No / Introduction: Potential-sensitive dyes have primarily been used to optically record action potentials (APs) in whole heart tissue. Using these dyes to record drug-induced changes in AP morphology of isolated cardiac myocytes could provide an opportunity to develop medium throughout assays for the pharmaceutical industry. Ideally, this requires that the dye has a consistent and rapid response to membrane potential, is insensitive to movement, and does not itself affect AP morphology. Materials and methods: We recorded the AP from isolated adult guinea-pig ventricular myocytes optically using di-8-ANEPPS in a single-excitation dual-emission ratiometric system, either separately in electrically field stimulated myocytes, or simultaneously with an electrical AP recorded with a patch electrode in the whole-cell bridge mode. The ratio of di-8-ANEPPS fluorescence signal was calibrated against membrane potential using a switch-clamp to voltage clamp the myocyte. Results: Our data show that the ratio of the optical signals emitted at 560/620 nm is linearly related to voltage over the voltage range of an AP, producing a change in ratio of 7.5% per 100mV, is unaffected by cell movement and is identical to the AP recorded simultaneously with a patch electrode. However, the APD90 recorded optically in myocytes loaded with di-8-ANEPPS was significantly longer than in unloaded myocytes recorded with a patch electrode (355.6 ± 13.5 vs. 296.2 ± 16.2ms; p< 0.01). Despite this effect, the apparent IC50 for cisapride, which prolongs the AP by blocking IKr, was not significantly different whether determined optically or with a patch electrode (91 ± 46 vs. 81 ± 20 nM). Discussion: These data show that the optical AP recorded ratiometrically using di-8- ANEPPS from a single ventricular myocyte accurately follows the action potential morphology. This technique can be used to estimate the AP prolonging effects of a compound, although di-8-ANEPPS itself prolongs APD90. Optical dyes require less technical skills and are less invasive than conventional electrophysiological techniques and, when coupled to ventricular myocytes, decreases animal usage and facilitates higher throughput assays.

This work is concerned with an attempt to clarify the role of cortisone in both the immune complement response and the progression of mouse leukemic tumor.

A implantação embrionária e a placentação em cobaios são caracterizadas pela presença trofoblastos que se destacam da placenta principal, semelhantes ao trofoblasto extra-viloso de humanos. Nestes animais ultrapassam os limites, e podem ser encontrados infiltrados no profundamente no endométrio e no em ambiente externo ultrapassando aos limites da parede uterina. A cobaia desenvolve uma importante estrutura fisiológica de troca materno-embrionária, denominada de placenta vitelina invertida, definidas como membrana fetal destituída parcial ou totalmente do revestimento trofoblástico que permite a exposição do endoderma extra-embrionário em contato direto com o tecido materno. Tais características denotam um mecanismo de controle da resposta imune materna distinta dos paradigmas estabelecidos na reprodução humana e de roedores, assim como ratos e camundongos. Sendo a mais intrigante, a destituição do trofoblasto como célula da interface-materno-fetal que controla a tolerância imune-materna.No presente trabalho, procurou-se estabelecer a organização da placenta vitelina de cobaios a partir da identificação das células que compõe esta membrana extra-embrionária e identificar em que momento ocorre à remoção das células trofoblásticas, e a subseqüente forma de interação das células da placenta vitelina na interface com o tecido materno. Para tanto foram utilizados cobaias fêmeas com idade gestacional conhecida, sacrificadas para coleta de segmentos uterinos nos períodos iniciais da gestação e destinados ao processamento histotógico de embebição em parafina. Na ausência de marcadores celulares específicos conhecidos para cobaios, foram realizados testes prospectivos com reações: citoquímicas de PAS e azul de toluidina (AT; um painel de lectinas biotinadas com afinidade específica para diferentes açúcares; e imunocitoquímica para citoqueratina. As reações realizadas com PAS e AT não identificaram populações celulares com marcação seletiva. Contudo dentre as lectinas tetadas, a Erytrina cristagali lectin (ECL) apresentou reação altamente seletiva para a população de trofoblasto mural (TM) que se origina do trofectoderma, mantendo esta reatividade ao longo da gestação. Esta marcação permitiu avaliar temporal e espacialmente o destino destas células que ao longo da gestação eram mantidas como monocamada de TM revestindo externamente a placenta vitelina e, portanto, não expondo