Inhibition Of Juvenile Hormone Biosynthesis Research Articles

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.

Synthesis, bioactivity and functional evaluation of linker-modified allatostatin analogs as potential insect growth regulators

Abstract Insect growth regulators play an important role in integrated pest management strategies. The FGLa–allatostatins (ASTs) are a family of neuropeptides that can inhibit juvenile hormone (JH) biosynthesis by the corpora allata (CA) of Diploptera punctata in vitro, are regarded as insect growth regulator candidates. In the search for new potential mimics and to explore the effect of linker length on inhibiting JH biosynthesis, a series of AST analogs were synthesized by modifying the linker of K24, which was found to have a significant effect on JH biosynthesis in vitro in our previous study. Functional evaluation demonstrated that all the target compounds can activate the Dippu-AstR, albeit with different potencies. Analog L6 with the longest linker (n = 5), exhibited not only a promising effect on inhibition of JH biosynthesis both in vitro and in vivo, but also good activity in inhibiting basal oocyte growth. Structure–activity relationships (SAR) studies showed that longer linkers provided greater contribution to activity.

Identification and functional characterization of FGLamide-related allatostatin receptor in Rhodnius prolixus

FGLamide-related ASTs (FGLa/ASTs) are a family of brain/gut peptides with numerous physiological roles, including inhibition of juvenile hormone (JH) biosynthesis by the corpora allata and inhibition of visceral muscle contraction. FGLa/ASTs mediate their effects by binding to a rhodopsin-like G-protein coupled receptor that is evolutionarily related to the vertebrate galanin receptor. Here we determine the cDNA sequence encoding FGLa/AST receptor (FGLa/AST-R) from the Chagas disease vector, Rhodnius prolixus (Rhopr-FGLa/AST-R), determine its spatial expression pattern using quantitative PCR and functionally characterize the receptor using a heterologous assay. Our expression analysis indicates that Rhopr-FGLa/AST-R is highly expressed in the central nervous system. The receptor is also expressed in various peripheral tissues including the dorsal vessel, midgut, hindgut and reproductive tissues of both males and females, suggesting a role in processes associated with feeding and reproduction. The possible involvement of Rhopr-FGLa/ASTs in the inhibition of JH biosynthesis is also implicated due to presence of the receptor transcript in the R. prolixus corpora cardiaca/corpora allata complex. The functional assay showed that various Rhopr-FGLa/ASTs activate the receptor, with EC50 values for the response in the nanomolar range. Moreover, Rhopr-FGLa/AST-R can couple with Gq alpha subunits and cause an increase in intracellular calcium concentration. Lastly, we tested various FGLa/AST analogs in our heterologous assay. These compounds also activated the receptor and thus have the potential to serve as insect growth regulators and aid in pest control.

Families of allatoregulator sequences: a 2011 perspective1This review is part of a virtual symposium on recent advances in understanding a variety of complex regulatory processes in insect physiology and endocrinology, including development, metabolism, cold hardiness, food intake and digestion, and diuresis, through the use of omics technologies in the postgenomic era.

Three different peptide families have been named “allatostatins” (ASTs), based on their initial purifications which were based on their ability to inhibit juvenile hormone (JH) biosynthesis. These include (i) a family of peptides that have a consensus C-terminal sequence Y/FXFGL-NH2; (ii) a family of peptides with a conserved C-terminal sequence W(X)6W-NH2; and(iii) a family of peptides with C-terminal sequence PISCF, some of which are C-terminally-amidated. Each allatostatin family has functions distinct and apart from the inhibition of JH biosynthesis. A peptide family known as the “allatotropins” serve to stimulate JH biosynthesis. This family of peptides also has been proven to exert multiple effects dependent on the species in question. Genome and peptidome projects are uncovering new members of these families and it is clear that these structures are not just confined to Insecta but are found in a range of invertebrates. The receptors for these neuropeptides have been identified and tested experimentally for specific ligand binding. The Y/FXFGLa-ASTs exert their action through galanin-like receptors, W(X)6Wa-ASTs through a sex peptide-binding receptor, and PISCF-ASTs through somatostatin-like receptors. These receptors are conserved through evolutionary time and are being identified in numerous invertebrates by way of genome projects.

