Absorption and blood clearance of labelled carotenoids ([14C]astaxanthin, [3H]Canthaxanthin and [3H]zeaxanthin) in mature female rainbow trout (Oncorhynchus mykiss)

• 1.1. Using one force-fed meal, eight mature female rainbow trout received [¹⁴C]astaxanthin ([¹⁴C]Ax) with [³H]canthaxanthin ([³H]Cx; N = 3) or with [³H]zeaxanthin ([³H]Zx; N = 5).
• 2.2. Approximately 200 μl of blood were collected via caudal puncture every 24 hr for 4 days. After 96 hr, the fish were killed and pyloric caeca (P.C.) from the duodenal intestine (D.I.) section, ileal intestine (I.I.), and posterior intestine (P.I.) were dissected out.
• 3.3. In the blood, Ax levels were higher than Cx followed by Zx levels.
• 4.4. This corresponds to their respective absorption by the trout as was confirmed by their relative concentrations in P.C., I.I. and P.I.
• 5.5. However, blood clearance was similar for all three compounds. [¹⁴C]Phoenicoxanthin ([¹⁴C]Px) was detected as a reduced metabolite of [¹⁴C]Ax in all gut sections.

     

Effect of inorganic phosphate on synthesis of 5-phosphoribosyl-1-pyrophosphate in cultured plant cells

• 1.1. Inorganic phosphate (Pi) was absorbed rapidly by suspension-cultured cells of Catharanthus roseus which had previously been cultured in Pi-free Murashige Skoog medium.
• 2.2. The intracellular levels of ATP, ADP and 5-phosphoribosyl-l-pyrophosphate (PRPP) increased markedly during the 24 hr which followed the addition of Pi (1.25mM).
• 3.3. Availability of PRPP in vivo, estimated by the measurement of nucleotide synthesis from [8-¹⁴C]adenine, was also increased by addition of Pi.
• 4.4. Only a 20% increase in the maximum catalytic activity of PRPP synthetase was observed in extracts of cells, prepared 24 hr after addition of Pi.
• 5.5. In contrast to results for mammalian PRPP synthetase, the activity of PRPP synthetase, partially purified from Catharanthus roseus, was inhibited by concentration of Pi greater than 5mM.
• 6.6. The mechanisms involved in the increased availability of PRPP and the synthesis of adenine nucleotides in the plant cells cultured in Pi-containing medium are discussed.

     

Time-dependent cadmium-neurotoxicity in rat brain synaptosomal plasma membranes

• 1.1. The modulation of lipid dynamics and lipid protein interactions were studied in rat brain synaptosomal plasma membranes (SPM) up to 24 hr after exposure to cadmium (Cd).
• 2.2. The activity of acetylcholinesterase and adenylate cyclase showed a considerable decrease after 6 hr of Cd exposure, followed by a progressive increase up to 24 hr.
• 3.3. SPM chemiluminescence showed a maximum decrease at 12 hr, demonstrating a considerable increase in lipid peroxidation.
• 4.4. SPM of Cd-exposed animals showed a statistical significant increase in fluorescence anisotropy parameter [(r₀/r) — 1]⁻¹ at 18 and 24 hr compared to SPM of the control, indicating a decrease of membrane fluidity.

     

Studies on compartmentation of uridine 5'-triphosphate during synthesis of cytoplasmic and plastid ribosomal RNA's in Euglena gracilis

• 1.1. Compartmentation of uridine 5'-triphosphate (UTP) was studied during synthesis of cytoplasmic ribosomal RNA (cyt-rRNA) and plastid ribosomal RNA (pl-rRNA) in photoorganotrophically grown cells of Euglena gracilis Z.
• 2.2. Using the approach of Wiegers et al. (1976) the steady state specific radioactivity of UTP was compared with that ofcyt-20S rRNA, cyt-25S rRNA, pl-16S rRNA and pl-23S rRNA under low and at 100-fold higher specific radioactivity of exogenously fed pHl-uracil.
• 3.3. The equal steady state specific radioactivities of all rRNAs at both feeding conditions argue against compartmentation of UTP during their synthesis.
• 4.4. At high specific radioactivity of exogenous [³H]-uracil the salvage-derived labelled UMP was shown to be diluted 15,000-fold by unlabelled UMP formed de novo, whereas this dilution factor was 100-fold lower at low specific radioactivity of [³H]-uracil indicating inhibition of the de novo synthesis of UMP.
• 5.5. Transport is suggested of uridine nucleotides into chloroplasts by the 15-fold higher specific radioactivity of intracellular [³H]-uracil than that of UTP as well as UMP residues in pl-rRNA.