as células do endoderma parietal ou visceral ao ecido materno. Pelo acompanhamento do desenvolvimento embrionário nos cortes seriados, foi constatada no interior do blastocisto a organização de duas massas celulares internas em pólos opostos desde a fase de pré-eclosão. Uma das massas celulares constituída de embrioblastos que dará origem aos os folhetos embrionários nas fases subseqüentes, enquanto a outra formada as células tronco trofoblásticas precursora do cone ectoplacentário (CE). A cavidade da blastocele que separa estas duas massas celulares tem a sua parede revestida pelo endoderma parietal em fase tardia, após a formação da cavidade amniótica. Estes achados demonstram a pecularidade da embriogênese no cobaio, diferente daquelas descritas para humanos e outros roedores, não permitem analogias diretas, o que pode ter contribuído para o equívoco na descrição clássica da organização e constituição da placenta vitelina invertida de constituição córion-amniótica. Isto é, o trofoblasto participa da organização da placenta vitelina inicial e permanece na membrana âmnion-córion-vitelina perfazendo todo o limite do embrião ao longo da gestação. Portanto a hipótese da placenta vitelina parcial ou totalmente invertida baseada na descrição clássica em cobaios é decorrente da interpretação equivocada da embriogênese destes animais. / The guinea pig embryo implantation and placentation is characterized by trophoblast cells detaching from the main placenta in a similar way of human extra-villous trophobasts that deeply intrude inside the endometrium and sometimes also found outside the uterine wall. Furthermore, this animal also develops inverted yolk sac placenta defined as fetal membrane partially or fully devoided of trophoblast sheet that allows extra-embryonic endoderma direct exposition to the maternal environment. These characteristics denote a distinct control mechanism of maternal immune response from the established paradigm for human and rodents (rat and mouse) reproduction, being most intriguing the depriving of trophoblast as cells of maternal-fetal interface regulating the maternal immune tolerance. The present work aimed to establish the organization of guinea pig yolk sac based on identification of cell populations composing this membrane and identification if, or, when the trophoblast cells are removed from and subsequent interaction way of yolk sac cell in interface with maternal tissue. It was used pregnant guinea pig sacrificed on established gestational day to collect uterine fragments on early pregnancy stage and processed by conventional paraffin embedding. Due to absence of known specific cell markers for guinea pig, was performed the prospective evaluation using PAS and toluidine blue (TB) cytochemistry and a screening using a panel of biotinylated lectin specific for different sugars and, anti-cytokeratin. The PAS and TB staining did not identify any specific cell population, however, among the lectins used, Erytrina cristagali lectin (ECL) showed high selective labeling to the trophoblast cells originated from the trophectoderm that was kept through the gestational period. This reaction pattern was useful to evaluate chronologically and topologically the fate of this cell and confirmed the constancy of these cells layering the yolk sac placenta in contact with maternal tissue and therefore, endodermal cells were not exposed to maternal environment. Evaluation of embryo development step by step in the serial sections showed the presence of two inner cell mass in opposite sites inside the pre-hatched blastocyst. One of this, was formed with embryoblast that latter will originate the embryonic sheets and the other formed with trophoblast stem cells (ST) will originate the ectoplacental-cone. The wall of blastocele cavity separate these two inner cell mass was initially covered by a single ECL positive mural trophoblast and only later after the amniotic cavity is formed the extraembyonic endodermal cells migrate from the embryonic sheets to cover internally the blastocele cavity to organize the yolk sac placenta. These findings show the peculiarity of guinea pig embryogenesis, quite different from those described for human and rodents and therefore, does not allow direct analogy and seems to contribute in the misunderstanding of classic description of inverted yolk sac placenta and its cellular organization. It means, the trophoblast cell participates in the early organization of yolk sac placenta and remains in chorioamniotic yolk sac fetal membrane constantly limiting the embryo surface in contact with maternal environment. Therefore, the hypothesis of complete or partially inverted yolk sac placenta seems to be a miss understanding of guinea pig embryogenesis.

