Electric organ polyamines and their effects on the acetylcholine receptor

1. The electric organ of Torpedo nobiliana contained putrescine (PUT), spermidine (SPD), spermine (SPM), and cadaverine (CAD). Traces of acetylated SPD and SPM were occasionaly seen. 2. Upon fractionation of the tissue by differential centrifugation, the polyamines (PA) were found predominantly in the soluble fraction. The postsynaptic membrane fraction, containing a high concentration of acetylcholine receptor (AChR), was proportionally enriched in SPM. The molar ratio of SPM to AChR was approximately two in these membranes. 3. The effect of exogeneous PA on AChR function was studied by two methods: carbamoylcholine (CCh)-dependent 86Rb+ influx into receptor-rich membrane vesicles and [alpha-125I]bungarotoxin (Bgt) binding to the AChR. 4. SPM inhibited both ion influx and the rate of Bgt binding at concentrations above 1 mM, and therefore it appears to act as a competitive antagonist of the AChR. 5. At submicromolar concentrations, and only after preincubation with the receptor-rich membrane, SPM and PUT increased the ion influx by about 20% over control values. 6. Preincubation with 100 nM SPM did not affect the equilibrium binding of iodinated toxin or the rate of toxin binding, and therefore SPM was not uncovering new receptors. 7. By measuring the initial rate of toxin binding after different periods of preincubation with 1 microM CCh, the rate of the slow phase of receptor desensitization was determined. This rate was not changed by 100 nM SPM. 8. Although these results suggest that at low concentrations SPM is a positive modulator of the AChR, the precise mechanism of action is not determined yet.     

Pure MnTBAP selectively scavenges peroxynitrite over superoxide: Comparison of pure and commercial MnTBAP samples to MnTE-2-PyP in two models of oxidative stress injury,an SOD-specific Escherichia coli model and carrageenan-induced pleurisy

MnTBAP is often referred to as an SOD mimic in numerous models of oxidative stress. We have recently reported that pure MnTBAP does not dismute superoxide, but commercial or poorly purified samples are able to perform O₂^·?dismutation with low-to-moderate efficacy via non-innocent Mn-containing impurities. Herein, we show that neither commercial nor pure MnTBAP could substitute for SOD enzyme in a SOD-deficient Escherichia coli model, whereas MnTE-2-PyP-treated SOD-deficient E. coli grew as well as a wild-type strain. This SOD-specific system indicates that MnTBAP does not act as an SOD mimic in vivo. In another model, carrageenan-induced pleurisy in mice, inflammation was evidenced by increased pleural fluid exudate and neutrophil infiltration and activation: these events were blocked by 0.3 mg/kg MnTE-2-PyP and, to a slightly lesser extent, by 10 mg/kg of either MnTBAP. Also, 3-nitrotyrosine formation, an indication of peroxynitrite existence in vivo, was blocked by both compounds; again MnTE-2-PyP was 33-fold more effective. Pleurisy model data indicate that MnTBAP exerts some protective actions in common with MnTE-2-PyP, which are not O₂^·? related and can be fully rationalized if one considers that the common biological role shared by MnTBAP and MnTE-2-PyP is related to their reduction of peroxynitrite and carbonate radical, the latter arising from ONOOCO₂ adduct. The log k_cat (O₂^·?) value for MnTBAP is estimated to be about 3.16, which is ~ 5 and ~ 6 orders of magnitude smaller than the SOD activities of the potent SOD mimic MnTE-2-PyP and Cu,Zn-SOD, respectively. This very low value indicates that MnTBAP is too inefficient at dismuting superoxide to be of any biological impact, which was confirmed in the SOD-deficient E. coli model. The peroxynitrite scavenging ability of MnTBAP, however, is only ~ 2.5 orders of magnitude smaller than that of MnTE-2-PyP and is not significantly affected by the presence of the SOD-active impurities in the commercial MnTBAP sample (log k_red (ONOO^?) = 5.06 for pure and 4.97 for commercial sample). The reduction of carbonate radical is equally fast with MnTBAP and MnTE-2-PyP. The dose of MnTBAP required to yield oxidative stress protection and block nitrotyrosine formation in the pleurisy model is > 1.5 orders of magnitude higher than that of MnTE-2-PyP, which could be related to the lower ability of MnTBAP to scavenge peroxynitrite. The slightly better protection observed with the commercial MnTBAP sample (relative to the pure MnTBAP) could arise from its impurities, which, by scavenging O₂^·?, reduce consequently the overall peroxynitrite and secondary ROS/RNS levels. These observations have profound biological repercussions as they may suggest that the effect of MnTBAP observed in numerous studies may conceivably relate to peroxynitrite scavenging. Moreover, provided that pure MnTBAP is unable to dismute superoxide at any significant extent, but is able to partially scavenge peroxynitrite and carbonate radical, this compound may prove valuable in distinguishing ONOO^?/CO₃^·? from O₂^·? pathways.     