Allatostatin-like receptors influence behaviour in Drosophila melanogaster and Caenorhabiditis elegans.

Event Abstract Back to Event Allatostatin-like receptors influence behaviour in Drosophila melanogaster and Caenorhabiditis elegans. William Bendena1*, Christine Wang1 and Ian Chin-Sang1 1 Queen's University Kingston, Department of Biology, Canada Neuropeptides and their action on receptors are involved in regulating various behaviours of many organisms. With the arrival of sequenced invertebrate genomes many new invertebrate neuropeptides as well as their G-protein coupled receptors (GPCRs) have been identified. In Drosophila melanogaster GPCR expression studies, generally in mammalian cells, have matched neuropeptides to their cognate receptor(s). Apart from the matching of receptor with ligand, very little is known about signal transduction pathways or how activation of the receptor may regulate animals physiology or behaviour. The first allatostatin family of neuropeptides found in invertebrates (PheGlyLeu-ASTs) share a conserved C-terminal-sequence of Tyr/PheXaaPheGlyLeu-NH2. In insects, depending on the insect PheGlyLeu-AST function varies. This neuropeptide family may contribute to several physiological mechanisms such as inhibition of juvenile hormone biosynthesis, inhibition of muscle contraction, myoendorine regulation, neuromodulation, and regulation of enzymatic activities. Two allatostatin receptors were first identified in Drosophila (Dar1 and Dar2) and orthologs have been identified in a variety of other invertebrates. The role of each receptor to a particular phyisological function or behaviour is unknown. As a comparative approach to understanding the contribution of allatostatin receptors to behaviour(s) we identified two allatostatin/galanin-like GPCRs in the nematode C. elegans.A deletion mutant of a C.elegans receptor that resembles Drosophila Dar-1 lost roaming behaviour with increased pivoting which impairs their ability to travel long distances on food. Animals with a mutant receptor gain fat. In the absence of food the mutant has normal foraging/roaming behaviour. With these observations we have assessed whether an RNAi reduction of Dar-1 in Drosophila leads to altered foraging/roaming behaviour in larvae on versus off food. Keywords: allatostatin, Behavior, foraging, G protein coupled receptors, Neuropeptides Conference: NASCE 2011: The inaugural meeting of the North American Society for Comparative Endocrinology, Ann Arbor, United States, 13 Jul - 16 Jul, 2011. Presentation Type: Invited Symposium Topic: General neuroendocrinology Citation: Bendena W, Wang C and Chin-Sang I (2011). Allatostatin-like receptors influence behaviour in Drosophila melanogaster and Caenorhabiditis elegans.. Front. Endocrinol. Conference Abstract: NASCE 2011: The inaugural meeting of the North American Society for Comparative Endocrinology. doi: 10.3389/conf.fendo.2011.04.00109 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 28 Jul 2011; Published Online: 09 Aug 2011. * Correspondence: Dr. William Bendena, Queen's University Kingston, Department of Biology, Ontario, Canada, william.bendena@queensu.ca Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers William Bendena Christine Wang Ian Chin-Sang Google William Bendena Christine Wang Ian Chin-Sang Google Scholar William Bendena Christine Wang Ian Chin-Sang PubMed William Bendena Christine Wang Ian Chin-Sang Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Biochemical, Molecular, and Functional Characterization of PISCF-Allatostatin, a Regulator of Juvenile Hormone Biosynthesis in the Mosquito Aedes aegypti