     

Phosphoinositides and inositol phosphates in Discopyge tschudii electrocyte membranes

• 1.1. The metabolism of inositol (Ins)-containing phospholipids and inositol phosphates has been studied by following the incorporation and distribution of myo-[³ H]Ins in metabolically active electrocytes from the electric ray Discopyge tschudii.
• 2.2. The apparent initial rate of myo-[³H]Ins incorporation into total phosphoinositides was ca 8.2 fmol/mg protein/hr. Phosphatidylinositol (Ptdlns) displayed the highest levels of labelling. Lithium inhibited this incorporation probably by limiting the recycling of myo-[³H]Ins from [³H]Ins-monophosphate.
• 3.3. The formation of water-soluble products of phosphoinositides between 7 and 24 hr was 4.1 ± 0.2, 0.4 ± 0.2 and 3.0 ± 1.0 fmol/μmmol total lipid phosphorus for myo-[³H]InsP, -InsP₂ and Ins-P₃ respectively.
• 4.4. Lithium ions are shown to modulate phosphoinositide synthesis and Ins-phosphate accumulation. Ins-mono-, bis- and tris-phosphate production was enhanced 5-, 3- and 2-fold by Li +.
• 5.5. The above results suggest the participation of a C-type phospholipase and of Li-sensitive phosphatases in the modulation of phosphoinositide metabolism in the electrocyte.

     

Metabolism of benzo[a]pyrene and persistence of DNA adducts in the brown bullhead (Ictalurus nebulosus)

• 1.1. The in vitro metabolism of [³H]benzo[a]pyrene (BP) and [¹⁴C]benzo[a]pyrene-7,8-dihydrodiol (BP-7,8-diol) by liver of brown bullhead (Ictalurus nebulosus) was characterized, as was the formation and persistence of BP-DNA adducts in vivo.
• 2.2. Compared to rat liver microsomes, bullhead liver microsomes produced relatively larger amounts of BP-7,8-diol (predominantly the [−] enantiomer) and smaller amounts of BP-4,5-diol.
• 3.3. BP phase I metabolites were efficiently converted by freshly isolated bullhead hepatocytes to conjugates, predominantly glucuronides.
• 4.4. BP-7,8-diol was metabolized by hepatocytes 4-fold more rapidly than was BP and was converted to approximately equal amounts of glucuronides, glutathione conjugates and sulfates.
• 5.5. BP-DNA adducts formed in bullhead liver with a lag time of several days and maximum adduct formation at 25–30 days. The major adduct was anti-BPDE-deoxyguanosine.

     

Characterization of the Melanoplus sanguinipes hemolymph after infection with Beauveria bassiana or wounding

• 1.1. The phenoloxidase activity, protein and carbohydrate levels were studied for 24 hr in the hemolymph of the migratory grasshopper, Melanoplus sanguinipes after artificial wounding of the insect cuticle or the injection of Beauveria bassiana conidia.
• 2.2. Injection or wounding induced a primary response and phenoloxidase activity was found to increase within 10–60 min. The values for phenoloxidase activity in viable B. bassiana-injected insects exhibited a secondary response, i.e., an increase 24 hr after injection.
• 3.3. In wounded insects and those injected with inactivated conidia, the phenoloxidase activity receded after the initial increase and remained at low levels.
• 4.4. Protein concentrations in the hemolymph increased immediately after infection and wounding and returned to basal levels during the course of the experiment.
• 5.5. Injection of viable B. bassiana resulted in a gradual increase in the protein concentrations between 12 and 24 hr.
• 6.6. There was no apparent change in the carbohydrate levels in either B. bassiana-infected or wounded insects.
• 7.7. These results are discussed in relation to their possible role(s) and interrelationships in the immune response to infection or wounding. Furthermore, we suggest that a “factor” is released after mechanical injury of the integument.

