Regulation of glycolytic intermediates in cockroach fat body by the corpus cardiacum

Glycolytic intermediates and related metabolites were measured in the fat body of the American cockroach (Periplaneta americana) to locate the rate-limiting reactions that regulate glycolysis during the action of the corpus cardiacum (CC) in vitro.

Acetylene inhibition technique underestimates in situ denitrification rates in intact cores of freshwater sediment

Abstract Denitrification in intact sediment cores was measured by the acetylene inhibition technique and compared with the nitrate flux between water and sediment. Less than half of the nitrate-N consumed by the sediment could be recovered as nitrous oxide-N. The low recovery rate of nitrous oxide from intact sediment cores indicated losses of nitrous oxide by diffusion down to nitrate-free sediment layers, where reduction of nitrous oxide may take place. In sediment slurries 100% of nitrate-N could be recovered as nitrous oxide-N as long as the nitrate concentration in the liquid phase was above 10 μM. Nitrous oxide added to nitrate-free sediment slurries was reduced regardless of whether acetylene was present or not. Therefore denitrification may be significantly underestimated by this method.     

A minimum mechanism for Na+−Ca++ exchange: Net and unidirectional Ca++ fluxes as functions of ion composition and membrane potential

Summary Both simultaneous and consecutive mechanisms for Na⁺–Ca⁺⁺ exchange are formulated and the associated systems of steady-state equations are solved numerically, and the net and unidirectional Ca⁺⁺ fluxes computed for a variety of ionic and electrical boundary conditions. A simultaneous mechanism is shown to be consistent with a broad range of experimental data from the squid giant axon, cardiac muscle and isolated sarcolemmal vesicles. In this mechanism, random binding of three Na⁺ ions and one Ca⁺⁺ on apposing sides of a membrane are required before a conformational change can occur, translocating the binding sites to the opposite sides of the membranes. A similar (return) translocation step is also permitted if all the sites are empty. None of the other states of binding can undergo such translocating conformational changes. The resulting reaction scheme has 22 reaction steps involving 16 ion-binding intermediates. The voltage dependence of the equilibrium constant for the overall reaction, required by the 31 Na⁺Ca⁺⁺ stoichiometry was obtained by multiplying and dividing, respectively, the forward and reverse rate constants of one of the translocational steps by exp(–FV/2RT). With reasonable values for the membrane density of the enzyme (120 sites m²) and an upper limit for the rate constants of both translocational steps of 10⁵·sec^–1, satisfactory behavior was obtainable with identical binding constants for Ca⁺⁺ on the two sides of the membrane (10⁶ m ^–1), similar symmetry also being assumed for the Na⁺ binding constant (12 to 60m ^–1). Introduction of order into the ion-binding process eliminates behavior that is consistent with experimental findings.     

Regionally selective alterations in enzymatic activities and metabolic fluxes during thiamin deficiency

To further elucidate the molecular basis of the selective damage to various brain regions by thiamin deficiency, changes in enzymatic activities were compared to carbohydrate flux through various pathways from vulnerable (mammillary bodies and inferior colliculi) and nonvulnerable (cochlear nuclei) regions after 11 or 14 days of pyrithiamin-induced thiamin deficiency. After 11 days,large decreases (–43 to –59%) in transketolase (TK) occurred in all 3 regions; 2-ketoglutarate dehydrogenase (KGDHC) declined (–45%), but only in mammillary bodies; pyruvate dehydrogenase (PDHC) was unaffected. By day 14, TK remained reduced by 58%–66%; KGDHC was now reduced in all regions (–48 to –55%); PDHC was also reduced (–32%), but only in the mammillary bodies. Thus, the enzyme changes did not parallel the pathological vulnerability of these regions to thiamin deficiency.¹⁴CO₂ production from¹⁴C-glucose labeled in various positions was utilized to assess metabolic flux. After 14 days, CO₂ production in the vulnerable regions declined severely (–46 to 70%) and approximately twice as much as those in the cochlear nucleus. Also by day 14, the ratio of enzymatic activity to metabolic flux increased as much as 56% in the vulnerable regions, but decreased 18 to 30% in the cochlear nuclei. These differences reflect a greater decrease in flux than enzyme activities in the two vulnerable regions. Thus, selective cellular responses to thiamin deficiency can be demonstrated ex vivo, and these changes can be directly related to alterations in metabolic flux. Since they cannot be related to enzymatic alterations in the three regions, factors other than decreases in the activity of these TPP-dependent enzymes must underlie selective vulnerability in this model of thiamin deficiency.Abbreviations KGDHC 2-ketoglutarate dehydrogenase complex EC 1.2.4.2., EC 2.3.1.61, EC 1.6.4.3. - PDHC pyruvate dehydrogenase complex EC 1.2.4.2., EC 2.3.1.12, EC 1.6.4.3 - TK transketolase (EC 2.2.1.1) - TPP thiamin pyrophosphate     

