Indomethacin antagonizes the ethanol-induced suppression of breathing activity but not the suppression of brain activity in the near-term fetal sheep

The effect of indomethacin on the ethanol-induced suppression of fetal breathing movements, low-voltage electrocortical (ECoG) activity, and electro-ocular (EOG) activity was studied in the near-term fetal sheep. Ten conscious instrumented pregnant ewes (between 129 and 131 days of gestation; term, 147 days) received 1-h maternal intravenous infusion of 1 g ethanol/kg total body weight and simultaneous fetal treatment with either indomethacin (2 mg/kg fetal body weight/h) (n = 5) or an equivalent volume of phosphate buffer (n = 5) intravenously for 9 h. Fetal ECoG activity, EOG activity, and fetal breathing movements were monitored continuously over the experimental periods. In animals treated with ethanol and buffer (n = 5), fetal breathing movements were suppressed for 8 h and low-voltage ECoG and EOG activity was suppressed for 2 h below preinfusion levels. In animals treated with ethanol and indomethacin (n = 5), fetal breathing movements were elevated for 13 h but low-voltage ECoG and EOG activity remained suppressed for 3 h below preinfusion levels. The data suggests that indomethacin can antagonize the ethanol-induced suppression of fetal breathing movements, but does not alter the ethanol-induced suppression of ECoG or EOG activity.     

Light Affects Flagellar Agglutinability in Chlamydomonas eugametos by Modification of the Agglutinin Molecules

The effect of light on the sexual competence of a light-sensitive mating type minus strain (mt⁻) of Chlamydomonas eugametos obtained by crossing a light-sensitive mating type plus strain (mt⁺) with a light-insensitive mt⁻ strain is described. As previously demonstrated for the mt⁺ parent, this study of one of the mt⁻ offspring shows that (a) a light-sensitive mechanism affects flagellar agglutinability in a rapid process that does not require protein synthesis; (b) only the activity of the flagellar agglutinins (glycoproteins responsible for agglutination) is susceptible to light while agglutinins on the cell body surface are not affected by light. We further demonstrate that (a) membrane vesicles naturally released from nonagglutinable dark gametes remain inactive. Extracts of these vesicles also remain inactive even though they contain agglutinin-like components; (b) inactive mt⁻ agglutinin is present in extracts of flagella from nonagglutinable dark gametes by comparison of its chromatographic, electrophoretic, and immunogenic properties with those of active agglutinin. When purified of all other flagellar proteins, it remains inactive; (c) a monoclonal antibody directed against the sexual agglutination site of the mt⁻ agglutintin discriminates between active and inactive agglutinins when present in a native state on the flagellar surface, but is unable to discriminate between them when they are denatured in sodium dodecyl sulfate-electrophoresis gels and blotted onto nitrocellulose. Taken collectively these observations suggest that light activation involves the chemical modification of the agglutinins in situ on the flagellar surface.     

Endogenous oscillator controls surface density of mating receptors in Chlamydomonas eugametos

Summary We describe a circadian rhythm in the surface density of receptors that play a dominant role in the mating process of the unicellular green alga Chlamydomonas eugametos.These receptors — called agglutinins — are large glycoproteins extrinsically bound to the membrane of gamete flagella. We found circadian fluctuations in their density. Since inhibition of protein synthesis affected the agglutinin density without a lag period at any time,we conclude that the density was dependent on de novo synthesis and that the fluctuations in density are caused by circadian oscillations in the rate of agglutinin synthesis. This phenomenon evidently underlies the pronounced endogenous rhythm in mating competence that we described previously (Demets et al. 1987). Finally, we speculate on the nature of the time keeping mechanism that is generating these rhythmic events.     

Differential effects of eicosapentaenoic acid and oleic acid on lipid synthesis and secretion by HepG2 cells

The effects of eicosapentaenoic acid and oleic acid on lipid synthesis and secretion by HepG2 cells were examined to identify fatty acid specific changes in lipid metabolism that might indicate a basis for the hypolipidemic effect attributed to eicosapentaenoic acid and related n-3 fatty acids. Cellular glycerolipid synthesis, as determined by [3H]glycerol incorporation, increased in a concentration-dependent manner in cells incubated 4 h with either eicosapentaenoic acid or oleic acid at concentrations between 10 and 300 microM. [3H]Glycerol-labeled triglyceride was the principal lipid formed and increased approximately fourfold with the addition of 300 microM oleic acid or eicosapentaenoic acid. Both fatty acids also produced a 20-40% increase in the total cellular triglyceride mass. Although both fatty acids increased triglyceride synthesis to similar extents, eicosapentaenoic acid-treated cells secreted 40% less [3H]glycerol-labeled triglyceride than cells fed oleic acid. Cellular synthesis of [3H]glycerol-labeled phosphatidylethanolamine and phosphatidylcholine was also reduced by 40% and 30%, respectively, in cells given eicosapentaenoic acid versus cells given oleic acid. Similar results were obtained in determinations of radiolabeled oleic acid and eicosapentaenoic acid incorporation. At a fatty acid concentration of 300 microM, incorporation of radiolabeled eicosapentaenoic acid into cellular triglycerides was greater than the incorporation obtained with radiolabeled oleic acid, while the reverse relationship was observed for the formation of phosphatidylcholine from the same fatty acids. Eicosapentaenoic acid is as potent as oleic acid in inducing triglyceride synthesis but eicosapentaenoic acid is a poorer substrate than oleic acid for phospholipid synthesis. The intracellular rise in de novo-synthesized triglyceride in eicosapentaenoic acid-treated cells without corresponding increases in triglyceride secretion suggests that eicosapentaenoic acid is less effective than oleic acid in promoting the transfer of de novo-synthesized triglyceride to nascent very low density lipoproteins.     

