Growth, oxygen consumption, and protein and RNA synthesis rates in the yolk sac larvae of the African catfish (Clarias gariepinus)

Rapidly growing African catfish yolk sac larvae were investigated during the first 22 h after hatching. Body compartment protein concentration increased fourfold yet oxygen consumption remained constant (mean=21.3 +/- 3.2 nmol O2 mg(-1) protein h(-1)), suggesting fast growth results mainly from yolk sac protein absorption. The protein synthesis rates at 1-2 and 5-6 h also equaled the highest conceivable rates of muscle protein synthesis; 11.6-11.9% and 7.4-7.9% day(-1), respectively. Therefore the corresponding energetic costs of protein synthesis were almost the theoretical minimum; 13.0 +/- 1.7-16.3 +/- 2.8 micromol O2 mg(-1) protein synthesised. Total protein synthesis expenditure (74.5-77.7 micromol O2 g(-1) protein h(-1)) was also less than other yolk sac larvae. These protein synthesis rates resulted from high RNA concentrations (113.2 +/- 3.4 microg RNA mg(-1) protein) and were also correlated with RNA translational efficiency. High translational efficiency (1 h; 1.2+/-0.1 mg protein synthesised microg(-1) RNA day(-1)) equaled high synthesis rate (36.8 +/- 5.4 microg RNA microg(-1) DNA day(-1)) and both declined over 22 h. This investigation suggests rapid growth combines growth efficiency and compensatory energy partitioning. This accommodates the ontogenetic and phylogenetic standpoints imposed by energy budget limitations.     

Seasonal variations in nitrate reductase activity and internal N pools in intertidal brown algae are correlated with ambient nitrate concentrations

Nitrogen metabolism was examined in the intertidal seaweeds Fucus vesiculosus, Fucus serratus, Fucus spiralis and Laminaria digitata in a temperate Irish sea lough. Internal NO(3) (-) storage, total N content and nitrate reductase activity (NRA) were most affected by ambient NO(3) (-), with highest values in winter, when ambient NO(3) (-) was maximum, and declined with NO(3) (-) during summer. In all species, NRA was six times higher in winter than in summer, and was markedly higher in Fucus species (e.g. 256 +/- 33 nmol NO(3) (-) min(-1) g(-1) in F. vesiculosus versus 55 +/- 17 nmol NO(3) (-) min(-1) g(-1) in L. digitata). Temperature and light were less important factors for N metabolism, but influenced in situ photosynthesis and respiration rates. NO(3) (-) assimilating capacity (calculated from NRA) exceeded N demand (calculated from net photosynthesis rates and C : N ratios) by a factor of 0.7-50.0, yet seaweeds stored significant NO(3) (-) (up to 40-86 micromol g(-1)). C : N ratio also increased with height in the intertidal zone (lowest in L. digitata and highest in F. spiralis), indicating that tidal emersion also significantly constrained N metabolism. These results suggest that, in contrast to the tight relationship between N and C metabolism in many microalgae, N and C metabolism could be uncoupled in marine macroalgae, which might be an important adaptation to the intertidal environment.     

The effect of seasonality on oxidative metabolism in Nacella (Patinigera) magellanica

We studied the seasonal variation on aerobic metabolism and the response of oxidative stress parameters in the digestive glands of the subpolar limpet Nacella (P.) magellanica. Sampling was carried out from July (winter) 2002 to July 2003 in Beagle Channel, Tierra del Fuego, Argentina. Whole animal respiration rates increased in early spring as the animals spawned and remained elevated throughout summer and fall (winter: 0.09+/-0.02 micromol O2 h-1 g-1; summer: 0.31+/-0.06 micromol O2 h-1 g-1). Oxidative stress was assessed at the hydrophilic level as the ascorbyl radical content/ascorbate content ratio (A./AH-). The A./AH- ratio showed minimum values in winter (3.7+/-0.2 10(-5)AU) and increased in summer (18+/-5 10(-5) AU). A similar pattern was observed for lipid radical content (122+/-29 pmol mg-1 fresh mass [FW] in winter and 314+/-45 pmol mg-1 FW in summer), iron content (0.99+/-0.07 and 2.7+/-0.6 nmol mg-1 FW in winter and summer, respectively) and catalase activity (2.9+/-0.2 and 7+/-1 U mg-1 FW in winter and summer, respectively). Since nitrogen derived radicals are thought to be critically involved in oxidative metabolism in cells, nitric oxide content was measured and a significant difference in the content of the Fe-MGD-NO adduct in digestive glands from winter and summer animals was observed. Together, the data indicate that both oxygen and nitrogen radical generation rates in N. (P.) magellanica are strongly dependent on season.     

