Franck Couillaud

Anti-juvenile effect of neuroparsin A, a neuroprotein isolated from locust corpora cardiaca

Abstract Neuroparsin A, a sulfur-containing protein synthesized by the medial part of the brain of Locusta and transported to the corpora cardiaca (CC) via the nervi corporis cardiaci I (Girardie et al. , 1987), was satisfactorily isolated using electro-elution. A specific immune serum against electro-eluted neuroparsin A was generated. On serial histological sections of the brain treated with the immune serum, only the median neurosecretory cells [stained in blue following the double staining Victoria blue-paraldehyde fuchsin (Al type)] were revealed using peroxidase-antiperoxidase technique. Inactivation of neuroparsin A by antigen-antibody complex formation following injections of immune serum induced green pigmentation, intermediary forms and precocious sexual maturation. These symptoms also follow juvenile hormone (JH) injections. Injections of immune serum antineuroparsin A or the electro-eluted neuroparsin A produced opposite effects on oocyte growth but had no effect on the rate of JH biosynthesis evaluated by radiochemical assay. The neurohormone neuroparsin A could be the median humoral inhibiting factor of the JH system which was previously demonstrated (Girardie, 1966, 1967) in the central area of the pars intercerebralis.

Immunological evidence for an allatostatin-like neuropeptide in the central nervous system of Schistocerca gregaria, Locusta migratoria and Neobellieria bullata.

Methanolic brain extracts of Locusta migratoria inhibit in vitro juvenile hormone biosynthesis in both the locust L. migratoria and the cockroach Diploptera punctata. A polyclonal antibody against allatostatin-5 (AST-5) (dipstatin-2) of this cockroach was used to immunolocalize allatostatin-5-like peptides in the central nervous system of the locusts Schistocerca gregaria and L. migratoria and of the fleshfly Neobellieria bullata. In both locust species, immunoreactivity was found in many cells and axons of the brain-retrocerebral complex, the thoracic and the abdominal ganglia. Strongly immunoreactive cells were stained in the pars lateralis of the brain with axons (NCC II and NCA I) extending to and arborizing in the corpus cardiacum and the corpora allata. Although many neurosecretory cells of the pars intercerebralis project into the corpus cardiacum, only 12 of them were immunoreactive and the nervi corporis cardiaci I (NCC I) and fibers in the nervi corporis allati II (NCA II) connecting the corpora allata to the suboesophageal ganglion remained unstained. S. gregaria and L. migratoria seem to have an allatostatin-like neuropeptide present in axons of the NCC II and the NCA I leading to the corpus cardiacum and the corpora allata. All these data suggest that in locusts allatostatin-like neuropeptides might be involved in controlling the production of juvenile hormone by the corpora allata and, perhaps, some aspects of the functioning of the corpus cardiacum as well. However, when tested in a L. migratoria in-vitro juvenile hormone-biosynthesis assay, allatostatin-5 did not yield an inhibitory or stimulatory effect. There is abundant AST-5 immunoreactivity in cell bodies of the fleshfly N. bullata, but none in the CA-CC complexes. Apparently, factors that are immunologically related to AST-5 do occur in locusts and fleshflies but, the active protion of the peptide required to inhibit JH biosynthesis in locusts is probably different from that of AST-5.

Image-guided, noninvasive, spatiotemporal control of gene expression

Spatiotemporal control of transgene expression is of paramount importance in gene therapy. Here, we demonstrate the use of magnetic resonance temperature imaging (MRI)-guided, high-intensity focused ultrasound (HIFU) in combination with a heat-inducible promoter [heat shock protein 70 (HSP70)] for the in vivo spatiotemporal control of transgene activation. Local gene activation induced by moderate hyperthermia in a transgenic mouse expressing luciferase under the control of the HSP70 promoter showed a high similarity between the local temperature distribution in vivo and the region emitting light. Modulation of gene expression is possible by changing temperature, duration, and location of regional heating. Mild heating protocols (2 min at 43°C) causing no tissue damage were sufficient for significant gene activation. The HSP70 promoter was shown to be induced by the local temperature increase and not by the mechanical effects of ultrasound. Therefore, the combination of MRI-guided HIFU heating and transgenes under control of heat-inducible HSP promoter provides a direct, noninvasive, spatial control of gene expression via local hyperthermia.

Activity of disconnected corpora allata in Locusta migratoria: Juvenile hormone biosynthesis in vitro and physiological effects in vivo

Abstract Severance of nervi corporis allati I (NCA I) in day-1 adult female Locusta migratoria resulted in a significant decrease and a loss of the characteristic pattern of juvenile hormone biosynthesis by the corpora allata as determined by radiochemical assay. This decrease in the rate of juvenile hormone biosynthesis was not reflected in basal oocyte growth. The lengths of the oocytes were the same in NCA-transectioned and in the sham-operated females. The effect of severance of both NCA I and NCA II on juvenile hormone biosynthesis and ovarian maturation was similar to the effect of NCA I severance only. Rate of juvenile hormone biosynthesis by corpora allata of fourth-instar larvae exhibited a maximum of activity in the middle of the stadium. The severance of NCA I early in the stadium resulted in a very low rate of juvenile hormone biosynthesis and a disappearance of this peak. In NCA I-transectioned larvae, the duration of the stadium was significantly increased although larvae moulted into normal fifth instar.

A cDNA, from Agrotis ipsilon, that encodes the pheromone biosynthesis activating neuropeptide (PBAN) and other FXPRL peptides.

