad av ett enzym som heter Na+/K+-ATPas som produceras i en α-subenhet av två isoformer av gener som heter α1a och α1b. Enzym aktiviteten ökar under smoltifieringen och är en stark indikator på att processen sker. Målet med denna studie var att undersöka en sjövandrande stam av atlantlax och se hur enzymaktiviteten utvecklas i jämförelse med gen expressionen av mRNA som kodar för produktionen av enzymet. Studien genomfördes vid fiskodlingen i Brattfors, Värmland där prover togs vid fyra tillfällen. Enzymaktiviteten jämfördes mellan två grupper av lax där en grupp fick full matranson 100 % och en grupp fick 15 % matranson. Senare jämfördes enzymaktiviteten för 100 % gruppen med gen expressionen inom samma grupp. Hypotesen var att tillgängligheten på mat skulle påverka smoltifieringen och att 15 % gruppen skulle ha en snabbare ökning i jämförelse med 100 % gruppen. Det skulle också vara en viss korrelation mellan enzymaktivitet och gen expression. Na+/K+-ATPas enzym aktiviteten visade inga större skillnader mellan grupperna förutom vid sista provtagningen. Båda grupperna hade en stor ökning från den andra till den tredje provtagningen med den högsta aktiviteten på 3.16 µmol ADP/mg/h. Detta följdes av ett fall i aktivitet vid sista provtagningen. Gen expressionen visade en snabb ökning av α1b genen över studien med en nedgång vid sista provtagningen och α1a hade en konstant men mindre ökning från första till sista provtagningen. Jämförelsen mellan enzymaktivitet och gen expression visade på en svag korrelation. Det fanns en skillnad i gen expression mellan studier gjorda på anadrom lax och sjövandrande lax i denna studie. Detta kan förklaras av att de olika livsstrategierna spelar en betydande roll i hur generna uttrycks.</p>

During the smoltification the Atlantic salmon (Salmo salar) develop different adaptations to survive in oceanic environment. One of the most important adaptations is the ability to excrete the surplus of salt through the gills. The excretion is controlled by an enzyme called Na+/K+-ATPase which is produced in an α-subunit by two gene isoforms called α1a and α1b. Enzyme activity is increasing during the smoltification process and is a strong indicator for that the process is taking place. The aim of this study was to investigate a landlocked strain of Atlantic salmon and see how the enzyme activity is developing in comparison with the gene expression of the mRNA that is coded for the enzyme. The study was carried out between March and May in the hatchery in Brattfors, Värmland. Fish were sampled at four occasions. The enzyme activity was compared between two groups of salmon where one group had full ration of food, 100% and the other group had a 15% food ration. The enzyme activity for the 100% group was then compared with the gene expression from the same group. The hypothesis was that food availability should effect smoltification and that the 15% group would have a faster increase in activity compared with the 100% group. There should also be some correlation between enzyme activity and gene expression. Na+/K+-ATPase enzyme activity showed no major differences between the groups except for a significant difference at the last sampling. Both groups had a large increase in activity from the second to the third sampling with a peak on 3.16 µmol ADP/mg/h at most. This was followed by a drop in activity at the last sampling date. The gene expression showed a fast increase of the α1b gene over the study with drop in the last sampling and the α1a gene had a constant increase from the first to the last sampling. The comparison with enzyme activity and gene expression showed a weak correlation. Compared with studies done on anadromous salmon and the land locked salmon in this study had a different development in gene expression. This could be explained that the different life strategies play an important role how the genes are expressed. / Under smoltifieringen utvecklar atlantlaxen (Salmo salar) olika anpassningar för att överleva i havsmiljö. En av de vikigaste anpassningarna är att utsöndra överskott av salt via gällarna. Exkretionen är kontrollerad av ett enzym som heter Na+/K+-ATPas som produceras i en α-subenhet av två isoformer av gener som heter α1a och α1b. Enzym aktiviteten ökar under