RESUMO
High sugar consumption is a risk factor for metabolic disturbances leading to memory impairment. Thus, rats subject to high sucrose intake (HSu) develop a metabolic syndrome and display memory deficits. We now investigated if these HSu-induced memory deficits were associated with metabolic and electrophysiological alterations in the hippocampus. Male Wistar rats were submitted for 9 weeks to a sucrose-rich diet (35% sucrose solution) and subsequently to a battery of behavioral tests; after sacrifice, their hippocampi were collected for ex vivo high-resolution magic angle spinning (HRMAS) metabolic characterization and electrophysiological extracellular recordings in slices. HSu rats displayed a decreased memory performance (object displacement and novel object recognition tasks) and helpless behavior (forced swimming test), without altered locomotion (open field). HRMAS analysis indicated a similar hippocampal metabolic profile of HSu and control rats. HSu rats also displayed no change of synaptic transmission and plasticity (long-term potentiation) in hippocampal Schaffer fibers-CA1 pyramid synapses, but had decreased amplitude of long-term depression in the temporoammonic (TA) pathway. Furthermore, HSu rats had an increased density of inhibitory adenosine A1 receptors (A1R), that translated into a greater potency of A1R in Schaffer fiber synapses, but not in the TA pathway, whereas the endogenous activation of A1R in HSu rats was preserved in the TA pathway but abolished in Schaffer fiber synapses. These results suggest that HSu triggers a hippocampal-dependent memory impairment that is not associated with altered hippocampal metabolism but is probably related to modified synaptic plasticity in hippocampal TA synapses.
Assuntos
Dieta/efeitos adversos , Sacarose Alimentar/toxicidade , Hipocampo/fisiopatologia , Transtornos da Memória/etiologia , Transtornos da Memória/fisiopatologia , Animais , Modelos Animais de Doenças , Emoções/fisiologia , Desamparo Aprendido , Locomoção/fisiologia , Potenciação de Longa Duração/fisiologia , Depressão Sináptica de Longo Prazo/fisiologia , Masculino , Atividade Motora/fisiologia , Ratos Wistar , Receptor A1 de Adenosina/metabolismo , Reconhecimento Psicológico/fisiologia , Sinapses/fisiologia , Transmissão Sináptica/fisiologiaRESUMO
BACKGROUND AND PURPOSE: Both cannabinoid CB1 and adenosine A2A receptors (CB1 receptors and A2A receptors) control synaptic transmission at corticostriatal synapses, with great therapeutic importance for neurological and psychiatric disorders. A postsynaptic CB1 -A2A receptor interaction has already been elucidated, but the presynaptic A2A receptor-mediated control of presynaptic neuromodulation by CB1 receptors remains to be defined. Because the corticostriatal terminals provide the major input to the basal ganglia, understanding the interactive nature of converging neuromodulation on them will provide us with novel powerful tools to understand the physiology of corticostriatal synaptic transmission and interpret changes associated with pathological conditions. EXPERIMENTAL APPROACH: Pharmacological manipulation of CB1 and A2A receptors was carried out in brain nerve terminals isolated from rats and mice, using flow synaptometry, immunoprecipitation, radioligand binding, ATP and glutamate release measurement. Whole-cell patch-clamp recordings were made in horizontal corticostriatal slices. KEY RESULTS: Flow synaptometry showed that A2A receptors were extensively co-localized with CB1 receptor-immunopositive corticostriatal terminals and A2A receptors co-immunoprecipitated CB1 receptors in these purified terminals. A2A receptor activation decreased CB1 receptor radioligand binding and decreased the CB1 receptor-mediated inhibition of high-K(+) -evoked glutamate release in corticostriatal terminals. Accordingly, A2A receptor activation prevented CB1 receptor-mediated paired-pulse facilitation and attenuated the CB1 receptor-mediated inhibition of synaptic transmission in glutamatergic synapses of corticostriatal slices. CONCLUSIONS AND IMPLICATIONS: Activation of presynaptic A2A receptors dampened CB1 receptor-mediated inhibition of corticostriatal terminals. This constitutes a thus far unrecognized mechanism to modulate the potent CB1 receptor-mediated presynaptic inhibition, allowing frequency-dependent enhancement of synaptic efficacy at corticostriatal synapses.
