GENÉTICA DA CONSERVAÇÃO DE Cabralea canjerana (Vell.) Mart. (Meliaceae) EM FRAGMENTOS FLORESTAIS DE MATA ATLÂNTICA NA APA FERNÃO DIAS

A Mata Atlântica é o bioma brasileiro mais severamente afetado pela fragmentação ambiental. A análise da estruturação da diversidade genética, assim como de características demográficas e quantitativas, permite inferir parâmetros populacionais importantes para os programas de conservação de espécies ameaçadas. Cabralea canjerana ssp. canjerana (Meliaceae) é uma espécie arbórea dioica considerada modelo para estudos de conservação da Mata Atlântica. Cento e oitenta e três indivíduos de oito subpopulações de C. canjerana foram coletados em fragmentos florestais na Área de Proteção Ambiental (APA) Fernão Dias, no Sul do Estado de Minas Gerais, Brasil. Utilizando marcadores microssatélites, a diversidade genética foi estimada e contrastada com medidas quantitativas e medidas geográficas dessas oito subpopulações. Elevados níveis de diversidade genética foram encontrados. Uma porção pequena, porém significativa, da variância genética total está estruturada entre as populações (θ = 0,053), que foram estruturadas em dois grupos distintos. As estimativas dos níveis de diversidade genética nas populações localizadas acima de 1.800 m de altitude foram maiores, corroborando a importância dessas populações na manutenção da diversidade genética. A densidade populacional observada também foi maior nos fragmentos de maior altitude (r = 0,849; p-valor = 0,007). Não houve correlação significativa entre as variáveis fenotípicas (altura dos indivíduos e diâmetro a 1,50 m acima do solo) e as medidas de diversidade genética. Esforços conservacionistas para que aumentem o fluxo gênico entre esses fragmentos florestais devem ser estimulados, principalmente entre os fragmentos de baixa altitude e os fragmentos de elevada altitude. É comprovado que populações situadas em elevadas altitudes possuem maior número de indivíduos por hectare e, portanto, maiores índices de diversidade genética. Essas áreas podem ser tratadas como repositório de diversidade genética, e sua manutenção é de extrema importância. O Código Florestal brasileiro determina que essas áreas acima de 1.800 m de altitudes sejam consideradas Áreas de Proteção Permanente (APP).

Internal ambience of bee colonies submitted to strengthening management by adding broods

This study aimed to investigate the ideal brood area to be introduced for strengthening Apis mellifera colonies, considering variations in the hive's internal ambience. Therefore, eight colonies were equipped with data loggers for recording variations in temperature and humidity inside the hives. Then, these colonies were split in four treatments involving the swapping of four, three, two and one brood comb between strong and weak colonies. Growth in the amount of brood and food stores to the four colony strengthening treatments was assessed counting the comb cells to record changes in the brood and food area. The colonies tended to restore their normal thermoregulation within four hours after they had been manipulated and the hive cover had been closed, which quickly increased brood area in the receiving colonies. Results showed that adding up to three brood combs constitute an important alternative for strengthening Apis mellifera colonies because it did not interfere with nest thermoregulation, speeded up population growth in weak colonies and did not affect the development of strong colonies.

Intestinal lesions in pigs affected with postweaning multisystemic wasting syndrome

Samples of mesenteric lymph nodes and intestines from 79 unthrifty 3- to 5-month-old postweaning pigs, confirmed as naturally affected with postweaning multisystemic wasting syndrome (PMWS), were studied. Pigs originated from 12 farms in southern Brazil and were selected on the basis of clinical signs and/or gross lesions suggestive of enteric disorder. Lymphohistiocytic infiltrates of varying intensity were associated with anti-porcine circovirus type 2 (anti-PCV2) immunostaining (IS) in samples of intestines and mesenteric lymph nodes from all pigs. Although most findings were similar to those described in PCV2-associated enteritis, anti-PCV2 IS in association with depletion of the goblet cell mucin stores (24 pigs), diffuse ileal villous atrophy and fusion (18 pigs), and dilatation of the lymphatic vessels (11 pigs) combined or not with lymphangitis were also observed. PCV2 antigen was immunohistochemically demonstrated in the cytoplasm and nuclei from intralesional epithelial cells, histiocytes, and endothelial-like cells in intestinal tissues. Together these findings imply an association with PCV2. The presence of co-infections by Lawsonia intracellularis, Brachyspira spp., Mycobacterium spp., Salmonella spp., rotavirus, parvovirus, coronavirus and enteric calicivirus with PCV2 in the intestinal lesions was investigated.

