Resistance of Amaranthus retroflexus to acetolactate synthase inhibitor herbicides in Brazil

When in competition with cotton, Amaranthus retroflexus can cause high yield losses. Due to the limited availability of selective herbicides registered for post emergence control of this weed, the same herbicides have been used repeated times over the last few years, which may have selected resistant biotypes. Biotypes of A. retroflexus collected from the main areas of cotton cultivation in Brazil were submitted to dose-response trials, by applying the herbicides trifloxysulfuron-sodium and pyrithiobac-sodium in doses equivalent to 0, ¼, ½, 1, 2 and 4 times the recommended rates. Resistance to ALS inhibitors was confirmed in biotypes of A. retroflexus. Biotype MS 2 from Mato Grosso do Sul, was cross-resistant to both trifloxysulfuron-sodium and pyrithiobac-sodium, while biotype MS 1 was resistant to trifloxysulfuron-sodium only. Likewise, singular and cross resistance was also confirmed in biotypes from Goiás (GO 3, GO 4 and GO 6), in relation to trifloxysulfuron­sodium and pyrithiobac-sodium. One biotype from Mato Grosso (MT 13) was not resistant to any of the ALS inhibitors evaluated in this work.

MULTIPLE RESISTANCE OF Sagittaria montevidensis BIOTYPES TO ACETOLACTATE SYNTHASE AND PHOTOSYSTEM II INHIBITING HERBICIDES

ABSTRACT The objective of this research was to evaluate the occurance of multiple resistance of Sagittaria montevidensis (SAGMO) biotypes to acetolactate synthase (ALS) and photosystem II (PSII) inhibiting herbicides through dose-response experiments. The experiment was conducted in a greenhouse from October 2012 to March 2013, in Pelotas, RS. The experimental design was completely randomized, with four replications. Treatments were arranged in a triple factorial design: two biotypes of S. montevidensis(SAGMO 35 - susceptible to herbicides and SAGMO 32 - suspected to be multiple resistance to ALS and PSII inhibiting herbicides), four herbicides (penoxsulam, (imazethapyr+imazapic), bentazon and saflufenacil) and 8 rates of these herbicides (1/32x, 1/16x, 1/8x, 1/4x, 1/2x, 0x, 1x, 2x, 4x, 8x, 16x, 32x and 64x). SAGMO 32 biotype presented high levels of resistance to penoxsulam, (imazethapyr+imazapic) and bentazon. For a 50% reduction in dry matter of the resistant biotype rate of 138 and 2.46 times higher than the label required for the susceptible biotype of the herbicides (imazethapyr+imazapic) and bentazon, respectively, are required. Saflufenacil may be used successfully to controlSagittaria montevidensis resistant in irrigated rice.

Disaccharidase levels in normal epithelium of the small intestine of rats with iron-deficiency anemia

Iron-deficiency anemia is the nutritional deficiency most frequently occurring throughout the world, which manifests as a complex systemic disease involving all cells, affecting enzyme activities and modifying protein synthesis. In view of these considerations, the objective of the present study was to determine the effects of iron-deficiency anemia on disaccharidases and on the epithelial morphokinetics of the jejunal mucosa. Newly weaned male Wistar rats were divided into 4 groups of 10 animals each: C6w received a standard ration containing 36 mg elemental iron per kg ration for 6 weeks; E6w received an iron-poor ration (5-8 mg/kg ration) for 6 weeks; C10w received an iron-rich ration (36 mg/kg ration) for 10 weeks; E10w received an iron-poor ration for 6 weeks and then an iron-rich ration (36 mg/kg) for an additional 4 weeks. Jejunal fragments were used to measure disaccharidase content and to study cell proliferation. The following results were obtained: 1) a significant reduction (P<0.001) of animal weight, hemoglobin (Hb), serum iron and total iron-binding capacity (TIBC) in group E6w as compared to C6w; reversal of the alterations in Hb, serum iron and TIBC with iron repletion (E10w = C10w); animal weights continued to be significantly different in groups E10w and C10w. 2) Sucrase and maltase levels were unchanged; total and specific lactase levels were significantly lower in group E6w and this reduction was reversed by iron repletion (E10w = C10w). 3) The cell proliferation parameters did not differ between groups. On the basis of these results, we conclude that lactase production was influenced by iron deficiency and that this fact was not related to changes in cell population and proliferation in the intestinal mucosa

Constitutive nitric oxide synthase (cNOS) activity in Langerhans islets from streptozotocin diabetic rats