Assuntos
Ácido Glutâmico/metabolismo , Receptor A2A de Adenosina/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Receptores Pré-Sinápticos/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Encéfalo/metabolismo , Encéfalo/fisiologia , Masculino , Camundongos Knockout , Ratos Wistar , Sinapses/metabolismo , Transmissão SinápticaRESUMO
Acute toxicity test (LD-50) using toxic shock syndrome toxin (TSST-1) was tested in BALB/c, C57BL/6 and Swiss mice. Animals (n = 10) were intraperitoneally injected with TSST-1 (0.01-10.0µg/mouse) followed 4h later by potentiating dose of lipopolysaccharide (75.0µg of LPS - E. coli O111:B4) and cumulative mortality was recorded over 72h. Control animals received either TSST-1 or LPS alone. The data were submitted to qui-Square test and acute toxicity test was calculated by probit analysis (confidence limits expressed as µg toxin/kg). BALB/c mice was the most sensitive (20.0µg/kg, 95 percent confidence limits: 9.0-92.0) followed by C57BL/6 (38.5µg/kg, 95 percent confidence limits: 9.11- 401.6). Data from Swiss mice was not conclusive, indicating only low sensitivity. Selection of the animal model and standardization of the experiment are fundamental for the development of serum neutralization tests used for final quality control of vaccine production.
A toxicidade aguda (DL-50) da toxina da síndrome do choque tóxico (TSST-1) foi testada em linhagens de camundongos BALB/c, C57BL/6 e Suíça. Os animais (n=10) inoculados intraperitoneal com doses crescentes de toxina (0,01 - 10,0µg/animal) receberam 4h após 75µg de LPS (E. coli O111: B4). A toxicidade aguda (DL50) foi observada por um período de 72h e os dados submetidos ao teste de qui- quadrado. Os resultados e os limites de confiança foram expressos em µg de toxina/kg. A linhagem BALB/c apresentou maior sensibilidade (20µg/kg - limite de confiança a 95 por cento entre 9,0- 92,0), seguida da C57BL/6 (38,5µg/kg - limite de confiança a 95 por cento entre 9,11 - 401,6). A amplitude dos limites de confiança deve-se à natureza da toxina, ao mecanismo de ação, a via de inoculação e ao animal utilizado. A seleção do modelo animal e a padronização do experimento são fundamentais para o desenvolvimento de testes de soro neutralização para fins de controle de qualidade do processo de produção de vacinas.
Assuntos
Animais , Experimentação Animal , Choque Séptico/induzido quimicamente , Camundongos , Modelos Animais , Testes de Toxicidade Aguda/análiseRESUMO
Diabetic encephalopathy is a recognized complication of untreated diabetes resulting in a progressive cognitive impairment accompanied by modification of hippocampal function. The purinergic system is a promising novel target to control diabetic encephalopathy since it might simultaneously control hippocampal synaptic plasticity and glucose handling. We now tested whether streptozotocin-induced diabetes led to a modification of extracellular ATP homeostasis and density of membrane ATP (P2) receptors in the hippocampus, a brain structure involved in learning and memory. The extracellular levels of ATP, evaluated in the cerebrospinal fluid, were reduced by 60.4+/-17.0% in diabetic rats. Likewise, the evoked release of ATP as well as its extracellular catabolism was also decreased in hippocampal nerve terminals of diabetic rats by 52.8+/-10.9% and 38.7+/-6.5%, respectively. Western blot analysis showed that the density of several P2 receptors (P2X(3,5,7) and P2Y(2,6,11)) was decreased in hippocampal nerve terminals. This indicates that the synaptic ATP signaling is globally depressed in diabetic rats, which may contribute for diabetes-associated decrease of synaptic plasticity. In contrast, the density of P2 receptors (P2X(1,2,5,6,7) and P2Y(6) but not P2Y(2)) increased in whole hippocampal membranes, suggesting an adaptation of non-synaptic P2 receptors to sense decreased levels of extracellular ATP in diabetic rats, which might be aimed at preserving the non-synaptic purinergic signaling.
