Corpus Cardiacum-corpus Allatum Complex Research Articles

MicroRNA let-7 is required for hormonal regulation of metamorphosis in the silkworm, Bombyx mori

The heterochronic microRNA let-7, which was first identified in Caenorhabditis elegans, controls the timing of developmental programs, and let-7 triggers the onset of the juvenile-adult transition in bilaterians. The expression of let-7 is strongly induced during the last larval stage of C. elegans and is highly expressed in the late last instar larvae/nymphs of the fly Drosophila melanogaster and the cockroach Blattella germanica. In the silkworm Bombyx mori, the expression of let-7 remarkably increases in the corpus cardiacum-corpus allatum complex (CC-CA) at the beginning of the last larval instar and is maintained at high levels during this instar. To determine the biological function of let-7 in B. mori, we generated a let-7 knockout line and a transgenic UAS-let-7 line. The let-7 knockout larvae were developmentally arrested in the prepupal stage and became pupal-adult intermediates after apolysis. When let-7 was ubiquitously overexpressed under the transcriptional control of an Actin3-GAL4 driver, developmental timing and growth of larvae were severely impaired in the penultimate (L4) instar, and these larvae underwent precocious metamorphosis from L4. Furthermore, our results showed that reception and signaling of ecdysteroids and juvenile hormones (JHs) normally occurred in the absence of let-7, whereas the biosynthesis of ecdysone and JHs were affected by disruption and overexpression of let-7. Together, the present study demonstrates that let-7 is required for the coordination of the biosynthesis of ecdysone and JH to ensure the developmental transition during the metamorphosis of B. mori.

Photoperiodic Response in the Pars Intercerebralis Neurons, Including Plast-MIP Neurons, in the Brown-Winged Green Bug, Plautia stali.

Many insects in temperate regions avoid environmental adversity for reproduction, and thus enter reproductive diapause according to photoperiod. This reproductive diapause is induced by inhibition of juvenile hormone biosynthesis in the corpus allatum. Some neuropeptides that have an effect on juvenile hormone biosynthesis have been detected in insect brains. Thus, the reproductive diapause may be photoperiodically regulated by these juvenile hormones-controlling neuropeptides. However, there is limited understanding of how the neurons expressing these neuropeptides respond to the photoperiod and control the peptide release accordingly. Here, we performed electrophysiological analyses in the pars intercerebralis (PI) of Plautia stali, where juvenile hormone inhibitory neuropeptides, Plautia stali myoinhibitory peptides (Plast-MIPs) are expressed. We found that the large neurons in the PI showed very high firing activity under diapause-inducing short day conditions. Neurotracer staining revealed that all recorded neurons projected to the nervus corporis cardiaci 1, which is known to be connected to the corpus cardiacum-corpus allatum complex. Finally, we determined how many of the large PI cells expressed Plast-MIP by single cell reverse transcription PCR. About half of large PI neurons coexpressed Plast-Mip and other neuropeptides, Diuretic hormone 44 and insulin-like peptide 1. The remaining cells only expressed Diuretic hormone 44 and insulin-like peptide 1. The present results suggested that large PI neurons, including Plast-MIP neurons, have enhanced activity under short day conditions, which may increase Plast-MIP release to the corpus cardiacum-corpus allatum complex and thus contribute to reproductive diapause.

Evidence for a new juvenile hormone in a stink bug, Plautia stali

The biosynthetic activity of the corpus cardiacum-corpus allatum (CC-CA) complexes taken from adults of a stink bug, Plautia stali, was stimulated by adding precursors of JH III, farnesoic acid or farnesol, to the incubation medium. This stimulation was mainly attributed to an increase of a product with an Rf value of 0.5 on thin layer chromatography (TLC) sheets. A comparison of the TLC profile of the CA products of P. stali with that of M. domestica indicated that the major fraction of CA products in the bug was neither JH III nor JHB 3. A bioassay system was established using last instar nymphs of P. stali to test the JH activity of the CA products: last instar nymphs implanted with CC-CA complexes taken from reproductively active adults or topically applied with synthetic JHs or JH analogues were reared until the following moult and their forewing and scutellum lengths relative to pronotum width were recorded as well as the number of antennal segments. After CA implantations, the tested bugs had reduced forewing and scutellum lengths and had the 4-segmented antennae characteristic of nymphs, instead of the 5-segmented, adult type antennae. JH applications caused similar juvenilizing effects in a dose dependent manner. This assay system revealed that the JH activity of the CA products was limited to the TLC fraction at an Rf value of 0.5 where the major CA product released in the presence of farnesoic acid or farnesol was found. From these results, it was suggested that the JH produced by P. stali is different from any known JHs.