Energetics of wet-suit diving in Japanese male breath-hold divers

The present study was undertaken to investigate energy balance in professional male breath-hold divers in Tsushima Island, Japan. In 4 divers, rectal (Tre) and mean skin (Tsk) temperatures and rate of O2 consumption (VO2) were measured during diving work in summer (27 degrees C water) and winter (14 degrees C water). Thermal insulation and energy costs of diving work were estimated. In summer, comparisons were made of subjects clad either in wet suits (protected) or in swimming trunks (unprotected), and in winter, they wore wet suits. The average Tre in unprotected divers decreased to 36.4 +/- 0.2 degrees C at the end of 1-h diving work, but in protected divers it decreased to 37.2 +/- 0.3 degrees C in 2 h in summer and to 36.9 +/- 0.1 degree C in 1.5 h in winter. The average Tsk of unprotected divers decreased to 28.0 +/- 0.6 degrees C in summer and that of protected divers decreased to 32.9 +/- 0.5 degrees C in summer and 28.0 +/- 0.3 degrees C in winter. Average VO2 increased 190% (from 370 ml/min before diving to 1,070 ml/min) in unprotected divers in summer, but in protected divers it rose 120% (from 360 to 780 ml/min) in summer and 110% (from 330 to 690 ml/min) in winter. Overall thermal insulation (tissue and wet suit) calculated for protected divers was 0.065 +/- 0.006 degree C X kcal-1 X m-2 X h-1 in summer and 0.135 +/- 0.019 degree C X kcal-1 X m-2 X h-1 in winter.(ABSTRACT TRUNCATED AT 250 WORDS)     

Stimulation of 2',5'-oligoadenylate synthetase activity in sheep endometrium during pregnancy, by intrauterine infusion of ovine trophoblast protein-1, and by intramuscular administration of recombinant bovine interferon-alpha I1.

In Expt 1, activity of 2',5'-oligoadenylate (2',5'-A) synthetase in endometrium collected on Day 16 (oestrus is Day 0) from the uterine horn ipsilateral to the corpus luteum was greater (P less than 0.001) for pregnant (135.5 +/- 1.72 nmol/mg protein/h) than for cyclic ewes (58.5 +/- 0.99 nmol/mg protein/h). In pregnant ewes, activity of 2',5'-A synthetase in endometrium collected from the contralateral uterine horn (119.5 +/- 1.72 nmol/mg protein/h) did not differ from that of the ipsilateral horn. In Expt 2, three ovariectomized ewes were treated with progesterone for 10 days and then with oestrogen for 2 days. Activity of 2',5'-A synthetase on Day 13 was 18% greater (P less than 0.10) in endometrium collected from the uterine horn receiving infusions of 30 micrograms ovine trophoblast protein-1 (oTP-1) twice a day on Days 10, 11 and 12(57.7 +/- 0.22 nmol/mg protein/h) than from the uterine horn receiving control infusions of serum protein (SP; 48.8 +/- 0.22 nmol/mg protein/h). In Expt 3, activity of 2',5'-A synthetase on Day 15 was not significantly greater in endometrium collected from the uterine horn of cyclic ewes receiving infusions of 30 micrograms oTP-1 twice a day on Days 12, 13 and 14 (46.5 +/- 0.37 nmol/mg protein/h) than in endometrium from the uterine horn receiving infusions of SP (38.2 +/- 0.37 nmol/mg protein/h). When results of Expt 2 and Expt 3 were combined, intrauterine infusion of oTP-1 increased (P less than 0.05) activity of 2',5'-A synthetase in endometrium by 20%.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of level and origin of rumen degradable nitrogen on rumen microbial growth and nitrogen utilisation efficiency of animals fed maize silage at maintenance.

