Effect of some vertebrate and invertebrate hormones on the population growth, mictic female production, and body size of the marine rotifer Brachionus plicatilis Müller

Eight vertebrate and invertebrate hormones were screened for theireffect on population growth, mictic female production, and body size of themarine rotifer Brachionus plicatilis. Growth hormone (GH) or human chorionicgonadotropin (HCG) at 0.0025–25 I.U. ml^–1 andestradiol-17 (E₂), triiodothyronine (T_{_3}),20-hydroxyecdysone (20-HE), 5-hydroxytryptamine (5-HT), gamma-aminobutyricacid (GABA) or juvenile hormone (JH) at 0.05–50 mg l^–1were added to 5-ml of Nannochloropsis oculata suspension (7×10⁶ cells ml^–1). From an initial densityof 1 individual ml^–1, rotifers were cultured with hormones for48 hours in 22 ppt seawater at 25 °C, in darkness.Rotifers were counted and classified into female types and transferred to anew algal food suspension without hormone every other day until day 8 whenbody size was measured. Population growth was significantly higher intreatments exposed to GABA (50 mg l^–1), GH (0.0025 and 0.025I.U. ml^–1), HCG (0.25 and 2.5 I.U. ml^–1), and5-HT (5 mg l^–1). E₂ caused a decrease inpopulation growth, whereas JH, 20HE, and T₃ had no effect.Mictic female production was significantly higher at 0.05 and 0.5 mgl^–1 JH and 0.05 and 5 mg l^–1 5HT. GH (0.0025 and0.025 I.U. ml^–1), E₂ (50 mg l^–1),GABA (0.5, 5 and 50 mg l^–1), and 20-HE (0.05 mgl^–1) treatments had significantly higher mictic femaleproduction only on day 8, 6, 4, and 6, respectively. T₃ andhCG had no effect on mictic female production. Lorica length increased by9.6% and 4.4% in rotifers treated with JH (0.05 mgl^–1) and GABA (5 mg l^–1), respectively. Correspondingly, lorica width increased by 8.9% and 2.6% inthese treatments. In comparison, 20-HE-, T₃-, and HCG-treatedrotifers were smaller (3.9–8.2%) and GH, 5-HT andE₂ had no effect on rotifer body size.     

Pharmacokinetic Analysis of Free Radicals by in vivo BCM (Blood Circulation Monitoring)-ESR Method

In pharmacokinetic studies, a variety of analytical method including radioisotopic detection and HPLC (high performance liquid chromatography) has been used. In the present investigation, we developed in vivo BCM (Blood Circulation Monitoring)-ESR method, which is a new technique with a conventional X-band ESR spectrometer for observing stable free radicals in the circulating blood of living rats under anaesthesia. Both 5-(PROXYL derivatives) and 6-(TEMPO derivatives) membered nitroxide spin probes with various types of substituent functional group were used. After physicochemical properties of the spin probes such as hyperfine coupling constant (A-value), g-value and partition coefficient as well as chemical stability of the compounds in the fresh blood were obtained, the in vivo BCM-ESR method was performed in normal rats. Several pharmacokinetic parameters such as half-life of the probes, distribution volume, total body clearance and mean residence time were obtained and discussed in terms of their chemical structures. In addition, clearance of a spin probe was related to the urine concentration. The BCM-ESR method was found to be very useful to observe free radicals at the real time. By time-dependent ESR signal decay of spin probes, pharmacokinetic parameters were obtained.     

Lung tissue behavior in the mouse during constriction induced by methacholine and endothelin-1

