NMR Studies of Pi-Containing Extracellular and Cytoplasmic Compartments in Brain

Abstract: The inorganic phosphate (P_i) NMR peak in brain has an irregular shape, which suggests that it represents more than a single homogeneous pool of P_i. To test the ability of the Marquardt-Levenberg (M-L) nonlinear curve fit algorithm software (Peak-Fit) to separate multiple peaks, locate peak centers, and estimate peak heights, we studied simulated P_i spectra with defined peak centers, areas, and signal-to-noise (S/N) ratios ranging from ∞ to 5.8. As the S/N ratio decreased below 15, the M-L algorithm located peak centers accurately when they were detected; however, small peaks tended to grow smaller and disappear, whereas the amplitudes of larger peaks increased. We developed an in vitro three-compartment model containing a mixture of P_i buffer, phosphocreatine, phosphate diester, and phosphate monoester (PME), portions of which were adjusted to three different pHs before addition of agar. Weighed samples of each buffered gel together with phospholipid extract and bone chips were placed in an NMR tube and covered with mineral oil. Following baseline correction, it was possible to separate the P_i peaks arising from the three compartments with different pH values if each peak made up 10–35% of total P_i area. In vivo, we identified the plasma compartment by intraarterial infusion of P_i. It was assumed that intracellular compartments contained high-energy phosphates and took up glucose. Based on these assumptions we subjected the brains to complete ischemia and observed that P_i compartments at pH 6.82, 6.92, 7.03, and 7.13 increased markedly in amplitude. If the brain cells took up and phosphorylated 2-deoxyglucose (2-DG), 2-DG-6-phosphate (2-DG-6-P) would appear in the PME portion of the spectrum ionized according to pH_i. Four 2-DG-6-P peaks with calculated pH values of 6.86, 6.94, 7.04, and 7.15 did appear in the spectrum, thereby confirming that the four larger P_i peaks represented intracellular spaces.     

ATPase, GTPase, and RNA binding activities associated with the 206-kilodalton protein of turnip yellow mosaic virus.

The 206-kDa protein of turnip yellow mosaic virus belongs to an expanding group of proteins containing a domain which includes the consensus nucleotide binding site GxxxxGKS/T. A portion of this protein (amino acids [aa] 916 to 1259) was expressed in Escherichia coli and purified by affinity chromatography to near homogeneity. In the absence of any other viral factors, it exhibited ATPase and GTPase activities in vitro. A mutant protein with a single amino acid substitution in the consensus nucleotide binding site (Lys-982 to Ser) exhibited only low levels of both activities, implying that Lys-982 is important for nucleoside triphosphatase activity. The protein also possessed nonspecific RNA binding capacity. Deletion mutants revealed that an N-terminal domain (aa 916 to 1061) and a C-terminal domain (aa 1182 to 1259) participate in RNA binding. The results presented here provide the first experimental evidence that turnip yellow mosaic virus encodes nucleoside triphosphatase and RNA binding activities.     

The fate of immunoglobulin G fed to larvae of Ostrinia nubilalis

Since plants can be transformed genetically to produce functional antibodies, an immunological approach may be developed for controlling their arthropod pests. Specific antibodies would protect plants from arthropods if they could gain access to the pest antigen in sufficient amounts such that the normal function of the antigen is disrupted. In order to study the fate of ingested antibodies in the body of the European corn borer (ECB), Ostrinia nubilalis (Hübner), (Lepidoptera: Pyralidae), we fed the larvae on serum-containing diet. When larvae were fed on the serum-containing diet for various lengths of time between 12 and 96 h, no significant differences were noted in the immunoglobulin G (IgG) concentration in their body. Immediately after the larvae stopped feeding, the concentrations of the IgG in their midgut was about one half that of the diet itself, but it decreased significantly after 6 h and again after 18 h (about 3 and 10 fold, respectively). Immediately after the larvae stopped feeding, the concentration of the IgG in their hemolymph was about 1/500 that in the diet. The concentration of IgG in the hemolymph of ECB larvae was influenced directly by the titer of antibodies in their diet. During the first 6 h after the larvae stop feeding the concentration of IgG in their hemolymph did not decrease significantly; however, it did so after 18 h (about 6 fold). The possibility that specific antibodies will gain access to antigens in the ECB body is discussed.     