The response of isoprene emission rate and photosynthetic rate to photon flux and nitrogen supply in aspen and white oak trees

Isoprene is the primary biogenic hydrocarbon emitted from temperate deciduous forest ecosystems. The effects of varying photon flux density (PFD) and nitrogen growth regimes on rates of isoprene emission and net photosynthesis in potted aspen and white oak trees are reported. In both aspen and oak trees, whether rates were expressed on a leaf area or dry mass basis, (1) growth at higher PFD resulted in significantly higher rates of isoprene emission, than growth at lower PFD, (2) there is a significant positive relationship between isoprene emission rate and leaf nitrogen concentration in both sun and shade trees, and (3) there is a significant positive correlation between isoprene emission rate and photosynthetic rate in both sun and shade trees. The greater capacity for isoprene emission in sun leaves was due to both higher leaf mass per unit area and differences in the biochemical and/or physiological properties that influence isoprene emission. Positive correlations between isoprene emission rate and leaf nitrogen concentration support the existence of mechanisms that link leaf nitrogen status to isoprene synthase activity. Positive correlations between isoprene emission rate and photosynthesis rate support previous hypotheses that isoprene emission plays a role in protecting photosynthetic mechanisms during stress.     

Effect of maximal arm exercise on skin blood flux in the paralyzed lower limbs in persons with spinal cord injury

The purpose of the present study was to examine the effect of maximal arm exercise on the skin blood circulation of the paralyzed lower limbs in persons with spinal cord injury (PSCI). Eight male PSCI with complete lesions located between T3 and L1 performed graded maximal arm-cranking exercise (MACE) to exhaustion. The skin blood flux at the thigh (SBFT) and that at the calf (SBFC) were monitored using laser-Doppler flowmeter at rest and for 15 s immediately after the MACE. The subject's mean peak oxygen uptake and peak heart rate was 1.41 ± 0.22 1·min⁻¹ and 171.6 ± 19.2 beats·min⁻¹, respectively. No PSCI showed any increase in either SBFT or SBFC after the MACE, when compared with the values at rest. These results suggest that the blood circulation of the skin in the paralyzed lower limbs in PSCI is unaffected by the MACE.     

