Distribution of the a2, a3, and a5 nicotinic acetylcholine receptor subunits in the chick brain

Nicotinic acetylcholine receptors (nAChRs) are ionotropic receptors comprised of a and ß subunits. These receptors are widely distributed in the central nervous system, and previous studies have revealed specific patterns of localization for some nAChR subunits in the vertebrate brain. In the present study we used immunohistochemical methods and monoclonal antibodies to localize the a2, a3, and a5 nAChR subunits in the chick mesencephalon and diencephalon. We observed a differential distribution of these three subunits in the chick brain, and showed that the somata and neuropil of many central structures contain the a5 nAChR subunit. The a2 and a3 subunits, on the other hand, exhibited a more restricted distribution than a5 and other subunits previously studied, namely a7, a8 and ß2. The patterns of distribution of the different nAChR subunits suggest that neurons in many brain structures may contain several subtypes of nAChRs and that in a few regions one particular subtype may determine the cholinergic nicotinic responses

Possible involvement of A1 receptors in the inhibition of gonadotropin secretion induced by adenosine in rat hemipituitaries in vitro

We investigated the participation of A1 or A2 receptors in the gonadotrope and their role in the regulation of LH and FSH secretion in adult rat hemipituitary preparations, using adenosine analogues. A dose-dependent inhibition of LH and FSH secretion was observed after the administration of graded doses of the R-isomer of phenylisopropyladenosine (R-PIA; 1 nM, 10 nM, 100 nM, 1 µM and 10 µM). The effect of R-PIA (10 nM) was blocked by the addition of 8-cyclopentyltheophylline (CPT), a selective A1 adenosine receptor antagonist, at the dose of 1 µM. The addition of an A2 receptor-specific agonist, 5-N-methylcarboxamidoadenosine (MECA), at the doses of 1 nM to 1 µM had no significant effect on LH or FSH secretion, suggesting the absence of this receptor subtype in the gonadotrope. However, a sharp inhibition of the basal secretion of these gonadotropins was observed after the administration of 10 µM MECA. This effect mimicked the inhibition induced by R-PIA, supporting the hypothesis of the presence of A1 receptors in the gonadotrope. R-PIA (1 nM to 1 µM) also inhibited the secretion of LH and FSH induced by phospholipase C (0.5 IU/ml) in a dose-dependent manner. These results suggest the presence of A1 receptors and the absence of A2 receptors in the gonadotrope. It is possible that the inhibition of LH and FSH secretion resulting from the activation of A1 receptors may have occurred independently of the increase in membrane phosphoinositide synthesis.

Evidence for the expression of native Mycobacterium tuberculosis phospholipase C: recognition by immune sera and detection of promoter activity

The genome of Mycobacterium tuberculosis H37Rv contains three contiguous genes (plc-a, plc-b and plc-c) which are similar to the Pseudomonas aeruginosa phospholipase C (PLC) genes. Expression of mycobacterial PLC-a and PLC-b in E. coli and M. smegmatis has been reported, whereas expression of the native proteins in M. tuberculosis H37Rv has not been demonstrated. The objective of the present study was to demonstrate that native PLC-a is expressed in M. tuberculosis H37Rv. Sera from mice immunized with recombinant PLC-a expressed in E. coli were used in immunoblots to evaluate PLC-a expression. The immune serum recognized a 49-kDa protein in immunoblots against M. tuberculosis extracts. No bands were visible in M. tuberculosis culture supernatants or extracts from M. avium, M. bovis and M. smegmatis. A 550-bp DNA fragment upstream of plc-a was cloned in the pJEM12 vector and the existence of a functional promoter was evaluated by detection of ß-galactosidase activity. ß-Galactosidase activity was detected in M. smegmatis transformed with recombinant pJEM12 grown in vitro and inside macrophages. The putative promoter was active both in vitro and in vivo, suggesting that expression is constitutive. In conclusion, expression of non-secreted native PLC-a was demonstrated in M. tuberculosis.

