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Biomedical subjects

L I Gilbert

Publications and source records attributed to L I Gilbert.

157 records · Page 9Linked to original sources
PubMed1965

Regulation of cockroach fat-body metabolism by the corpus cardiacum in vitro.

Incubation in vitro of young, adult, male Leucophaea maderae fat body with extracts of corpora cardiaca or intact corpora cardiaca results in stimulation of oxygen consumption but reduction in carbon dioxide evolved from carbohydrate. The carbohydrate is preferentially used for trehalose synthesis, and the endogenous metabolism of the fat body appears to be supported by increased lipid utilization. A hormone from the corpus cardiacum is most likely responsible for these effects and may act at two points, at least, in the glycolytic pathway.

Adipose Tissue↗
PubMedDate not supplied

Control of ecdysteroidogenesis: activation and inhibition of prothoracic gland activity.

The ecdysteroid hormones, mainly 20-hydroxyecdysone (20E), play a pivotal role in insect development by controlling gene expression involved in molting and metamorphosis. In the model insect Manduca sexta the production of ecdysteroids by the prothoracic gland is acutely controlled by a brain neurohormone, prothoracicotropic hormone (PTTH). PTTH initiates a cascade of events that progresses from the influx of Ca2+ and cAMP generation through phosphorylation of the ribosomal protein S6 and S6-dependent protein synthesis, and concludes with an increase in the synthesis and export of ecdysteroids from the gland. Recent studies indicate that S6 phosphorylation probably controls the steroidogenic effect of PTTH by gating the translation of selected mRNAs whose protein products are required for increased ecdysteroid synthesis. Inhibition of S6 phosphorylation prevents an increase in PTTH-stimulated protein synthesis and subsequent ecdysteroid synthesis. Two of the proteins whose translations are specifically stimulated by PTTH have been identified, one being a beta tubulin and the other a heat shock protein 70 family member. Current data suggest that these two proteins could be involved in supporting microtubule-dependent protein synthesis and ecdysone receptor assembly and/or function. Recent data also indicate that the 20E produced by the prothoracic gland feeds back upon the gland by increasing expression and phosphorylation of a specific USP isoform that is a constituent of the functional ecdysone receptor. Changes in the concentration and composition of the ecdysone receptor complex of the prothoracic gland could modulate the gland's potential for ecdysteroid synthesis (e.g. feedback inhibition) by controlling the levels of enzymes or other proteins in the ecdysteroid biosynthetic pathway.

Amino Acid Sequence↗
PubMedDate not supplied

Insect prothoracic glands: a role for cyclic AMP in the stimulation of alpha-ecdysone secretion.

The prothoracic glands of the tobacco hornworm, Manduca sexta, were studied to determine if cyclic AMP is involved in the regulation of alpha-ecdysone secretion. Culturing glands in the presence of the phosphodiesterase inhibitors, aminophylline and 1-methyl-3-isobutylxanthine, caused a greater than 2-fold stimulation of ecdysone secretion while cyclic AMP alone was ineffective. Based on a dose-response analysis, 1-methyl-3-isobutylxanthine was 200 times more potent than aminophylline. Measurements of endogenous prothoracic gland cyclic AMP during the fifth larval instar demonstrated that dramatically increased levels preceded the increase in in vitro ecdysone-secretory ability. The data suggest that cyclic AMP may act as a second messenger in the stimulation of prothoracic gland alpha-ecdysone secretion by the prothoracicotropic brain hormone.

Aminophylline↗
PubMedDate not supplied

Purification and characterization of a juvenile hormone binding protein from the hemolymph of the fourth instar tobacco hornworm, Manduca sexta.

A protein which binds the insect juvenile hormone has been isolated from the hemolymph of the fourth instar tobacco hornworm, Manduca sexta (Lepidoptera). Bioassay and chemical characterization of the bound ligand from the purified binding protein indicates that this molecule is the primary macromolecule responsible for juvenile hormone transport in the hemolymph of this insect. The juvenile hormone binding protein has been purified using gel filtration, ion exchange chromatography and preparative polyacrylamide gel electrophoresis. The protein is a single polypeptide chain of about 28,000 daltons with a sedimentation coefficient of 2.2S and an isoelectric point of 5.0. Binding analysis using a hydroxyapatite batch assay indicates that the juvenile hormone binding protein has one binding site with a Ka of 1.2 times 10(7) M-1 at 4 degrees C.

Animals↗
PubMedDate not supplied

Juvenile hormone synthesis in vitro by larval and pupal corpora allata of Manduca sexta.

The synthesis of juvenile hormone I (JH I) and juvenile hormone III (JH III) in vitro by isolated corpora allata (CA) and brain-corpora cardiaca-corpora allata complexes (BR-CC-CA) from fourth and fifth instar larvae and pupae of Manduca sexta was assessed by JH I and JH III radioimmunoassays of incubation medium. Fluctuations in synthesis occurred at specific stages during larval-pupal development. These fluctuations temporally approximated those of previously determined hemolymph titers of JH I and III for this developmental period, except for the synthesis of JH III during the prepupal period, when the JH III hemolymph titer was negligible but JH III synthesis in vitro increased to the highest rate observed. The overall agreement between CA activity and the JH hemolymph titers suggests that the in vitro system supports physiological gland activity and that changes in biosynthesis contribute significantly to changes in the titers. A comparison of JH I and JH III synthesis by isolated CA with that by BR-CC-CA suggests that the BR-CC can differentially modulate the synthesis of these two homologs. The significance of these findings with regard to the possible endocrine function of JH III and the proposed neuroendocrine control of the CA is discussed.

