The Distribution of Primary Nitric Oxide Synthase- and Parvalbumin- Immunoreactive Afferents in the Dorsal Funiculus of the Lumbosacral Spinal Cord in a Dog

1. The aim of the present study was to examine the distribution of unmyelinated, small-diameter myelinated neuronal nitric oxide synthase immunoreactive (nNOS-IR) axons and large-diameter myelinated neuronal nitric oxide synthase and parvalbumin-immunoreactive (PV-IR) axons in the dorsal funiculus (DF) of sacral (S1–S3) and lumbar (L1–L7) segments of the dog. 2. nNOS and PV immunohistochemical methods were used to demonstrate the presence of nNOS-IR and PV-IR in the large-diameter myelinated, presumed to be proprioceptive, axons in the DF along the lumbosacral segments. 3. Fiber size and density of nNOS-IR and PV-IR axons were used to compartmentalize the DF into five compartments (CI–CV). The first compartment (CI) localized in the lateralmost part of the DF, containing both unmyelinated and small-diameter myelinated nNOS-IR axons, is homologous with the dorsolateral fasciculus, or Lissauer tract. The second compartment (CII) having similar fiber organization as CI is situated more medially in sacral segments. Rostrally, in lower lumbar segments, CII moves more medially, and at upper lumbar level, CII reaches the dorsomedial angle of the DF and fuses with axons of CIV. CIII is the largest in the DF and the only one containing large-diameter myelinated nNOS-IR and PV-IR axons. The largest nNOS-IR and PV-IR axons of CIII (8.0–9.2 μm in diameter), presumed to be stem Ia proprioceptive afferents, are located in the deep portion of the DF close to the dorsal and dorsomedial border of the dorsal horn. The CIV compartment varies in shape, appearing first as a small triangular area in S3 and S2 segments, homologous with the Philippe–Gombault triangle. Beginning at S1 level, CIV acquires a more elongated shape and is seen throughout the lumbar segments as a narrow band of fibers extending just below the dorsal median septum in approximately upper two-thirds of the DF. The CV is located in the basal part of the DF. In general, CV is poor in nNOS-IR fibers; among them solitary PV-IR fibers are seen. 4. The analysis of the control material and the degeneration of the large- and medium-caliber nNOS-IR fibers after unilateral L7 and S1 dorsal rhizotomy confirmed that large-caliber nNOS-IR and and PV-IR axons, presumed to be proprioceptive Ia axons, and their ascending and descending collaterals are present in large number in the DF of the lumbosacral intumescence. However, in the DF of the upper lumbar segments, the decrease in the number of nNOS-IR and PV-IR fibers is quite evident.     

Treatment of osteoporosis with a modified zeolite shows beneficial effects in an osteoporotic rat model and a human clinical trial

The severity of osteoporosis in humans manifests in its high incidence and by its complications that diminish quality of life. A societal consequence of osteoporosis is the substantial burden that it inflicts upon patients and their families. Several bone-modifying drugs have been prescribed to patients with osteoporosis. However, evidence for their anti-fracture efficacy remains inconclusive. To the contrary, long-term use of anti-osteoporotic drugs such as bisphosphonates and Denosumab, an RANKL inhibitor, have resulted in adverse events. We now present an alternative and adjuvant approach for treatment of osteoporosis. The data derive from in vivo studies in an ovariectomized rat model and from a randomized double blind, placebo-controlled human clinical study. Both studies involved treatment with Panaceo Micro Activation (PMA)-zeolite-clinoptilolite, a defined cation exchange clinoptilolite, which clearly improved all bone histomorphometric parameters examined from ovariectomized animals, indicative for increased bone formation. Moreover, intervention with PMA-zeolite-clinoptilolite for one year proved safe in humans. Furthermore, patients treated with PMA-zeolite-clinoptilolite showed an increase in bone mineral density, an elevated level of markers indicative of bone formation, a significant reduction in pain, and significantly improved quality of life compared with patients in the control (placebo) group. These encouraging positive effects of PMA-zeolite-clinoptilolite on bone integrity and on osteoporosis warrant further evaluation of treatment with PMA-zeolite-clinoptilolite as a new alternative adjuvant therapy for osteoporosis.     