Padronização da técnica de nanopartícula de ouro não modificada (AuNPs) para detecção de Actinobacillus pleuropneumoniae em pulmões de suínos

Testes diagnósticos baseados na detecção de ácidos nucleicos sem amplificação prévia através da utilização de nanopartículas de ouro (AuNPs) têm sido descritos para várias enfermidades. Este trabalho teve como objetivo desenvolver uma técnica de AuNPs não modificada para detecção de Actinobacillus pleuropneumoniae (App). Utilizaram-se 70 amostras de pulmão de suínos, 17 sem lesão e 53 com lesões características de pneumonia, objetivando a detecção de App. O oligonucleotídeo utilizado foi baseado no gene ApxIV. O teste de AuNPs apresentou sensibilidade de 93,8% e especificidade de 84,6% quando comparado com a detecção pela PCR. Os resultados mostraram boa concordância entre os testes de AuNPs e a PCR, sendo que a técnica pode ser utilizada como alternativa aos testes convencionais, já que é de fácil e rápida execução e não exige infraestrutura e mão de obra especializada.

Resistência de populações de capim-colchão (Digitaria ciliaris) aos herbicidas inibidores da Acetil Co-A Carboxilase

O objetivo do presente trabalho foi avaliar as populações com suspeita de ocorrência de biótipos resistentes de Digitaria ciliaris aos herbicidas inibidores da AcetilCoA Carboxilase (ACCase), por meio de curvas de dose-resposta, bem como estabelecer o grau de resistência cruzada aos herbicidas cicloexanodionas (CHD) e ariloxifenoxipropionatos (APP) desses biótipos. O experimento foi conduzido em casa de vegetação, utilizando-se quatro populações com suspeita de resistência (R1, R2, R3 e R4) e uma população suscetível (S). O delineamento experimental adotado foi o de blocos ao acaso, com quatro repetições, sendo os tratamentos resultado da interação fatorial entre cinco populações, três herbicidas (fluazifop-p-butil, sethoxydim e tepraloxydim) e oito doses de herbicidas (0C, 0,06C, 0,125C, 0,5C, 1C, 2C, 4C e 10C), em que C é a dose comercial recomendada para cada produto. Foram realizadas avaliações de porcentagem de controle aos 28 dias após a aplicação (DAA). A partir dos resultados obtidos, conclui-se que as populações R1, R2, R3 e R4 apresentaram-se como biótipos resistentes aos herbicidas inibidores da ACCase, com diferentes níveis de resistência cruzada aos herbicidas com esse mecanismo de ação.

Molecular Basis for Resistance to Fluazifop-P-Butyl in Itchgrass (Rottboellia cochinchinensis) from Costa Rica

Rottboellia cochinchinensis is an annual grass weed species known as itchgrass, or "caminadora" in America´s Spanish speaking countries, and has become a major and troublesome weed in several crops. The application of fluazifop-P-butyl at recommended rates (125 g a.i. ha-1) was observed to be failing to control itchgrass in a field in San José, Upala county, Alajuela province, Costa Rica. Plants from the putative resistant R. cochinchinensis population survived fluazifop-P-butyl when treated with 250 g a.i. ha-1 (2X label rate) at the three- to four-leaf stage under greenhouse conditions. PCR amplification and sequencing of partial carboxyl transferase domain (CT) of the acetyl-CoA carboxylase (ACCase) gene were used to determine the molecular mechanism of resistance. A single non-synonymous point mutation from TGG (susceptible plants) to TGC (putative resistant plants) that leads to a Trp-2027-Cys substitution was found. This Trp-2027-Cys mutation is known to confer resistance to all aryloxyphenoxyproprionate (APP) herbicides to which fluazifop-P-butyl belongs. To the best of our knowledge, this is the first report of fluazifop-P-butyl resistance and a mutation at position 2027 for a Costa Rican R. cochinchinensis population.

