Carbon Isotope Ratios (13C/12C) in Fractions of Field-Grown Grape

The carbon isotope ratio (δ¹³C%) in leaves and berries of Vitis vinifera L. was determined during the fruiting season. The δ¹³C was from ?24.6 to ?26.3 in total leaf tissue, while it is from ?22.7 to ?24.3 in the aqueous extract of leaves. The δ¹³C in the berries was more stable, around ?24, with no change during ripening. Malic and tartaric acid, isolated from the berries at the onset of ripening, showed a δ¹³C of ?23.5 and ?23.0 respectively. It was concluded that malic acid accumulated in the berries at the onset of ripening is not formed directly through β-carboxylation.     

Testing the branch autonomy theory: a 13C/14C double-labelling experiment on differentially shaded branches

The impact of a heterogeneous within‐crown light environment on carbon allocation was investigated on young walnut trees trained on two branches: one left in full sunlight, the other shaded until leaf fall resulting in 67% reduction in photosynthetically active radiation. In September, the two branches were separately labelled with ¹⁴CO₂ and ¹³CO₂, respectively, so that the photosynthates from each branch could be traced independently at the same time. Although some carbon movements could be detected within 5 d in both directions (including from the shaded branch to the sun branch), between‐branch carbon movements were very limited: approximately 1% of the diurnal net assimilation of a branch. At this time of the year branch autonomy was nearly total, leading to increased relative respiratory losses and a moderate growth deficit in the shaded branch. The ratio of growth to reserve storage rate was only slightly affected, indicating that reserves acted not as a mere buffer for excess C but as an active sink for assimilates. In winter, branch autonomy was more questionable, as significant amounts of carbon were imported into both branches, possibly representing up to 10% of total branch reserves. Further within‐plant carbon transfers occurred in spring, which totally abolished plant autonomy, as new shoots sprouted on each branch received significantly more C mobilized from tree‐wide reserves than from local, mother‐branch located reserves. This allowed great flexibility of tree response to environment changes at the yearly time scale. As phloem is considered not functional in winter, it is suggested that xylem is involved as the pathway for carbohydrate movements at this time of the year. This is in agreement with other results regarding sugar exchanges between the xylem vessels and the neighbouring reserve parenchyma tissues.     

Production of [14C]Acetylcholine and [14C]Carbon Dioxide from [U–14C]Glucose in Tissue Prisms from Aging Rat Brain

Abstract: Production of [¹⁴C]acetylcholine and ¹⁴CO₂ was examined by using tissue prisms from neocortex, hippocampus, and striatum from rats aged approximately 5 months, 13 months, and 27 months. [¹⁴C]Acetylcholine synthesis in the striatum showed highly significant decreases with age for measurements in the presence of both 5 m m - and 31 m m -K⁺, contrasting with the lack of significant change in ¹⁴CO₂ production in this region. The neocortex and hippocampus showed only small changes, especially when comparison was made between 13-month and senescent animals. Measurements of the release of [¹⁴C]acetylcholine and influence of atropine on this release confirmed the relative stability with age of the cholinergic system in the neocortex.     

UPTAKE OF [14C]ASPARTIC ACID AND [14C]GLUTAMIC ACID BY RETINAL SYNAPTOSOMAL FRACTIONS

Abstract— Uptake systems for [¹⁴C]aspartate and [¹⁴C]glutamate were characterized in two distinct synaptosomal fractions solated from rabbit retina. The P, synaptosomal fraction was highly enriched in large photoreceptor cell synaptosomes but contained very few conventional sized synaptosomes from amacrine, horizontal or bipolar cells. In contrast, the P₂ synaptosomal fraction contained numerous conventional sized synaptosomes and was virtually free of photoreceptor cell synaptosomes. Both synaptosomal fractions took up [¹⁴C]aspartate and [¹⁴C]glutamate with high affinity [ K _m= 1–2μM). Uptake characteristics were similar to those described for high affinity uptake systems in brain synaptosomes, i.e. saturation kinetics; temperature and Na⁺ dependence. Although the presence of a high affinity uptake system is not a definitive criterion for demonstration of functional neurotransmitter systems, it is an important and necessary prerequisite and can thus be considered as supportive evidence for the involvement of asparate and glutamate in neurotransmission in rabbit retina.     