Aedes aegypti PISCF-allatostatin or allatostatin-C (Ae-AS-C) was isolated using a combination of high performance liquid chromatography and enzyme-linked immunosorbent assay (ELISA). The matrix-assisted laser desorption/ionization time-of-flight (TOF) mass spectrum of positive ELISA fractions revealed a molecular mass of 1919.0 Da, in agreement with the sequence qIRYRQCYFNPISCF, with bridged cysteines. This sequence was confirmed by matrix-assisted laser desorption/ionization tandem TOF/TOF mass spectrometry analysis. The corresponding Ae-AS-C cDNA was amplified by PCR, and the sequence of the peptide was confirmed. An in vitro radiochemical assay was used to study the inhibitory effect of synthetic Ae-AS-C on juvenile hormone biosynthesis by the isolated corpora allata (CA) of adult female A. aegypti. The inhibitory action of synthetic Ae-AS-C was dose-dependent; with a maximum at 10(-9) m. Ae-AS-C showed no inhibitory activity in the presence of farnesoic acid, an immediate precursor of juvenile hormone, indicating that the Ae-AS-C target is located before the formation of farnesoic acid in the pathway. The sensitivity of the CA to inhibition by Ae-AS-C in the in vitro assay varied during the adult life; the CA was most sensitive during periods of low synthetic activity. In addition, the levels of Ae-AS-C in the brain were studied using ELISA and reached a maximum at 3 days after eclosion. These studies suggest that Ae-AS-C is an important regulator of CA activity in A. aegypti.

Inhibition of juvenile hormone biosynthesis in mosquitoes: effect of allatostatic head factors, PISCF- and YXFGL-amide-allatostatins

We investigated the role of head factors and allatostatins (ASs) on the regulation of juvenile hormone (JH) synthesis in female adult mosquito. The biosynthetic activity of the Aedes aegypti corpora allata (CA) in vitro was inhibited by factors present in the head. Disconnecting the CA from the brain resulted in a significant increase in the rate of JH biosynthesis. Inhibition was not dependent on intact nervous connections; co-incubation of CA with brains or brain extracts resulted in a significant decrease of JH biosynthesis. This inhibitory effect of brain extracts was reversible and heat stable; extracts lost the inhibitory activity after proteinase K digestion suggesting a peptidic structure. In a first attempt to elucidate the nature of this inhibitory factor, we tested in our CA in vitro system the effect of members of two families of allatostatins already described in mosquitoes. Anopheles gambiae PISCF-allatostatin (homolog to Manduca PISCF-allatostatin) significantly inhibited JH synthesis, while Ae. aegypti YXFGL-amide-allatostatins (homologs to cockroach YXFGL-amide-allatostatins) did not affect JH synthesis. These results represent the first description of an allatostatic effect of PISCF-allatostatins outside the Lepidoptera.

Inhibition of juvenile hormone biosynthesis in Gryllus bimaculatus by Glycosmis pentaphylla leaf compounds

The EtOAc fraction of Glycosmis pentaphylla leaf extract inhibits the juvenile hormone III-biosynthesis in vitro of corpora allata from 3 day old females of the field cricket Gryllus bimaculatus. The bioactive compound which is responsible for this activity was identified as the quinazolone alkaloid arborine. This alkaloid showed also a larvicidal activity against the mosquito Culex quinquefasciatus.

Inactivation of Dip-Allatostatin 5 by Membrane Preparations from the CockroachDiploptera punctata

Incubation of Dip-AST 5 (Asp-Arg-Leu-Tyr-Ser-Phe-Gly-Leu-NH2) with membrane preparations of midgut, hindgut, brain, or corpora allata (CA) results in its inactivation in terms of the inhibition of juvenile hormone biosynthesis. Dip-AST 5 is initially cleaved at Gly7-Leu8to yield the N-terminal heptapeptide (Asp-Arg-Leu-Tyr-Ser-Phe-Gly). At supraphysiological concentration, the half-life of Dip-AST 5 varied from 24 min by membrane preparations of brain to approximately 53 min following incubation with midgut membrane preparations. At more physiological concentrations (nanomolar), Dip-AST 5 was still initially cleaved to yield the inactive N-terminal heptapeptide with a half-life ranging from 23 min with brain membrane preparations to 85 min with membrane preparations of midgut. The fact that Dip-AST 5 is rapidly degraded to an inactive product by membrane preparations or whole tissues (CA) indicates that Dip-AST 5 has a different metabolic fate in tissue preparations than in diluted hemolymph (Garsideet al.,1997). These findings demonstrate that the degradation of allatostatins by tissue preparations ofD. punctatamay play an important role in the termination of their ability to inhibit juvenile hormone biosynthesis by the CA and/or to modulate muscle activity in the hindgut.