Assuntos
Diabetes Mellitus Experimental/fisiopatologia , Hipocampo/fisiopatologia , Receptores Purinérgicos P2/fisiologia , Transdução de Sinais/fisiologia , Trifosfato de Adenosina/líquido cefalorraquidiano , Trifosfato de Adenosina/metabolismo , Animais , Western Blotting , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/fisiopatologia , Eletroforese em Gel de Poliacrilamida , Espaço Extracelular/metabolismo , Hipocampo/metabolismo , Homeostase/fisiologia , Imuno-Histoquímica , Masculino , Terminações Nervosas/metabolismo , Ratos , Ratos WistarRESUMO
BACKGROUND AND PURPOSE: Inhibitory CB(1) cannabinoid receptors and excitatory TRPV(1) vanilloid receptors are abundant in the hippocampus. We tested if two known hybrid endocannabinoid/endovanilloid substances, N-arachidonoyl-dopamine (NADA) and anandamide (AEA), presynapticaly increased or decreased intracellular calcium level ([Ca(2+)](i)) and GABA and glutamate release in the hippocampus. EXPERIMENTAL APPROACH: Resting and K(+)-evoked levels of [Ca(2+)](i) and the release of [(3)H]GABA and [(3)H]glutamate were measured in rat hippocampal nerve terminals. KEY RESULTS: NADA and AEA per se triggered a rise of [Ca(2+)](i) and the release of both transmitters in a concentration- and external Ca(2+)-dependent fashion, but independently of TRPV(1), CB(1), CB(2), or dopamine receptors, arachidonate-regulated Ca(2+)-currents, intracellular Ca(2+) stores, and fatty acid metabolism. AEA was recently reported to block TASK-3 potassium channels thereby depolarizing membranes. Common inhibitors of TASK-3, Zn(2+), Ruthenium Red, and low pH mimicked the excitatory effects of AEA and NADA, suggesting that their effects on [Ca(2+)](i) and transmitter levels may be attributable to membrane depolarization upon TASK-3 blockade. The K(+)-evoked Ca(2+) entry and Ca(2+)-dependent transmitter release were inhibited by nanomolar concentrations of the CB(1) receptor agonist WIN55212-2; this action was sensitive to the selective CB(1) receptor antagonist AM251. However, in the low micromolar range, WIN55212-2, NADA and AEA inhibited the K(+)-evoked Ca(2+) entry and transmitter release independently of CB(1) receptors, possibly through direct Ca(2+) channel blockade. CONCLUSIONS AND IMPLICATIONS: We report here for hybrid endocannabinoid/endovanilloid ligands novel dual functions which were qualitatively similar to activation of CB(1) or TRPV(1) receptors, but were mediated through interactions with different targets.