Nitric oxide synthase activity was measured in Langerhans islets isolated from control and streptozotocin diabetic rats. The activity of the enzyme was linear up to 150 µg of protein from control rats and was optimal at 0.1 µM calcium, when it was measured after 45 min of incubation at 37oC in the presence of 200 µM arginine. Specific activity of the enzyme was 25 x 10-4 nmol [3H]citrulline 45 min-1 mg protein-1. Streptozotocin diabetic rats exhibited less enzyme activity both in total pancreas homogenate and in isolated Langerhans islets when compared to control animals. Nitric oxide synthase activity measured in control and diabetic rats 15 days after the last streptozotocin injection in the second group of animals corresponded only to a constitutive enzyme since it was not inhibited by aminoguanidine in any of the mentioned groups. Hyperglycemia in diabetic rats may be the consequence of impaired insulin release caused at least in part by reduced positive modulation mediated by constitutive nitric oxide synthase activity, which was dramatically reduced in islets severely damaged after streptozotocin treatment.

Changes in the TBARs content and superoxide dismutase, catalase and glutathione peroxidase activities in the lymphoid organs and skeletal muscles of adrenodemedullated rats

Thiobarbituric acid reactant substances (TBARs) content, and the activities of glucose-6-phosphate dehydrogenase (G6PDh), citrate synthase (CS), Cu/Zn- and Mn-superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX) were measured in the lymphoid organs (thymus, spleen, and mesenteric lymph nodes (MLN)) and skeletal muscles (gastrocnemius and soleus) of adrenodemedullated (ADM) rats. The results were compared with those obtained for sham-operated rats. TBARs content was reduced by adrenodemedullation in the lymphoid organs (MLN (28%), thymus (40%) and spleen (42%)) and gastrocnemius muscle (67%). G6PDh activity was enhanced in the MLN (69%) and reduced in the spleen (28%) and soleus muscle (75%). CS activity was reduced in all tissues (MLN (75%), spleen (71%), gastrocnemius (61%) and soleus (43%)), except in the thymus which displayed an increment of 56%. Cu/Zn-SOD activity was increased in the MLN (126%), thymus (223%), spleen (80%) and gastrocnemius muscle (360%) and was reduced in the soleus muscle (31%). Mn-SOD activity was decreased in the MLN (67%) and spleen (26%) and increased in the thymus (142%), whereas catalase activity was reduced in the MLN (76%), thymus (54%) and soleus muscle (47%). It is particularly noteworthy that in ADM rats the activity of glutathione peroxidase was not detectable by the method used. These data are consistent with the possibility that epinephrine might play a role in the oxidative stress of the lymphoid organs. Whether this fact represents an important mechanism for the establishment of impaired immune function during stress remains to be elucidated.

Evidence for the presence of a kininogen-like species in a case of total deficiency of low and high molecular weight kininogens

Low and high molecular weight kininogens (LK and HK), containing 409 and 626 amino acids with masses of ~65 and 120 kDa after glycosylation, respectively, are coded by a single gene mapped to the human chromosome 3 by alternative splicing of the transcribed mRNA. The NH2-termini Glu1-Thr383 region, identical in LK and HK, contains bradykinin (BK) moieties Arg363-Arg371. LK, HK and their kinin products Lys-BK and BK are involved in several biologic processes. They are evolutionarily conserved and only 7 patients, all apparently normal, have been reported to lack them. In one of these patients (Williams' trait), a codon mutation (Arg178 ® stop) has been blamed for the absence of LK and HK. However, using Western blots with 2 monoclonal anti-HK antibodies, one that recognizes the region common to LK and HK and the other that recognizes only HK, I detected ~110-kDa bands in the plasma of this LK/HK-deficient patient vs ~120-kDa bands in normal human and ape plasmas. With polyclonal anti-Lys-BK antibody, which strongly detects BK cleaved at its COOH-terminus in purified HK, I detected ~110-kDa bands in the normal and the deficient plasmas. Western blots with a monoclonal anti-prekallikrein (PK) antibody showed that surface activation of PK and distribution of PK activation products, both dependent on HK, were similar in these plasmas. These findings suggest that a mutant gene yielded a kininogen-like species possibly involving aberrant mRNA splicing - structurally different from normal HK, but apparently with the capacity to carry out seemingly vital HK functions.