A cDNA encoding the prohormone of the pheromone biosynthesis activating neuropeptide (PBAN) in the moth Agrotis ipsilon was isolated. The cDNA contains 834 nucleotides, coding for a 193-amino acid protein that exhibits 89% identity with PBAN prohormones of other moths. The prohormone contains five potential peptides belonging to the FXPRL family. The peptide corresponding to the Bombyx mori diapause hormone exhibits an extra residue, and the C-terminal leucine is replaced by an isoleucine, introducing a new type of variability in this family of peptides. Northern blot analysis revealed expression in suboesophagal ganglion complexes. Constitutive heterologous expression of Agi-PBAN cDNA in yeast, using three different antibodies, did not produce PBAN-immunoreactive material.

Gene expression and gene therapy imaging

The fast growing field of molecular imaging has achieved major advances in imaging gene expression, an important element of gene therapy. Gene expression imaging is based on specific probes or contrast agents that allow either direct or indirect spatio-temporal evaluation of gene expression. Direct evaluation is possible with, for example, contrast agents that bind directly to a specific target (e.g., receptor). Indirect evaluation may be achieved by using specific substrate probes for a target enzyme. The use of marker genes, also called reporter genes, is an essential element of MI approaches for gene expression in gene therapy. The marker gene may not have a therapeutic role itself, but by coupling the marker gene to a therapeutic gene, expression of the marker gene reports on the expression of the therapeutic gene. Nuclear medicine and optical approaches are highly sensitive (detection of probes in the picomolar range), whereas MRI and ultrasound imaging are less sensitive and require amplification techniques and/or accumulation of contrast agents in enlarged contrast particles. Recently developed MI techniques are particularly relevant for gene therapy. Amongst these are the possibility to track gene therapy vectors such as stem cells, and the techniques that allow spatiotemporal control of gene expression by non-invasive heating (with MRI guided focused ultrasound) and the use of temperature sensitive promoters.

Role of juvenile hormone biosynthesis in dominance status and reproduction of the bumblebee, Bombus terrestris

SummaryIn Bombus terrestris females, dominant status is correlated with high levels of juvenile hormone (JH) biosynthesis and rapid oöcyte maturation. Queenright workers, which are inhibited by the dominant queen, complete the cycle of oöcyte maturation while exhibiting a continuous low rate of JH production, but their egglaying is inhibited. Measurements in foundress queens suggest that the low JH titer during oögenesis is probably not responsible for the inhibition of egg-laying. Queenless workers, kept individually, are not activated either for JH production or oöcyte maturation. In groups of three queenless workers, a dominance order becomes established and high rates of JH synthesis are observed in the dominant egg-laying workers, with low rates in subordinated workers. In groups of founder queens, also, a dominance order becomes established and results in a reduced rate of JH production in subordinated females.

Inhibition of male corpora allata activity and sexual pheromone responsiveness in the black cutworm, Agrotis ipsilon, by the hypocholesterolemic agent, fluvastatin

Male sexual behavior of the black cutworm Agrotis ipsilon is controlled by corpora allata. Allatectomies performed on day-1 and day-3 males inhibited the typical sexual behavior in day-4 males when exposed to female pheromone in a wind tunnel. Both JH-III and JH-III acid were able to restore male sexual behavior, suggesting that corpora allata act through their endocrine activity. We demonstrated that corpora allata incubated in vitro produced only the acid form of JH-III and JH-II. Fluvastatin, an HMG-CoA reductase inhibitor, inhibited JH acid biosynthesis by the corpora allata when fluvastatin was added to incubation medium or injected in males 4 h before the bioassay. However, endocrine activity of CA resumed 5–6 h after injection, indicating that the effect of fluvastatin was temporary. Fluvastatin injection also induced temporary inhibition of male responsiveness.

Control of C-16 juvenile hormone biosynthesis in active corpora allata of female African locusts

Summary Corpora allata from 8-day-old female Locusta migratoria, during the phase of yolk deposition, exhibit high rates of C-16 juvenile hormone (JH) biosynthesis. The effect of different potential factors which may be involved in the regulation of corpora allata activity is reported. The biosynthetic activity of corpora allata was determined by radiochemical assay. In maturing females, no changes in corpora allata activity are detected during one daily cycle. Starvation reduces JH biosynthesis only 3 days after the beginning of the food deprivation. Suppression of the median neurosecretory material by electrocoagulation of the internal cardiaca tract (TCC-I) does not disturb JH biosynthesis whereas the transection of the allata I nerve fibres (NCA-I) or the electrocoagulation of the lateral neurosecretory pericarya results in a rapid decline of JH biosynthesis. These data indicate that the median and lateral allatotropins are different, and that only the lateral neurosecretory material exerts an allatos...

Regulation of juvenile hormone titer in African locust

In the African locust, the titer of C-16 JH in hemolymph (determined by GC-MS), reflects the rate of hormone biosynthesis (determined by RCA) in normal adult females. Severance of the nervi corporis allati-I (NCA-I) results in a low C-16 JH biosynthesis without affecting physiological events dependent on JH. In NCA-I-transectioned animals, JH titer is higher than in control locusts. JH catabolism does not seem to be involved in this high titer of hormone associated with a very low rate of JH production. In sham-operated females, the bulk of injected [3H]C-16 JH quickly disappeared from the hemolymph but JH was retained in the bloodless body. After severance of the NCA-I, the remaining radiolabeled JH in the hemolymph increased. These results suggest the role of tissue JH-binding or of JH excretion in regulating its level in the locust.