Allatostatin-immunoreactive neurons projecting to the corpora allata of adult Diploptera punctata.

A monoclonal antibody against allatostatin I was used to demonstrate the allatostatin-immunoreactive pathways between the brain and the corpus cardiacum-corpus allatum complex in the adult cockroach Diploptera punctata. The antibody was two to three orders of magnitude more sensitive to allatostatin I than to the other four known members of the allatostatin family. Whole and sectioned brains in which immunoreactivity was localized with horseradish peroxidase-H2O2-diaminobenzidine reaction showed strongly immunoreactive cells in the pars lateralis of the brain with axons leading to and arborizing in the corpus cardiacum and the corpus allatum. Although many neurosecretory cells of the pars intercerebralis project to the corpora allata only, four strongly immunoreactive cells were evident here (two pairs on either side), and these did not project to the corpus cardiacum and corpus allatum but rather terminated within the protocerebrum in areas in which lateral cells also formed arborizations. Immunoreactivity was found in many other cells in the brain, especially in the tritocerebrum.

Immunocytochemical localization and immunochemical characterization of an insulin-related peptide in the insect Leucophaea maderae

Immunocytochemical tests with eight monoclonal antibodies against either bovine or human insulin and seven polyclonal antibodies against bovine insulin were carried out to determine the presence of insulin-like neuropeptides in the brain and affiliated neuroendocrine structures of the insect Leucophaea maderae. Reaction products identified in the brain, subesophageal ganglion, and corpus cardiacum-corpus allatum complex indicate the presence of materials resembling mammalian insulins in its antigenic properties. The immunostaining observed with monoclonal antibodies appears to indicate the occurrence of an insulin-related peptide that shows sequential similarities with parts of both the A- and B-chains of mammalian insulin molecules. These suppositions are supported by the results of dot-blot and two-site time-resolved immunofluorometric assay (TRI-IFMA) screenings of fractions of Leucophaea tissue extracts obtained by chromatography. The polyclonal antibodies yielded reaction products in some of the same areas and in additional parts of the neuroendocrine system not visualized by the monoclonal antibodies. Immunoreaction was observed in the following areas: the pars intercerebralis of the protocerebrum, the nervi corporis cardiaci I transporting insulin-like material to the corpus cardiacum, the dorsolateral protocerebral area and the optic lobes, the deutocerebrum, the tritocerebrum, and the subesophageal ganglion. In addition, smaller cell bodies with immunoreactive deposits occur at the border between proto- and deuto-cerebrum, and in the central area of the protocerebrum. The distribution of reactive material in the corpus cardiacum-corpus allatum complex after use of both groups of antibodies was the same.(ABSTRACT TRUNCATED AT 250 WORDS)

Hormonal regulation of carbohydrate metabolism in the blowfly Phormia terraenovae

The corpus cardiacum-corpus allatum complex of the blowfly, Phormia terraenovae, contains a factor causing activation of glycogen phosphorylase and an increase of the total carbohydrate concentration in the haemolymph. Trehalose which contributes about 90% of the blowfly's normal blood sugar is preferentially elevated upon gland injection. Hypertrehalosaemia is also induced when blowfly gland extract is injected into the American cockroach, Periplaneta americana, as a test insect. Separation of a methanolic corpus cardiacum extract from blowflies on reversed-phase high-performance liquid chromatography results in only one distinct absorbance peak having a fair hypertrehalosaemic activity in cockroaches and activating blowfly abdominal glycogen phosphorylase. The material elutes slightly later than the hypertrehalosaemic hormone II (Pea-CAH-II) of P. americana. There is no evidence for a second hypertrehalosaemic factor or for antagonistically acting molecules. Stress induced by shaking blowflies at various frequencies results in an activation of glycogen phosphorylase and blood carbohydrate elevation; both actions are unaffected by the age of the experimental insects (tested up to 7 days after imaginal ecdysis). Flight (1 h) causes a 50% decrease in the glycogen content of the blowfly and a 2-fold increase in haemolymph carbohydrate levels. Release of the hypertrehalosaemic material of P. terraenovae is also shown by collecting and purifying blood from stressed blowflies, and injecting the peptide fraction into test cockroaches, which then unequivocally show a hypertrehalosaemic response.

