Glomerular ultrastructure of the trout,Salmo gairdneri: Effects of angiotensin II and adaptation to seawater

Summary The effect of angiotensin infusion on the glomerular ultrastructure of freshwater- and seawater-adapted rainbow trout, Salmo gairdneri, has been examined by scanning and transmission electron microscopy. Adaptation of trout to seawater resulted in epithelial podocyte flattening, primary process broadening and apparent loss of foot processes in almost all glomeruli, features which were uncommon in freshwater-adapted trout. Similar changes were induced by infusion of freshwater-adapted animals with angiotensin, suggesting that the renin-angiotensin system plays a role in the modification of glomerular epithelial ultrastructure. Adaptation of trout to seawater also reduced glomerular diameter, but infusion of freshwater-adapted animals with angiotensin did not mirror this effect. Infusion of angiotensin into seawater-adapted animals increased the overall thickness of glomerular basement membrane by increasing the lamina rara interna and lamina densa. This did not occur when freshwater-adapted fish were either infused with angiotensin or adapted to seawater. These findings suggest that other humoral systems are involved in the control of glomerular diameter and basement membrane thickness as part of an integrated response to increased environmental salinity.     

Isolation ofAcholeplasma oculi from human amniotic fluid in early pregnancy

Acholeplasmas have been isolated from a variety of animals, insects, and plants, but onlyAcholeplasma laidlawii has previously been found in humans. We have isolatedAcholeplasma oculi in pure culture from the amniotic fluid of a woman at 19 weeks of gestation. The organism was positively identified by growth inhibition, epi-immunofluorescence, and arbutin hydrolysis. Demonstration of organisms directly in amniotic fluid by DNA fluorochrome and immunofluorescence staining provided additional evidence that the isolate was genuine and not a medium contaminant. The remainder of the pregnancy was unremarkable, and a full-term male infant was delivered without complications. Even though there is some evidence possibly associatingA. oculi with various diseases in livestock, the prevalence and significance ofA. oculi in humans has not been determined.     

A Saccharomyces cerevisiae genomic plasmid bank based on a centromere-containing shuttle vector

A set of genomic plasmid banks was constructed using the centromere-containing yeast shuttle vector YCp50. The centromere-containing vector is useful for the isolation of genes that are toxic to yeast when present in high copy number. Fourteen independent banks were prepared each with an average representation of two to three times the yeast genome. Any individual plasmid from a given bank is guaranteed to be of independent origin from plasmids obtained from each of the other banks. The banks were constructed from three different size classes of DNA fragments that resulted from varying conditions of partial digestion with Sau3A. This avoided the bias caused by differential sensitivity of sites to cleavage with Sau3A. Insert DNA is sufficiently large that most genes will be present in the set of plasmid banks at a frequency of about 0.1%.     

Biochemical studies of soluble atrial natriuretic peptide (ANP) receptors from rat olfactory bulb and vascular smooth muscle cells

1. Aim. The biochemical characteristics of atrial natriuretic peptide receptors (ANP-R) derived from rat vascular smooth muscle (A-10 cell line) and central nervous system (CNS; olfactory bulb) tissue were compared. 2. Method and Results. ANP-Rs from each source were solubilized with 40 to 65% efficiency utilizing the nonionic detergent Lubrol-PX. Upon solubilization, the ANP-R from each source maintained the ability to bind 125I-ANP (99-126) with a high affinity; Scatchard analysis indicated that the VSMC ANP-R displayed a Kd for the radioligand of approximately 10 pM, whereas the olfactory receptor possessed a Kd of about 165 pM. The Bmax values for the soluble VSMC and olfactory ANP-Rs were 285 and 30 fmol/mg protein, respectively. Competition binding studies indicated that the VSMC ANP-R bound ANP(99-126), ANP(103-126), and ANP(103-123) with similar affinities, whereas the olfactory ANP-R was much more sensitive to changes in the COOH-terminal structure of the competing peptide. The soluble ANP-Rs from VSMC and olfactory were chromatographically indistinguishable on phenyl-, DEAE-, and wheat germ agglutinin-agarose columns. However, the ANP-Rs could be distinguished using GTP-agarose; the olfactory ANP-R was capable of binding to the resin, whereas the VSMC ANP-R was not. 3. Conclusions. Coupled with other studies, these data suggest that the A10 VSMC ANP-R observed in this study may not be coupled to guanylate cyclase and may represent a receptor serving a clearance function, whereas a significant proportion of the olfactory CNS ANP-R appears to be associated with GTP-binding proteins, likely particulate guanylate cyclase, and probably represents a coupled form of the receptor.     