Assuntos
Ácidos Araquidônicos/farmacologia , Cálcio/metabolismo , Dopamina/análogos & derivados , Ácido Glutâmico/metabolismo , Hipocampo/efeitos dos fármacos , Alcamidas Poli-Insaturadas/farmacologia , Ácido gama-Aminobutírico/metabolismo , Animais , Dopamina/farmacologia , Endocanabinoides , Fluorometria , Hipocampo/metabolismo , Masculino , Ratos , Ratos Wistar , Receptor CB1 de Canabinoide/fisiologia , Receptor CB2 de Canabinoide/fisiologia , Receptores Dopaminérgicos/fisiologia , Canais de Cátion TRPV/fisiologiaRESUMO
The subsynaptic distribution of kainate receptors is still a matter of much debate given its importance to understand the way they influence neuronal communication. Here, we show that, in synapses of the rat hippocampus, presynaptic kainate receptors are localized within the presynaptic active zone close to neurotransmitter release sites. The activation of these receptors with low concentrations of agonists induces the release of [(3)H]glutamate in the absence of a depolarizing stimulus. Furthermore, this modulation of [(3)H]glutamate release by kainate is more efficient when compared with a KCl-evoked depolarization that causes a more than two-fold increase in the intra-terminal calcium concentration but no apparent release of [(3)H]glutamate, suggesting a direct receptor-mediated process. Using a selective synaptic fractionation technique that allows for a highly efficient separation of presynaptic, postsynaptic and non-synaptic proteins we confirmed that, presynaptically, kainate receptors are mainly localized within the active zone of hippocampal synapses where they are expected to be in a privileged position to modulate synaptic phenomena.
Assuntos
Hipocampo/metabolismo , Receptores Pré-Sinápticos/metabolismo , Animais , Western Blotting , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Sinalização do Cálcio/fisiologia , Ácido Glutâmico/metabolismo , Hipocampo/ultraestrutura , Imuno-Histoquímica , Masculino , Proteínas do Tecido Nervoso/metabolismo , Ratos , Ratos Wistar , Receptores de Ácido Caínico/fisiologia , Sinaptossomos/metabolismo , Sinaptossomos/ultraestruturaRESUMO
Adenosine is a neuromodulator that controls neurotransmitter release through inhibitory A1 and facilitatory A2A receptors. Although both adenosine receptor-mediated inhibition and facilitation of glutamate release have been observed, it is not clear whether both A1 and A2A receptors are located in the same glutamatergic nerve terminal or whether they are located on different populations of these terminals. Thus, we have tested if single pyramidal glutamatergic neurons from the hippocampus simultaneously expressed A1 and A2A receptor mRNA and if A1 and A2A receptors were co-localized in hippocampal glutamatergic nerve terminals. Single cell PCR analysis of visually identified pyramidal neurons revealed the simultaneous presence of A1 and A2A receptor mRNA in four out 16 pyramidal cells possessing glutamatergic markers but not GABAergic or astrocytic markers. Also, A1 and A2A receptor immunoreactivities were co-localized in 26 +/- 4% of nerve terminals labeled with antibodies against vesicular glutamate transporters type 1 or 2, i.e. glutamatergic nerve terminals. This indicates that glutamatergic neurons in the hippocampus co-express A1 and A2A receptors and that these two receptors are co-localized in a subset of glutamatergic nerve terminals.
Assuntos
Ácido Glutâmico/metabolismo , Hipocampo/metabolismo , Terminações Nervosas/metabolismo , Células Piramidais/metabolismo , Receptor A1 de Adenosina/biossíntese , Receptor A2A de Adenosina/biossíntese , Animais , Astrócitos/metabolismo , Hipocampo/citologia , Hipocampo/ultraestrutura , Imuno-Histoquímica , Masculino , Proteínas de Membrana Transportadoras/metabolismo , Terminações Nervosas/ultraestrutura , Células Piramidais/ultraestrutura , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína Vesicular 1 de Transporte de Glutamato , Proteína Vesicular 2 de Transporte de GlutamatoRESUMO