Inhibition of carbachol-induced inositol phosphate accumulation in the embryonic retina promoted by kainate and veratridine

In the present study, we report that low concentrations of the glutamate ionotropic agonist kainate decreased the turnover of [3H]-phosphoinositides ([3H]-InsPs) induced by muscarinic receptors in the chick embryonic retina. When 100 µM carbachol was used, the estimated IC50 value for kainate was 0.2 µM and the maximal inhibition of ~50% was obtained with 1 µM or higher concentrations of the glutamatergic agonist. Our data also show that veratridine, a neurotoxin that increases the permeability of voltage-sensitive sodium channels, had no effect on [3H]-InsPs levels of the embryonic retina. However, 50 µM veratridine, but not 50 mM KCl, inhibited ~65% of the retinal response to carbachol. While carbachol increased [3H]-InsPs levels from 241.2 ± 38.0 to 2044.5 ± 299.9 cpm/mg protein, retinal response decreased to 861.6 ± 113.9 cpm/mg protein when tissues were incubated with carbachol plus veratridine. These results suggest that the accumulation of phosphoinositides induced by activation of muscarinic receptors can be inhibited by the influx of Na+ ions triggered by activation of kainate receptors or opening of voltage-sensitive sodium channels in the chick embryonic retina.

Annual changes in serum calcium and inorganic phosphate levels and correlation with gonadal status of a freshwater murrel, Channa punctatus (Bloch)

Adult Channa punctatus murrels of both sexes (60-80 g) were collected locally from Ramgarh Lake during the second week of every month (10 individuals of each sex/month) throughout the year. Blood samples were collected and analyzed for serum calcium and phosphate levels by the methods of Trinder (1960) and Fiske and Subbarow (1925), respectively. Gonads were fixed to judge the state of maturation of the fish. Males exhibited no change in serum calcium levels throughout the year in correlation with testicular maturation. However, serum phosphate levels exhibited a rise in correlation with the increased gonadosomatic index. Females showed marked seasonal changes in serum calcium and phosphate levels which were associated with ovarian maturation (vitellogenesis).

Erythrocyte glucose-6-phosphate dehydrogenase activity assay and affinity for its substrate under "physiological" conditions

Glucose-6-phosphate dehydrogenase (G6PD) activity and the affinity for its substrate glucose-6-phosphate were investigated under conditions similar to the physiological environment in terms of ionic strength (I: 0.188), cation concentration, pH 7.34, and temperature (37oC). A 12.4, 10.4 and 21.4% decrease was observed in G6PD B, G6PD A+ and G6PD A- activities, respectively. A Km increase of 95.1, 94.4 and 95.4% was observed in G6PD B, G6PD A+ and G6PD A-, respectively, leading to a marked decrease in affinity. In conclusion, the observation of the reduced activity and affinity for its natural substrate reflects the actual pentose pathway rate. It also suggests a much lower NADPH generation, which is crucial mostly in G6PD-deficient individuals, whose NADPH availability is poor.

Vascular changes after cardiopulmonary bypass and ischemic cardiac arrest: roles of nitric oxide synthase and cyclooxygenase

Cardiac surgery involving ischemic arrest and extracorporeal circulation is often associated with alterations in vascular reactivity and permeability due to changes in the expression and activity of isoforms of nitric oxide synthase and cyclooxygenase. These inflammatory changes may manifest as systemic hypotension, coronary spasm or contraction, myocardial failure, and dysfunction of the lungs, gut, brain and other organs. In addition, endothelial dysfunction may increase the occurrence of late cardiac events such as graft thrombosis and myocardial infarction. These vascular changes may lead to increased mortality and morbidity and markedly lengthen the time of hospitalization and cost of cardiac surgery. Developing a better understanding of the vascular changes operating through nitric oxide synthase and cyclooxygenase may improve the care and help decrease the cost of cardiovascular operations.

Neuronal and endothelial nitric oxide synthase gene knockout mice

Targeted disruption of the neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS) genes has led to knockout mice that lack these isoforms. These animal models have been useful to study the roles of nitric oxide (NO) in physiologic processes. nNOS knockout mice have enlarged stomachs and defects in the inhibitory junction potential involved in gastrointestinal motility. eNOS knockout mice are hypertensive and lack endothelium-derived relaxing factor activity. When these animals are subjected to models of focal ischemia, the nNOS mutant mice develop smaller infarcts, consistent with a role for nNOS in neurotoxicity following cerebral ischemia. In contrast, eNOS mutant mice develop larger infarcts, and show a more pronounced hemodynamic effect of vascular occlusion. The knockout mice also show that nNOS and eNOS isoforms differentially modulate the release of neurotransmitters in various regions of the brain. eNOS knockout mice respond to vessel injury with greater neointimal proliferation, confirming that reduced NO levels seen in endothelial dysfunction change the vessel response to injury. Furthermore, eNOS mutant mice still show a protective effect of female gender, indicating that the mechanism of this protection cannot be limited to upregulation of eNOS expression. The eNOS mutant mice also prove that eNOS modulates the cardiac contractile response to ß-adrenergic agonists and baseline diastolic relaxation. Atrial natriuretic peptide, upregulated in the hearts of eNOS mutant mice, normalizes cGMP levels and restores normal diastolic relaxation.