     

Changes in high-energy phosphate metabolism in the water scorpion,Ranatra chinensis,under cold water-warm water stress

• 1.1. To evaluate changes in high-energy phosphate metabolism in the water scorpion (Ranatra chinensis) under restraint and cold water-warm water stresses, in vivo [³¹P]NMR spectra were obtained.
• 2.2. Under restraint stress, arginine phosphate (Arg-P) decreased by 10% after 1 hr and remained at that level thereafter, while β-ATP showed negligible changes over 6 hr.
• 3.3. As the water temperature gradually increased or decreased, the relative concentration of Arg-P decreased due to enzyme regulation.
• 4.4. Repeated cold water-warm water stress, which consisted of repeated 15 min exposures to cold water (5°C) followed by 15 min exposures to warm water (30°C) caused distinct decreases in Arg-P and β-ATP concentration. These decreases were dependent on the frequency of exposure.
• 5.5. Phosphomonoesters (PME) increased not only with restraint stress but also with cold water-warm water stress.
• 6.6. The effect of cold water-warm water stress on high-energy phosphate metabolism was greater than that of restraint stress.

     

Preparation and characterization of biotinylated probes for the β-interferon receptor

• 1.1. Recently we described the isolation of the β-interferon receptor [Zhang et al. (1986) J. biol. Chem. 261, 8017–8021]. A highly purified product was obtained but in low quantities.
• 2.2. The use ofbiotinylated β-interferon as a ligand represents an alternate approach to receptor isolation.
• 3.3. We have prepared and characterized the derivatives N-(biotinyl)- and N-(biotinyl-ϵ-aminocaproyl)-recombinant human [Ser¹⁷-interferon β (B- and BC-recHulFNβ).
• 4.4. Biotin incorporation does not result in any loss of antiviral activity, demonstrating the recognition of the derivative by the cell receptor.
• 5.5. The biotinylated recHuIFNβ binds specifically and reversibly to succinoylavidin or guanidine thiocyanate-stripped succinoylavidin linked to a Sepharose matrix.
• 6.6. Comparison of the competition curves obtained with [¹⁴C]biotin and [³H]biotinyl recHuIFN, in the presence of increasing concentrations of biotin suggests that the IFN moiety of the derivative has little effect on the affinity of biotin for avidin.
• 7.7. Biotinylated recHuIFNβ derivatives represent useful probes for the β-IFN receptor.

     

Possible involvement of adenylylation in the modification of a 26 kDa protein in rat parotid acinar cells

• 1.1. Adenylylation, a posttranslational modification of proteins, was investigated in saponin-permeabilized acinar cells of the rat parotid gland.
• 2.2. When cells were incubated with [2,8-³H]ATP, several proteins, including a 26 kDa protein in the particulate fraction, were labeled.
• 3.3. Upon incubation of cells with [α-³²P]ATP in the presence of cAMP and 3-isobutyl-lmethylxanthine, ³²P-labeling of the 26 kDa protein was observed.
• 4.4. After treatment with snake venom phosphodiesterase, [³²P]AMP was released from the 26kDa protein. Such release was not observed when cells were labeled with [γ-³²P]ATP.
• 5.5. The ³²P-labeling pattern of proteins with [α-³²P]ATP was clearly different from that with [adenylate-³²P]NAD⁺.
• 6.6. The results suggest that the 26 kDa protein is one of the adenylylation substrates in rat parotid acinar cells.

     

In vitro substrate utilization for lipid synthesis in liver explants from hyperthyroid chickens

• 1.1. Indian River male broiler chickens growing from 7 to 28 days of age were fed diets containing 12, 18, 24 and 30% protein + 0 or 1 mg triiodothyronine (T₃)/kg of diet to study energetic costs of lipogenesis and the use of various substrates for in vitro lipogenesis.
• 2.2. De novo lipid and CO₂ production were determined in the presence of [1-¹⁴C]pyruvate, [2-¹⁴q]pyruvate, [3-¹⁴C]pyruvate, [2-¹⁴C]acetate and [U-¹⁴C]alanine.
• 3.3. Oxygen consumption was determined in mitochondrial preparations to estimate the energetic costs in expiants synthesizing lipid.
• 4.4. Radiolabeled CO₂ derived from [1-¹⁴C]pyruvate was used as an estimate of coenzyme A availability in liver expiants. Lipids derived from [2-¹⁴C]pyruvate, [2-¹⁴C]acetate and [U-¹⁴C]alanine estimate relative substrate efficiency.
• 5.5. Labeled CO₂ production from [1-¹⁴C]pyruvate was greatest in that group fed a 12% protein diet and least in the group fed a 30% protein diet.
• 6.6. In addition, T₃ increased CO₂ production from [1-¹⁴C]pyruvate.
• 7.7. The production of ¹⁴CO₂ from the second carbon of pyruvate or acetate was increased by T₃.
• 8.8. The low-protein diet (12% protein) increased (P <0.05) lipogenesis.
• 9.9. Adding T₃ to the diets decreased carbon flux into lipid from all substrates, but increased CO₂ production from all substrates without changing stage 3 and 4 respiration rates in mitochondrial preparations.
• 10.10. These observations imply that coenzyme A availability may have regulated de novo lipogenesis in the present study.
• 11.11. It was also concluded that previously noted effects of T₃ on intermediary metabolism may involve metabolic pathways that do not involve changes in mitochondrial function.