Adenosine A1 receptor-mediated inhibition of in vitro prolactin secretion from the rat anterior pituitary

In previous studies, we demonstrated biphasic purinergic effects on prolactin (PRL) secretion stimulated by an adenosine A2 agonist. In the present study, we investigated the role of the activation of adenosine A1 receptors by (R)-N6-(2-phenylisopropyl)adenosine (R-PIA) at the pituitary level in in vitro PRL secretion. Hemipituitaries (one per cuvette in five replicates) from adult male rats were incubated. Administration of R-PIA (0.001, 0.01, 0.1, 1, and 10 µM) induced a reduction of PRL secretion into the medium in a U-shaped dose-response curve. The maximal reduction was obtained with 0.1 µM R-PIA (mean ± SEM, 36.01 ± 5.53 ng/mg tissue weight (t.w.)) treatment compared to control (264.56 ± 15.46 ng/mg t.w.). R-PIA inhibition (0.01 µM = 141.97 ± 15.79 vs control = 244.77 ± 13.79 ng/mg t.w.) of PRL release was blocked by 1 µM cyclopentyltheophylline, a specific A1 receptor antagonist (1 µM = 212.360 ± 26.560 ng/mg t.w.), whereas cyclopentyltheophylline alone (0.01, 0.1, 1 µM) had no effect. R-PIA (0.001, 0.01, 0.1, 1 µM) produced inhibition of PRL secretion stimulated by both phospholipase C (0.5 IU/mL; 977.44 ± 76.17 ng/mg t.w.) and dibutyryl cAMP (1 mM; 415.93 ± 37.66 ng/mg t.w.) with nadir established at the dose of 0.1 µM (225.55 ± 71.42 and 201.9 ± 19.08 ng/mg t.w., respectively). Similarly, R-PIA (0.01 µM) decreased (242.00 ± 24.00 ng/mg t.w.) the PRL secretion stimulated by cholera toxin (0.5 mg/mL; 1050.00 ± 70.00 ng/mg t.w.). In contrast, R-PIA had no effect (468.00 ± 34.00 ng/mg t.w.) on PRL secretion stimulation by pertussis toxin (0.5 mg/mL; 430.00 ± 26.00 ng/mg t.w.). These results suggest that inhibition of PRL secretion after A1 receptor activation by R-PIA is mediated by a Gi protein-dependent mechanism.

Negative correlation between phospholipase and esterase activity produced by Fusarium isolates

Fusarium species have emerged as one of the more outstanding groups of clinically important filamentous fungi, causing localized and life-threatening invasive infections with high morbidity and mortality. The ability to produce different types of hydrolytic enzymes is thought to be an important virulence mechanism of fungal pathogens and could be associated with the environment of the microorganism. Here, we have measured the production of two distinct lipolytic enzymes, phospholipase and esterase, by sixteen Fusarium isolates recovered from the hospital environment, immunocompromised patients’ blood cultures, foot interdigital space scrapings from immunocompromised patients, and foot interdigital space scrapings from immunocompetent patients (4 isolates each). Fourteen of these 16 isolates were identified asFusarium solani species complex (FSSC) and two were identified as F. oxysporum species complex (FOSC). Some relevant genus characteristics were visualized by light and electron microscopy such as curved and multicelled macroconidia with 3 or 4 septa, microconidia, phialides, and abundant chlamydospores. All Fusarium isolates were able to produce esterase and phospholipase under the experimental conditions. However, a negative correlation was observed between these two enzymes, indicating that a Fusarium isolate with high phospholipase activity has low esterase activity and vice versa. In addition, Fusarium isolated from clinical material produced more phospholipases, while environmental strains produced more esterases. These observations may be correlated with the different types of substrates that these fungi need to degrade during their nutrition processes.