Storage substances in the androgametogenesis and mature pollen grain of Ilex paraguariensis St. Hil. (Aquifoliaceae)

Storage substances such as starch grains, proteins and lipids were studied during the male gametogenesis and in the mature pollen grain of Ilex paraguariensis St. Hil. (Aquifoliaceae). There are two cycles of amylogenesis and amylolyse. The first cycle lasts until the vacuolated stage when the starch is hydrolyzed and amorphous proteins are stored inside the single vacuole. The next cycle begins after mitosis with the formation of the vegetative and generative cells. At this point, the young vegetative cell stores many starch grains that are bigger than in the first cycle. During the maturation of the male gametophyte, the starch is hydrolyzed and it is absent in the mature pollen grain. Small lipid droplets surround the young generative cell after the mitosis of the androspore and are dispersed in the vegetative cytoplasm during its maturity. The relationship between the pollen storage substances and the ontogeny of the layers in the sporoderm, formation of the generative cell, and the male germ unit were discussed.

Control of the phosphorylation of the astrocyte marker glial fibrillary acidic protein (GFAP) in the immature rat hippocampus by glutamate and calcium ions: possible key factor in astrocytic plasticity

The present review describes recent research on the regulation by glutamate and Ca2+ of the phosphorylation state of the intermediate filament protein of the astrocytic cytoskeleton, glial fibrillary acidic protein (GFAP), in immature hippocampal slices. The results of this research are discussed against a background of modern knowledge of the functional importance of astrocytes in the brain and of the structure and dynamic properties of intermediate filament proteins. Astrocytes are now recognized as partners with neurons in many aspects of brain function with important roles in neural plasticity. Site-specific phosphorylation of intermediate filament proteins, including GFAP, has been shown to regulate the dynamic equilibrium between the polymerized and depolymerized state of the filaments and to play a fundamental role in mitosis. Glutamate was found to increase the phosphorylation state of GFAP in hippocampal slices from rats in the post-natal age range of 12-16 days in a reaction that was dependent on external Ca2+. The lack of external Ca2+ in the absence of glutamate also increased GFAP phosphorylation to the same extent. These effects of glutamate and Ca2+ were absent in adult hippocampal slices, where the phosphorylation of GFAP was completely Ca2+-dependent. Studies using specific agonists of glutamate receptors showed that the glutamate response was mediated by a G protein-linked group II metabotropic glutamate receptor (mGluR). Since group II mGluRs do not act by liberating Ca2+ from internal stores, it is proposed that activation of the receptor by glutamate inhibits Ca2+ entry into the astrocytes and consequently down-regulates a Ca2+-dependent dephosphorylation cascade regulating the phosphorylation state of GFAP. The functional significance of these results may be related to the narrow developmental window when the glutamate response is present. In the rat brain this window corresponds to the period of massive synaptogenesis during which astrocytes are known to proliferate. Possibly, glutamate liberated from developing synapses during this period may signal an increase in the phosphorylation

Partial recovery of erythrocyte glycogen in diabetic rats treated with phenobarbital