Techniques in molecular biology

Book Review

Techniques in molecular biologyJ.M. Walker and W. Gaastra (Eds.), vol. 2. London: Croom Helm, 1987. iv + 332 pages. £14.95. ISBN 0-7099-3673-7     

Chemical evolution and the origin of life

This bibliography is the seventeenth annual supplement to the comprehensive bibliography on the same subject which was published in Space Life Sci. 2(1970), 225-295; 3(1972), 293-304; 4(1973), 309-329 and in Origins of Life 5(1974), 505-527; 6(1975), 285-300; 7(1976), 75-85; 8(1977), 59-66; 9(1978), 67-74; 10(1980), 69-87; 10(1980), 379-404; 11(1981), 273-288; 12(1982), 93-118; 13(1983), 61-80; 15(1984), 55-69; 17(1987), 171-184; 17(1987), 185-206.     

Temporal changes in the egglaying behaviour of the Hessian fly

Responses of mated female Hessian flies were investigated by analysing the behaviour of individual flies in wheat and oats. The behavioural repertoire of such females included: flying, alighting on leaves, arching of the body so that the tip of the abdomen touched the leaf surface, antennation, movements of the tip of the abdomen across the leaf at right angles to leaf veins, sitting with the ovipositor straight but still extended, and sitting with the ovipositor telescoped into the body. After alighting, females on wheat showed a higher frequency of transitions from arching to antennation and a lower frequency of transitions from arching to abdomen straight than females on oats. During the first 5 min of observations, individuals released into arenas with wheat arched and antennated 2–3 times more frequently than females released into oats. Time allotted to behaviours also differed; during the first 5 min, females in wheat spent 50 percent more time arching, whereas females in oats spent 50 percent more time sitting. Females in wheat also stayed longer and laid 4 times more eggs than females in oats. Temporal changes in egglaying were monitored by quantifying hourly rates of egglaying in no-choice assays for several hours following mating at 9:00 am. During the first and second hours post-mating, egglaying occurred infrequently. However, during the third hour post-mating (11:00 am to noon) females on wheat laid 5 times more eggs than females on oats. Rates of egglaying decreased on wheat but increased on oats during the fourth hour, and then during the fifth hour, decreased on both wheat and oats. Changes in egglaying responses were also evident when behaviours of individual females were measured 1–3 h vs. 3–7 h post-mating. Females deprived of host plants and released into wheat or oats later in the day showed higher frequencies of arching and antennation and laid more eggs before leaving the arena.

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Résumé Les réactions de femelles préalablement accouplées de Mouches de Hesse ont été examinées en analysant le comportement de femelles isolées sur blé et sur avoine. Le répertoire comportemental de ces femelles comprenait: le vol, l'atterrissage sur feuille, la flexion du corps de sorte que l'extrémité de l'abdomen touchât la surface de la feuille, l'antennation, les mouvements de l'extrémité de l'abdomen sur la feuille à angle droit des nervures, le repos avec la tarière droite et encore dévaginée, le repos avec la tarière télescopée à l'intérieur du corps. Sur blé plus que sur avoine, les femelles après atterrissage ont présenté une fréquence plus élevée de passage de la flexion à l'antennation que de la flexion à l'abdomen droit. Durant les 5 premières minutes d'observation, les individus libérés dans des enceintes avec blé fléchirent et antennèrent 2 à 3 fois plus que ceux libérés sur avoine. Les durées des différentes séquences différaient aussi: sur blé, pendant les 5 premières minutes, les femelles passèrent plus de 50% du temps à fléchir, tandis que sur avoine elles passèrent plus de 50% du temps en repos. Les femelles restèrent aussi plus longtemps sur les feuilles de blé et y pondirent 4 fois plus d'oeufs que sur avoine.Les femelles de M. destructor ont montré une plasticité du seuil d'acceptation. Pendant les premières heures de ponte, elles ont été très sélectives et refusèrent, ou ne pondirent que quelques oeufs sur avoine, mais acceptèrent volontiers le blé. La discrimination s'est poursuivie tant que les femelles ont eu accès au blé en même temps qu'à l'avoine. Cependant, quand les femelles ont été privées de blé pendant plusieures heures, l'acceptation de l'avoine a augmenté. Cet accroissement de l'acceptation a eu lieu à peu près au moment où les femelles sur blé pondaient leurs derniers oeufs.