Estudo das latências e amplitudes dos potenciais evocados auditivos de média latência em indivíduos audiologicamente normais

Estudo de coorte contemporânea com corte transversal. O Potencial Evocado Auditivo de Média Latência (PEAML) é gerado entre 10 e 80ms e possui múltiplos geradores, com maior contribuição da região tálamo-cortical. O estabelecimento de critérios de normalidade para os valores de latência e amplitude é necessário para uso clínico. OBJETIVOS: Analisar a latência e amplitude do PEAML em indivíduos sem alterações audiológicas, e verificar a confiabilidade da amplitude Pa-Nb. MATERIAL E MÉTODO: Foram coletados os PEAML de 25 indivíduos durante o ano de 2005 e analisados os componentes Na, Pa, Nb para cada orelha testada (A1 e A2), e posicionamento de eletrodo (C3 e C4). RESULTADOS: Observou-se diferença estatisticamente significante entre os valores médios de latência para C3A1 e C4A1 com relação aos componentes Na e Pa, não sendo encontrada esta diferença para o componente Nb e valores médios das amplitudes Na-Pa e Pa-Nb. CONCLUSÃO: Foram estabelecidos os valores das médias e desvios padrão para os parâmetros latência e amplitude dos componentes Na, Pa, Nb, e Na-Pa e Pa-Nb, nas condições C3A1, C4A1, C3A2, C4A2, proporcionando os parâmetros para a análise e interpretação deste potencial.

Avaliação da função cognitiva da aprendizagem em crianças com distúrbios obstrutivos do sono

Os distúrbios obstrutivos do sono são relativamente freqüentes na população pediátrica, porém o impacto da perda do sono na aprendizagem e função cognitiva não está bem estabelecido. OBJETIVO: Avaliar se pacientes com distúrbios obstrutivos do sono apresentam alteração de aprendizagem, memória e atenção. CASUÍSTICA E MÉTODO: Foram avaliadas 81 crianças de 6 a 12 anos de idade, divididas em 3 grupos: grupo SAHOS (n=24), grupo Ronco Primário (n=37) e grupo Controle (n=20), através de testes de aprendizagem (Teste de Rey) e cognitivos (Dígito, Código, Cancelamento de Letras e Símbolos). Todas as crianças realizaram polissonografia. RESULTADOS: O grupo SAHOS (n=24) e o grupo Ronco Primário (n=37) apresentaram diferença estatisticamente significante nas variáveis A1 (p=0,001) do Teste de Rey quando comparados ao grupo controle. O grupo Ronco Primário apresentou ainda diferenças nas variáveis A2, A4, AT e A6 do Teste de Rey (p=0,020; p=0,05; p=0,004; p=0,05, respectivamente) em relação ao grupo controle (n=20). CONCLUSÃO: Crianças com distúrbios obstrutivos do sono apresentam piores resultados no teste de aprendizagem e memória (Teste de Rey), principalmente o grupo RP, quando este é comparado ao grupo SAHOS. Os testes de atenção apresentam resultados semelhantes entre os grupos.

Volumes de substratos comerciais, solo e composto orgânico afetando a formação de mudas de maracujazeiro-amarelo em diferentes ambientes de cultivo

Este trabalho teve como objetivo avaliar o crescimento de mudas de maracujazeiro- amarelo em três diferentes tamanhos de recipientes, em três condições de cultivo protegido, utilizando seis diferentes substratos. Foram empregados três ambientes de cultivo: (A1) estufa em arco, coberta de filme de polietileno de 150 μm, abertura zenital e tela termorrefletora de 50% sob o filme; (A2) viveiro agrícola de tela de monofilamento, com 50% de sombra; e (A3) viveiro agrícola de tela termorrefletora, com 50% de sombra. Foram testados três volumes (V1 = xx cm³ -sacolas de polietileno de 7,5 x 11,5 cm; V2 = xx cm33 - sacolas de polietileno 10,0 x 16,5 cm; e V3 = xx cm33 -sacolas de polietileno de 15,0 x 21,5 cm) e seis substratos (S1 = solo; S2 = Plantmax®; S3 = vermiculita; S4 = fibra de coco; S5 = fibra de coco chips; e S6 = Organosuper®, composto orgânico comercial). O delineamento experimental foi inteiramente casualizado, em esquema de parcelas subsubdivididas, em que os ambientes foram as parcelas, os recipientes de diferentes volumes as subparcelas e os substratos as subsubparcelas. Aos 50 dias após a semeadura, foram medidos a altura das plantas, o comprimento da raiz, a massa de matéria seca da parte aérea e das raízes. A partir das massas de matéria seca determinaram-se a relação massa de matéria seca de parte aérea e raiz e massa de matéria seca total. A estufa foi o melhor ambiente quando se utilizou o recipiente de XX cm33 o qual proporcionou mudas maiores, com maior biomassa seca aérea, radicular e total. A vermiculita foi o melhor substrato, porém o solo adubado é uma alternativa menos dispendiosa para a região.

