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  • 1
    ISSN: 1432-0878
    Schlagwort(e): Ecdysteroid receptor ; Cytochemistry ; Central nervous system ; Prothoracic gland ; Manduca sexta (Insecta)
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Medizin
    Notizen: Summary Brains and subesophageal ganglia from day 3.5 fifth stadium larvae of Manduca sexta were incubated in vitro with 4 nM tritiated ponasterone A, a 20-hydroxyecdysone analog, to determine whether uptake and specific binding of ecdysteroids occur at a cellular level. These tissues, which were taken just prior to the commitment peak in the hemolymph-ecdysteroid titer, showed saturable uptake of 3H-ponasterone A after 40–60 min of incubation. Uptake was blocked by the addition of 400 nM unlabelled ponasterone A, or of 500 nM or 1000 nM 20-hydroxyecdysone. RH 5849, a synthetic 20-hydroxyecdysone agonist with a long half-life, for which ecdysteroid receptors have low affinity, also reduced ponasterone A uptake at a concentration of 10 μM. Autoradiographs of 4 μm sections of brains revealed distinct nuclear concentrations of silver grains over cell populations in the pars intercerebralis, pars lateralis, and ventral tritocerebrum. Nuclear labelling was also found in many small cells around the mushroom bodies and the neuropil, and between the inner and outer larval optic lobes. Nuclear labelling of cells in the subesophageal ganglion was observed in the fronto-medial and lateral regions, in small cells around the neuropil, and caudally in a few large neurons. In addition to cells with nuclear labelling, both brains and ganglia at this development stage contained cells with exclusively cytoplasmic or both nuclear and cytoplasmic labelling. None of these apparent binding sites were observed in the competition experiments, suggesting that the binding is specific.
    Materialart: Digitale Medien
    Bibliothek Standort Signatur Band/Heft/Jahr Verfügbarkeit
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  • 2
    Digitale Medien
    Digitale Medien
    New York, NY [u.a.] : Wiley-Blackwell
    Archives of Insect Biochemistry and Physiology 30 (1995), S. 149-164 
    ISSN: 0739-4462
    Schlagwort(e): corpora allata ; adenylyl cyclase ; CAMP ; calcium ; calmodulin ; biogenic amines ; dopamine ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Quelle: Wiley InterScience Backfile Collection 1832-2000
    Thema: Biologie
    Notizen: An assay was developed with which to study basic characteristics of an adenylyl cyclase in the corpora allata (CA) of the tobacco hornworm, Manduca sexta. The assay used glands collected and frozen at -80°C, to circumvent the problem of tissue availability. With this protocol for storage of tissue, less than 25% of the enzyme activity in fresh tissue was lost. Substances such as sodium fluoride (NaF) and Gpp(NH)p (a non-hydrolyzable GTP analog), which typically stimulate the adenylyl cyclases in other insect tissues, increased enzyme activity several-fold. There was a progressive decrease in the capacity of the CA adenylyl cyclase to be stimulated by NaF during the fifth stadium, suggesting a possible developmental change in the capacity of the associated G protein to be stimulated by NaF. The calcium/calmodulin (CaM) dependence of adenylyl cyclase activity was also investigated. The results demonstrated that addition of up to 10-4 M calcium to assays of enzyme activity in whole gland homogenates of both larval (day O) and prepupal (day 6) CA resulted in only a slight increase in the activity of the enzyme over basal rates in the presence of the calcium chelator EGTA. However, addition of as little as 5 m̈M CaM in the presence of 10-4 to 10-3 M calcium increased adenylyl cyclase activity three to five-fold. A similar stimulation was obtained with washed membrane preparations of day 0 and day 6 glands, but required a substantially higher concentration of CaM. Results demonstrated that the CA possess a calcium/CaM-dependent adenylyl cyclase from day 0 through day 6. A preliminary investigation of the effect of two biogenic amines on the CA adenylyl cyclase revealed that enzyme activity was not affected by octopamine, but a stage-specific effect was obtained with dopamine. Concentrations of 10-6 and 10-7 M stimulated enzyme activity in hornogenates of day 0 glands but inhibited activity in homogenates of day 6 CA. © 1995 Wiley-Liss, Inc.
    Zusätzliches Material: 8 Ill.
    Materialart: Digitale Medien
    Bibliothek Standort Signatur Band/Heft/Jahr Verfügbarkeit
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