Allatostatin-like-immunoreactive neurons of the tobacco hornworm, Manduca sexta, and isolation and identification of a new neuropeptide related to cockroach allatostatins.

The YXFGLamide C-terminus serves to define most members of a family of structurally related neuropeptides, the YXFGLamides. These peptides have been identified from the nervous system of various insects and include the allatostatins of cockroaches and crickets, the schistostatins of locusts, and the callatostatins of blowflies. The YXFGLamides have been shown to have various functions, including inhibition of juvenile hormone biosynthesis in cockroaches and crickets and inhibition of contraction of certain insect visceral muscles. We wanted to know if these peptides occur in Manduca sexta and what functions they might have. A new peptide, AKSYNFGLamide, was isolated and identified from M. sexta and has been named "lepidostatin-1"; this is the first YXFGLamide to be found in a lepidopteran, and there are indications that additional YXFGLamides occur in M. sexta. An antiserum to cockroach allatostatins (YXFGLamides) was shown to recognize lepidostatin-1 of M. sexta and was used to map YXFGLamide-immunoreactive neurons in larvae. Because immunoreactive interneurons were found to form an extensive neuropil, YXFGLamides probably function as neuromodulators in M. sexta. Neuroendocrine cells in the brain, abdominal ganglia, and their respective neurohemal organs were YXFGLamide immunoreactive and appear to release YXFGLamides as neurohormones. Immunoreactivity to YXFGLamides and M. sexta diuretic hormone were found to be colocalized and appear to be coreleased in these neuroendocrine cells, indicating that YXFGLamides may be involved in regulation of fluid transport. Innervation of the corpora allata by YXFGLamide-immunoreactive processes was very sparse, suggesting that this innervation does not play an important role in allatostasis. Many thoracic motor neurons were YXFGLamide immunoreactive, suggesting that YXFGLamides may have a myomodulatory or myotrophic function in larvae. However, this immunoreactivity disappeared early in metamorphosis and did not reappear in the adult. The YXFGLamide-immunoreactive neurons in the terminal abdominal ganglion were found to innervate the hindgut, indicating that YXFGLamides may be involved in the control of the rate of myogenic contractions of the larval hindgut.

Role of second messengers in the regulation of juvenile hormone production in insects, with particular emphasis on calcium and phosphoinositide signaling

Juvenile hormone (JH) biosynthesis is regulated by both peptidergic and aminergic inputs. Depending upon the species and developmental stage, these signals may be either inhibitory or stimulatory (allatostatins and allatotropins). Signal transduction within corpus allatum cells occurs by way of known second messengers, including the cyclic nucleotides, Ca2+, and the phosphoinositides. We review the role of signal transduction in the regulation of JH biosynthesis in the locust, Locusta migratoria, the hornworm, Manduca sexta, and the cockroach, Diploptera punctata. Differences in signal transduction between the species reflect in part differences in the signals which regulate the corpus allatum. For example, JH biosynthesis in locusts appears to be regulated principally by allatotropic signals, whereas in the cockroach allatostatic signals appear to predominate. In M. sexta, both types of signals appear to function and are stage-dependent. This may reflect the complex nature of the juvenile hormone products in this species, whereas in locusts and cockroaches only JH III is produced. Modulation of intracellular calcium (CA2+) levels by drugs (ionophores, Ca2+ chelators and releasing agents, CA2+ channel blockers) or depolarizing agents including current injection dramatically alters JH biosynthesis, as do agents that activate the phosphoinositide pathway. Modulation of intracellular cAMP levels is associated with changes in JH production, with increases in JH biosynthesis occurring as a consequence of the stimulation of the adenylate cyclase system in locusts but with an inhibition of JH biosynthesis in cockroaches. The situation in M. sexta is more complex and appears to be both stage- and product-dependent. The effects of second messengers in the three species differ, probably as a consequence of the differences in the peptide signals that regulate JH biosynthesis and in the intrinsic state of the corpora allata (either switched on or switched off, in the absence of external signals). © 1996 Wiley-Liss, Inc.