Isoenergetic maize silage diets, fed at maintenance to 24 suckling cows (Exp. 1) and to 3 (Exp. 2) or 4 (Exp. 3) rumen fistulated sheep, were compared with a urea and controlled release NPN diet (Exps. 1 and 2) and with a protein-N supplemented diet (Exp. 3). Supplementation increased blood urea concentrations (44.7 +/- 22.3 vs. 97.6 +/- 23.7 mg urea-N x L(-1)) (Exp. 1) and renal urea excretion (2.5 +/- 1.1 vs. 7.6 +/- 1.8 g urea-N x d(-1)) (Exps. 2 and 3), whereas blood allantoin concentrations (286.7 +/- 77.0 micromol x L(-1)) (Exp. 1) and renal excretion of purine derivatives (357.6 +/- 90.7 mg purine-N x d(-1)) (Exps. 2 and 3) were not affected, indicating additional N supplementation did not improve rumen microbial growth. However, some deficiency of rumen degradable N might occur in non supplemented diets as suggested by the reduced rumen NH3-N and RNA concentrations (868 +/- 270 vs. 1466 +/- 466 mg RNA x kg(-1) rumen contents).     

Oxidation of benzene to phenol, catechol, and 1,2,3-trihydroxybenzene by toluene 4-monooxygenase of Pseudomonas mendocina KR1 and toluene 3-monooxygenase of Ralstonia pickettii PKO1

Aromatic hydroxylations are important bacterial metabolic processes but are difficult to perform using traditional chemical synthesis, so to use a biological catalyst to convert the priority pollutant benzene into industrially relevant intermediates, benzene oxidation was investigated. It was discovered that toluene 4-monooxygenase (T4MO) of Pseudomonas mendocina KR1, toluene 3-monooxygenase (T3MO) of Ralstonia pickettii PKO1, and toluene ortho-monooxygenase (TOM) of Burkholderia cepacia G4 convert benzene to phenol, catechol, and 1,2,3-trihydroxybenzene by successive hydroxylations. At a concentration of 165 microM and under the control of a constitutive lac promoter, Escherichia coli TG1/pBS(Kan)T4MO expressing T4MO formed phenol from benzene at 19 +/- 1.6 nmol/min/mg of protein, catechol from phenol at 13.6 +/- 0.3 nmol/min/mg of protein, and 1,2,3-trihydroxybenzene from catechol at 2.5 +/- 0.5nmol/min/mg of protein. The catechol and 1,2,3-trihydroxybenzene products were identified by both high-pressure liquid chromatography and mass spectrometry. When analogous plasmid constructs were used, E. coli TG1/pBS(Kan)T3MO expressing T3MO formed phenol, catechol, and 1,2,3-trihydroxybenzene at rates of 3 +/- 1, 3.1 +/- 0.3, and 0.26 +/- 0.09 nmol/min/mg of protein, respectively, and E. coli TG1/pBS(Kan)TOM expressing TOM formed 1,2,3-trihydroxybenzene at a rate of 1.7 +/- 0.3 nmol/min/mg of protein (phenol and catechol formation rates were 0.89 +/- 0.07 and 1.5 +/- 0.3 nmol/min/mg of protein, respectively). Hence, the rates of synthesis of catechol by both T3MO and T4MO and the 1,2,3-trihydroxybenzene formation rate by TOM were found to be comparable to the rates of oxidation of the natural substrate toluene for these enzymes (10.0 +/- 0.8, 4.0 +/- 0.6, and 2.4 +/- 0.3 nmol/min/mg of protein for T4MO, T3MO, and TOM, respectively, at a toluene concentration of 165 microM).     

Developmental changes of gamma-aminobutyric acid (GABA) and putative amino acid neurotransmitters in the organotypic culture of newborn mouse cerebellum

The quantitative changes and metabolism of GABA and putative amino acid neurotransmitters during early developmental stages in the organotypic culture of newborn mouse cerebellum were examined by using the high-performance liquid chromatograph (HPLC) technique. D-[U-14C]Glucose was used as a precursor of amino acids. To analyze amino acid neurotransmitters, explants were incubated for 4 weeks under standard conditions. The amount of GABA linearly increased from 8.7 +/- 1.3 nmol/mg protein (2 days in vitro, 2 DIV) to 26.5 +/- 6.1 nmol/mg protein (15 DIV) and was saturated after that (24.0 +/- 3.6 nmol/mg protein at 30 DIV). During the period of GABA increase, the capability for GABA synthesis from [14C]glucose increased rapidly from 3.03 +/- 0.67 nCi/mg protein (2 DIV, 3 h incubation) to 9.32 +/- 1.34 nCi/mg protein (15 DIV, 3 h incubation). In the case of glutamic acid, a putative neurotransmitter of granule cell parallel fibers in the cerebellum, the amount in explants was nearly constant during incubation, in contrast with the fact that the amount in vivo gradually increased. However, the capability for glutamic acid synthesis from [14C]glucose increased from 10.80 +/- 3.01 nCi/mg protein (2 DIV, 1 h incubation) to 27.62 +/- 4.71 nCi/mg protein (22 DIV, 1 h incubation). In the case of taurine, found in abundance in fetal brain and supposed to play a specific role in the development and maturation of the central nervous system, the amount in explants decreased from 139.8 +/- 4.0 nmol/mg protein (2 DIV) to 54.0 +/- 0.8 nmol/mg protein (30 DIV).(ABSTRACT TRUNCATED AT 250 WORDS)     

Lipoprotein lipase activity is stimulated by insulin and dexamethasone in cardiomyocytes from diabetic rats.