Electrophysiological recordings were used to investigate the effects of ATP analogues on theta-burst-induced long-term potentiation (LTP) in rat hippocampal slices. alpha,beta-Methylene ATP (alpha,beta-MeATP; 20 microM) decreased LTP from 36+/-9% to 17+/-5%, an effect prevented by adenosine A(1) receptor blockade in accordance with the localised catabolism of ATP analogues into adenosine, leading to adenosine A(1) receptor activation. Thus, to probe the role of extracellular ATP, all experiments were performed with the A(1) receptor selective antagonist, 1,3-dipropyl-8-cyclopentylxanthine (50 nM). In these conditions, alpha,beta-MeATP or 5'-adenylylimido-diphosphate (beta,gamma-ImATP; 20 microM) facilitated LTP by 120%, an effect prevented by the P2 receptor antagonists, pyridoxalphosphate-6-azophenyl-2'-4'-disulphonic acid (PPADS; 20 microM) or suramin (75 microM), as well as by the P2X(1/3)-selective antagonist 8-(benzamido)naphthalene-1,3,5-trisulfonate (10 microM). The facilitations of LTP by either alpha,beta-MeATP or beta,gamma-ImATP (20 microM) were also prevented by both 4-(2-[7-amino-2-(2-furyl(1,2,4)-triazolo(2,3a)-(1,3,5)triazin-5-yl-amino]ethyl)phenol (50 nM) or 7-2(-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c] pyrimidine (50 nM), antagonists of facilitatory adenosine A(2A) receptors, were occluded by the A(2A) receptor agonist, CGS 21680 (10 nM) and were prevented by the protein kinase C inhibitor, chelerythrine (6 microM) and unaffected by the protein kinase A inhibitor, H89 (1 microM). Furthermore, beta,gamma-ImATP (20 microM) enhanced [(3)H]adenosine outflow from rat hippocampal slices by nearly 150%, an effect prevented by PPADS (20 microM) or suramin (75 microM). The adenosine transport inhibitors, nitrobenzylthioinosine (5 microM) and dipyridamole (10 microM) also prevented beta,gamma-ImATP (20 microM)-induced [(3)H]adenosine outflow and facilitation of LTP. These results suggest that ATP analogues facilitate LTP through P2 receptor activation that mainly triggers adenosine release leading to the activation of adenosine A(2A) receptors.
Assuntos
Trifosfato de Adenosina/análogos & derivados , Adenosina/metabolismo , Hipocampo/metabolismo , Potenciação de Longa Duração , Receptor A2A de Adenosina/metabolismo , Receptores Purinérgicos P2/metabolismo , Adenosina/antagonistas & inibidores , Trifosfato de Adenosina/antagonistas & inibidores , Trifosfato de Adenosina/metabolismo , Trifosfato de Adenosina/fisiologia , Animais , Técnicas de Cultura de Células , Eletrofisiologia , Potenciais Pós-Sinápticos Excitadores , Hipocampo/fisiologia , Masculino , Proteína Quinase C/metabolismo , Proteína Quinase C/fisiologia , Ratos , Ratos Wistar , Receptor A2A de Adenosina/fisiologia , SinaptossomosRESUMO
Increasing prominence is being given to the use of best current evidence in clinical practice and health services and programme management decision-making. The role of information in evidence-based practice (EBP) is discussed, together with questions of how advanced information systems and technology (IS&T) can contribute to the establishment of a broader perspective for EBP. The author examines the development, validation and use of a variety of sources of evidence and knowledge that go beyond the well-established paradigm of research, clinical trials, and systematic literature review. Opportunities and challenges in the implementation and use of IS&T and knowledge management tools are examined for six application areas: reference databases, contextual data, clinical data repositories, administrative data repositories, decision support software, and Internet-based interactive health information and communication. Computerized and telecommunications applications that support EBP follow a hierarchy in which systems, tasks and complexity range from reference retrieval and the processing of relatively routine transactions, to complex "data mining" and rule-driven decision support systems.