Nagase, Takahide, Hirotoshi Matsui, Tomoko Aoki, YasuyoshiOuchi, and Yoshinosuke Fukuchi. Lung tissue behavior in the mouseduring constriction induced by methacholine and endothelin-1. J. Appl. Physiol. 81(6):2373-2378, 1996.Recently, mice have been extensively used toinvestigate the pathogenesis of pulmonary disease because appropriatemurine models, including transgenic mice, are being increasinglydeveloped. However, little information about the lung mechanics of miceis currently available. We questioned whether lung tissue behavior andthe coupling between dissipative and elastic processes, hysteresivity(), in mice would be different from those in the other species. Toaddress this question, we investigated whether tissue resistance (Rti)and  in mice would be affected by varying lung volume, constrictioninduced by methacholine (MCh) and endothelin-1 (ET-1), andhigh-lung-volume challenge during induced constriction. From measuredtracheal flow and tracheal and alveolar pressures in open-chest ICRmice during mechanical ventilation [tidal volume = 8 ml/kg,frequency (f) = 2.5 Hz], we calculated lung resistance(RL), Rti, airway resistance(Raw), lung elastance (EL),and  (=2fRti/EL). Underbaseline conditions, increasing levels of end-expiratory transpulmonarypressure decreased Raw and increased Rti. The administration ofaerosolized MCh and intravenous ET-1 increasedRL, Rti, Raw, andEL in a dose-dependent manner.Rti increased from 0.207 ± 0.010 to 0.570 ± 0.058 cmH₂O ^· ml^{1 ·} safter 10⁷ mol/kg ET-1(P < 0.01). After inducedconstriction, increasing end-expiratory transpulmonary pressuredecreased Raw. However,  was not affected by changing lung volume,constriction induced by MCh and ET-1, or high-lung-volume challengeduring induced constriction. These observations suggest that1)  is stable in mice regardlessof various conditions, 2) Rti is animportant fraction of RL andincreases after induced constriction, and3) mechanical interdependence mayaffect airway smooth muscle shortening in this species. In mammalianspecies, including mice, analysis of  may indicate that both Rti andEL essentially respond to asimilar degree.

     

A 570-kb DNA Sequence of the Escherichia coli K-12 Genome Corresponding to the 28.0-40.1 min Region on the Linkage Map

The 569,750 base pair sequence corresponding to the 28.0–40.1min region on the genetic map of Escherichia coli K-12 (W3110)was determined. This region includes the replication terminusregion and contained at least 549 potential open reading frames.Among them, 160 (29%) were previously reported, 174 (32%) werehomologous to other known genes, 102 (18%) were identical orsimilar to hypothetical genes registered in databases, and theremaining 113 (21%) did not show a significant similarity toany other gene. Of interest was the finding of a large numberof genes and gene clusters in andnear the replication terminationregion which had been thought to be genetically silent. Thoseincludeda cluster of genes for fatty acid ß-oxidation,the third copy of the pot (spermidine/putrescine transport system)gene cluster, the second dpp (dipeptide transport system) operon,the second dsm (anaerobic dimethyl sulfoxide reductase) operon,a cluster of fim (fimbrial) genes anda DNA helicase-like genewith a high molecular weight. In addition, we found the dnaC-and dnaT-like genes in the cryptic prophage, Rac, anda numberof genes originated probably from plasmids.     

Nucleotide Sequence of a 28-kb Mouse Genomic Region Comprising the Imprinted Igf2 Gene

The mouse insulin-like growth factor II gene (Igf2) is physicallylinked to the insulin II gene (Ins2) and both are subject totissue-specific genomic imprinting. The paternal-specific expressionof Igf2 has been associated with hypermethylation of some CpGsites in the 5' flanking region and in the body of the gene.As a first step in analyzing the structural features of thisimprinted locus, we here report the complete nucleotide sequenceof Igf2, including all introns and the intergenic region adjacentto Ins2. This 28-kb segment of mouse chromosome 7 exhibits 80%overall identity with the corresponding rat sequence and hasa high GC content of 52%. In addition to the known CpG islandwithin the second Igf2 promoter, another island was identifiedapproximately 2 kb 5' to the first exon. Other features of thislocus include a 35-fold tandem repeat of an 11-bp sequence thatoverlaps Igf2 pseudo-exon 2, and a B2 repeat element in theintergenic region between Ins2 and Igf2. The GC-richness andthe presence of CpG islands associated with tandem repeats arecommon features of imprinted genes and thus may play a rolein the imprinting mechanism.     

Synthesis, solution structure, binding activity, and cGMP activation of human guanylin and its disulfide isomer

Guanylin is a recently isolated peptide consisting of 15 amino acid residues with four cysteines, which may form two intramolecular disulfide bridges, and stimulates intestinal membrane guanylate cyclase. The position of the disulfide linkages of guanylin was predicted from its structural similarity to a heat stable enterotoxin which is thought to be responsible for secretory diarrhoea. Both guanylin, with disulfide positions 4–12 and 7–15, and its disulfide isomer, with disulfides positions 4–15 and 7–12, were chemically synthesized by the solid-phase method and purified. Two specific disulfides were selectively formed and confirmed by sequencing, mass spectrometry and high-performance liquid chromatography in combination with enzymatic cleavage. The structure of both isomers has been investigated in solution by ¹H nuclear magnetic resonance spectroscopy. Guanylin exists as a mixture of two stable conformations which have compact spiral structures, from comparison with literature data. In contrast, the disulfide isomer of guanylin shows only a single conformation with an elongated curved plate-like structure. Binding assays were performed using labelled guanylin with membranes obtained from rat jejunum. Both disulfide isomers were investigated by the cGMP assay. Both binding and cGMP assays indicated that the relevant form of disulfide bridges in the intact guanylin was as predicted.     