Type 1 diabetes mellitus reduces lipoprotein lipase (LPL) activity in the heart. The diabetic phenotype of decreased LPL activity in freshly isolated cardiomyocytes persisted after overnight culture (16 h). Total cellular LPL activity was 311+/-56 nmol oleate released x h(-1) x mg(-1) cell protein in diabetic cultured cardiomyocytes compared with 661+/-81 nmol oleate released x h(-1) x mg(-1) cell protein for control cultured cells. Diabetes also resulted in lower heparin-releasable (HR) LPL activity compared with control cells (111+/-25 vs. 432+/-63 nmol x h(-1) x mg(-1) cell protein). In kinetic experiments, the reduction in total cellular LPL and HR-LPL activities in cultured cells from diabetic hearts was due to a decrease in maximal velocity, with no change in apparent Km for substrate (triolein). LPL activity in primary cultures of cardiomyocytes from control rats is stimulated by the combination of insulin (Ins) and dexamethasone (Dex). Overnight treatment of cultured cardiomyocytes from diabetic rats with Ins+Dex elicited an 84% increase in cellular LPL activity (to 572+/-65 nmol x h(-1) x mg(-1) cell protein) and a 194% increase in HR-LPL activity (to 326+/-46 nmol x h(-1) x mg(-1) cell protein). This stimulation occurred at subnanomolar concentrations of the hormones, but neither hormone was effective alone. The amount of immunoreactive LPL protein mass in cultured cardiomyocytes from diabetic hearts was unchanged by Ins+Dex treatment. Addition of oleic acid (60 microM) to the overnight culture medium inhibited the already reduced HR-LPL activity in diabetic cultured cells by 73% (to 30+/-4 nmol x h(-1) x mg(-1) cell protein). The presence of oleic acid also reduced hormone-stimulated HR-LPL activity. Increasing the glucose concentration in the culture medium to 26 mM had no effect on total cellular LPL or HR-LPL activities.     

Saturation mutagenesis of Burkholderia cepacia R34 2,4-dinitrotoluene dioxygenase at DntAc valine 350 for synthesizing nitrohydroquinone, methylhydroquinone, and methoxyhydroquinone

Saturation mutagenesis of the 2,4-dinitrotoluene dioxygenase (DDO) of Burkholderia cepacia R34 at position valine 350 of the DntAc alpha-subunit generated mutant V350F with significantly increased activity towards o-nitrophenol (47 times), m-nitrophenol (34 times), and o-methoxyphenol (174 times) as well as an expanded substrate range that now includes m-methoxyphenol, o-cresol, and m-cresol (wild-type DDO had no detectable activity for these substrates). Another mutant, V350M, also displays increased activity towards o-nitrophenol (20 times) and o-methoxyphenol (162 times) as well as novel activity towards o-cresol. Products were synthesized using whole Escherichia coli TG1 cells expressing the recombinant R34 dntA loci from pBS(Kan)R34, and the initial rates of product formation were determined at 1 mM substrate by reverse-phase high-pressure liquid chromatography. V350F produced both nitrohydroquinone at a rate of 0.75 +/- 0.15 nmol/min/mg of protein and 3-nitrocatechol at a rate of 0.069 +/- 0.001 nmol/min/mg of protein from o-nitrophenol, 4-nitrocatechol from m-nitrophenol at 0.29 +/- 0.02 nmol/min/mg of protein, methoxyhydroquinone from o-methoxyphenol at 2.5 +/- 0.6 nmol/min/mg of protein, methoxyhydroquinone from m-methoxyphenol at 0.55 +/- 0.02 nmol/min/mg of protein, both methylhydroquinone at 1.52 +/- 0.02 nmol/min/mg of protein and 2-hydroxybenzyl alcohol at 0.74 +/- 0.05 nmol/min/mg of protein from o-cresol, and methylhydroquinone at 0.43 +/- 0.1 nmol/min/mg of protein from m-cresol. V350M produced both nitrohydroquinone at a rate of 0.33 nmol/min/mg of protein and 3-nitrocatechol at 0.089 nmol/min/mg of protein from o-nitrophenol, methoxyhydroquinone from o-methoxyphenol at 2.4 nmol/min/mg of protein, methylhydroquinone at 1.97 nmol/min/mg of protein and 2-hydroxybenzyl alcohol at 0.11 nmol/min/mg of protein from o-cresol. The DDO variants V350F and V350M also exhibited 10-fold-enhanced activity towards naphthalene (8 +/- 2.6 nmol/min/mg of protein), forming (1R,2S)-cis-1,2-dihydro-1,2-dihydroxynaphthalene. Hence, mutagenesis of wild-type DDO through active-site engineering generated variants with relatively high rates toward a previously uncharacterized class of substituted phenols for the nitroarene dioxygenases; seven previously uncharacterized substrates were evaluated for wild-type DDO, and four novel monooxygenase-like products were found for the DDO variants V350F and V350M (methoxyhydroquinone, methylhydroquinone, 2-hydroxybenzyl alcohol, and 3-nitrocatechol).     