Metabolism of [14C]ss-glucose and [14C]-acetate by lettuce embryos during early germination

The metabolism of ['⁴C]-labelled glucose and acetate has been investigated during the early germination - before radicle emergence - of lettuce ( Lactuca sativa L., cv. Val d'Orge) embryos. Similar amounts of radioactivity from both substrates were evolved as C., or incorporated into organic acids, amino acids and proteins. A large part of the [¹⁴C]-glucose was also incorporated into sucrose and polysaecharides, and a small part into the glycerol moiety of lipids. Acetate was massively incorporated into lipids, and only slightly into neutral compounds. These results show that both glucose and acetate can be utilized as respiratory substrates during early germination of lettuce embryos. Various biosynthetic pathways leading to amino acids, proteins, polysaecharides and lipids are active during this period.     

Transport of Kinetin-814C in Petioles

Several differences in the translocation pattern of radioactive kinetin in plant petioles were determined. Radioactivity from kinetin-8-¹⁴C (Kn*) moved from donor agar blocks through petioles of bean and cocklebur but not of cotton. There was no difference in basipetal or acropetal movement of radioactivity from Kn* in cocklebnr petioles, but there was in bean petioles. In bean petioles this movement was preferentially basipetal, but it was influenced by the age of the petiole and by the presence of added indoleactic acid. The combination treatment accelerated the basipetal movement of radioactivity from Kn* in young bean petioles and not in old ones. All data is based on radioactivity translocated into receiver agar blocks which were assayed individually in a liquid scintillation spectrometer. The results show that plant species, direction of transport, age of tissue, and presence of IAA can all influence the translocation of Kn* in petioles.     

Natural 13C and 15N abundance of field-collected fungi and their ecological implications

The natural abundance of ¹³C and ¹⁵N was measured in basidiocarps of at least 115 species in 88 genera of ectomycorrhizal, wood-decomposing and litter-decomposing fungi from Japan and Malaysia. The natural abundance of ¹³C and ¹⁵N was also measured in leaves, litter, soil and wood from three different sites. ¹⁵N and ¹³C were enriched in ectomycorrhizal and wood-decomposing fungi, respectively, relative to their substrates. Ectomycorrhizal and wood-decomposing fungi could be distinguished on the basis of their δ¹³C and δ¹⁵N signatures. Although there was high variability in the isotopic composition of fungi, the following isotope- enrichment factors (ε, mean±SD) of the fungi relative to substrates were observed:
ε_{ectomycorrhizal fungi/litter} = 6.1±0.4‰¹⁵N
ε_{ectomycorrhizal fungi/wood} = 1.4±0.8‰¹³C
ε_{wood-decomposing fungi/wood} = −0.6±0.7‰¹⁵N
ε_{wood-decomposing fungi/wood} = 3.5±0.9‰¹³C
The basis of isotope fractionation in C metabolism from wood to wood-decomposing fungus is discussed.     

Acquisition and within-plant allocation of 13C and 15N in CO2-enriched Quercus robur plants

We assessed the effects of doubling atmospheric CO₂ concentration, [CO₂], on C and N allocation within pedunculate oak plants (Quercus robur L.) grown in containers under optimal water supply. A short-term dual ¹³CO₂ and ¹⁵NO₃^? labelling experiment was carried out when the plants had formed their third growing flush. The 22-week exposure to 700 μl l^?1 [CO₂] stimulated plant growth and biomass accumulation (+53% as compared with the 350 μl l^?1 [CO₂] treatment) but decreased the root/shoot biomass ratio (-23%) and specific leaf area (-18%). Moreover, there was an increase in net CO₂ assimilation rate (+37% on a leaf dry weight basis; +71% on a leaf area basis), and a decrease in both above- and below-ground CO₂ respiration rates (-32 and -26%, respectively, on a dry mass basis) under elevated [CO₂]. ¹³C acquisition, expressed on a plant mass basis or on a plant leaf area basis, was also markedly stimulated under elevated [CO₂] both after the 12-h ¹³CO₂ pulse phase and after the 60-h chase phase. Plant N content was increased under elevated CO₂ (+36%), but not enough to compensate for the increase in plant C content (+53%). Thus, the plant C/N ratio was increased (+13%) and plant N concentration was decreased (-11%). There was no effect of elevated [CO₂] on fine root-specific ¹⁵N uptake (amount of recently assimilated ¹⁵N per unit fine root dry mass), suggesting that modifications of plant N pools were merely linked to root size and not to root function. N concentration was decreased in the leaves of the first and second growing flushes and in the coarse roots, whereas it was unaffected by [CO₂] in the stem and in the actively growing organs (fine roots and leaves of the third growth flush). Furthermore, leaf N content per unit area was unaffected by [CO₂]. These results are consistent with the short-term optimization of N distribution within the plants with respect to growth and photosynthesis. Such an optimization might be achieved at the expense of the N pools in storage compartments (coarse roots, leaves of the first and second growth flushes). After the 60-h ¹³C chase phase, leaves of the first and second growth flushes were almost completely depleted in recent ¹³C under ambient [CO₂], whereas these leaves retained important amounts of recently assimilated ¹³C (carbohydrate reserves?) under elevated [CO₂].     