A new variant of Autographa californica nuclear polyhedrosis virus

A variant of the baculovirus, Autographa californica nuclear polyhedrosis virus, has been isolated in Idaho during an epizootic disease in a field population of A. californica. Genotypic characterization indicates that the virus is distinct from variants previously characterized. Analysis of five clones, derived by plaque purification in cell culture, indicates relative homogeneity of the original virus isolate. Further exploration of the factors involved in natural genetic variability of baculoviruses is appropriate.     

Constitutively active rhodopsin mutants causing night blindness are effectively phosphorylated by GRKs but differ in arrestin-1 binding

The effects of activating mutations associated with night blindness on the stoichiometry of rhodopsin interactions with G protein-coupled receptor kinase 1 (GRK1) and arrestin-1 have not been reported. Here we show that the monomeric form of WT rhodopsin and its constitutively active mutants M257Y, G90D, and T94I, reconstituted into HDL particles are effectively phosphorylated by GRK1, as well as two more ubiquitously expressed subtypes, GRK2 and GRK5. All versions of arrestin-1 tested (WT, pre-activated, and constitutively monomeric mutants) bind to monomeric rhodopsin and show the same selectivity for different functional forms of rhodopsin as in native disc membranes. Rhodopsin phosphorylation by GRK1 and GRK2 promotes arrestin-1 binding to a comparable extent, whereas similar phosphorylation by GRK5 is less effective, suggesting that not all phosphorylation sites on rhodopsin are equivalent in promoting arrestin-1 binding. The binding of WT arrestin-1 to phospho-opsin is comparable to the binding to its preferred target, P-Rh*, suggesting that in photoreceptors arrestin-1 only dissociates after opsin regeneration with 11-cis-retinal, which converts phospho-opsin into inactive phospho-rhodopsin that has lower affinity for arrestin-1. Reduced binding of arrestin-1 to the phospho-opsin form of G90D mutant likely contributes to night blindness caused by this mutation in humans.     

Cerebral leucine uptake and protein synthesis in the near-term ovine fetus: relation to fetal behavioral state

To investigate quantitatively how sweating and cutaneous blood flow responses at the onset of dynamic exercise are affected by increasing exercise intensity in mildly heated humans, 18 healthy male subjects performed cycle exercise at 30, 50, and 70% of maximal O2 uptake (VO2 max) for 60 s in a warm environment. The study was conducted in a climatic chamber with a regulated ambient temperature of 35 degrees C and relative humidity of 50%. The subjects rested in the semisupine position in the chamber for 60 min, and then sweating rate (SR) and skin blood flow were measured during cycle exercise at three different intensities. Changes in the heart rate, rating of perceived exertion, and mean arterial blood pressure were proportional to increasing exercise intensity, whereas esophageal and mean skin temperatures were essentially constant throughout the experiment. The SR on the chest, forearm, and thigh, but not on the palm, increased significantly with increasing exercise intensity (P < 0.05). The mean SR of the chest, forearm, and thigh increased 0.05 mg.cm-2.min-1 with an increase in exercise intensity equivalent to 10% VO2 max. On the other hand, the cutaneous vascular conductance (CVC) on the chest, forearm, and palm decreased significantly with increasing exercise intensity (P < 0.05). The mean CVC of the chest and forearm decreased 5.5% and the CVC on the palm decreased 8.0% with an increase in exercise intensity equivalent to 10% VO2 max. In addition, the reduction in CVC was greater on the palm than on the chest and forearm at all exercise intensities (P < 0.01). We conclude that nonthermal sweating and cutaneous blood flow responses are exercise intensity dependent but directionally opposite at the onset of dynamic exercise in mildly heated humans. Furthermore, cutaneous blood flow responses to increased exercise intensity are greater in glabrous (palm) than in nonglabrous (chest and forearm) skin.