Assuntos
Medicina Baseada em Evidências , Sistemas de Informação , Sistemas de Apoio a Decisões Clínicas , Difusão de Inovações , Humanos , Serviços de Informação , InternetRESUMO
The use of ELISA to determine antisnake venom potency of horse immune sera should provide benefits of costs and reproducibility compared to in vivo assays. In the present investigation we evaluated the correlation between ELISA antibody levels and in vivo neutralization assays. For the indirect ELISA method, 0.016 micrograms/well of Bothrops jararaca or Crotalus durissus terrificus venom were used to coat the plates and 100 microliters/well of each sample of antibothropic or anticrotalic venom sera were used at 1:10,000 dilution. Sheep anti-horse IgG conjugated to peroxidase was added and the substrate H2O2/o-phenylenediamine produced the color that was read at 492 nm. A correlation coefficient of r = 0.97 was found for anticrotalic venom antibodies and no significant correlation was observed for antibothropic venom sera using 16 serum samples from immunized horses. However, when three antibothropic venom sera showing high in vivo neutralization potency and low absorbance in ELISA or high absorbance values and low in vivo protection were not included in the correlation analysis the coefficient value was r = 0.88. The correlation coefficient did not improve for all 16 antibothropic sera when a partially purified Bothrops jararaca venom fraction was used to coat the ELISA plates. The results indicate that ELISA could be used to determine the neutralizing potency of anticrotalic venom sera. For the antibothropic venom sera further studies are needed.
Assuntos
Antivenenos/fisiologia , Bothrops , Venenos de Crotalídeos/imunologia , Crotalus , Ensaio de Imunoadsorção Enzimática , Testes de Neutralização , Animais , Soros Imunes/imunologiaRESUMO
The use of ELISA to determine antisnake venom potency of horse immune sera should provide benefits of cost and reproducibility compared to in vivo assays. In the present investigation we evaluated the correlation between ELISA antibody levels and in vivo neutralization assays. For the indirect ELISA method, 0.016 µg/well of Bothrops jararaca or Crotalus durissus terrificus venom were used to coat the plates and 100 µl/well of each sample of antibothropic or anticrotalic venom sera were used at 1:10,000 dilution. Sheep anti-horse IgG conjugated to peroxidase was added and the substrate H202/o-phenylenediamine produced the color that was read at 492 nm. A correlation coefficient of r=0.97 was found for anticrotalic venom antibodies and no significant correlation was observed for antibothropic venom sera using 16 serum samples from immunized horses. However, when three antibothropic venom sera showing high in vivo neutralization...
Assuntos
Animais , Antivenenos/fisiologia , Ensaio de Imunoadsorção Enzimática , Testes de Neutralização , Venenos de Crotalídeos/imunologia , Bothrops , Crotalus , Soros Imunes/imunologiaRESUMO
Owing to the diffusion of computers--personal as well as highly sophisticated models--and of modern data bank systems and more accessible programming methods that no longer require hiring professional programmers to develop a variety of applications, administrators are increasingly using this new technology as a basis for making more realistic decisions. Good information systems are of great importance in the management of local health systems; however, in order for them to be truly useful, data collection must be homogeneous so that the data may be subsequently consolidated and compared. In this connection the authors present a model consisting of four stages: strategic planning; analysis of needs; allocation of resources; and selection of alternatives for consideration by project teams and suppliers of equipment and programs.
Assuntos
Atenção à Saúde/organização & administração , Sistemas de Informação AdministrativaAssuntos
Linfoma não Hodgkin/patologia , Linfoma/patologia , Neoplasias Gástricas/patologia , Adulto , Diagnóstico Diferencial , Feminino , Gastrectomia , Gastroscopia , Humanos , Linfoma/diagnóstico , Linfoma/terapia , Masculino , Pessoa de Meia-Idade , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/terapiaRESUMO
Opportunistic systemic fungal infections are more frequent than generally realized. Increased awareness and a high index of suspicion of fungal super-infection in the presence of sepsis is required to bring about recognition and therapy. The intravenous catheter is an important portal of entry or may act as a foreign body favoring localization of a septic process. In its presence, fungemia must be guarded against. Whenever an intravenous catheter is removed, its tip should be cultured. Removal alone may be a critical item in therapy. In febrile patients, in whom the course of fever is not established, frequent blood cultures with attention directed specifically at fungi should be obtained. Fungi are not easily isolated and identified and only by requesting special attention from the microbiologist can the diagnosis be established in the average institutional laboratory in time to permit appropriate therapy. Since available therapeutic measures are strikingly effective when instituted early, awareness and alertness on the part of the clinician constitute the key to cure.