Erythrocytes may play a role in glucose homeostasis during the postprandial period. Erythrocytes from diabetic patients are defective in glucose transport and metabolism, functions that may affect glycogen storage. Phenobarbital, a hepatic enzyme inducer, has been used in the treatment of patients with non-insulin-dependent diabetes mellitus (NIDDM), increasing the insulin-mediated glucose disposal. We studied the effects of phenobarbital treatment in vivo on glycemia and erythrocyte glycogen content in control and alloxan-diabetic rats during the postprandial period. In control rats (blood glucose, 73 to 111 mg/dl in femoral and suprahepatic veins) the erythrocyte glycogen content was 45.4 ± 1.1 and 39.1 ± 0.8 µg/g Hb (mean ± SEM, N = 4-6) in the femoral artery and vein, respectively, and 37.9 ± 1.1 in the portal vein and 47.5 ± 0.9 in the suprahepatic vein. Diabetic rats (blood glucose, 300-350 mg/dl) presented low (P<0.05) erythrocyte glycogen content, i.e., 9.6 ± 0.1 and 7.1 ± 0.7 µg/g Hb in the femoral artery and vein, respectively, and 10.0 ± 0.7 and 10.7 ± 0.5 in the portal and suprahepatic veins, respectively. After 10 days of treatment, phenobarbital (0.5 mg/ml in the drinking water) did not change blood glucose or erythrocyte glycogen content in control rats. In diabetic rats, however, it lowered (P<0.05) blood glucose in the femoral artery (from 305 ± 18 to 204 ± 45 mg/dl) and femoral vein (from 300 ± 11 to 174 ± 48 mg/dl) and suprahepatic vein (from 350 ± 10 to 174 ± 42 mg/dl), but the reduction was not sufficient for complete recovery. Phenobarbital also stimulated the glycogen synthesis, leading to a partial recovery of glycogen stores in erythrocytes. In treated rats, erythrocyte glycogen content increased to 20.7 ± 3.8 µg/g Hb in the femoral artery and 30.9 ± 0.9 µg/g Hb in the suprahepatic vein (P<0.05). These data indicate that phenobarbital activated some of the insulin-stimulated glucose metabolism steps which were depressed in diabetic erythrocytes, supporting the view that erythrocytes participate in glucose homeostasis

Dose-dependent effects of 17-ß-estradiol on pituitary thyrotropin content and secretion in vitro

We studied the basal and thyrotropin-releasing hormone (TRH) (50 nM) induced thyrotropin (TSH) release in isolated hemipituitaries of ovariectomized rats treated with near-physiological or high doses of 17-ß-estradiol benzoate (EB; sc, daily for 10 days) or with vehicle (untreated control rats, OVX). One group was sham-operated (normal control). The anterior pituitary glands were incubated in Krebs-Ringer bicarbonate medium, pH 7.4, at 37oC in an atmosphere of 95% O2/5% CO2. Medium and pituitary TSH was measured by specific RIA (NIDDK-RP-3). Ovariectomy induced a decrease (P<0.05) in basal TSH release (normal control = 44.1 ± 7.2; OVX = 14.7 ± 3.0 ng/ml) and tended to reduce TRH-stimulated TSH release (normal control = 33.0 ± 8.1; OVX = 16.6 ± 2.4 ng/ml). The lowest dose of EB (0.7 µg/100 g body weight) did not reverse this alteration, but markedly increased the pituitary TSH content (0.6 ± 0.06 µg/hemipituitary; P<0.05) above that of OVX (0.4 ± 0.03 µg/hemipituitary) and normal rats (0.46 ± 0.03 µg/hemipituitary). The intermediate EB dose (1.4 µg/100 g body weight) induced a nonsignificant tendency to a higher TSH response to TRH compared to OVX and a lower response compared to normal rats. Conversely, in the rats treated with the highest dose (14 µg/100 g body weight), serum 17-ß-estradiol was 17 times higher than normal, and the basal and TRH-stimulated TSH release, as well as the pituitary TSH content, was significantly (P<0.05) reduced compared to normal rats and tended to be even lower than the values observed for the vehicle-treated OVX group, suggesting an inhibitory effect of hyperestrogenism. In conclusion, while reinforcing the concept of a positive physiological regulatory role of estradiol on the TSH response to TRH and on the pituitary stores of the hormone, the present results suggest an inhibitory effect of high levels of estrogen on these responses

Outward potassium current oscillations in macrophage polykaryons: extracellular calcium entry and calcium-induced calcium release