Animals↗
PubMedDate not supplied

Metabolism in vitro of cholesterol and 25-hydroxycholesterol by the larval prothoracic glands of Manduca sexta.

The prothoracic glands in vitro convert 25-hydroxycholesterol (25C) to 25-hydroxy-7-dehydrocholesterol (7d25C) and to ecdysteroids at a greater rate than cholesterol (C) is converted to ecdysteroids via 7-dehydrocholesterol (7dC). Mediated via a cytochrome P450 most probably located in the endoplasmic reticulum (ER), both intact and extensively homogenized prothoracic glands, as well as crude subcellular fractions, were able to 7,8-dehydrogenate 25C to 7d25C eight-fold more efficiently than they could convert C to 7dC. However, less than a two-fold difference was observed in the subsequent monooxygenase mediated conversion of these two intermediates formed in situ into ecdysteroids, mainly ecdysone (E) and 2-deoxyecdysone (2dE) and/or their 3-dehydroderivatives. When 7dC, and particularly 7d25C, were made directly available to these tissue preparations, their conversion to ecdysteroids greatly exceeded that of the in situ conversion of either C or 25C, via 7dC or 7d25C, respectively. Indeed, there was an eight-fold increase in the VMAX for 25C dehydrogenation by homogenized glands relative to the dehydrogenation of C. Most important, however, was the 1000-fold increase in the VMAX observed for the direct production of E from emulsified 7d25C by gland homogenates relative to E production from 25C via 7d25C synthesized in situ. Thus, it is apparent that even after the rapid and efficient conversion of 25C to 7d25C within the ER, the subsequent rate of conversion of this intermediate to E is greatly retarded relative to that observed following the direct incubation of emulsified 7d25C with gland homogenates. These differential kinetics of direct and indirect 7d25C incorporation into E are interpreted as evidence for the existence of a barrier to the efficient translocation of the delta 5,7-sterol intermediates from the ER to another site where the subsequent, uncharacterized initial conversions leading to ecdysteroids take place. On the basis of studies on mammalian adrenal cortical steroidogenesis, this site is postulated to be the inner membrane/matrix of the mitochondria. The present data support the hypothesis that the translocation of both 7dC and 7d25C, first from the site of their probable synthesis within the ER membranes, next through the cytosol to the outer mitochondrial membrane, and then across the intramitochondrial aqueous space to the inner membrane/matrix compartment, may be analogous to the translocation in the adrenal cortex of ER-derived C, first to the plasma membrane and/or to the outer mitochondrial membrane and then to the inner mitochondrial membrane/matrix for P450scc-mediated conversion into pregnenolone.

Animals↗
PubMedDate not supplied

Differential incorporation of cholesterol and cholesterol derivatives into ecdysteroids by the larval ring glands and adult ovaries of Drosophila melanogaster: a putative explanation for the l(3)ecd1 mutation.

Studies in vitro revealed that intact ring glands of Drosophila melanogaster convert tritiated cholesterol (C) and 25-hydroxycholesterol (25C) via 7-dehydrocholesterol (7dC) and 7-dehydro-25-hydroxycholesterol (7d25C), respectively, to ecdysone (E) and 2-deoxyecdysone (2dE), while both intact and homogenized ovaries synthesize only 2dE from these precursors. Emulsified 7d25C was incorporated directly into ecdysteroids by these tissue preparations at a much greater rate than was 7d25C made in situ from 25C. To probe the basis of the biochemical defect in the ecdysteroid deficient conditional mutant ecdysoneless (ecd1), the differential incorporation into ecdysteroids of C (via 7dC), and particularly of 25C (via 7d25C), was measured relative to that observed after the incubation of 7d25C directly with both wild type and mutant tissues in vitro at 30 degrees C, the restrictive temperature. Both C and 25C were equally 7,8-dehydrogenated in situ to 7dC or 7d25C, respectively, by both wild type and mutant tissues at 30 degrees C. However, the rate of subsequent conversion of either of these delta 5,7-sterol intermediates synthesized in situ to ecdysteroids was reduced an average of 50% in the mutant tissues relative to the wild type. Yet, when emulsified 7d25C was incubated directly with either the wild type or mutant tissues at the restrictive temperature, the amplified rate of conversion of the freely available 7d25C to ecdysteroid by these tissues was identical. These data suggest that the defect in ecd1 tissue-mediated ecdysteroidogenesis does not involve a "hit" on any of the enzymes involved in either the 7,8-dehydrogenation of C or 25C or in the subsequent oxidation of 7d25C or 7dC to ecdysteroid. Rather, the mutation appears to affect the expression of a gene governing the translocation of delta 5,7-sterol intermediates from the subcellular compartment where they are synthesized and/or stored to the site of subsequent oxidation to ecdysteroid.

Animals↗
PubMedDate not supplied

Alterations in ecdysone content during the post-embryonic development of Chironomus thummi: correlations with chromosomal puffing.

The ecdysone titer of larvae and pupae of Chironomus thummi was determined by radioimmune assay (RIA) and revealed a concentration of about 150 ng/g fresh weigth in late third instar larvae and a peak of more than 450 ng/g fresh weight just preceding pupation. The ecdysone titer curve shows a high degree of correlation with previously observed puffing activity at the ecdysone sensitive chromosomal site IIIdl. Further, beta-ecdysone is the only endogenous ecdysone detected by high pressure liquid chromatography during larval-pupal development. It was also observed that developmentally arrested Chironomus contain less ecdysone than "normal" larvae.

Animals↗