A Reexamination of Correlations of Amino Acids with Particular Secondary Structures

Using the data from Protein Data Bank the correlations of primary and secondary structures of proteins were analyzed. The correlation values of the amino acids and the eight secondary structure types were calculated, where the position of the amino acid and the position in sequence with the particular secondary structure differ at most 25. The diagrams describing these results indicate that correlations are significant at distances between −9 and 10. The results show that the substituents on Cβ or Cγ atoms of amino acid play major role in their preference for particular secondary structure at the same position in the sequence, while the polarity of amino acid has significant influence on α-helices and strands at some distance in the sequence. The diagrams corresponding to polar amino acids are noticeably asymmetric. The diagrams point out the exchangeability of residues in the proteins; the amino acids with similar diagrams have similar local folding requirements. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Volatile constituents of Erodium cicutarium (L.) L’ Hérit. (Geraniaceae)

Essential oils from Erodium cicutarium were obtained by hydrodistillation (samples consisting of entire plants (ec1), leaves and stems (ec2)) and analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC/MS), resulting in a total of 177 components being identified. The essential oils were of a very similar chemical composition and consisted mainly of aliphatic compounds and their derivatives. Fatty acids and fatty acid derived compounds were the most common, 51.3% (ec1) and 60.1% (ec2), followed by carotenoid derived compounds, 12.6% (ec1) and 20.2% (ec2), and then terpenoids, 14.9% (ec1) and 14.2% (ec2). The main constituents in the oils were hexadecanoic acid, 22.8% (ec2) and 35.9% (ec1) and hexahydrofarnesyl acetone, 10.8% (ec2) and 11.6% (ec1). The results obtained differ markedly from those previously reported for the same species.     

Nouveautés de la flore halophile du Littoral croate

Summary The archipelago opposite Senj (Croatian Coast) is very exposed to the Bora hurricanes accompanied by air dryness, salt spray, and periodic coast icing. Therefore the xerohalophytic flora of this area is very interesting. Four new taxa from there are described:Astragalus glacialis n. spec. subsp.glacialis s. s. and subsp.uraganicus n. subsp.,Allium horvatii n. spec., andPinus nigra Arn. subsp.croatica n. subsp., a stenoendemic relict near extinction on the isle of Prvi.     

The contribution of the melanin pathway to overall body pigmentation during ontogenesis of Periplaneta americana

The most prominent colors observed in insects are black or brown, whose production is attributed to the melanin pathway. At present, though, the contribution of this pathway to overall body pigmentation throughout ontogenesis is still lacking. To address this question we examined the roles of 2 key melanin genes (TH and DDC), in embryonic and postembryonic development of the American cockroach, Periplaneta americana. Our results show that the melanin pathway does not contribute to the light brown coloration observed in the first nymphs. However, the dark brown coloration in mid nymphs and adults is produced solely from the melanin pathway. In addition, the DDC RNAi results reveal that it is dopamine melanin, not DOPA melanin, acts as the main contributor in this process. Overall, present study provides a new insight into insect pigmentation suggesting that genetic mechanisms of coloration can change during ontogenesis. Future studies of additional basal insect lineages will be required to assess in more details the generality of this phenomenon.     

Cutting edge: CD8+ T cell priming in the absence of NK cells leads to enhanced memory responses

It is uncertain whether NK cells modulate T cell memory differentiation. By using a genetic model that allows the selective depletion of NK cells, we show in this study that NK cells shape CD8(+) T cell fate by killing recently activated CD8(+) T cells in an NKG2D- and perforin-dependent manner. In the absence of NK cells, the differentiation of CD8(+) T cells is strongly biased toward a central memory T cell phenotype. Although, on a per-cell basis, memory CD8(+) T cells generated in the presence or the absence of NK cells have similar functional features and recall capabilities, NK cell deletion resulted in a significantly higher number of memory Ag-specific CD8(+) T cells, leading to more effective control of tumors carrying model Ags. The enhanced memory responses induced by the transient deletion of NK cells may provide a rational basis for the design of new vaccination strategies.     

Immobilization of lipase from Candida rugosa on Sepabeads(®): the effect of lipase oxidation by periodates

The objective of this paper was the investigation of a suitable Sepabeads^? support and method for immobilization of lipase from Candida rugosa. Three different supports were used, two with amino groups, (Sepabeads^? EC-EA and Sepabeads^? EC-HA), differing in spacer length (two and six carbons, respectively) and one with epoxy group (Sepabeads^? EC-EP). Lipase immobilization was carried out by two conventional methods (via epoxy groups and via glutaraldehyde), and with periodate method for modification of lipase. The results of activity assays showed that lipase retained 94.8% or 87.6% of activity after immobilization via epoxy groups or with periodate method, respectively, while glutaraldehyde method was inferior with only 12.7% of retention. The immobilization of lipase, previously modified by periodate oxidation, via amino groups has proven to be more efficient than direct immobilization of lipase via epoxy groups. In such a way immobilized enzyme exhibited higher activity at high reaction temperatures and higher thermal stability.     

Effects of clonidine preemptive analgesia on acute postoperative pain in abdominal surgery

Preemptive analgesia refers to blockade of afferent nerve fibers before a painful stimulus, which prevents or reduces subsequent pain even beyond the effect of the block. The aim of the study was to compare the effect of clonidine used before and at the end of operation on pain control in abdominal surgery. A total of 77 patients admitted for colorectal surgery were randomly classified into three groups: epidural clonidine before operation, epidural clonidine at the end of operation, and control group. After the operation on patient demand, analgesia with boluses of epidural morphine was instituted. The parameters of postoperative pain level using VAS score (visual analog scale), sedation and analgesics consumption were determined as outcome measures at 1, 2, 6, and 24 h of the operation. Clonidine administered before operation provided lowest pain scores at 6 and 24 h (p < 0.05). Clonidine administered at the end of operation had low pain scores at 1 and 2 h, with a significant pain breakthrough thereafter (6.93 +/- 1.66 at 6 h and 4.04 +/- 2.39 at 24 h) compared with the group administered clonidine before operation (3.60 +/- 2.94 and 3.71 +/- 1.82). Clonidine administered before operation provided less sedation (p < 0.05) and a significantly lower use of analgesics (p < 0.05). Blockade of nociceptive stimulus using the centrally acting alpha2-adrenergic agonist clonidine before the onset of pain stimulus resulted in reduced pain levels, sedation and analgesic requirement.