O treinamento físico (TF) melhora a resposta imune de indivíduos saudáveis e traz benefícios para o paciente asmático, mas seu papel na resposta alérgica é desconhecido. Objetivo: Avaliar o papel do TF de moderada intensidade na inflamação pulmonar alérgica crônica. Métodos: 54 cobaias, divididas em 4 grupos: grupo controle (C) (não sensibilizados e não treinados), grupo OVA (sensibilizados à ovalbumina (OVA) e não treinados), grupo treinamento físico (TF) (não sensibilizados e submetidos a um TF), e grupo OVA+TF (sensibilizados à OVA e submetidos a um TF). A sensibilização à OVA teve duração de 8 semanas e o programa de TF de 6 semanas iniciando 15 dias após o início da sensibilização. Cada grupo foi dividido em 2 subgrupos. No primeiro foi avaliada a inflamação pulmonar e os níveis de óxido nítrico exalado (NOex) e no segundo, a hiperresponsividade brônquica à metacolina (Mch). Resultados: A sensibilização à OVA induziu a um aumento da densidade de eosinófilos e linfócitos, expressão de IL(interleucina)-4 e IL-13 e na espessura do músculo liso na via aerea assim como espessura do epitélio comparado aos animais não-sensibilizados (p<0,05). Os animais do grupo OVA+TF apresentaram uma redução da densidade de eosinófilos, linfócitos, IL-4 e IL-13 comparado com o grupo OVA (p<0,05). Nem a sensibilização crônica a OVA ou TF influenciaram a expressão das citocinas Th1 (IL-2 e IFN-) ou a expressão das citocinas regulatórias (IL-10 e IL-1-ra) e nos níveis de NOex. Os grupos que realizaram TF tiveram aumento na espessura do epitélio quando comparados com grupos não-treinados embora não há diferença entre os grupos na avaliação da hiperresponsividade brônquica. Conclusão: Nossos resultados sugerem que o TF reduz a inflamação alérgica sem modificar a hiperresponsividade brônquica e o remodelamento das vias aéreas / Background: Aerobic training (TF) has a positive effects on health subjects and bring benefits on the immune system of asthmatic patients. However, its role on allergic immune response remains poorly understood. Objective: To evaluate the effects of TF in chronic allergic inflammation. Methods: Fiftyfour animals, divided in 4 groups: non-trained and non-sensitized (C), nonsensitized and aerobic exercise (TF), ovalbumin sensitized and non-trained (OVA), and sensitized and aerobic exercise (OVA+TF). OVA or saline sensitization was performed during 8 weeks. TF was performed in a treadmill during 6 weeks beginning in the 3rd week of sensitization. Each group were divided in two groups. In the first one, it was evaluated airway inflammation and levels of exhaleted oxide nitric (NOex), on the second, airway hyperresponsiveness to a methacholine (Mch). Results: OVA sensitization induced an increase in the eosinophils and lymphocytes counting, expression of IL-4 and IL-13 and the amount of airway smooth muscle and epithelium thickness compared to non-sensitized animals (p<0.05). Sensitized animals submitted to TF presented a reduction in the eosinophil and lymphocyte counting, expression of IL-4 and IL-13 compared with OVA group (p<0.05) but not OVA-induced changes in airway remodeling (p>0.05). Neither OVA nor TF induced any difference in the expression of Th1 (IL-2 and IFN-) and regulatory cytokine (IL-10 and IL1-ra) and the levels of NOex. Trained groups presented an increase in epithelium thickness as compared to the nontrained groups however we did not find difference between groups on hyperresponsiveness avaliation. Conclusion: Our results suggest that TF reduces allergic airway inflammation without changes in bronchial hyperresponsiveness and airway remodeling