Metabolic Fluxes Between [14C]2-Deoxy-D-Glucose and [14C]2-Deoxy-D-Glucose-6-Phosphate in Brain In Vivo

The rates of the phosphorylation and dephosphorylation of 2-deoxyglucose were measured in rat brain in vivo using tracer kinetic techniques. The rate constant for each reaction was estimated from two separate experiments with different protocols for tracer administration. Tracer amounts of [1-14C]2-deoxyglucose (1 microCi) were injected through the internal carotid artery (intraarterial experiment), or through the atrium (intravenous experiment). Brains were sampled by freeze-blowing at various times after the injection. In the intraarterial experiment, the rate constant for the forward reaction from 2-deoxyglucose to 2-deoxyglucose phosphate was calculated by dividing the initial rate of 2-deoxyglucose phosphate production by the 2-deoxyglucose content in brain. The rate constant for the reverse reaction from 2-deoxyglucose phosphate to 2-deoxyglucose was calculated from the decay constant of 2-deoxyglucose phosphate. The rate constants estimated were 10.1 +/- 1.4%/min (SD) and 3.00 +/- 0.01%/min (SD), respectively, for the forward and reverse reactions. In the intravenous experiment, rate constants for both reactions were estimated by compartmental analysis. By fitting data to program SAAM-27, the rate constants for the forward and reverse reactions were estimated as 11.4 +/- 0.4%/min (SD) and 5.1 +/- 0.4%/min (SD), respectively. The rate constants determined were compared to those for the reactions between glucose and glucose-6-phosphate, estimated previously from labeled glucoses. It is concluded that the rate of glucose utilization measured by the 2-deoxyglucose method reflects the rate of the hexokinase reaction and not the rate of glucose utilization or brain energy utilization.     

Studies on the uptake of (14C) amino acids derived from both dietary (14C) protein and dietary (14C) amino acids by rainbow trout, Salmo gairdneri Richardson

Previous studies have shown that rainbow trout fed on diets containing whole protein have superior growth rates compared to fish fed on diets of similar amino acid composition but containing a high proportion of free amino acids. The influence of several nutritional factors on the uptake of radioactivity from food pellets containing either [U-^I4C] protein or [U-¹⁴C] amino acids into the systemic blood of trout has been investigated. The time taken for radioactivity in the free amino acid fraction of blood to reach a peak after a meal containing [U-¹⁴C] protein had been given was much shorter, and the level of radioactivity in the blood higher, in trout with almost empty stomachs than in fish with almost full stomachs; uptake of radioactivity into blood amino acids was also more rapid and reached much higher concentrations when pellets containing [U-¹⁴C] amino acids were fed than when [U-¹⁴C] protein was fed. Incorporation of radioactivity into blood protein continued for a much longer period and reached higher levels when a pellet containing [U-¹⁴C] protein was fed than when a pellet containing [U-¹⁴C] amino acids was fed. Previous dietary history (low or high protein intake) did not appear to affect the rate of absorption of amino acids from either protein or free amino acid pellets. The uptake rates from pellets containing free amino acids could be slowed by mixing the dietary amino acids with albumin. The distribution, postabsorption, of radioactivity in the different fractions of blood and liver suggested that incorporation of carbon residues into glycogen and lipid from an amino acid diet was greater than from a protein diet. The converse was true of incorporation of radioactivity into tissue protein.     