A vitamin-free minimal synthetic medium forCryptococcus neoformans

The use of a simple synthetic medium is essential for study on the growth and physiology ofCryptococcus neoformans. In the present study, a minimal synthetic liquid medium (MSM) was tested for the growth of 23C. neoformans strains. This medium contained a low concentration of glucose, ammonium sulphate and inorganic salts with a pH value of 4.5, but no amino acids or vitamins. The strains were starved for 4 days to eliminate nutrients which might have been carried over from their pre-culture medium. Then, they were inoculated in the MSM at an initial OD of 0.020 at 550 nm and incubated at 37 °C for 20 days. Cell growth was generally monitored daily by measuring the absorbance at 550 nm. The medium supported the growth of the strains tested and gave an average final OD of 0.500. The results obtained indicate thatC. neoformans may be autotrophic with respect to vitamins and in particular to thiamine. The MSM medium is easy to prepare and store. It is highly reproducible and useful for studies on the growth and physiology ofC. neoformans.     

Effect of Macrolide Antibiotics on Macrophage Functions

Macrolide antibiotics have a variety of actions other than antimicrobial activities. Recently, it has been suggested that macrolide antibiotics act as immunomodulators. In this study, we evaluated the effects of macrolide antibiotics on macrophage functions. For the macrophage, we used the mouse macrophage cell line J774.1. The following effects of macrolide antibiotics on macrophage functions were evaluated: the effect of macrolide antibiotics on macrophage growth; the phagocytosis of beads; cytocidal activity against Candida albicans; and chemotaxis to lipopolysaccharide (LPS). Macrolide antibiotics except for azithromycin significantly stimulated the growth of the macrophage. In addition, pretreatment with macrolide antibiotics except for roxithromycin significantly stimulated the macrophage phagocytosis of beads, macrophage chemotaxis to LPS, and macrophage cytocidal activity against Candida albicans. These results suggest that macrolide antibiotics stimulate macrophage functions.     

Use of a surface plasmon resonance biosensor for the determination of binding constants of transiently expressed recombinant antibody to its antigen

Summary By using a commercially available surface plasmon resonance (SPR) biosensor, the values of the association rate constant (kass), dissociation rate constant (kdiss), and association constant (KA = kass / kdiss) for binding to the antigens were determined. They were almost the same for the recombinant antibody expressed in COS cells, CHO cells, and mouse hybridoma cells. The system of transient expression of the recombinant antibody (Ab) in COS cells and SPR analysis of the supernatant should be useful for rapid expression and evaluation of the binding ability of large numbers of engineered Abs.     

σ1 Receptor Subtype Is Involved in the Facilitation of Cortical Dopaminergic Transmission in the Rat Brain

Our previous studies have shown that three sigma () receptor ligands, (+)-N-allylnormetazocine ((+)-SKF-10,047), (±)-pentazocine and 1,3-di(2-tolyl)guanidine (DTG) differently regulated the dopamine (DA) transmission in the rat brain. In the present study, we attempted to clarify the role of ₁ receptor subtype in the regulation of DA transmission using a novel and selective ₁ receptor agonist, 1-(3,4-dimethoxyphenethyl)-4-(3-phenylpropyl)piperazine dihydrochloride (SA4503) in the rat brain. Acute administration of SA4503 (1.0 mg/kg, p.o.) significantly increased DA and 3,4-dihydroxyphenylacetic acid (DOPAC) levels in the rat frontal cortex, but not in the other six regions, hippocampus, striatum, midbrain, cerebellum, medulla/pons and hypothalamus. The increase of cortical DA level elicited by SA4503 was fully reversed by N,N-dipropyl-2-(4-methoxy-3-(2-phenylethoxy)phenyl)ethylamine (NE-100) (0.25 mg/kg, p.o.), a putative ₁ receptor antagonist. In addition, SA4503 (1.0 mg/kg, p.o.) showed an increase of cortical L-3,4-dihydroxyphenylalanine (L-DOPA) accumulation under the inhibition of dopa decarboxylase activity with m-hydrobenzylhydrazine (NSD-1015), suggesting that SA4503 has activated the cortical DA synthesis rate. These results suggest that the ₁ receptor subtype plays an important role in the facilitation of cortical DA transmission. In addition, this phenomenon is partially involved in the augmentation of DA synthesis rate.