A aferição da pressão intraocular (PIO) é essencial para o exame oftálmico. Objetivou-se estabelecer os valores da curva diária da PIO de porquinhos da índia de diferentes faixas etárias com tonômetro de rebote. A PIO foi aferida às 6, 12, 18 e 24 horas utilizando o tonômetro de rebote (Tonovet®). Os porquinhos da índia foram subdivididos em dois grupos com 10 animais no grupo I e 4 animais no grupo II, considerando as idades, designados por GI (animais entre 2 a 3 meses de idade) e GII (animais entre 2 a 3 anos de idade). Previamente, foram realizados teste da lágrima de Schirmer, prova da fluoresceína, biomicroscopia com lâmpada de fenda e oftalmoscopia indireta em todos animais. O valor médio da pressão intraocular foi de 12,0 ± 1,83 mmHg. Foram encontradas diferenças significativas entres as idades, sendo que os valores médios para o grupo I foram 11,63 ± 0,29 mmHg, e para o grupo II foram 12,82 ± 0,45mmHg (P= 0,0295). Não foram encontradas diferenças entre ambos os olhos (P= 0,7454). Não foram encontradas diferenças significativas nos valores da PIO relacionadas ao sexo (P= 0,1858). Os valores da PIO em porquinhos da índia tiveram alteração nas horas avaliadas. Com base nos resultados obtidos foi possível concluir que os valores da PIO em porquinhos da índia da sofrem alterações ao longo do dia. Os menores valores de pressão intraocular ocorreram às 18 horas. / The measuring of intraocular pressure (IOP) it is essential for the ophthalmic evaluation; the aim of this study was establish the values of the IOP diurnal curve of guinea pigs with different ages using the rebound tonometer (Tonovet®). The animals were divided according to the age in two groups, Group I (GI) constituted with 10 guinea pigs aged between 2 -3 moths, and Group II (GII), 4 animals with 2 -3 years of old. Previously the Schirmer tear test, fluorescein test, biomicroscopy with slit lamp, and direct optalmoscopy were performed on all animals. The men value of IOP was 12.0±0.34 mmHg. Statistical difference were found between ages (mean GI 11.64 ± 0.29 mmHg, and GII 12.83 ± 0.45mmHg), did not have statistical difference related with gender (P= 0.1858) or between eyes (P= 0.7454). In conclusion the guinea pigs had alteration of IOP true the day, the values of IOP are higher in the morning period, decrease true the day and at the night period haves an elevation again.

A simple high pressure liquid chromatography procedure was used to determine 2,4-D and pronamide exposure in spraymen and their dermal absorption and excretion in guinea pigs. Results of dermal application of these herbicides to guinea pigs demonstrated a strong correlation between the applied dermal dose and the urinary residue excretion over the dosage range tested. As the dosage was increased, the urinary excretion of residues was also increased. However, the excretion of 2,4-D amine mixture following dermal treatment of guinea pigs was prolonged as compared to that of pronamide. Residue levels of these compounds were also determined to estimate skin contamination after sampling by filter pads attached to the clothing and arms of agricultural spraymen. Residues in the workers' urine before and after exposure were also determined. Average exposure values of 44.93 mg/hr/man for 2,40D and 0.83 mg/hr/man for pronamide were extrapolated from residue values obtained from analyzing the pads. Little correlation was found between the measured residues from exposed subjects and residues quantified in their urine samples.