smoltifieringen och är en stark indikator på att processen sker. Målet med denna studie var att undersöka en sjövandrande stam av atlantlax och se hur enzymaktiviteten utvecklas i jämförelse med gen expressionen av mRNA som kodar för produktionen av enzymet. Studien genomfördes vid fiskodlingen i Brattfors, Värmland där prover togs vid fyra tillfällen. Enzymaktiviteten jämfördes mellan två grupper av lax där en grupp fick full matranson 100 % och en grupp fick 15 % matranson. Senare jämfördes enzymaktiviteten för 100 % gruppen med gen expressionen inom samma grupp. Hypotesen var att tillgängligheten på mat skulle påverka smoltifieringen och att 15 % gruppen skulle ha en snabbare ökning i jämförelse med 100 % gruppen. Det skulle också vara en viss korrelation mellan enzymaktivitet och gen expression. Na+/K+-ATPas enzym aktiviteten visade inga större skillnader mellan grupperna förutom vid sista provtagningen. Båda grupperna hade en stor ökning från den andra till den tredje provtagningen med den högsta aktiviteten på 3.16 µmol ADP/mg/h. Detta följdes av ett fall i aktivitet vid sista provtagningen. Gen expressionen visade en snabb ökning av α1b genen över studien med en nedgång vid sista provtagningen och α1a hade en konstant men mindre ökning från första till sista provtagningen. Jämförelsen mellan enzymaktivitet och gen expression visade på en svag korrelation. Det fanns en skillnad i gen expression mellan studier gjorda på anadrom lax och sjövandrande lax i denna studie. Detta kan förklaras av att de olika livsstrategierna spelar en betydande roll i hur generna uttrycks.

Apoptosis is a special form of cell death that if non-functional may lead to diseases such as cancer. A reduction of the intracellular potassium ion (K+) content is necessary for activating enzymes important for the execution of apoptosis. Pharmacological modulation of K+ fluxes to reduce intracellular K+ in cancer cells might therefore force the cells into apoptosis and decrease tumour cell mass. Human malignant pleural mesothelioma (MPM) is a form of cancer often caused by asbestos exposure. Although asbestos has been banned in the Western World, the incidence of MPM is expected to increase. Cisplatin is the first-line chemotherapy for MPM, but acquired resistance to the drug is a clinical problem. This thesis is mainly based on work with the human malignant pleural mesothelioma cell line (P31 wt) and a cisplatin-resistant sub-line (P31 res). The aim was to first characterize K+ fluxes in P31 wt and P31 res cells, and then manipulate them in order to reduce intracellular K+ and induce apoptosis with K+ manipulation alone or in combination with cisplatin. Characterization of K+ fluxes in P31 wt cells showed that: 1) ouabain, a digitalis-like drug, and specific blocker of the Na+, K+, ATPase pump, effectively inhibited K+ uptake, 2) bumetanide, a diuretic, and an inhibitor of the Na+, K+, 2Cl-¬-cotransporter, had a transient effect on K+ uptake, and 3) the antifungal drug amphotericin B stimulated K+ efflux. In order to determine intracellular K+ content, the potassium-binding fluorescent probe PBFI-AM was used in a 96-well plate assay. After a 3-h incubation with ouabain, with or without bumetanide, combined with amphotericin B, the intracellular K+ content was reduced in P31 wt cells but not in P31 res cells. Ouabain induced apoptosis in both P31 wt and P31 res cells. P31 res cells were sensitized to cisplatin by ouabain, since 10 mg/L cisplatin in combination with ouabain induced about the same percentage of apoptotic cells as 40 mg/L cisplatin. Apoptosis was executed via caspase-3 activation in both P31 wt and P31 res cells. Amphotericin B enhanced ouabain-induced apoptosis in P31 wt cells via caspase-9 activation, with increased caspase-3 activation and DNA fragmentation as consequences. Ouabain-induced apoptosis in P31 res cells was executed via increased expression of pro-apoptotic Bak. The combination of cisplatin with ouabain and amphotericin B was stressful to both P31 wt and P31 res cells, since SAPK/JNK a known factor in stress-induced apoptosis was activated. In conclusion, K+ flux manipulation with clinical used drugs can induce apoptosis per se and also enhance cisplatin-induced apoptosis in P31 wt and P31 res cells.