The activity of cathepsin D in saliva of cystic fibrosis patients

Cystic fibrosis (CF) is genetically determined illness, which is caused by the mutation in the CFTR gene. CFTR protein is also expressed in epithelial cells of parotid glands, therefore parotid glands are also affected in CF patients. Cathepsin D is one of the proteolitic cascade enzymes. Physiological wearing out result in occurrence of trace quantities of this enzyme in serum and body fluids, including saliva. Among different enzymes, saliva contains cathepsin D (CTSD, EC 3.4.23.5). The aim of this study was to determine cathepsin D activity in mixed saliva in cystic fibrosis patients and healthy controls. The study was performed in a group of 26 CF patients (10F, 16M). The results obtained in CF group was compared with the results of thirty healthy subjects (12F, 14M). From each subject 8 ml of mixed saliva was obtained: before and after the stimulation of saliva excretion using paraffin pledgets. Protein and glycoprotein content was assessed using Winzler's method. Protein concentration in controls and CF group before stimulation of excretion was 1.15+/-0.714 mg/mL and 1.54+/-0.925 mg/mL. After stimulation protein concentration in saliva has lowered to 0.88+/-0.77 mg/mL in CF group and 1.24+/-1.213 mg/mL in controls. Glycoprotein concentration in controls and in CF group was respectively: before stimulation 1.08+/-0.271 mg/mL and 1.05+/-0.344 mg/mL; after stimulation 0.92+/-0.292 mg/mL and 0.86+/-0.283 mg/mL. The activity of CTSD in controls was 45.9+/-24.98 Tyr nmol/mL/4h before stimulation and 109.3+/-56.94 Tyr nmol/mL/4h after stimulation of excretion. In CF group CTSD activity before stimulation was 134.5+/-81.80 Tyr nmol/mL/4h and after stimulation 134.4+/-62.18 Tyr nmol/mL/4h. Comparing the CTSD activity in both groups statistically significant difference has been revealed in samples collected before stimulation of excretion (p=0.013). The activity of cathepsin D in saliva of cystic fibrosis patient is significantly higher than in healthy controls before the stimulation of excretion with paraffin pledgets.     

Body protein stores and isotopic indicators of N balance in female reindeer (Rangifer tarandus) during winter

We studied bred and unbred female reindeer (Rangifer tarandus tarandus) during 12 wk of winter when ambient temperatures were low and nitrogen (N) demand for fetal growth is highest in pregnant females. Animals were fed a complete pelleted diet ad lib. that contained 2.54% N in dry matter that was 80% +/- 2% (X +/- SD) digestible. Female reindeer lost 64% +/- 14% of body fat but gained 34% +/- 11% of lean mass from 10 wk prepartum to parturition. These changes were equivalent to average balances of -14.14 +/- 2.35 MJ d(-1) and 10 +/- 3 g N d(-1). Blood cells, serum, and urine declined in (15)N/(14)N in late winter as body protein was gained from the diet. Blood cells of newborn calves were more enriched in (15)N and (13)C than that of their mothers, indicating the deposition of fetal protein from maternal stores. To quantify pathways of N flow in reindeer, N balance was measured by confining animals to cages for 10 d at 4 wk from parturition. N balance was inversely related to (15)N/(14)N in urea-N but not related to (15)N/(14)N of blood cells, creatinine, and feces. The proportion of urea-N derived from body protein increased above 0.46 as N balance fell below -200 mg N kg(-0.75) d(-1). Proportions of urea-N from body protein were -0.01 +/- 0.21 in pregnant females before and after caging and were consistent with average body protein gain in winter. Storage of protein allows reindeer and caribou to tolerate diets that are low in N without impairing fetal development.     