Outward current oscillations associated with transient membrane hyperpolarizations were induced in murine macrophage polykaryons by membrane depolarization in the absence of external Na+. Oscillations corresponded to a cyclic activation of Ca2+-dependent K+ currents (IKCa) probably correlated with variations in intracellular Ca2+ concentration. Addition of external Na+ (8 mM) immediately abolished the outward current oscillations, suggesting that the absence of the cation is necessary not only for their induction but also for their maintenance. Oscillations were completely blocked by nisoldipine. Ruthenium red and ryanodine reduced the number of outward current cycles in each episode, whereas quercetin prolonged the hyperpolarization 2- to 15-fold. Neither low molecular weight heparin nor the absence of a Na+ gradient across the membrane had any influence on oscillations. The evidence suggests that Ca2+ entry through a pathway sensitive to Ca2+ channel blockers is elicited by membrane depolarization in Na+-free medium and is essential to initiate oscillations, which are also dependent on the cyclic release of Ca2+ from intracellular Ca2+-sensitive stores; Ca2+ ATPase acts by reducing intracellular Ca2+, thus allowing slow deactivation of IKCa. Evidence is presented that neither a Na+/Ca2+ antiporter nor Ca2+ release from IP3-sensitive Ca2+ stores participate directly in the mechanism of oscillation

Increase in skeletal muscle protein content by the ß-2 selective adrenergic agonist clenbuterol exacerbates hypoalbuminemia in rats fed a low-protein diet

This investigation examined how the nutritional status of rats fed a low-protein diet was affected when the animals were treated with the ß-2 selective agonist clenbuterol (CL). Males (4 weeks old) from an inbred, specific-pathogen-free strain of hooded rats maintained at the Dunn Nutritional Laboratory were used in the experiments (N = 6 rats per group). CL treatment (Ventipulmin, Boehringer-Ingelheim Ltd., 3.2 mg/kg diet for 2 weeks) caused an exacerbation of the symptoms associated with protein deficiency in rats. Plasma albumin concentrations, already low in rats fed a low-protein diet (group A), were further reduced in CL rats (A = 25.05 ± 0.31 vs CL = 23.64 ± 0.30 g/l, P<0.05). Total liver protein decreased below the level seen in either pair-fed animals (group P) or animals with free access to the low-protein diet (A = 736.56 ± 26 vs CL = 535.41 ± 54 mg, P<0.05), whereas gastrocnemius muscle protein was higher than the values normally described for control (C) animals (C = 210.88 ± 3.2 vs CL = 227.14 ± 1.7 mg/g, P<0.05). Clenbuterol-treated rats also showed a reduction in growth when compared to P rats (P = 3.2 ± 1.1 vs CL = -10.2 ± 1.9 g, P<0.05). This was associated with a marked decrease in fat stores (P = 5.35 ± 0.81 vs CL = 2.02 ± 0.16 g, P<0.05). Brown adipose tissue (BAT) cytochrome oxidase activity, although slightly lower than in P rats (P = 469.96 ± 16.20 vs CL = 414.48 ± 11.32 U/BAT x kg body weight, P<0.05), was still much higher than in control rats (C = 159.55 ± 11.54 vs CL = 414.48 ± 11.32 U/BAT x kg body weight, P<0.05). The present findings support the hypothesis that an increased muscle protein content due to clenbuterol stimulation worsened amino acid availability to the liver and further reduced albumin synthesis causing exacerbation of hypoalbuminemia in rats fed a low-protein diet.

Signal transduction induced in Trypanosoma cruzi metacyclic trypomastigotes during the invasion of mammalian cells