Most vertebrates suffer permanent damage after minutes of anoxia. Many insects however, have part of their life cycle in anoxia or constant hypoxia, such as during their egg-hatching phase, by living as deep burrowers, or at high altitudes. Insects are able to survive in anoxia from hours to days, or even months by developing various strategies through evolution. For example, the locusts (Locusta migratoria) enter a reversible coma during anoxia that is associated with an arrest of ventilation, and a reinitiation of ventilation when returned to normoxia. This coma is correlated with a surge in the concentration of extracellular potassium ions ([K+]o), and recovery from this reversible coma is dependent on re-establishing the functional [K+]o. Prior exposure to a sublethal heatshock (HS)-preconditioning grants locusts a temporary resilience to anoxia; however, the molecular mechanisms of this protection are still unclear. This project investigated the effects of HS-preconditioning on locusts’ ventilation, the total enzymatic activity of the Na+/K+-ATPase, as well as its distribution within the metathoracic ganglion and tested the hypothesis that HS-preconditioning alters locusts’ ventilation and increases the totally Na+/K+-ATPase activity and its concentration within neuronal membranes. I recorded electromyograms of locusts’ ventilatory motor patterns in the presence and absence of anoxic coma by placing a copper wire electrode on ventilatory muscles 161 or 173 in control and HS-preconditioned animals. In addition, I studied the enzymatic activity of the Na+/K+-ATPase using a pyruvate kinase/lactate dehydrogenase assay and the localization of the Na+/K+-ATPase using immunohistochemistry in control and HS-preconditioned locusts at different stages of coma. I found that the ventilatory cycle period was decreased and the ventilatory muscle burst duration was increased after recovery from anoxic coma in HS-preconditioned locusts. I also found that anoxia did not affect the activity or the localization of the Na+/K+-ATPase. However, HS-preconditioning increased the total activity of the Na+/K+-ATPase and the localization of the Na+/K+-ATPase within the neuronal membranes. From this project, I concluded that HSpreconditioning affected locusts’ ventilatory motor pattern after recover from anoxia and increased the total activity and the neuronal membrane localization of the Na+/K+-ATPase. / Thesis (Master, Biology) -- Queen's University, 2011-09-26 13:14:48.472

Neurons perform complex computations, communications and precise transmissions of information in the form of action potentials (APs). The high level of heterogeneity and complexity at all levels of organization within a neuron and the functional requirement of highly permeable cell membranes leave neurons exposed to damage when energy levels are insufficient for the active maintenance of ionic gradients. When energy is limiting the ionic gradient across a neuron’s cell membrane risks being dissipated which can have dire consequences. Other researchers have advocated “generalized channel arrest” and/or “spike arrest” as a means of reducing the neuronal permeability allowing neurons to adjust the demands placed on their electrogenic pumps to lower levels of energy supply. I investigated the consequences of hypoxia on the propagation of a train of APs down the length of a fast conducting axon capable of transmitting APs at very high frequencies. Under normoxic conditions I found that APs show conduction velocities and instantaneous frequencies nearly double that of neurons experiencing energy limiting hypoxic conditions. I show that hypoxia affects AP conduction differently for different lengths of axon and for APs of different instantaneous frequencies. Action potentials of high instantaneous frequency in branching lengths of axon within ganglia were delayed more significantly than those in non-branching lengths contained within the connective and fail preferentially in branching axon. I found that octopamine attenuates the effects of hypoxia on AP propagation for the branching length of axon but has no effect on the non-branching length of axon. Additionally, for energetically stable cells, application of the anti-diabetic medication metformin or the hyperpolarization-activated cyclic nucleotide-gated (HCN) channel blocker ZD7288 resulted in a reduced performance similar to that seen in neurons experiencing energetic stress. Furthermore both metformin and ZD7288 affect the shape of individual APs within an AP train as well as the original temporal sequence of the AP train, which encodes behaviourally relevant information. I propose that the reduced performance observed in an energetically compromised cell represents an adaptive mechanism employed by neurons in order to maintain the integrity of their highly heterogeneous and complex organization during periods of reduced energy supply. / Thesis (Master, Biology) -- Queen's University, 2011-10-07 14:41:46.972