Nitrogen fixation and primary production in Western Mediterranean

Nitrogen fixation, nitrate assimilation and primary production ((13)C/(15)N method) were investigated during one year and half in the northwestern Mediterranean Sea. Nitrogen fixation was detectable all over the year with rates ranged from 2 to 17 nmol N l(-1) d(-1)(d). Highest values being obtained during spring associated with the phytoplankton bloom. High rates (4-8 nmol N l(-1) d(-1)(d)) were also measured during summer, when primary productivity was very low. Then, diazotrophy process supplies significant new nitrogen during summer oligotrophic periods. This new nitrogen input can balance the annual nitrogen biogeochemical budget in the Mediterranean Sea and should explain the high nitrate/phosphate ratio observed in deep waters.     

Taurine supplementation induces multidrug resistance protein 2 and bile salt export pump expression in rats and prevents endotoxin-induced cholestasis

The effect of oral taurine supplementation on endotoxin-induced cholestasis was investigated in rat liver. At 12h following lipopolysaccharide (LPS) injection (4mg/kg body weight i.p.) bile flow and bromosulfophthalein (BSP) and taurocholate (TC) excretion were determined in the perfused liver and the expression of the canalicular transporters multidrug resistance protein 2 (Mrp2) and bile salt export pump (Bsep) was analyzed. Injection of LPS induced a significant decrease of bile flow ( 2.2+/-0.2 microl/g liver wet weight/min vs 3.3+/-0.1 microl/g liver wet weight in controls), biliary BSP excretion (10.8+/-2.2 nmol/g/min vs 21.0+/-3.8 nmol/g/min), and biliary TC excretion (114+/-23 nmol/g/min vs 228+/-8 nmol/g/min). These effects were due to transporter retrieval from the canalicular membrane and downregulation of Mrp2 and Bsep expression. In taurine-supplemented rats bile flow was 30% higher than that in untreated rats and the expression of Mrp2 and Bsep protein was increased two- to threefold. In taurine-supplemented rats there was no significant reduction of bile flow or of BSP and TC excretion at 12h following LPS injection. This protective effect of taurine was due to higher Mrp2 and Bsep protein levels compared to nonsupplemented LPS-treated rats, whereas relative Mrp2 retrieval from the canalicular membrane induced by LPS was not significantly different. LPS-induced tumor necrosis factor alpha and interleukin-1beta release were lower in taurine-fed rats; however, downregulation of Mrp2 and Bsep expression by LPS was delayed but not prevented. The data show that oral supplementation of taurine induces Mrp2 and Bsep expression and may prevent LPS-induced cholestasis.     

Increased cathepsin D-like activity in cortex, tubules, and glomeruli isolated from rats with experimental nephrotic syndrome.

We have examined the activity and distribution of cathepsin D (EC 3.4.23.5), a major renal lysosomal endoproteinase, in the various anatomical and functional areas of normal rat kidney. Cathepsin D-like activities (delta A280/h per mg of protein) in normal rat tissues were: cortex, 0.78 +/- 0.05, n = 37; medulla, 0.62 +/- 0.03, n = 12; papilla, 0.63 +/- 0.04, n = 12; tubules, 0.74 +/- 0.04, n = 28; glomeruli, 0.59 +/- 0.03, n = 28; and liver, 0.41 +/- 0.02, n = 28. Enzyme activity was maximal at pH 3.0-3.5 and inhibited more than 90% by pepstatin (6.7 micrograms/ml), suggesting that the enzyme is cathepsin D. In subsequent experiments we measured cathepsin D-like activity in cortex, tubules and glomeruli isolated from rats with puromycin aminonucleoside (PAN)-induced nephrotic syndrome. Treated animals (15 mg of PAN/100g body wt., intraperitoneally) developed proteinuria beginning 4 days after injection and exceeding 900 mg/24h on day 9. In two separate experiments involving 52 animals we observed a significant increase in cathepsin D-like activity in cortex (+82.7%), tubules (+109.6%) and glomeruli (+54.7%) isolated from PAN-treated rats killed during marked proteinuria (day 9, mean total urinary protein excretion: 937 +/- 94 mg/24h). This increase was observed whether the activity was expressed per mg of DNA or per mg of protein. Increased cathepsin D-like activity was first observed in cortex and tubules coincident with the onset of proteinurea (day 4, mean total urinary protein excretion: 112 +/- 23 mg/24h). In contrast with the significant elevation of renal cathepsin D-like activity, the activity (nmol/h per mg of protein) of alpha-L-fucosidase (EC 3.2.1.51), a non-proteolytic enzyme, was markedly decreased in the identical samples used for the measurement of cathepsin D-like activity: cortex (-46.4%); tubules (-46.1%); and glomeruli (-38.5%). In addition to changes in renal enzyme activities, PAN-treated rats excreted large amounts of cathepsin D-like activity in their urine (beginning on day 3) compared with nearly undetectable cathepsin D-like activity in the urine from control rats. The significant increases in glomerular and tubular cathepsin D activity may reflect an important role for this enzyme in the pathophysiology associated with PAN-induced nephrotic syndrome.     