     

Amino acid synthesis during hyperosmotic stress in penaeus aztecus postlarvae

• 1.1. Glycine, proline, and taurine are the quantitatively most important amino acid osmolytes in Penaeus aztecus postlarvae.
• 2.2. Taurine dominates the amino acid pool in low salinity, while proline dominates the amino acid pool at higher salinities.
• 3.3. Although not major contributors to the pool, glutamate and alanine are constitutively synthesized from [¹⁴C]glucose and [¹⁴C]glutamate under constant salinity and under hyperosmotic stress treatments.
• 4.4. Proline synthesis from [¹⁴C]-precursors is apparent under constant high (but not low) salinity and is significantly induced by hyperosmotic stress.
• 5.5. No appreciable glycine synthesis was observed from [¹⁴C]glucose or [¹⁴C]glutamate under any experimental conditions.

     

Selective effect of melatonin on the proliferation of lymphoid cells

• 1.1. The present study was designed to investigate the effect of melatonin on the proliferation of normal lymphocytes and certain T-lymphomas and myelomas under in vitro conditions.
• 2.2. The results revealed that administration of 200 μM melatonin inhibited significantly the incorporation of [³H]thymidine into both normal mouse and human lymphocytes and T-lymphoblastoid cell lines.
• 3.3. On the contrary, melatonin provoked an increase of myeloma cell proliferation.
• 4.4. The influence of melatonin on hybridoma cell lines was negligible.
• 5.5. Collectively, these data demonstrated that the chief pineal indole affect selectively the processes of lymphoblastoid cell growth.

     

Synthesis of reserved polysaccharide from wax esters accumulated as the result of anaerobic energy generation in Euglena gracilis returned from anaerobic to aerobic conditions

• 1.1. Euglena gracilis SM-ZK (a non-photosynthetic mutant), cultured in Koren-Hutner medium, containing glucose, malate and glutamate as the main nutrients, were incubated anaerobiosis for 24 hr, and then returned to aerobic conditions. Wax esters, which were synthesized from paramylon (the reserved polysaccharide) for ATP generation under anaerobiosis (wax ester fermentation) were promptly degraded immediately after the cells were replenished with sufficient O₂. A large part (about 70%) of the decomposed wax esters were converted back to paramylon.
• 2.2. When cells were fed with [1–14C]acetate or [U-¹⁴C]acetate immediately after transfer from anaerobic to aerobic conditions, radioactivity incorporated into paramylon in the cells fed with [U-¹⁴C]acetate was about 1.5-times as high as that with [1-¹⁴C]acetate, proposing that glyoxylate cycle participates in the conversion from wax esters to paramylon.
• 3.3. Paramylon synthesis from [1-¹⁴C]acetate was considerably activated by anaerobic preincubation of cells for several hours.
• 4.4. Isocitrate lyase and malate synthase occurred in cells cultured in Koren-Hutner medium, but the activities were obviously lower than those in cells grown on ethanol. These enzymes were not induced by the anaerobic preincubation.

     

Sublethal effects of hypoxia in the abalone (Haliotis rufescens) as measured by in vivo31P NMR spectroscopy

• 1.1. Effects of hypoxia were investigated in red abalones (Haliotis rufescens) using a flow-through exposure system and in vivo³¹P NMR spectroscopy.
• 2.2. Following seawater acclimation, abalones were exposed to air for 1 hr, then seawater for 2.5 hr to check recovery; parallel controls were performed without air exposure.
• 3.3. In foot muscle, hypoxia produced a decrease in phosphoarginine concentration and intracellular pH, an increase in inorganic monophosphate concentration, and no change in that of ATP; upon resubmergence, all effects generally recovered.
• 4.4. The changes induced by hypoxia during normal tidal changes are consistent with the blockage of mitochondrial oxidative phosphorylation.
• 5.5. Use of in vivo NMR allows measurement of the biochemical effects of natural stress factors in live, intact aquatic organisms in the laboratory.