Deslizamento de rodados de tratores de pneus em diferentes operações agrícolas

Nos tratores agrícolas, o deslizamento dos rodados ocorre por diversos fatores, entre eles o esforço de tração necessário para deslocar determinado implemento e o tipo de superfície que está em contato com a banda de rodagem dos pneus. Este trabalho tem o objetivo de avaliar o deslizamento do rodado de pneus de um trator em diferentes sistemas de preparo do solo na cultura do milho, no município de Bambuí, Estado de Minas Gerais. O experimento foi conduzido a campo, na safrinha outonal de 2011, na Fazenda Varginha, do Instituto Federal Minas Gerais - campus Bambuí, em um Latossolo Vermelho distroférrico típico. O experimento foi montado em esquema de parcelas sub-subdivididas, tendo, nas parcelas, as operações, nas subparcelas as marchas e, nas sub-subparcelas, as rotações, no delineamento em blocos casualizados, com três repetições, totalizando 81 unidades experimentais. Os fatores e níveis estudados foram (PC) preparo convencional, (CM) cultivo mínimo e (PD) plantio direto. Para cada tratamento, foram utilizadas três marchas (A2, A3 e B1), em três rotações (1600, 1900 e 2200 rpm). Pelos resultados, conclui-se que, no preparo convencional, o maior índice de deslizamento ocorre na operação de semeadura; no cultivo mínimo, na operação de escarificação e, no plantio direto, na operação de semeadura.

The measurement by serological means of the impact of the Hong Kong/68 influenza virus on a population

In order to obtain evidence on the size of the impact of the Hong Kong/68 variant of influenza A2 virus on the population of São Paulo, Brazil, serum samples taken in 1967 before this variant appeared and during successive years after it appeared were examined for their antibody content. Haemagglutination-inhibition tests performed on a total of 2726 serum samples from adults showed a sharp decrease in 1969 of the proportion of sera without antibody to the Hong Kong/68 variant and a corresponding mercase in the proportion with high titres. It was concluded that about three-quarters of the adult population became infected at some time after the variant appeared, the majority in the first year of prevalence.

A eficácia do antiveneno botrópico-crotálico na neutralização das principais atividades do veneno de Bothrops jararacussu

A mionecrose é um dos efeitos causados pelo veneno de Bothrops jararacussu. Uma miotoxina com homologia estrutural à fosfolipase A2 (PLA2), mas sem atividade enzimática, foi isolada desse veneno. O veneno de Crotalus durissus terrificus apresenta também atividade miotóxica, que vem sendo atribuída à crotoxina e à PLA2 (crotoxina B), o componente básico do complexo crotoxina. O veneno de Bothrops jararacussu apresenta três proteínas, que têm identidade imunológica com a PLA2 da crotoxina. O presente trabalho comparou a eficiência dos antivenenos polivalentes comerciais produzidos pelo Instituto Butantan - o antiveneno botrópico (AB) e o antiveneno botrópico-crotálico (AB/C) - na neutralização das atividades letal, hemorrágica, coagulante e miotóxica do veneno de B. jararacussu. Os dois antivenenos neutralizaram de maneira semelhante a atividade hemorrágica, mas o AB/C foi três vezes mais potente que o AB em neutralizar a ação miotóxica e duas vezes mais potente na neutralização da letalidade e na ação coagulante do veneno de B. jararacussu. Os dados sugerem que a utilização do AB/C pode ser vantajosa no tratamento de pacientes picados por serpentes dessa espécie

Heparin-antivenom association: differential neutralization effectiveness in Bothrops atrox and Bothrops erythromelas envenoming