Screening for acetylcholinesterase inhibitory activity in cyanobacteria of the genus Nostoc

Fifty-four cyanobacterial strains of the genus Nostoc from different habitats were screened for acetylcholinesterase inhibitory activity. Water-methanolic extracts from freeze-dried biomasses were tested for inhibitory activity using Ellman's spectrophotometric method. Acetylcholinesterase inhibitory activity higher than 90% was found in the crude extracts of Nostoc sp. str. Lukesova 27/97 and Nostoc ellipsosporum Rabenh. str. Lukesova 51/91. Extracts from Nostoc ellipsosporum str. Lukesova 52/91 and Nostoc linckia f. muscorum (Ag.) Elenk. str. Gromov, 1988, CALU-980 inhibited AChE activity by 84.9% and 65.3% respectively. Moderate AChE inhibitory activity (29.1-37.5%) was found in extracts of Nostoc linckia Roth. str. Gromov, 1962/10, CALU-129, Nostoc muscorum Ag. str. Lukesova 127/97, Nostoc sp. str. Lhotsky, CALU-327 and Nostoc sp. str. Gromov, CALU-998. Extracts from another seven strains showed weak anti-AChE activities. The active component responsible for acetylcholinesterase inhibition was identified in a crude extract of Nostoc sp. str. Lukesova 27/97 using HPLC and found to occur in one single peak.     

Stage-specific distribution of oxidative radicals and antioxidant enzymes in the midgut of Leptinotarsa decemlineata

The titers of reactive oxygen species (ROS) represented by superoxide anion and general peroxides, and the activities of antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), are regulated in the midgut of the Colorado potato beetle (CPB) relative to the gut compartment, developmental stage, and food intake. ROS concentration is low in the potato leaves but it is very high in their digest in insect's anterior midgut. It is proposed that intensive ROS production in this gut region is linked to the processing of allelochemicals. SOD and CAT activities, low oxygen tension, and unidentified redox systems that maintain a slightly reducing milieu in the midgut lumen (pe+pH=6.95 declining to 5.36), obviously contribute to the decrease of ROS concentration along the gut length to a minimum in the wall of posterior midgut region. SOD and CAT activities are higher in the potato leaves than in the midgut tissues but the role of plant enzymes in ROS elimination within the gut lumen remains to be shown. A lower level of ROS and a higher antioxidant potential in the adult than in the larval midgut indicate stage specificity in the management of oxidative stress. The antioxidant defense is high in the diapausing adults that contain no detectable superoxide and about ten times less peroxides than the reproducing adults.     

Stimulatory effects of bioamines norepinephrine and dopamine on locomotion of Pyrrhocoris apterus (L.): is the adipokinetic hormone involved?

In the present paper we studied the effects of five biogenic amines - norepinephrine, dopamine, octopamine, serotonin and histamine - on the locomotory activity and mobilization of lipids in the adult females of the firebug, Pyrrhocoris apterus (L.). We tested the hypothesis (1) whether the stimulation of walking activity in the bugs injected with the bioamines is associated also with their hyperlipaemic effects, like in the case of adipokinetic hormones (AKHs), and (2) whether these effects are direct or mediated through a release of the AKHs into the hemolymph. The results demonstrated that all five tested biogenic amines mobilized the fat body lipids, but only norepinephrine and dopamine were capable to enhance the walking activity simultaneously with an elevation of the lipid level in the hemolymph. Those two amines had no effect on the level of AKHs in CNS, but modulated the AKHs level in hemolymph: norepinephrine increased it, while dopamine decreased it. The results indicate an apparent feedback between AKH characteristics and dopamine and norepinephrine actions occurring in this insect species. While the stimulatory effects of norepinephrine on lipid mobilization and walking activity could involve the release of bug's own AKHs, dopamine probably employs an independent stimulatory pathway.     