A new Sphagesaurus (Mesoeucrocodylia: Notosuchia) from the Upper Cretaceous of Monte Alto City (Bauru Group,Brazil), and a revision of the Sphagesauridae

Oxygen consumption and ammonia excretion rates were assessed for Terebratulina retusa (L.) held under 3 different regimes of temperature and food availability. These were: 5.6?C, no food (cold, starved); 5.8?C, food present (cold, fed) and 10.7?C food present (warm, fed), which simulated winter conditions, summer conditions and an intermediate treatment. Regressions of oxygen consumption on ash‐free dry weight (AFDW) had slopes which were not significantly different from each other and ranged from 0.953 to 0.999. A common slope of 0.976 was calculated and intercepts based on the common slope used to compare oxygen consumption in each treatment. The rise from cold, starved conditions to warm, fed was 24.5 per cent and this was significant (P < 0.05). Other differences were not significant (P > 0.05) but the cold, fed result was 12.6 per cent higher than cold, starved. Therefore feeding and temperature probably account for equivalent proportions of the rise in metabolism from winter to summer. Ammonia production data were much more variable. Excretion rates of a 50 mg AFDW individual (in ng‐at NH₃‐N.h^‐1) were as follows: cold, starved: 30.2 cold, fed: 7.1; and warm, fed: 22.9. Oxygen to nitrogen (O:N) ratios reflected these results. Mean O:N ratios were: cold, starved: 8.0; cold, fed: 42.4; warm, fed: 16.3. This shows that the simulated winter group relied heavily on protein to fuel their metabolism, the simulated summer group were less dependent on protein and the intermediate group probably used lipids and carbohydrates to fuel metabolic demands. This possibly reflected a trade off between food supply and increased metabolism from treatment to treatment, demonstrating a flexibility which could have been a contributing factor in the ecological tolerance and geological longevity of some brachiopods.     

A metabolic theory of ecology applied to temperature and mass dependence of N and P excretion by common carp

Our study used a metabolic theory of ecology (MTE) to explore scaling of metabolic rates by body size and temperature, and to predict nutrient excretion by common carp (Cyprinus carpio). At high biomasses, common carp have negative impacts on water quality, and one mechanism is excretion of the nutrients N and P. We measured whole-body and mass-specific excretion rates during summer and winter for fish of different sizes (wet mass range 28–1,196 g) to produce an allometric scaling model capable of predicting excretion at different temperatures. We found positive relationships between both dissolved and total nutrient concentrations and fish wet mass in summer and winter, with greater excretion rates in summer (mean water temperature 24.2°C) than in winter (mean water temperature 9.2°C). Mass-specific excretion rates decreased with increasing fish size, consistent with the MTE, and the temperature-adjusted model explained more variation for N excretion than for P. The proportion of dissolved nutrients (NH₄ and PO₄) to total nutrients increased with increasing fish size. The significance of these models is that they can be used to predict population-based nutrient excretion by common carp when thermal history, fish density and size distribution in a water body are known.     

