Improved high-affinity binding of 3H-apomorphine with a subcellular membrane fraction of mammalian brain tissue

The binding of low concentrations of [³H](?)apomorphine to preparations of calf and rat forebrain tissue was evaluated. Fractionation of crude homogenates to prepare a membrane fraction (P₄) of striatal or caudate homogenates increased the proportion of saturable to total binding from 33% to over 80%, and increased the apparent density of binding sites from 94 to 681 fmol/mg protein. Binding in calf caudate P₄ tissue was protein-dependent and optimal at pH = 7.0 to 7.5, and T = 20 to 25°C; at higher temperatures tissue binding sites appeared to degrade. The half-time of association and dissociation at 22°C were, respectively, 14.0 and 18.5 min; equilibration was complete in 60 min. Kinetic characteristics of high-affinity binding obtained from association and dissociation constants and from saturation isotherms were similar (K_d = 2.1 to 3.4 nM). The pharmacology of competition for ³H-APO suggests selectivity for dopamine-agonist interactions. These results indicate that the P₄ membrane preparation may be useful for the evaluation of dopamine-agonist binding sites or “receptors.”     

Developmental regulation of phenylalanine hydroxylase activity in Drosophila melanogaster

Herein we demonstrate that Drosophila larvae possess a synthetic activity capable of converting phenylalanine to tyrosine. This system is readily extractable and displays many characteristics of phenylalanine hydroxylase systems described in other organisms, the most notable being that a tetrahydropteridine is required for full expression of activity. The level of phenylalanine hydroxylase activity present in the organism varies with the stage of development: from an undetected level of activity at the first larval instar, there is a rapid increase in phenylalanine hydroxylase activity which reaches a peak at the time of puparium formation, after which there is a rapid decrease again to an undetected level.     

Patterns of protein synthesis following heat shock in pupae of Drosophila melanogaster

Pupae of Drosophila melanogaster were heat-shocked under conditions required to induce phenocopies in more than 90% of the flies that subsequently emerge. The effects of these treatments on protein synthesis in two tissues (thoracic epithelium and brain) were followed for several hours after the heat treatments. Results from pulse-labeling and protein separations on sodium dodecylsulfate (SDS) acrylamide gels showed a virtually complete cessation of protein synthesis immediately after the shock, followed by a noncoordinate resumption of the starting pattern. Similar experiments following double heat shocks demonstrated a more rapid resumption of synthesis of heat shock proteins after two successive heat treatments than after a single one.     

Specific protection from phenocopy induction by heat shock

Mild heat treatments applied to whole animals or cell cultures of Drosophila prior to lethal heat shocks result in increased survival and protection against phenocopy induction. The optimal condition for the preliminary mild heat treatment is that which induces the synthesis of heat-shock proteins but does not turn off the protein synthesis that is in progress. Recovery of protein synthesis but not RNA synthesis following a drastic heat shock is much enhanced by the pretreatments. The results suggest that the protection for survival and against phenocopy induction is due to storage of messenger RNA.     

A new method of in situ hybridization

A new method for gene mapping at the chromosome level using in situ hybridization and scanning electron microscopy is described and has been applied to mapping the rRNA genes of Drosophila melanogaster. Biotin is covalently attached to Drosophila rRNA via a cytochrome c bridge at a ratio of one cytochrome-biotin per 130 nucleotides by a chemical procedure. Polymethacrylate spheres with a diameter of ca. 60 nm are prepared by emulsion polymerization and are covalently attached to the protein avidin at a ratio of 5–20 avidins per sphere. The biotin-labeled rRNA is hybridized to denatured DNA in a chromosome squash. Upon incubation with a sphere solution, some of the biotin sites become labeled with spheres because of the strong non-covalent interaction between biotin and avidin. The chromosome squash is examined in the scanning electron microscope (SEM). Polymer spheres, which are visible in the SEM, are observed to label the nucleolus, where the rRNA genes are located.Contribution number 5121 from the Department of Chemistry.     

Massenentwicklungen von Algen und Wasserlinsen in der Talsperre Bleiloch (Thüringen) — gegenwärtige Situation,Ursachenanalyse und erste Ergebnisse zu deren Verringerung Algal Blooms and Duckweed Covers in the Bleiloch Reservoir (Thuringia) — Actually Situation,Causal Analysis and First Results of its Overcome

The effects of a decreased sewage release from a cellulose mill on the hypertrophic Bleiloch reservoir were studied. The waste water treatment reduced the concentration of yellow substances in the reservoir and resulted in a more light-coloured waterbody. It is estimated that the changes in the light attenuation in the Bleiloch reservoir after 1989 can be described as eutrophication. Selected physical, chemical, and biological parameters from 1994 show the effect of changes. Three large pelagic enclosures (diameter 10 m, length 20 m) were installed in the reservoir to assess effects of artifical mixing on phytoplankton biomass and composition. Two circulation techniques have been used in the enclosures (relase of compressed air at the bottom of the enclosure and mechanical mixing, where water was pumped from the surface downward). The investigation demonstrate that the magnitude of seasonal peaks of chlorophyll were depressed in the mixed enclosures during circulation. The results were discussed in comparison with a reference enclosure without mixing and the waterbody of the reservoir. Cryptophytes and other flagellates achieved dominance during mixing in the “air”-enclosure, while diatoms were the most abundant species in the „pump”︁-enclosure. The development of filamentous cyanobacteria, which are significant indicators of eutrophication, was disturbed in the enclosures. They were outcompeted by cryptophytes, movable chlorophytes and diatoms. Both mechanical pump and compressed air can be used to control excessive algal growth. Further studies on the dynamics of phytoplankton to artifical mixing are suggested. The results of further studies should design an in situ experiment in the reservoir to reduce algal growth by light limitation. Further on a dense duckweed cover spreads over the surface of the Bleiloch reservoir, which influences the light regime in the waterbody. Therefore factors influencing the duckweed growth were analysed. Special attention was focused on the influence of the pH, consequences of the mixing in the waterbody and the role of resting stages.     

Optimal deep brain stimulation of the subthalamic nucleus—a computational study

Deep brain stimulation (DBS) of the subthalamic nucleus, typically with periodic, high frequency pulse trains, has proven to be an effective treatment for the motor symptoms of Parkinson’s disease (PD). Here, we use a biophysically-based model of spiking cells in the basal ganglia (Terman et al., Journal of Neuroscience, 22, 2963–2976, 2002; Rubin and Terman, Journal of Computational Neuroscience, 16, 211–235, 2004) to provide computational evidence that alternative temporal patterns of DBS inputs might be equally effective as the standard high-frequency waveforms, but require lower amplitudes. Within this model, DBS performance is assessed in two ways. First, we determine the extent to which DBS causes Gpi (globus pallidus pars interna) synaptic outputs, which are burstlike and synchronized in the unstimulated Parkinsonian state, to cease their pathological modulation of simulated thalamocortical cells. Second, we evaluate how DBS affects the GPi cells’ auto- and cross-correlograms. In both cases, a nonlinear closed-loop learning algorithm identifies effective DBS inputs that are optimized to have minimal strength. The network dynamics that result differ from the regular, entrained firing which some previous studies have associated with conventional high-frequency DBS. This type of optimized solution is also found with heterogeneity in both the intrinsic network dynamics and the strength of DBS inputs received at various cells. Such alternative DBS inputs could potentially be identified, guided by the model-free learning algorithm, in experimental or eventual clinical settings. Action Editor: Steven J. Schiff Xiao-Jiang Feng and Eric Shea-Brown contributed equally to this work.