Penetration of Trypanosoma cruzi into mammalian cells depends on the activation of the parasite's protein tyrosine kinase and on the increase in cytosolic Ca2+ concentration. We used metacyclic trypomastigotes, the T. cruzi developmental forms that initiate infection in mammalian hosts, to investigate the association of these two events and to identify the various components of the parasite signal transduction pathway involved in host cell invasion. We have found that i) both the protein tyrosine kinase activation, as measured by phosphorylation of a 175-kDa protein (p175), and Ca2+ mobilization were induced in the metacyclic forms by the HeLa cell extract but not by the extract of T. cruzi-resistant K562 cells; ii) treatment of parasites with the tyrosine kinase inhibitor genistein blocked both p175 phosphorylation and the increase in cytosolic Ca2+ concentration; iii) the recombinant protein J18, which contains the full-length sequence of gp82, a metacyclic stage surface glycoprotein involved in target cell invasion, interfered with tyrosine kinase and Ca2+ responses, whereas the monoclonal antibody 3F6 directed at gp82 induced parasite p175 phosphorylation and Ca2+ mobilization; iv) treatment of metacyclic forms with phospholipase C inhibitor U73122 blocked Ca2+ signaling and impaired the ability of the parasites to enter HeLa cells, and v) drugs such as heparin, a competitive IP3-receptor blocker, caffeine, which affects Ca2+ release from IP3-sensitive stores, in addition to thapsigargin, which depletes intracellular Ca2+ compartments and lithium ion, reduced the parasite infectivity. Taken together, these data suggest that protein tyrosine kinase, phospholipase C and IP3 are involved in the signaling cascade that is initiated on the parasite cell surface by gp82 and leads to Ca2+ mobilization required for target cell invasion.

The brain decade in debate: I. Neurobiology of learning and memory

This article is a transcription of an electronic symposium in which some active researchers were invited by the Brazilian Society for Neuroscience and Behavior (SBNeC) to discuss the last decade's advances in neurobiology of learning and memory. The way different parts of the brain are recruited during the storage of different kinds of memory (e.g., short-term vs long-term memory, declarative vs procedural memory) and even the property of these divisions were discussed. It was pointed out that the brain does not really store memories, but stores traces of information that are later used to create memories, not always expressing a completely veridical picture of the past experienced reality. To perform this process different parts of the brain act as important nodes of the neural network that encode, store and retrieve the information that will be used to create memories. Some of the brain regions are recognizably active during the activation of short-term working memory (e.g., prefrontal cortex), or the storage of information retrieved as long-term explicit memories (e.g., hippocampus and related cortical areas) or the modulation of the storage of memories related to emotional events (e.g., amygdala). This does not mean that there is a separate neural structure completely supporting the storage of each kind of memory but means that these memories critically depend on the functioning of these neural structures. The current view is that there is no sense in talking about hippocampus-based or amygdala-based memory since this implies that there is a one-to-one correspondence. The present question to be solved is how systems interact in memory. The pertinence of attributing a critical role to cellular processes like synaptic tagging and protein kinase A activation to explain the memory storage processes at the cellular level was also discussed.

Effect of medroxyprogesterone acetate on thyrotropin secretion in adult and old female rats

Steroid hormones have been implicated in the modulation of TSH secretion; however, there are few and controversial data regarding the effect of progesterone (Pg) on TSH secretion. Medroxyprogesterone acetate (MPA) is a synthetic alpha-hydroxyprogesterone analog that has been extensively employed in therapeutics for its Pg-like actions, but that also has some glucocorticoid and androgen activity. Both hormones have been shown to interfere with TSH secretion. The objective of the present study was to investigate the effects of MPA or Pg administration to ovariectomized (OVX) rats on in vivo and in vitro TSH release and pituitary TSH content. The treatment of adult OVX rats with MPA (0.25 mg/100 g body weight, sc, daily for 9 days) induced a significant (P<0.05) increase in the pituitary TSH content, which was not observed when the same treatment was used with a 10 times higher MPA dose or with Pg doses similar to those of MPA. Serum TSH was similar for all groups. MPA administered to OVX rats at the lower dose also had a stimulatory effect on the in vitro basal and TRH-induced TSH release. The in vitro basal and TRH-stimulated TSH release was not significantly affected by Pg treatment. Conversely, MPA had no effect on old OVX rats. However, in these old rats, ovariectomy alone significantly reduced (P<0.05) basal and TRH-stimulated TSH release in vitro, as well as pituitary TSH content. The results suggest that in adult, but not in old OVX rats, MPA but not Pg has a stimulatory effect on TSH stores and on the response to TRH in vitro.