Epidermal growth factor in serum, urine, submandibular glands and kidneys of diabetic mice

Levels of epidermal growth factor (EGF) in serum were significantly decreased in streptozotocin (STZ)-diabetic mice (446 +/- 168 pg/ml after 1 week and 423 +/- 52 after 4 weeks vs 766 +/- 162 pg/ml in controls, P.002 and less than .001. respectively) and in genetically diabetic ob/ob mice (455 +/- 285 vs 962 +/- 453 pg/ml in nondiabetic ob/+ controls, P.043). The urinary excretion of EGF was significantly increased in STZ mice (104 +/- 53 vs 51 +/- 23 ng/h, P.013) but unchanged in ob/ob mice (33 +/- 9 vs 45 +/- 16 ng/h, P.134). However, when expressed per mg creatinine it was decreased in both cases: in STZ mice to 680 +/- 250 ng/mg at 1 week and 684 +/- 211 at 4 weeks vs 1250 +/- 303 ng/mg in controls (P less than .01); and in the ob/ob mice to 552 +/- 117 vs 1237 +/- 300 ng/mg in ob/+ controls (P less than .01). EGF content of the submandibular glands of STZ mice remained unchanged at 1 week (13.1 +/- 2.9 vs 11.0 +/- 1.8 micrograms/mg protein, P.170) but dropped by 4 weeks (4.7 +/- 1.2 micrograms/mg, P less than .001); in the ob/ob mice it was less than 20% that of controls (2.1 +/- 0.8 vs 12.2 +/- 3.6 micrograms/mg protein). In kidneys, the EGF content was not altered in either ob/ob (524 +/- 50 vs 571 +/- 33 pg/mg protein) or STZ mice (652 +/- 183 vs 665 +/- 80 pg/mg). The preproEGF mRNA level in STZ-treated mice was reduced after 4 weeks in submandibular glands but not in kidneys. The results show that diabetes affects EGF production, utilization and/or excretion in mice and that kidneys are spared from suppression of EGF synthesis that is pronounced in the submandibular glands.     

Determination of 7-methylguanine, N2,N2-dimethylguanosine, and pseudouridine in ultrafiltrated serum of healthy adults by high-performance liquid chromatography

7-Methylguanine (m7Gua), N2,N2-dimethylguanosine (m2(2)Guo), and pseudouridine (psi) are degradation products from RNA turnover and can be used as markers for the whole-body turnover of mRNA-cap, tRNA, and rRNA (in healthy individuals, urinary excretion of these catabolites follows a regular pattern; the relative molar ratio of psi:m7Gua:m2(2)Guo is approximately 100:19:6). HPLC methods were developed to measure serum concentrations of these RNA catabolites after deproteinization of the samples by ultrafiltration through microcollodion bags with a nominal exclusion Mr of 12,400. For healthy adults the following values (mean +/- SD) were found: psi, 2760 +/- 460 nmol/liter (n = 10); m7Gua, 129.7 +/- 24.0 nmol/liter (n = 13); m2(2)Guo, 31.0 +/- 3.7 nmol/liter (n = 9). The relative molar ratio of these substances in serum derived from our data is approximately 100:4.7:1.1. 7-Methylguanosine (m7Guo) added to serum is to a large extent converted to the corresponding free base, m7Gua, the form which is excreted in urine.     

Effect of monensin on the specific activity of ammonia production by ruminal bacteria and disappearance of amino nitrogen from the rumen.

When unadapted mixed ruminal bacteria (312 mg of protein per liter) were treated with monensin (5 mM) in vitro, the rates of ammonia production from enzymatic digests of casein, gelatin, and soy protein (0.5 g of N per liter) were decreased from 46 +/- 2 to 24 +/- 1, 20 +/- 1 to 7 +/- 1, and 40 +/- 2 to 18 +/- 2 nmol/mg of protein per min, respectively. Monensin also caused a decrease in ammonia production in vivo. Nonlactating dairy cows which were fed 0.56 kg of timothy hay 12 times per day had a steady-state ruminal ammonia concentration of 2.7 +/- 0.1 mM, and the ammonia concentration decreased to 1.2 +/- 0.2 mM when monensin (350 mg/day) was added to the diet. The decrease in ammonia production was associated with a 10-fold reduction (4.1 x 10(6) versus 4.2 x 10(5)/ml) in the most probable number of ammonia-producing ruminal bacteria that could use protein hydrolysate as an energy source. Monensin had little effect on the most probable number of carbohydrate-utilizing ruminal bacteria (6.5 versus 7.0 x 10(8)/ml). The addition of protein hydrolysates (560 g) to the rumen caused a rapid increase in the ammonia concentration, but this increase was at least 30% lower when the animals were fed monensin.(ABSTRACT TRUNCATED AT 250 WORDS)