Role of second messengers in the regulation of juvenile hormone production in insects, with particular emphasis on calcium and phosphoinositide signaling

Juvenile hormone (JH) biosynthesis is regulated by both peptidergic and aminergic inputs. Depending upon the species and developmental stage, these signals may be either inhibitory or stimulatory (allatostatins and allatotropins). Signal transduction within corpus allatum cells occurs by way of known second messengers, including the cyclic nucleotides, Ca2+, and the phosphoinositides. We review the role of signal transduction in the regulation of JH biosynthesis in the locust, Locusta migratoria, the hornworm, Manduca sexta, and the cockroach, Diploptera punctata. Differences in signal transduction between the species reflect in part differences in the signals which regulate the corpus allatum. For example, JH biosynthesis in locusts appears to be regulated principally by allatotropic signals, whereas in the cockroach allatostatic signals appear to predominate. In M. sexta, both types of signals appear to function and are stage-dependent. This may reflect the complex nature of the juvenile hormone products in this species, whereas in locusts and cockroaches only JH III is produced. Modulation of intracellular calcium (CA2+) levels by drugs (ionophores, Ca2+ chelators and releasing agents, CA2+ channel blockers) or depolarizing agents including current injection dramatically alters JH biosynthesis, as do agents that activate the phosphoinositide pathway. Modulation of intracellular cAMP levels is associated with changes in JH production, with increases in JH biosynthesis occurring as a consequence of the stimulation of the adenylate cyclase system in locusts but with an inhibition of JH biosynthesis in cockroaches. The situation in M. sexta is more complex and appears to be both stage- and product-dependent. The effects of second messengers in the three species differ, probably as a consequence of the differences in the peptide signals that regulate JH biosynthesis and in the intrinsic state of the corpora allata (either switched on or switched off, in the absence of external signals). © 1996 Wiley-Liss, Inc.

Effects of Diploptera punctata allatostatins on juvenile hormone biosynthesis and endogenous juvenile hormone III levels in virgin and mated female Periplaneta americana

Corpora allata from 4-day-old virgin and sexually mature female Periplaneta americana were tested for sensitivity to inhibition of juvenile hormone biosynthesis in vitro by four allatostatins from Diploptera punctata. Two of these allatostatins were also tested for their ability to lower juvenile hormone levels in vivo when injected in aqueous solution. In 4-day-old virgin females (showing comparatively low rates of juvenile hormone synthesis) corpora allata were sensitive to all four neuropeptides. Inhibition of juvenile hormone synthesis was roughly equivalent at the single dose tested (58–74% inhibition @ 10 −5 M). By contrast, corpora allata from mid-cycle sexually mature females (in which juvenile hormone synthesis approaches maximum rates) were sensitive only to D. punctata allatostatin 4. Investigation of the sensitivity of corpora allata to allatostatins 1 and 4 throughout the reproductive cycle revealed that at no stage did allatostatin 1 cause more than 25% inhibition of juvenile hormone synthesis. Allatostatin 4 however, caused a moderate inhibitory response early in the cycle (33–71%), rising to a highly significant response (81–97%) towards the end of maximal oocyte growth when endogenous (control) rates of juvenile hormone synthesis are naturally declining. The activities of the Diploptera allatostatins against Periplaneta corpora allata in vitro are matched by their actions in vivo. Injections of allatostatins 1 and 4 (100 μg/insect) into day-4 virgin P. americana were effective in bringing about a lowering of total body JH III levels 12 h post-injection. Similar injections of allatostatin 1 into mid-cycle mated females produced no apparent effect but injection of allatostatin 4 did result in a substantial reduction (50–70%) of endogenous total body JH III levels in these insects.