<p>Neuropeptide Y (NPY) is an abundant neurotransmitter in the nervous system and forms a family of evolutionarily related peptides together with peptide YY (PYY), pancreatic polypeptide (PP) and polypeptide Y (PY). These peptides are ligands to a family of receptors that mediate a wide range of physiological effects including stimulation of appetite. This work describes the molecular cloning of four novel NPY receptors.</p><p>In rat a receptor called PP1, later renamed Y₄, was cloned and characterized. It displays the highest amino acid sequence identity to the Y₁ receptor. Rat Y₄ differs extensively from human Y₄, cloned subsequently, in both pharmacological properties, tissue distribution, and amino acid sequence with only 75% identity. Rat and human Y₄are the most diverged orthologues in the NPY receptor family.</p><p>In guinea pig, the y₆ receptor gene was found to be a pseudogene with several frameshift mutations. The gene is a pseudogene in human and pig too, but seems to give rise to a functional receptor in mouse and rabbit. This unusual evolutionary situa- tion may be due to inactivation of the gene in a mammalian ancestor and then restoration of expression in mouse and rabbit, but perhaps more likely due to independent inactivations in guinea pig, human and pig.</p><p>In zebrafish, two new intronless receptor genes were cloned. Sequence comparisons suggest that both receptors are distinct from the mammalian receptors Y₁, Y₄ and y₆, hence they were named Ya and Yb. Chromosomal localization provides further support that Ya and Yb may be distinct subtypes. </p><p>The discoveries of the rat Y₄ and zebrafish Ya and Yb receptors were unexpected and show that the NPY receptor family is larger than previously thought.</p>

Neuropeptide Y (NPY) is an abundant neurotransmitter in the nervous system and forms a family of evolutionarily related peptides together with peptide YY (PYY), pancreatic polypeptide (PP) and polypeptide Y (PY). These peptides are ligands to a family of receptors that mediate a wide range of physiological effects including stimulation of appetite. This work describes the molecular cloning of four novel NPY receptors. In rat a receptor called PP1, later renamed Y4, was cloned and characterized. It displays the highest amino acid sequence identity to the Y1 receptor. Rat Y4 differs extensively from human Y4, cloned subsequently, in both pharmacological properties, tissue distribution, and amino acid sequence with only 75% identity. Rat and human Y4 are the most diverged orthologues in the NPY receptor family. In guinea pig, the y6 receptor gene was found to be a pseudogene with several frameshift mutations. The gene is a pseudogene in human and pig too, but seems to give rise to a functional receptor in mouse and rabbit. This unusual evolutionary situa- tion may be due to inactivation of the gene in a mammalian ancestor and then restoration of expression in mouse and rabbit, but perhaps more likely due to independent inactivations in guinea pig, human and pig. In zebrafish, two new intronless receptor genes were cloned. Sequence comparisons suggest that both receptors are distinct from the mammalian receptors Y1, Y4 and y6, hence they were named Ya and Yb. Chromosomal localization provides further support that Ya and Yb may be distinct subtypes. The discoveries of the rat Y4 and zebrafish Ya and Yb receptors were unexpected and show that the NPY receptor family is larger than previously thought.