Effect of larval fish and nutrient enrichment on plankton dynamics in experimental ponds

The hypotheses that larval fish density may potentially affect phytoplankton abundance through regulating zooplankton community structure, and that fish effect may also depend on nutrient levels were tested experimentally in ponds with three densities of larval walleye, Stizostedion vitreum (0, 25, and 50 fish m^–3), and two fertilizer types (inorganic vs organic fertilizer). A significant negative relationship between larval fish density and large zooplankton abundance was observed despite fertilizer types. Larval walleye significantly reduced the abundances of Daphnia, Bosmina, and Diaptomus but enhanced the abundance of various rotifer species (Brachionus, Polyarthra, and Keratella). When fish predation was excluded, Daphnia became dominant, but Daphnia grazing did not significantly suppress blue-green algae. Clearly, larval fish can be an important regulator for zooplankton community. Algal composition and abundance were affected more by fertilizer type than by fish density. Inorganic fertilizer with a high N:P ratio (20:1) enhanced blue-green algal blooms, while organic fertilizer with a lower N:P ratio (10:1) suppressed the abundance of blue-green algae. This result may be attributed to the high density of blue-green algae at the beginning of the experiment and the fertilizer type. Our data suggest that continuous release of nutrients from suspended organic fertilizer at a low rate may discourage the development of blue-green algae. Nutrient inputs at a low N:P ratio do not necessarily result in the dominance of blue-green algae.     

Significant extension in northern Australia of the known geographic range of the Shield Shrimp Triops australiensis (Crustacea: Notostraca)

The program BIOCLIM predicts the total geographic distribution of species, based upon the biogeoclimatic characteristics common to the localities at which they are known to occur. Field studies in the Northern Territory have located the Shield Shrimp Triops australiensis at localities substantially north of its known and predicted geographic distribution.     

Meteorological factors and the time of onset of chest pain in acute myocardial infarction

Analysis of the time of onset of chest pain in 2254 patients with a myocardial infarction admitted to a coronary care unit in Leicester during a 10-year period shows an association with temperature and humidity. During both the most cold and humid times of the year, the relationship is a strong one. A generalized linear model with a log link was used to fit the data and the backward elimination selection procedure suggested a humid, cold day might help to trigger the occurrence of myocardial infarction. In addition, cold weather was found to have a stronger effect on the male population while those men aged between 50 and 70 years were more sensitive to the effect of high humidity.     

Growth and carbon utilization by sprouted propagules of two species of submersed rooted aquatic plants grown in darkness

Hydrilla verticillata (L. f.) Royle tubers from monoecious plants andPotamogeton gramineus L. winter buds were sprouted and allowed to grow in the dark for 120 days. We measured plant length and counted the number of leaves at 2–3 day intervals.Hydrilla grew most rapidly during the first 16–17 days andPotamogeton grew most rapidly during the first 16–25 days. Measurement of propagule carbon content over time indicated that cessation of rapid growth coincided with depletion of tuber carbon by one-half forHydrilla. ForPotamogeton, growth was reduced after 16 to 25 days while the winter bud C half-life was 37 days. Calculations indicated thatHydrilla mobilized 49% andPotamogeton 39% of the initial propagule carbon to support growth. In a second experiment, in which plants were grown in substrate the plants grew taller and produced slightly more leaves per plant.Potamogeton removed from darkness after specified time periods, and allowed to grow for 21 days in a greenhouse recovered from 20–30 days in the dark. Similarly treatedHydrilla plants recovered from up to 80 days in the dark.Potamogeton had mobilized 79% of initial C by the time it was unable to recover from the dark treatment. Combined results for both species indicate that the majority of propagule C was utilized in the first 16 to 30 days following sprouting. In conjunction with an understanding propagule sprouting requirements, this information will be useful in the timing of application for management techniques. The U.S. Government right to retain a non-exclusive, royalty-free licence in and to any copyright is acknowledged. The U.S. Government right to retain a non-exclusive, royalty-free licence in and to any copyright is acknowledged.