Heparin, in some regions of Brazil has been used in the treatment of bothropic accidents, but the data found in the literature are inconclusive about its effectiveness. The venoms of Bothrops atrox and of B. erythromelas were characterized according to their biological activities. The capacity of heparin in neutralizing these activities was tested with doses of 3 and 6 IU in isolated form and associated to Antibothropic Serum (ABS). It was verified that heparin, in doses of 3 and 6 IU, was not effective in neutralizing the desfibrinating and edema-forming activities of B. atrox venom and the hemorrhagic and coagulant actions of both venoms. Heparin diminished the effectiveness of the ABS in the neutralization of the hemorrhagic and edema-forming activities of the B. atrox venom. However, heparin in the 6 IU dose was capable of neutralize the edema-forming of the B. erythromelas and increase the effectiveness of the ABS. Heparin also neutralized the phospholipasic A2 activity of B. atrox (14.3%) and B. erythromelas (28.0%) venoms. For B. erythromelas venom, the associated treatment, heparin and ABS, was more effective in the neutralization of its lethal activity.

Antifungal susceptibilities of clinical and environmental isolates of Cryptococcus neoformans in goiânia city, Goiás, Brasil

We evaluated the antifungal activities of amphotericin B, fluconazole, itraconazole and voriconazole in 70 Cryptococcus neoformans strains obtained from cerebrospinal fluid from AIDS patients and 40 C. neoformans strains isolated from the environment. Four clinical isolates were identified as C. neoformans var. gattii. The susceptibility test was done using a broth microdilution method according to NCCLS M27-A2. Range minimal inhibitory concentrations (MICs) for C. neoformans clinical isolates were 0.06-1.0 µg/mL for amphotericin B, 0.125-8 µg/mL for fluconazole, 0.03-0.5 µg/mL for itraconazole and 0.03-0.25 µg/mL for voriconazole. C. neoformans environmental isolates showed range MICs 0.015-0.125 µg/mL, 0.25-2.0 µg/mL, 0.007-0.125 µg/mL and 0.03-0.25 µg/mL for amphotericin B, fluconazole, itraconazole and voriconazole respectively. The MICs results obtained from clinical and environmental isolates showed similar pattern of susceptibility and no resistance has been found in our isolates.

Genotyping, serotyping and determination of mating-type of Cryptococcus neoformans clinical isolates from São Paulo State, Brazil

The basidiomycetous yeast Cryptococcus neoformans is an important fungal pathogen mainly in immunocompromised patients. In this study, 47 clinical isolates of C. neoformans from regions of São Paulo State were studied serologically by using the Crypto Check Iatron RM 304-K kit, their genetic diversity was estimated by PCR-fingerprinting with a microsatellite-specific sequence (GACA)4, RAPD with primer 6 (Amersham Pharmacia Biotech), PCR-restriction fragment length polymorphism (RFLP) analysis of the phospholipase B gene (PLB1) digested with AvaI and mating type analysis by PCR. All 47 strains isolated from HIV positive patients included in this study were serotype A and MATalpha. The majority of the isolates (45/47) were VNI and only two were VNII by PCR-fingerprinting and PCR-RFLP analysis. High degree of homogeneity was observed when (GACA)4 was used, being highly correlated (> 0.9). In contrast, the RAPD analysis was more heterogeneous with higher number of molecular profiles. By PCR-RFLP, no new molecular type was found, enhancing the suggestion that the differences based on conserved gene as PLB1, can be resultant of ongoing divergent evolution within the C. neoformans complex, into the current eight subtypes. Our results furnish new information on the molecular epidemiology of C. neoformans in the southeast region of Brazil.

Candiduria in a public hospital of São Paulo (1999-2004): characteristics of the yeast isolates

The study involved 100 yeast isolates, obtained from urine samples provided by a Public Pediatric Hospital of São Paulo, Brazil, from 1999 to 2004. The most frequent species was Candida albicans, followed by C. tropicalis, C. glabrata and C. parapsilosis. In regard to virulence, 97% of the isolates showed index 3 for proteinase and 63% index 2 for phospholipase. The most frequent killer biotypes were 511 and 888.