Biochemical diversity among sulfur-dependent, hyperthermophilic microorganisms

Abstract: Hyperthermophiles are a recently discovered group of microorganisms that grow at and above 90°C. They currently comprise over 20 different genera, and except for two novel bacteria, all are classified as Archaea. The majority of these organisms are obligately anaerobic heterotrophs that reduce elemental sulfur (S°) to H₂S. The best studied from a biochemical perspective are the archaeon, Pyrococcus furiosus , and the bacterium, Thermotoga maritima , both of which are saccharolytic. P. furiosus is thought to contain a new type of Entner-Doudoroff pathway for the conversion of carbohydrates ultimately to acetate, H₂ and CO₂. The pathway is independent of nicotinamide nucleotides and involves novel types of ferredoxin-linked oxidoreductases, one of which has tungsten, a rarely used element, as a prosthetic group. The only site of energy conservation is at the level of acetyl CoA, which in the presence of ADP and phosphate is converted to acetate and ATP in a single step. In contrast, T. maritima utilizes a conventional Embden-Meyerhof pathway for sugar oxidation. P. furiosus also utilizes peptides as a sole carbon and energy source. Amino acid oxidation is thought to involve glutamate dehydrogenase together with at least three types of novel ferredoxin-linked oxidoreductases which catalyze the oxidation of 2-ketoglutarate, aryl pyruvates and formaldehyde. One of these enzymes also utilizes tungsten. In P. furiosus , virtually all of the reductant that is generated during the catabolism of both carbohydrates and peptides is channeled to a cytoplasmic hydrogenase. This enzyme is now termed sulhydrogenase, as it reduces both protons to H₂ and S°(or polysulfide) to H₂S. S° reduction appears to lead to the conservation of energy in P. furiosus but not in T. maritima , although the mechanism by which this occurs is not known.     

Genetic manipulation of 6-phosphofructokinase in potato tubers

The aim of this work was to discover whether genetic manipulation of 6-phosphofructokinase [EC 2.7.1.11; PFK(ATP)] influenced the rate of respiration of tuber tissue of Solanum tuberosum L. Transgenic plants were produced that contained the coding sequence of the Escherichia coli pfkA gene linked to a patatin promoter. Expression of this chimaeric gene in tubers resulted in a 14to 21-fold increase in the maximum catalytic activity of PFK(ATP) without affecting the activities of the other glycolytic enzymes. Tubers, and aged disks of tuber tissue, from transformed plants showed no more than a 30% fall in the content of hexose 6-monophosphates; the other intermediates of glycolysis increased threeto eightfold. Fructose-2,6-bisphosphate was barely detectable in aged disks of transformed tubers. The relative rates of ¹⁴CO₂ production from [1-¹⁴C]-and [6-¹⁴C]-glucose supplied to disks of transformed and control tubers were similar. Oxygen uptake and CO₂ production by aged disks of transformed tubers did not differ significantly from those from control tubers. The same was true of CO₂ production, in air, and in nitrogen, for tuber tissue. It is concluded that PFK(ATP) does not dominate the control of respiration in potato tubers.Abbreviations Fru2,6bisP fructose-2,6-bisphosphate - FW freshweight - GUS -glucuronidase - PFK(ATP) 6-phosphofructokinase - PFK(PPi) pyrophosphate: fructose-6-phosphate 1-phosphotransferase     

Volatile anesthetics inhibit NMDA-stimulated45Ca uptake by rat brain microvesicles

We have previously shown that volatile anesthetics inhibit glutamate-stimulated [³H]MK-801 binding to the ionophore of NMDA receptor complexes in rat brain. In the present study, we examined the influence of enflurane and halothane on NMDA-stimulated⁴⁵Ca uptake by a microvesicle fraction isolated from rat brain. NMDA stimulated⁴⁵Ca uptake (30 sec) by rat brain microvesicles by up to 70% with an EC₅₀ of 1.4±0.5 M. The NMDA-stimulated⁴⁵Ca uptake was inhibited by MK-801 and D-AP-5 with IC₅₀'s of 10 M. Enflurane and halothane inhibited⁴⁵Ca uptake stimulated by 100 M NMDA by as much as 60–80% with IC₅₀'s of 0.2–0.3 mM, concentrations achieved during routine clinical use. Basal⁴⁵Ca uptake measured in the absence of agonist was not affected by the anesthetics. Glycine did not affect the level of NMDA-stimulated⁴⁵Ca uptake, but markedly reduced the inhibition of uptake caused by enflurane and halothane. Preincubation of microvesicles with NMDA resulted in a desensitization of NMDA-stimulated⁴⁵Ca uptake, with a t_1/2 of 20 sec. Enflurane and halothane diminished both the extent and rate of development of this desensitization, as did glycine. These findings support the idea that volatile anesthetic interference with neurotransmission at NMDA receptor complexes contributes to the development of the anesthetic state.