Inactivation of the Corpora Allata in the Final Instar of the Tobacco Hornworm,Manduca sexta, Requires Integrity of Certain Neural Pathways from the Brain

Neither the implantation of active CA nor treatment with O-ethyl,S-phenylphosphoramidothiolate (EPPAT), a potent inhibitor of the juvenile hormone esterase (JHE), prevented metamorphosis of final instar tobacco hornworms. However, a combination of the two treatments often blocked metamorphosis and caused the formation of supernumerary larvae or larval-pupal intermediates. So also, in conjunction with EPPAT treatment, unilateral severance of the medial nerve from the brain to the corpus cardiacum-corpus allatum complex often resulted in abnormal supernumerary or intermediate larval forms. Thus, clearance of JH from mature hornworm larvae prior to metamorphosis appears normally to depend on two mechanisms: (1) cessation of JH production by inhibition of the CA via the innervation of these glands, and (2) destruction of previously secreted existing JH via production of JHE. In the present experiments, each of these mechanisms appeared fully able to clear JH sufficiently to permit normal metamorphosis, because only simultaneous interruption of both mechanisms led to formation of supernumerary larvae. Acting in concert as they do late in larval life, these two mechanisms ensure the timely and thorough clearance of JH in preparation for metamorphosis.

Juvenile hormone biosynthesis by corpus cardiacum-corpus allatum complexes of larval Lymantria dispar

1. 1. A radiochemical assay was used to examine juvenile hormone (JH) synthesis and secretion in vitro by incubating two pairs of larval corpus cardiacum-corpus allatum complexes (CC-CA) from, Lymantria dispar, in 50 μl of osmotically balanced Grace's medium containing 1 μC1 [ 3H-methyl]-methionine for 6 hr. 2. 2. For CC-CA of fourth instar female larvae, maximal incorporation of 3H-methyl was 0.15 pmol/pr/hr between days 2 and 3. High pressure liquid chromatographic (HPLC) analysis suggested that the biosynthetic products are mainly JH III with a little JH II at times. 3. 3. For CC-CA of last instar female larvae, incorporation of 3H-methyl was 0.48 pmol/pr/hr at the beginning of the stadium and decreased to negligible levels by day 10. HPLC analysis suggested that CC-CA of last instar larvae produced only JH III. Volume increases in CA during the last instar were associated with declining activities of JH secretion. 4. 4. Comparisons of maximal rates of 3 H-methyl incorporation by each unit volume of CA revealed that in the last instar each unit volume (μm 3) of glandular tissue secreted 50% more JH than in the fourth instar.

Hyperglycaemic and myoactive factors in the corpora cardiaca of the mealworm, Tenebrio molitor