Effects of synthetic peptides on thylakoid phosphoprotein phosphatase reactions

A synthetic peptide analogue of the phosphorylation site of LHC II, when phosphorylated by thylakoid membranes, served as a substrate for the thylakoid phosphoprotein phosphatase. The phosphopeptide became dephosphorylated at a low rate, comparable to that of the 9 kDa phosphoprotein. Phospho-LHC II itself became dephosphorylated much more rapidly, at a rate unaffected by endogenous phosphorylation of the peptide. Endogenous phosphorylation of the peptide was also without effect on other thylakoid protein phosphorylation and dephosphorylation reactions. In contrast, dephosphorylation of many thylakoid phosphoproteins was inhibited by addition of a pure, chemically-synthesised phosphopeptide analogue of phospho-LHC II. This result suggests that at least one thylakoid phosphoprotein phosphatase exhibits a broad substrate specificity. The results indicate that any one of a number of amino acid sequences can give a phosphoprotein configuration that is recognised by a single phosphatase.     

Various chemical agents can induce premature chromosome condensation in Vicia faba

A number of chemical agents known to influence the key cell cycle regulatory factors were used to assess the requirements of hydroxyurea-treated root meristem cells of Vicia faba for premature condensation of chromosomes (PCC). These included caffeine and 2-aminopurine (inhibitors of ATM/ATR sensor kinases activated by DNA damage or stalled replication forks), inhibitors of protein kinases (staurosporine and wortmannin), inhibitors of protein phosphatases (sodium vanadate and calyculin A), and other compounds like 1,2-dioctyl-sn-glycerol, an activator of protein kinase C, 5-azacytidine, an inhibitor of DNA methyltransferase, dithiothreitol and N-etylmaleimide, capable to up- and down-regulate the activity of Cdc25 phosphatase. Cytological parameters used to evaluate quantitative aspects of PCC allowed us to discriminate various phenotypes of cells and, consistent with the extent of chromosomal fragmentation, to classify them as S- or G2-PCC. Two significant aspects relevant to the induction of premature mitosis in plants seem to emerge: one concerns the inverse relationship between the incidence of mitotic and PCC events, the other refers to the extent with which a variety of chemical agents may activate mechanisms that override the S-M replication checkpoint. 1,2-dioctyl-sn-glycerol, an activator of protein kinase C in animal cells proved extremely effective in stimulation of PCC, in spite of evident lack of molecular targets in plants.     

High-performance liquid chromatographic determination of ambroxol in human plasma

Ambroxol has been determined in biological fluids using a rapid and sensitive high-performance liquid chromatographic method. The samples prepared from plasma by liquid—liquid extraction were analysed on reversed-phase silica gel by competing-ion chromatography with ultraviolet detection. The method was applied to the determination of ambroxol levels in twelve healthy volunteers after oral administration of 90 mg of ambroxol in tablets of Mucosolvan and Ambrosan.     

Structures of ISC th4 transpososomes reveal the role of asymmetry in copy‐out/paste‐in DNA transposition

Copy‐out/paste‐in transposition is a major bacterial DNA mobility pathway. It contributes significantly to the emergence of antibiotic resistance, often by upregulating expression of downstream genes upon integration. Unlike other transposition pathways, it requires both asymmetric and symmetric strand transfer steps. Here, we report the first structural study of a copy‐out/paste‐in transposase and demonstrate its ability to catalyze all pathway steps in vitro. X‐ray structures of ISC th4 transposase, a member of the IS 256 family of insertion sequences, bound to DNA substrates corresponding to three sequential steps in the reaction reveal an unusual asymmetric dimeric transpososome. During transposition, an array of N‐terminal domains binds a single transposon end while the catalytic domain moves to accommodate the varying substrates. These conformational changes control the path of DNA flanking the transposon end and the generation of DNA‐binding sites. Our results explain the asymmetric outcome of the initial strand transfer and show how DNA binding is modulated by the asymmetric transposase to allow the capture of a second transposon end and to integrate a circular intermediate.