     

Blood volume recovery in hemorrhaged japanese quail

• 1.1. Blood volume and plasma biochemical changes and feed and water consumption in response to a hemorrhage by phlebotomy of 30% of the calculated total blood volume with and without replacement of blood volume with physiological saline were determined in juvenile male Coturnix coturnix japonica.
• 2.2. Plasma protein and osmolality decreased rapidly posthemorrhage and did not recover by 72 hr posthemorrhage.
• 3.3. Plasma glucose, Na⁺ and K⁺ increased within Ihr postphlebotomy. Plasma Na⁺ returned to nonphlebotomized levels within 6 hr postphlebotomy.
• 4.4. Saline replacement of blood volume resulted in hypervolemia within 3–5 min postphlebotomy.
• 5.5. Phlebotomized quail receiving no saline recovered blood volume to 0 hr (nonphlebotomized) levels within l hr postphlebotomy.

     

Contrast in adenine uptake by chicken and rabbit erythrocytes in vitro

• 1.1. Relative to rabbit erythrocytes, chicken red blood cells exhibit a much greater capacity to utilize [³H]adenine for nucleotide synthesis in vitro, even at 5°C and in the absence of added inorganic phosphate.
• 2.2. This difference is largely due to a higher concentration of phosphoribosylpyrophosphate and greater activity of adenine phosphoribosyltransferase in the avian cells. lli]3. The capacity of avian erythrocytes for utilization of guanine and hypoxanthine is several fold less than that of adenine.
• 3.4. The data are consistent with lower activity for hypoxanthine/guanine phosphoribosyltransferase than for adenine phosphoribosyltransferase in intact chicken erythrocytes.
• 4.5. The results indicate that reutilization of adenine by chicken erythrocytes may be physiologically significant.

     

Juvenile hormone biosynthesis by corpus cardiacum-corpus allatum complexes of larval Lymantria dispar

• 1.1. A radiochemical assay was used to examine juvenile hormone (JH) synthesis and secretion in vitro by incubating two pairs of larval corpus cardiacum-corpus allatum complexes (CC-CA) from, Lymantria dispar, in 50 μl of osmotically balanced Grace's medium containing 1 μC1 [³H-methyl]-methionine for 6 hr.
• 2.2. For CC-CA of fourth instar female larvae, maximal incorporation of ³H-methyl was 0.15 pmol/pr/hr between days 2 and 3. High pressure liquid chromatographic (HPLC) analysis suggested that the biosynthetic products are mainly JH III with a little JH II at times.
• 3.3. For CC-CA of last instar female larvae, incorporation of ³H-methyl was 0.48 pmol/pr/hr at the beginning of the stadium and decreased to negligible levels by day 10. HPLC analysis suggested that CC-CA of last instar larvae produced only JH III. Volume increases in CA during the last instar were associated with declining activities of JH secretion.
• 4.4. Comparisons of maximal rates of ³ H-methyl incorporation by each unit volume of CA revealed that in the last instar each unit volume (μm³) of glandular tissue secreted 50% more JH than in the fourth instar.

     

In vivo effect of Berenil on rat liver polyamine metabolism

• 1.1. Berenil, administered to rats in vivo, promoted a decrease in liver SAMDC activity, but an increase in ODC and SAT activity.
• 2.2. Its effect on ODC was completely prevented by cycloheximide, that on SAT only partially.
• 3.3. Berenil had no effect on ODC activity in adrenalectomized rats. Adrenergic antagonists counteracted the effect of Berenil on ODC activity.
• 4.4. Polyamine content was increased. The maximum modification was observed for putrescine and N¹-acetylspermidine.

     

PLA2 activity in rat liver nuclei

• 1.1. Subcellular fractions of rat liver were assayed for PLA₂ activity.
• 2.2. The PLA₂ assay measures the release of [³ H]oleic acid from phospholipids, using labeled E. coli as substrate.
• 3.3. Nuclear fractions contained PLA₂ activity, which was Ca²⁺ dependent and could not be explained from mitochondrial, microsomal or plasma membrane contamination.
• 4.4. The V_max value of nuclear PLA₂ is 0.30 ± 0.04 pmol oleic acid/min/mg protein; its K_m value is 0.86±0.12μM, similar to that of mitochondrial PLA₂.
• 5.5. We conclude that rat liver nuclei contain PLA₂ activity.