Inhibition of juvenile hormone biosynthesis and methyl farnesoate epoxidase activity by 1,5‐disubstituted imidazoles in the cockroach, Diploptera punctata

AbstractSeventeen substituted imidazoles were tested as inhibitors of juvenile hormone (JH) III synthesis by cockroach corpora allata in an in‐vitro radio‐chemical assay. Most of these 1,5‐disubstituted imidazoles were highly potent, with IC50 values of less than 100nM. The compounds differed in their ability to cause an accumulation of the precursor methyl farnesoate in the glands. Four of the imidazoles were tested by topical application to previtellogenic adult females, and all caused a significant inhibition of JH synthesis and an accumulation of intraglandular methyl farnesoate for at least three days after treatment. Methyl farnesoate epoxidase activity of homogenates of corpora allata was inhibited by the compounds TH‐14 and TH‐27. This P450‐dependent epoxidase activity was inhibited at less than 10 nM. The results show that the 1,5‐disubstituted imidazoles are powerful inhibitors of the last step of juvenile synthesis in this cockroach.

Signal transduction in the inhibition of juvenile hormone biosynthesis by allatostatins: roles of diacylglycerol and calcium

The effects of pharmacological agents that interfere with the 1,4,5-inositol trisphosphate (IP3)/diacylglycerol (DAG) pathway on juvenile hormone (JH) biosynthesis by corpora allata (CA) of the cockroach Diploptera punctata have been investigated. These effects were assessed in the presence of the inhibitory neuropeptides, allatostatins, with a view to elucidating the pathway for signal transduction in the inhibition of JH biosynthesis. Treatment of CA with inhibitors of DAG kinase to elevate the concentration of DAG within the CA cells, resulted in a significant, dose-dependent decrease in JH biosynthesis. Simultaneous treatment of glands with both DAG kinase inhibitors and allatostatins further enhanced this effect, suggesting that DAG is an intermediate in the allatostatin-induced inhibition of JH production. The inhibitory actions of the phorbol ester activator of PKC, PDBu, or of allatostatin on JH biosynthesis were partially blocked by pre-incubating the CA with PKC inhibitors. Treatment of CA with the calcium-mobilizing drug thapsigargin resulted in a significant stimulation in JH biosynthesis in glands from mated females producing JH at high rates. Thapsigargin was also able to reverse the effect of allatostatins in high-activity mated CA. This suggests an involvement of the other product of phosphoinositide hydrolysis, IP3, in the modulation of JH biosynthesis at specific developmental times and in glands showing specific levels of activity.

Structure-activity studies of allatostatin 4 on the inhibition of juvenile hormone biosynthesis by corpora allata: The importance of individual side chains and stereochemistry

The production of juvenile hormone III (JH III) by the corpora allata of the cockroach Diploptera punctata is regulated in part by peptides originating from the brain. One group of these peptides, termed allatostatins, reversibly inhibits the biosynthesis of JH in vitro. Allatostatin 4 (AST4: Asp-Arg-Leu-Tyr-Ser-Phe-Gly-Leu-amide) is the smallest member of the AST family yet defined and was used as the benchmark peptide for these initial structure-activity studies. Two initial analog series of AST4 were examined for the ability of each analog to inhibit JH biosynthesis by corpora allata in vitro. Each analog series consisted of analogs that contained a single amino acid change from the native AST4 sequence. The first series contained Ala replacement analogs and the second contained analogs with d-amino acid replacements. The first analog series used Ala replacements to help indicate which amino acid side chains were most important for inhibition of JH biosynthesis. The most important side chain appeared to be Leu 8 followed by Phe 6 and Tyr 4. Additionally, the d -mamino acid series suggested that a secondary structural element(s) at the C- terminus of AST4 could be important to the biological activity.

Assessment of the role of cyclic nucleotides in allatostatin-induced inhibition of juvenile hormone biosynthesis in Diploptera punctata

In an effort to identify the signal transduction mechanism associated with the inhibition of juvenile hormone (JH) biosynthesis by the neuropeptides allatostatins, levels of the cyclic nucleotides cAMP and cGMP were measured in corpora allata (CA) of virgin and mated Diploptera punctata females using radioimmunoassays. Treatment of isolated CA with varying concentrations of synthetic allatostatins 1, 2, 3 or 4 did not elicit significant changes in the levels of either cAMP or cGMP in any of the test glands, suggesting that these compounds do not act as second messengers for the four allatostatins tested. Simultaneous treatment of CA with allatostatin 4 and the adenylate cyclase activator forskolin did not increase the degree of inhibition of juvenile hormone biosynthesis relative to that obtained with forskolin (5 or 50 μM) alone. We interpret these results as lending further support to the suggestion that cyclic nucleotides do not play a role in the signal transduction of allatostatins 1–4 in cockroach CA.