     

Purification and characterization of a methionine-specific aminopeptidase from Salmonella typhimurium

An aminopeptidase specific for methionine (peptidase M) has been purified from wild-type and mutant Salmonella typhimurium strains. Recombinant peptidase M was also purified from Escherichia coli. These preparations were characterized with respect to their physicochemical properties using analytical ultracentrifugation, SDS/PAGE, isoelectric focusing, titration curve analysis, amino acid analysis, N-and C-terminal sequencing and various spectroscopic methods. Peptidase M activity is stimulated by Co2+, in agreement with previous studies using crude extracts of Salmonella. The purified preparations did not contain significant amounts of any metal. Enzymically important metal is loosely associated and lost during enzyme purification. Peptidase M was shown to contain seven free sulphydryl residues none of which are involved in either intra-or inter-molecular disulphide bonds. Most appear solvent-accessible as evidenced by their reactivity under native conditions. Limited modification of the sulphydryl residues with either iodoacetamide or 5,5'-dithiobis(2-nitrobenzoic acid) led to inactivation. Several cysteines were shown to be labelled to various degrees by peptide mapping of inactivated S-[14C]carboxymethylated protein. Whether cysteine modification affects enzymic activity directly (blocking an active site) or indirectly (by causing conformational change) remains to be established.     

Eimeria vermiformis and E. mitis: inhibition of development in vivo by cyclosporin A

Treatment of the host (Mus musculus, Gallus domesticus) with cyclosporin A during infection with Eimeria vermiformis or E. mitis resulted in a reduction in the numbers of oocysts passed in the feces and/or a delay in patency. The general immunosuppressive effects of the treatment were confirmed in chickens by monitoring their antibody responses to human erythrocytes and lymphoproliferative responses to phytohemagglutinin. Nevertheless, mice and chickens treated with cyclosporin A during a primary infection with E. vermiformis or E. mitis, respectively, were immune to subsequent challenge with these organisms. Thus, cyclosporin A did not interfere with priming. The antiparasite effect of the drug did not allow an evaluation of its effect on established immunity to the coccidia when it was administered at the time of challenge. In an exceptional treated chicken, however, delayed patency of the challenge infection was followed by the production of a number of oocysts similar to that found in unprimed animals. This suggests that the mechanisms of immunity to challenge may be susceptible to disruption by cyclosporin A.     

Adenosine, a cytoprotective autocoid: effects of adenosine on neutrophil plasma membrane viscosity and chemoattractant receptor display

At inflammatory sites neutrophils are stimulated to produce a variety of toxic agents, yet rarely harm the endothelium across which they migrate. We have recently found that endothelium releases adenosine which, acting via receptors on the surface of human neutrophils, inhibits generation of toxic metabolites by stimulated neutrophils but, paradoxically, promotes chemotaxis. Agents which diminish plasma membrane viscosity affect neutrophil function similarly, possibly by modulating chemoattractant receptor number or affinity. We therefore determined whether adenosine receptor agonists modulate neutrophil function by decreasing membrane viscosity and/or chaning the affinity of chemoattractant (N-fMet-Leu-Phe, FMLP) receptors. Surprisingly, 5′-(N-ethylcar☐amido)adenosine (NECA, 10 μM), the most potent agonist at neutrophil adenosine receptors, increased plasma membrane viscosity, as measured by fluorescence anisotropy of the plasma membrane specific probe 1-(4-trimethylaminophenyl)-6-diphenyl-1,3,5-hexatriene (TMA-DPH), in unstimulated neutrophils from a mean microviscosity of 1.67 ± 0.02 (S.E.) to 1.80 ± 0.02 (p < 0.001) while inosine (10 μM), a poor adenosine receptor agonist, had no effect (1.73 ± 0.04, p =n.s. vs. control, p < 0.01 vs. NECA). Adenosine receptor agonists increased plasma membrane viscosity in neutrophils with the same order of potency previously seen for inhibition of superoxide anion generation and enhancement of chemotaxis (NECA > adenosine = N⁶-phenylisopropyladenosine). The adenosine receptor antagonist 8-(p-sulfophenyl)theophylline reversed the effect of NECA on plasma membrane viscosity. Unlike other agents which modulate plasma membrane viscosity, NECA (10 μM) did not significantly change the number or affinity of [³H]FMLP binding sites on neutrophils. In contrast to the hypothesis of Yuli et al. these results indicate that occupancy of adenosine receptors on neutrophils increases plasma membrane viscosity without affecting chemoattractant receptor display.