A aferição da pressão intraocular (PIO) é essencial para o exame oftálmico. Objetivou-se estabelecer os valores da curva diária da PIO de porquinhos da índia de diferentes faixas etárias com tonômetro de rebote. A PIO foi aferida às 6, 12, 18 e 24 horas utilizando o tonômetro de rebote (Tonovet®). Os porquinhos da índia foram subdivididos em dois grupos com 10 animais no grupo I e 4 animais no grupo II, considerando as idades, designados por GI (animais entre 2 a 3 meses de idade) e GII (animais entre 2 a 3 anos de idade). Previamente, foram realizados teste da lágrima de Schirmer, prova da fluoresceína, biomicroscopia com lâmpada de fenda e oftalmoscopia indireta em todos animais. O valor médio da pressão intraocular foi de 12,0 ± 1,83 mmHg. Foram encontradas diferenças significativas entres as idades, sendo que os valores médios para o grupo I foram 11,63 ± 0,29 mmHg, e para o grupo II foram 12,82 ± 0,45mmHg (P= 0,0295). Não foram encontradas diferenças entre ambos os olhos (P= 0,7454). Não foram encontradas diferenças significativas nos valores da PIO relacionadas ao sexo (P= 0,1858). Os valores da PIO em porquinhos da índia tiveram alteração nas horas avaliadas. Com base nos resultados obtidos foi possível concluir que os valores da PIO em porquinhos da índia da sofrem alterações ao longo do dia. Os menores valores de pressão intraocular ocorreram às 18 horas. / The measuring of intraocular pressure (IOP) it is essential for the ophthalmic evaluation; the aim of this study was establish the values of the IOP diurnal curve of guinea pigs with different ages using the rebound tonometer (Tonovet®). The animals were divided according to the age in two groups, Group I (GI) constituted with 10 guinea pigs aged between 2 -3 moths, and Group II (GII), 4 animals with 2 -3 years of old. Previously the Schirmer tear test, fluorescein test, biomicroscopy with slit lamp, and direct optalmoscopy were performed on all animals. The men value of IOP was 12.0±0.34 mmHg. Statistical difference were found between ages (mean GI 11.64 ± 0.29 mmHg, and GII 12.83 ± 0.45mmHg), did not have statistical difference related with gender (P= 0.1858) or between eyes (P= 0.7454). In conclusion the guinea pigs had alteration of IOP true the day, the values of IOP are higher in the morning period, decrease true the day and at the night period haves an elevation again.

A aferição da pressão intraocular (PIO) é essencial para o exame oftálmico. Objetivou-se estabelecer os valores da curva diária da PIO de porquinhos da índia de diferentes faixas etárias com tonômetro de rebote. A PIO foi aferida às 6, 12, 18 e 24 horas utilizando o tonômetro de rebote (Tonovet®). Os porquinhos da índia foram subdivididos em dois grupos com 10 animais no grupo I e 4 animais no grupo II, considerando as idades, designados por GI (animais entre 2 a 3 meses de idade) e GII (animais entre 2 a 3 anos de idade). Previamente, foram realizados teste da lágrima de Schirmer, prova da fluoresceína, biomicroscopia com lâmpada de fenda e oftalmoscopia indireta em todos animais. O valor médio da pressão intraocular foi de 12,0 ± 1,83 mmHg. Foram encontradas diferenças significativas entres as idades, sendo que os valores médios para o grupo I foram 11,63 ± 0,29 mmHg, e para o grupo II foram 12,82 ± 0,45mmHg (P= 0,0295). Não foram encontradas diferenças entre ambos os olhos (P= 0,7454). Não foram encontradas diferenças significativas nos valores da PIO relacionadas ao sexo (P= 0,1858). Os valores da PIO em porquinhos da índia tiveram alteração nas horas avaliadas. Com base nos resultados obtidos foi possível concluir que os valores da PIO em porquinhos da índia da sofrem alterações ao longo do dia. Os menores valores de pressão intraocular ocorreram às 18 horas. / The measuring of intraocular pressure (IOP) it is essential for the ophthalmic evaluation; the aim of this study was establish the values of the IOP diurnal curve of guinea pigs with different ages using the rebound tonometer (Tonovet®). The animals were divided according to the age in two groups, Group I (GI) constituted with 10 guinea pigs aged between 2 -3 moths, and Group II (GII), 4 animals with 2 -3 years of old. Previously the Schirmer tear test, fluorescein test, biomicroscopy with slit lamp, and direct optalmoscopy were performed on all animals. The men value of IOP was 12.0±0.34 mmHg. Statistical difference were found between ages (mean GI 11.64 ± 0.29 mmHg, and GII 12.83 ± 0.45mmHg), did not have statistical difference related with gender (P= 0.1858) or between eyes (P= 0.7454). In conclusion the guinea pigs had alteration of IOP true the day, the values of IOP are higher in the morning period, decrease true the day and at the night period haves an elevation again.