Previous studies have shown that ischemia-reperfusion (I/R) injury is associated with cardiac dysfunction and depression in sarcolemmal Na+-K+-ATPase activity. This study was undertaken to evaluate the role of proteases in these alterations by subjecting rat hearts to different times of global ischemia, and reperfusion after 45 min of ischemia. Decreases in Na+-K+-ATPase activity at 60 min of global ischemia were associated with augmented activities of both calpain and MMPs and depressed protein content of β1- and β2-subunits, without changes in α1- and α2-subunits of the enzyme. However, reperfusion of ischemic heart produced depression in Na+-K+-ATPase activity, no change in the augmented calpain activity, but decreases in augmented MMP-2 activity and Na+-K+-ATPase content. MDL28170, a calpain inhibitor, was more effective in attenuating I/R-induced alterations than doxycycline, an MMP inhibitor. Incubation of control SL preparation with calpain, unlike MMP-2, depressed Na+-K+-ATPase activity and decreased α1, α2 and β2 without changes in β1. These results support the view that activation of calpain is involved in depressing Na+-K+-ATPase activity and degradation of its subunits in hearts subjected to I/R injury.

Previous studies have shown that ischemia-reperfusion (I/R) injury is associated with cardiac dysfunction and depression in sarcolemmal Na+-K+-ATPase activity. This study was undertaken to evaluate the role of proteases in these alterations by subjecting rat hearts to different times of global ischemia, and reperfusion after 45 min of ischemia. Decreases in Na+-K+-ATPase activity at 60 min of global ischemia were associated with augmented activities of both calpain and MMPs and depressed protein content of β1- and β2-subunits, without changes in α1- and α2-subunits of the enzyme. However, reperfusion of ischemic heart produced depression in Na+-K+-ATPase activity, no change in the augmented calpain activity, but decreases in augmented MMP-2 activity and Na+-K+-ATPase content. MDL28170, a calpain inhibitor, was more effective in attenuating I/R-induced alterations than doxycycline, an MMP inhibitor. Incubation of control SL preparation with calpain, unlike MMP-2, depressed Na+-K+-ATPase activity and decreased α1, α2 and β2 without changes in β1. These results support the view that activation of calpain is involved in depressing Na+-K+-ATPase activity and degradation of its subunits in hearts subjected to I/R injury.

Made available in DSpace on 2016-06-02T19:32:11Z (GMT). No. of bitstreams: 1 DissGHDP.pdf: 1446855 bytes, checksum: b44d0b337ad1c73932cf0a253c2bdad6 (MD5) Previous issue date: 2004-04-30 / Universidade Federal de Sao Carlos / Copper is an essential trace-element for all live organisms however, when its concentration is high in the aquatic environment, copper is potentially toxic. Most physiological changes caused by copper in fish has been analyzed after acute exposure (96h). The aim of this study was investigate the physiological responses of Prochilodus scrofa to stress, during acute copper exposition (24, 48, 72 and 96 hours) and recovery period after animal transference to clean water, and the time necessary to recovery after animals transference to clean water. The changes associated to fish transference to experimental systems were also evaluated. Juveniles Prochilodus scrofa were acclimated during 30 days and then fish were sampled to take blood for pH, hematocrit, plasma cortisol glucose and ions (Na+, Cl- e K+) analyses and specific Na+/K+-ATPase activity of gills. The remaining fish were divided in two groups: control (C) and exposed to copper (CU) and transferred to glass aquariums where was kept for 4 days. CU groups were exposed to copper (96h-CL50 = 29 µgCu