MECHANISMS FOR THE EFFLUX OF [14C]DOPA AND [14C]DOPAMINE FROM THE CSF OF RHESUS MONKEYS

—Clearance of [¹⁴C]DOPA and [¹⁴C]dopamine from CSF was investigated in anaesthetized rhesus monkeys (M. Mulatta) subjected to ventriculocisternal perfusion. The efflux coefficients, k_VE, at tracer concentrations (3–5 m ) in the perfusate were 0.0487 ml/min and 0.0325 ml/min for [¹⁴C]DOPA and [¹⁴C]dopamine, respectively. Carrier DOPA (10 mm ) in the perfusate decreased the efflux of [¹⁴C]DOPAsignificantly, but carrier dopamine had no appreciable effect on the clearance of [¹⁴C]dopamine. These findings suggest that DOPA is cleared from CSF in part by a saturable mechanism which may be located in the choroid plexus, whereas dopamine leaves the ventricular system by passive diffusion. Radioactivity in the caudate nucleus immediately adjacent to the perfused ventricle averaged 15.5 % and 12.6% of the radioactivity in the perfusates with [¹⁴C]DOPA or [¹⁴C]dopamine, respectively. These distribution percentages were similar to those found for various extracellular indicators after ventriculocisternal perfusion and may indicate that the efflux of intraventricularly-administered exogenous DOPA and dopamine occurs in part through extracellular channels.     

Beyond CO2-fixation by Rubisco – an interpretation of 13C/12C variations in tree rings from novel intra-seasonal studies on broad-leaf trees

Evidence is presented for a very specific, seasonally recurring tri‐phase carbon isotope pattern in tree rings of broad‐leaf deciduous tree species. It is derived from highly resolved intra‐annual measurements of ¹³C/¹²C ratios of wood and cellulose from tree rings of Fagus sylvatica, Populus nigra, Quercus petraea and Morus alba. Investigations on δ¹³C from buds and leaves of Fagus sylvatica revealed a similar tri‐phase δ¹³C pattern. At the very beginning of a growing season, the δ¹³C trend of tree rings and foliage shows a marked increase of up to 5‰. The maximum δ¹³C‐value of each vegetation period always occurs in young heterotrophic leaves shortly after bud burst and persistently in the early wood of each tree ring, when growth depends on carbon reserves. Thereafter, δ¹³C profiles represent the autotrophic stage of the leaves, which show different patterns of variation, by and large characterized by a decline. The minimum δ¹³C‐value always shows up in the late wood of each tree ring. At the very end of each tree ring δ¹³C‐values start rising again. This increase in δ¹³C marks the gradual switch‐over to storage‐dependent growth and can also be observed in senescent leaves. Seasonal changes of more than 4‰ were measured, whereas contiguous δ¹³C values rarely differed from each other by more than 0.3‰. This tri‐phase pattern cannot be explained by the common model of carbon isotope fractionation during photosynthesis. It appears to be primarily an indication of seasonal changes in down‐stream processes of the carbohydrate metabolism. Environmental influences on the carbon isotope fractionation during photosynthesis are presumably of secondary importance and expressed by certain peculiarities showing up during the autotrophic phase, i.e. the mid‐section of the seasonal δ¹³C pattern.     

13C and 15N in microarthropods reveal little response of Douglas-fir ecosystems to climate change

Understanding ecosystem carbon (C) and nitrogen (N) cycling under global change requires experiments maintaining natural interactions among soil structure, soil communities, nutrient availability, and plant growth. In model Douglas-fir ecosystems maintained for five growing seasons, elevated temperature and carbon dioxide (CO₂) increased photosynthesis and increased C storage belowground but not aboveground. We hypothesized that interactions between N cycling and C fluxes through two main groups of microbes, mycorrhizal fungi (symbiotic with plants) and saprotrophic fungi (free-living), mediated ecosystem C storage. To quantify proportions of mycorrhizal and saprotrophic fungi, we measured stable isotopes in fungivorous microarthropods that efficiently censused the fungal community. Fungivorous microarthropods consumed on average 35% mycorrhizal fungi and 65% saprotrophic fungi. Elevated temperature decreased C flux through mycorrhizal fungi by 7%, whereas elevated CO₂ increased it by 4%. The dietary proportion of mycorrhizal fungi correlated across treatments with total plant biomass (n= 4, r²= 0.96, P= 0.021), but not with root biomass. This suggests that belowground allocation increased with increasing plant biomass, but that mycorrhizal fungi were stronger sinks for recent photosynthate than roots. Low N content of needles (0.8–1.1%) and A horizon soil (0.11%) coupled with high C : N ratios of A horizon soil (25–26) and litter (36–48) indicated severe N limitation. Elevated temperature treatments increased the saprotrophic decomposition of litter and lowered litter C : N ratios. Because of low N availability of this litter, its decomposition presumably increased N immobilization belowground, thereby restricting soil N availability for both mycorrhizal fungi and plant growth. Although increased photosynthesis with elevated CO₂ increased allocation of C to ectomycorrhizal fungi, it did not benefit plant N status. Most N for plants and soil storage was derived from litter decomposition. N sequestration by mycorrhizal fungi and limited N release during litter decomposition by saprotrophic fungi restricted N supply to plants, thereby constraining plant growth response to the different treatments.     