Targeting and translocation of endothelial nitric oxide synthase

This review explores advances in our understanding of the intracellular regulation of the endothelial isoform of nitric oxide synthase (eNOS) in the context of its dynamically regulated subcellular targeting. Nitric oxide (NO) is a labile molecule, and may play important biological roles both within the cell in which it is synthesized and in its interactions with nearby cells and molecules. The localization of eNOS within the cell importantly influences the biological role and chemical fate of the NO produced by the enzyme. eNOS, a Ca2+/calmodulin-dependent enzyme, is subject to a complex pattern of intracellular regulation, including co- and post-translational modifications and interactions with other proteins and ligands. In endothelial cells and cardiac myocytes eNOS is localized in specialized plasmalemmal signal-transducing domains termed caveolae; acylation of the enzyme by the fatty acids myristate and palmitate is required for targeting of the protein to caveolae. Targeting to caveolae facilitates eNOS activation following receptor stimulation. In resting cells, eNOS is tonically inhibited by its interactions with caveolin, the scaffolding protein in caveolae. However, following agonist activation, eNOS dissociates from caveolin, and nearly all the eNOS translocates to structures within the cell cytosol; following more protracted incubations with agonists, most of the cytosolic enzyme subsequently translocates back to the cell membrane. The agonist-induced internalization of eNOS is completely abrogated by chelation of intracellular Ca2+. These rapid receptor-mediated effects are seen not only for "classic" eNOS agonists such as bradykinin, but also for estradiol, indicating a novel non-genomic role for estrogen in eNOS activation. eNOS targeting to the membrane is labile, and is subject to receptor-regulated Ca2+-dependent reversible translocation, providing another point for regulation of NO-dependent signaling in the vascular endothelium.

Effects of a neuronal nitric oxide synthase inhibitor on lipopolysaccharide-induced fever

It has been demonstrated that nitric oxide (NO) has a thermoregulatory action, but very little is known about the mechanisms involved. In the present study we determined the effect of neuronal nitric oxide synthase (nNOS) inhibition on thermoregulation. We used 7-nitroindazole (7-NI, 1, 10 and 30 mg/kg body weight), a selective nNOS inhibitor, injected intraperitoneally into normothermic Wistar rats (200-250 g) and rats with fever induced by lipopolysaccharide (LPS) (100 µg/kg body weight) administration. It has been demonstrated that the effects of 30 mg/kg of 7-NI given intraperitoneally may inhibit 60% of nNOS activity in rats. In all experiments the colonic temperature of awake unrestrained rats was measured over a period of 5 h at 15-min intervals after intraperitoneal injection of 7-NI. We observed that the injection of 30 mg/kg of 7-NI induced a 1.5oC drop in body temperature, which was statistically significant 1 h after injection (P<0.02). The coinjection of LPS and 7-NI was followed by a significant (P<0.02) hypothermia about 0.5oC below baseline. These findings show that an nNOS isoform is required for thermoregulation and participates in the production of fever in rats.

Zinc accumulation in phosphate granules of Ucides cordatus hepatopancreas

Amorphous phosphate granules are present in vertebrate and invertebrate organisms. The functions attributed to these structures depend on their mineral contents and organic matrix composition. In the present study we have determined zinc concentrations in the hepatopancreas of the crab Ucides cordatus from regions contaminated with zinc, and the elemental composition of hepatopancreal phosphate granules. Organisms were collected from the contaminated areas of Sepetiba Bay (SB) and Guanabara Bay (GB), and from a non-contaminated area, Ribeira Bay (RB). The first two sites are located near the metropolitan region of Rio de Janeiro city, Brazil. Atomic absorption spectroscopy (AAS) showed a significant difference (P<0.05) for zinc concentration in the hepatopancreas from organisms collected at the contaminated sites GB (210 ± 20 µg/g dry weight) and SB (181 ± 16 µg/g dry weight) compared to the non-contaminated site RB (76 ± 14 µg/g dry weight). Phosphate granules isolated from hepatopancreatic tissue were studied by electron diffraction (ED), energy dispersive X-ray analysis (EDX) and electron spectroscopic imaging (ESI). ED of granules presented no diffraction spots, indicating that these structures are in an amorphous state, while EDX of granules isolated from a contaminated area contained P, Ca and Zn. Mg, Cl and Fe were also found in some of the spectra. ESI showed that O, P and Ca were colocalized in the mineralized layers of most granules observed. The correlation between the results obtained by AAS and those obtained by microanalytical techniques suggests that the hepatopancreatic granules of U. cordatus may be related to the phenomenon of heavy metal retention.