The corpus cardiacum-corpus allatum complex of Tenebrio molitor contains factors causing hyperglycaemia, stimulating carbohydrate release from the fat body in vitro, and accelerating heart beat in the donor insect. However, no adipokinetic response can be demonstrated upon injection of extracts of this complex in larvae or young imagines. The carbohydrate elevation in the haemolymph is time- and dose-dependent, with a maximal effect about 90–180 min after injection. About 0.03–0.05 gland equivalents are needed to produce a significant increase in the larval and imaginal bioassays; maximal responses in both systems were reached with about 0.5 gland equivalents. Fat body treated in vitro with an extract of 0.5 gland equivalents releases about 130% more carbohydrate into 1 ml physiological saline after 1 h than paired tissue controls. The characteristic response of a semi-isolated adult heart to the endogenous myoactive factor(s) of the corpus cardiacum-allatum extract was an increase in heart beat frequency and a decrease in the amplitude of contraction. The response was dose-dependent and 0.05 gland equivalents were needed to exert a stimulatory effect on the myocardium. Fractionation of a methanolic extract of endocrine complex from Tenebrio molitor by RP-HPLC shows that the hyperglycaemic activity is confined to two regions. A myoactive factor appears to elute just after the second hyperglycaemic material, whereas a cardio-inhibitory factor has more hydrophobic properties. Synthetic neuropeptides adipokinetic hormone I and red pigment concentrating hormone injected as a dose of 8 pmol produce a weak hyperglycaemic effect, whereas myoactive factors I and II and hypertrehalosaemic factor II cause no hyperglycaemia. No adipokinetic response was shown upon injection of any of the neuropeptides tested. A very small stimulation of the adult heart beat frequency was observed upon administration of 20 pmol adipokinetic hormone I and myoactive factor I; all other peptides were without effect. In conclusion, the bioassays with Tenebrio appear to show some species-specificity and the hyperglycaemic/myoactive factors present in the corpus cardiacum-allatum neurosecretory complex may possess distinctive properties compared with adipokinetic hormone I, red pigment concentrating hormone, myoactive factors I and II, and hypertrehalosaemic factor II.

Gastrin/CCK-like immunoreactivity in the corpus cardiacum-corpus allatum complex of the cockroach Leucophaea maderae.

By use of immunocytochemistry, a gastrin/CCK-like material has been demonstrated in the corpus cardiacum-corpus allatum complex of the cockroach Leucophaea maderae. Reactivity toward gastrin and CCK with region-specific antisera suggests that the gastrin/CCK-like peptide of this insect contains the COOH-terminal tetrapeptide sequence which is common to gastrin and CCK, and that the material is more gastrin-like than CCK-like. The results indicate that, like other neuropeptides, the gastrin/CCK peptide family appeared early in evolution within neuronal elements, and that the COOH-terminal region of gastrin has been conserved during phylogeny.

The Hormonal Control of Diuresis in the Cabbage White Butterfly Pieris Brassicae

The diuresis which follows the pupal-adult ecdysis of Pieris brassicae is hormonally controlled. Use of the isolated Malpighian tubules as a bioassay shows the presence of substantial diuretic activity in homogenates of the brain and corpus cardiacum-corpus allatum complex. The hormone is probably produced in the brain and released from a storage site in the corpora cardiaca. The tubules of the butterfly are maximally responsive to the diuretic hormone at the time of eclosion.

Glucagon-like and insulin-like hormones of the insect neurosecretory system

Aqueous extracts of corpus cardiacum-corpus allatum complexes of the adult tobacco hornworm Manduca sexta produced both glycogenolysis and hypoglycaemia when injected into the larval form of the same species. Application of specific radioimmuno assays to similar extracts showed also that these gland complexes contain both glucagon-like and insulin-like peptides. Further, the partially purified immunoreactive peptides had the expected biological activities. The former decreased the glycogen content of the fatbody and the latter the circulating trehalose levels in recipient animals. These results suggest the existence of hormones in these invertebrates having both biological and structural similarities to vertebrate insulin and glucagon.

Hormonal control of larval coloration in the armyworm, Leucania separata

Leucania separata larvae show various degrees of darkening depending on the population density. A ligature applied behind the thorax of crowded or yellow solitary larvae caused black or reddish brown pigmentation in the anterior part after the larval ecdysis. Extirpation of the brain, the corpus cardiacumcorpus allatum complexes, or the suboesophageal ganglion reduced the degree of melanization in the crowded larvae, lack of the suboesophageal ganglion having a particularly striking effect. Transplantation of 3 complexes of brain-corpora cardiaca-corpora allata-suboesophageal ganglion induced intense black pigmentation in the isolated abdomens of crowded larvae and reddish brown pigmentation with some melanization in the isolated abdomens of yellow solitary larvae, though the melanization in the latter was weaker than in the former. Implantation of these organs or of the suboesophageal ganglia into yellow solitary larvae caused black and reddish brown pigmentation after a larval ecdysis. In the pieces of integument implanted into the body cavity of crowded larvae, melanization occurred after ecdysis, whereas it did not occur in most of the fragments implanted in yellow solitary larvae. Transplantation of corpora allata and other organs from solitary larvae or injection of juvenile hormone into crowded larvae did not inhibit melanization.