Action of a juvenile hormone analogue on the activity of Periplaneta americana corpora allata in vitro and on juvenile hormone III levels in vivo

The effects of the juvenile hormone (JH) analogue fenoxycarb (ethyl[2-(4-phenoxyphenoxy)-ethyl]carbamate) on the activity of corpora allata (CA) from adult female Periplaneta americana have been investigated. The in vitro biosynthesis of JH III by isolated CA was inhibited by about 85% in the presence of a high concentration (1 × 10 −4 M) of fenoxycarb. However, at lower concentrations (1 × 10 −6 M and 1 × 10 −8 M) no inhibition of JH biosynthesis was apparent. Topical treatment of adult female cockroaches with fenoxycarb (100 μg/insect) did not reduce the subsequent rate of JH III biosynthesis by CA in vitro. By contrast, the same treatment markedly reduced the titre of endogenous JH III in intact cockroaches. These results suggest that CA activity in adult female P. americana may be controlled by negative feedback, and that this system of control is dependent on the maintenance of contact between the CA and nervous or humoral factors in the intact insect. Alternatively, it is possible that treatment with fenoxycarb increases the rate at which endogenous JH is metabolized.

Effects of food and water availability, and of NCA-1 section, upon juvenile hormone biosynthesis and oöcyte development in adult female Periplaneta americana

The combined stimuli from feeding, drinking, mating and crowding are required for the highest rates of oöcyte development in maturing adult female Periplaneta americana. A graded series of “sexually suppressed” females can be produced by withholding one or more of these stimuli, and this stepwise retardation of ovarian development appears to be achieved by a progressive increase in corpus allatum restrain. It seems that all of these environmental cues are centrally integrated such that juvenile hormone-dependent processes can proceed at an appropriate pace. Water availability is evidently the most important factor. Water-deprived females are sexually unreceptive, and are found to have very low rates of juvenile hormone biosynthesis and ovarian development. This holds true even when they are provided with food. In contrast, 75% of starved females are sexually receptive if allowed free access to drinking water. At the same time they have enhanced corpus allatum activity, and show significant oöcyte growth. The mode of regulation of corpus allatum function in adult female P. americana appears to be significantly different to the model proposed for the cockroaches Leucophaea maderae and Diploptera punctata. Allatotropic signals may be more important than inhibitory signals in the former species. The glands continue to be moderately active in fed, mated female P. americana after NCA-1 section (although a major peak of corpus allatum activity is not obvious), and the rate of oöcyte development is not greatly reduced. However, NCA-1 mediated inhibition of juvenile hormone biosynthesis is less readily demonstrated. We could observe no enhancement of corpus allatum activity nor stimulation of oöcyte growth after unilateral NCA-1 section when the operation was performed on starved virgins, and the same result was found after bilateral NCA-1 section when starvation or virginity were separately enforced. A slight stimulation of juvenile hormone biosynthesis, together with a small increase in oöcyte development, could only be demonstrated after both NCA-1 were cut in starved virgins. We conclude that neurally mediated corpus allatum inhibition in has yet to be adequately verified, and that the available evidence does not contradict the theory that juvenile hormone biosynthesis in adult females could be regulated predominantly by chemicals released into the haemolymph.

Inhibition of juvenile hormone III biosynthesis in Periplaneta americana with the fungal metabolite compactin (ML-236B)

Incorporation of the fungal metabolite compactin into an in vitro corpora allata culture inhibited juvenile hormone III biosynthesis by the glands of adult female Periplaneta americana ( ID 50 ⋍ 1 × 10 −9 M). Topical treatment of adult females (100 μg compactin/insect) 12 hr before extirpation of corpora allata resulted in about 50% inhibition of juvenile hormone III biosynthesis when these glands were subsequently cultured in vitro. However, no inhibition of juvenile hormone biosynthesis was apparent when adult females were treated by injection.