L-1). Sampling was done 1, 2, 3 and 4 days of copper exposure. Surviving fish were then transferred to aquariums with flowing water without copper and sampling was done 1, 2, 7, 15 and 30 days of recovery. Blood sampling was taken for the same analyses above cited. Plasma cortisol concentration increased 300 a 400% in control and copper exposed groups related to acclimated fish, plasma glucose increased 800% only in the copper exposed groups during the exposure period and returned to normal levels in the 2nd day of recovery. In general, the changes in plasma ion (Na+, Cl- e K+) concentrations and pH were not significant in relation to those of acclimated fish however, significant changes were found between the values of control and copper exposed groups during copper exposure and recovery period. The activity of Na+/K+-ATPase of gills was lower in the CU group during the copper exposure period and on the 1st day of recovery. The hematocrit increased during metal exposure but on the 1st day of recovery reached the normal values. Although changes were found on the analyzed parameters, on the 7th days after the transference to clean water all parameters evidenced a tendency to recovery the physiological changes reaching the values of acclimated fish. The present study showed that the stress responses to copper exposure is complex and P. scrofa presents high susceptibility to handling evidencing the importance of comparative analyses involving acclimated and/or unstressed fish together with the controls and exposed fish to a given stressor. / O cobre é um elemento-traço essencial a todos os organismos, entretanto quando em alta concentração no meio aquático pode ser potencialmente tóxico. As alterações fisiológicas causadas pelo cobre em peixes têm sido analisadas, em geral, após exposições agudas (96h). O presente estudo teve como objetivo avaliar as alterações fisiológicas associadas ao estresse, durante o período de exposição aguda ao cobre e o período de recuperação em água sem a presença de cobre. As alterações provocadas pelo estresse associado à transferência dos animais para os sistemas experimentais também foram avaliadas. Prochilodus scrofa jovens foram aclimatados durante 30 dias e posteriormente animais foram amostrados e o sangue coletado para análise de pH e hematócrito sangüíneos, cortisol, glicose e íons (Na+, Cl- e K+) plasmáticos e determinação da atividade da enzima Na+/K+-ATPase das brânquias. Os animais remanescentes foram divididos em dois grupos controle (C) e exposto ao cobre (CU) e transferidos para aquários de vidro onde permaneceram durante 4 dias. O grupo CU foi exposto ao cobre (CL50-96h = 29 µgCu L-1). As amostragens foram efetuadas após 1, 2, 3 e 4 dias de exposição ao cobre. Os animais sobreviventes foram então transferidos para aquários com água corrente sem cobre e amostragens foram efetuadas após 1, 2, 7, 15 e 30 dias de recuperação. Amostras de sangue foram coletadas para as mesmas análises citadas acima. A concentração de cortisol plasmático aumentou 300 a 400% nos grupos controle e expostos ao cobre em relação aos animais aclimatados e a glicose plasmática aumentou 800% somente nos animais expostos ao cobre durante o período de exposição e retornaram aos níveis dos animais aclimatados no 2o dia de recuperação. As alterações nas concentrações dos íons plasmáticos (Na+, Cl- e K+) e pH, em geral, não foram significantes em relação aos animais aclimatados, mas alterações significativas foram observadas entre os grupos controle e exposto ao cobre durante a exposição ao cobre e recuperação. A atividade da enzima Na+/K+-ATPase nas brânquias foi menor no grupo CU durante a exposição ao cobre e 1o dia de recuperação. O hematócrito aumentou significativamente durante a exposição ao metal, mas após o primeiro dia de recuperação retornou aos valores normais. Embora tenha ocorrido alteração nos parâmetros analisados, após o sétimo dia de transferência para água sem cobre houve uma tendência de recuperação em todos os aspectos fisiológicos. Os resultados do presente estudo mostraram que a resposta ao estresse por exposição ao cobre é complexa e que P. scrofa apresenta a alta susceptibilidade à manipulação evidenciando a importância de estudos que envolvem comparações com animais considerados aclimatados ou não estressados e animais de grupos controle e expostos a um dado estressor.