Correlating δ13C and δ18O in cellulose of trees

We measured the carbon and oxygen isotopic composition of stem cellulose of Pinus sylvestris, Picea abies, Fagus sylvatica and Fraxinus excelsior. Several sites along a transect of a small valley in Switzerland were selected which differ in soil moisture conditions. At every site, six trees per species were sampled, and a sample representing a mean value for the period from 1940 to 1990 was analysed. For all species, the mean site δ¹³C and δ¹⁸O of stem cellulose are related to the soil moisture availability, whereby higher isotope ratios are found at drier sites. This result is consistent with isotope fractionation models when assuming enhanced stomatal resistance (thus higher δ¹³C of incorporated carbon) and increased oxygen isotope enrichment in the leaf water (thus higher δ¹⁸O) at the dry sites. δ¹⁸ O-δ¹³C plots reveal a linear relationship between the carbon and oxygen isotopes in cellulose. To interpret this relationship we developed an equation which combines the above-mentioned fractionation models. An important new parameter is the degree to which the leaf water enrichment is reflected in the stem cellulose. In the combined model the slope of the δ¹⁸O-δ¹³C plot is related to the sensitivity of the p_i/p_a of a plant to changing relative humidity.     

Metabolism of [14C]trans-zeatin and [14C]benzyladenine by letached yellow, green and variegated leaves of Schefflera

[8-¹⁴C]Benzyladenine (BA) and [8-¹⁴C] trans-zeatin (tZ) were fed through the petiole to mature, detached green, yellow and variegated leaves of Schefflera arboricola. Recovery of radioactivity from the plant material ranged between 4.2 and 22.1%. More radioactivity was recovered when tZ was applied compared to BA. Green leaves or the green parts of variegated leaves yielded more radioactivity than the yellow leaf material. BA was metabolized much faster than the endogenous cytokinin tZ. It would appear that while lower amounts of radioactivity were present in yellow leaves, as well as in yellow parts of variegated leaves, the rate of cytokinin metabolism was nevertheless faster. Metabolites that were formed to a greater extent in these yellow parts were the nucleotides of both cytokinins. Currently it is not known whether or not cytokinins influence chlorophyll and other pigment development in chimeric variegated leaves.     

Compartmentation of [14C]Glutamate and [14C]Glutamine Oxidative Metabolism in the Rat Hippocampus as Determined by Microdialysis

Abstract: Metabolic compartmentation of amino acid metabolism in brain is exemplified by the differential synthesis of glutamate and glutamine from the identical precursor and by the localization of the enzyme glutamine synthetase in glial cells. In the current study, we determined if the oxidative metabolism of glutamate and glutamine was also compartmentalized. The relative oxidation rates of glutamate and glutamine in the hippocampus of free-moving rats was determined by using microdialysis both to infuse the radioactive substrate and to collect ¹⁴CO₂ generated during their oxidation. At the end of the oxidation experiment, the radioactive substrate was replaced by artificial CSF, 2 min-fractions were collected, and the specific activities of glutamate and glutamine were determined. Extrapolation of the specific activity back to the time that artificial CSF replaced ¹⁴C-amino acids in the microdialysis probe yielded an approximation of the interstitial specific activity during the oxidation. The extrapolated interstitial specific activities for [¹⁴C]glutamate and [¹⁴C]glutamine were 59 ± 18 and 2.1 ± 0.5 dpm/pmol, respectively. The initial infused specific activities for [U-¹⁴C]glutamate and [U-¹⁴C]glutamine were 408 ± 8 and 387 ± 1 dpm/pmol, respectively. The dilution of glutamine was greater than that of glutamate, consistent with the difference in concentrations of these amino acids in the interstitial space. Based on the extrapolated interstitial specific activities, the rate of glutamine oxidation exceeds that of glutamate oxidation by a factor of 5.3. These data indicate compartmentation of either uptake and/or oxidative metabolism of these two amino acids. The presence of [¹⁴C]glutamine in the interstitial space when [¹⁴C]glutamate was perfused into the brain provided further evidence for the glutamate/glutamine cycle in brain.