Chiral 1,2-diacylglycerols in the haemolymph of the locust, Locusta migratoria.

A 1H-NMR method using chiral shift reagents was applied in the stereochemical analysis of the haemolymph 1,2-diacylglycerols of Locusta migratoria. Conversion of the 1,2-diacylglycerols into 1,2-diacetyl-3-tritylglycerols allowed the accurate determination of the enantiomeric purity, whereas direct trimethylsilylation of the unmodified or hydrogenated haemolymph 1,2-diacylglycerols proved to be less suitable because of signal broadening. In the haemolymph of Locusta, sn-1,2-diacylglycerols with a remarkably high optical purity were found to be present. In the resting locust, at least 96% of the haemolymph 1,2-diacylglycerols have the sn-1,2-configuration, in locusts in which the haemolymph diacylglycerol concentration was elevated by fat body triacylglycerol mobilization induced by flight activity or injection of adipokinetic hormone, over 97% of the 1,2-diacylglycerols is the sn-1,2-enantiomer. The few percent sn-2,3-enantiomer may not have been present initially. Positional distribution of the fatty acids in the fat body triacylglycerols and in the haemolymph sn-1,2-diacylglycerols obtained from locusts after a 2 h flight revealed nearly identical occupation of the sn-2-positions in both acylglycerols. The distribution patterns in the sn-1-position of the 1,2-diacylglycerols and the combined sn-1 and sn-3 positions of the triacylglycerols are compatible with the possible existence of a stereospecific sn-3-triacylglycerol lipase.

Insect apolipophorin III: interaction of locust apolipophorin III with diacylglycerol.

In the formation of low-density lipophorin (LDLp) by the loading of diacylglycerol onto high-density lipophorin (HDLp) in insect hemolymph, apolipophorin III (apoLp-III) plays an essential role by binding to the increasing surface of the expanding lipoprotein particle. The present data on the surface properties of apoLp-III from Locusta migratoria demonstrate a preferential interaction with diacylglycerol. Injection of apoLp-III underneath a diacylglycerol monolayer results in a rapid interaction with the lipid; interaction with a phosphatidylcholine monolayer was considerably less. Locust apoLp-III binds with high affinity (Kd = 7.9.10(-9) M) to 1,2-diacylglycerol, which is consistent with its function in the LDLp particle; affinity for phosphatidylcholine is considerably lower. While the molecular area of locust apoLp-III in a monolayer is 2080 A2/molecule at the collapse pressure, in mixed monolayers of apoLp-III and lipid, the mean molecular area is decreased. Deglycosylation of the apoLp-III did not affect its interfacial stability. ApoLp-III from the moth Manduca sexta, which we included for comparison, demonstrated a similar reduction in molecular area resulting from interaction with lipid. These data do not support the hypothesis that interaction of apoLp-III with a lipid surface will lead to doubling of the molecular area of the protein (Kawooya, J.K., Meredith, S.C., Wells, M.A., Kézdy, F.J. and Law, J.H. (1986) J. Biol. Chem. 261, 13588-13591). The area of locust apoLp-III of 12.9 A2/amino acid residue at the collapse pressure is consistent with monolayers of alpha-helical proteins; circular dichroic spectra confirm a high alpha-helix content. The surface properties of apoLp-III reported here enable a high surface concentration of diacylglycerol in the LDLp particle, allowing the lipoprotein to act as an efficient reutilizable lipid shuttle.

Lipoproteins act as a reusable shuttle for lipid transport in the flying death's-head hawkmoth, Acherontia atropos.

High-density lipophorin (HDLp), the major insect plasma lipoprotein in resting insects, has been postulated to function as a 'reusable shuttle' for lipid transport between tissues, capable of accepting or depositing lipids with maintenance of the structural properties of the particle. Injection of differentially radiolabeled HDLp into resting death's-head hawkmoths revealed that disappearance of the [14C]palmitate labeled lipid component of HDLp (principally diacyglycerol) was relatively quickly (half-life approx. 3 h), whereas turnover time of the apolipoproteins (marked with [14C]protein hydrolysate) was considerably longer (half-life approx. 26 h). These results strongly support the above proposal. To fuel long-distance flight, insects transport lipid in the hemolymph in the form of diacylglycerol-rich low-density lipophorin (LDLp) resulting from a conversion of HDLp to LDLp. By injection of differentially radiolabeled LDLp into flying hawkmoths we demonstrate for the first time in vivo that this mechanism of lipoprotein conversion also functions as a 'reusable shuttle'. While half-life of the lipid moiety of LDLp labeled with [14C]palmitate or [14C]glycerol (mainly diacylglycerol) during flight was only 43 and 94 min, respectively, turnover rate of its apolipoprotein moiety was considerably lower (half-life approx. 30 h). The results demonstrate the unique role of HDLp, i.e., the reversible conversion to LDLp, in lipid delivery to insect flight muscles.

Recombinant locust apolipophorin III: characterization and NMR spectroscopy.

Apolipophorin III (apoLp-III) from the locust Locusta migratoria is an exchangeable apolipoprotein that reversibly binds to lipoproteins. During lipid binding the protein has been proposed to undergo a major conformational change. To study the mechanism of lipid binding we have cloned and expressed recombinant protein in bacteria, permitting stable isotope enrichment for heteronuclear NMR spectroscopy and site-directed mutagenesis. The cDNA coding for apoLp-III was subcloned into the pET expression vector and transformed into Escherichia coli cells. Induction of expression resulted in the specific appearance of apoLp-III in the cell culture medium, indicating it escaped the bacteria without lysis. The protein was purified from the cell-free supernatant by reversed-phase HPLC, characterized and compared to the natural protein isolated from locust hemolymph. SDS-PAGE revealed the recombinant protein has a molecular mass of approximately 17 kDa, similar to that of deglycosylated natural apoLp-III. Monoclonal antibodies were used to detect recombinant apoLp-III in the cells as well as in cell-free medium of induced bacterial cultures. Amino acid sequencing and analysis confirmed the identity of the recombinant protein as L. migratoria apoLp-III. Circular dichroism spectroscopy of recombinant and natural apoLp-III showed similar spectra, both displaying high contents of alpha-helical secondary structure. Denaturation studies of lipid-free apoLp-III with guanidine hydrochloride showed that both proteins have similar denaturation midpoints and DeltaG values indicating similar protein stability. The natural and recombinant protein were functional in lipoprotein binding assays. Using recombinant protein, uniformly and specifically labeled with 15N-amino acids, two dimensional 1H-15N heteronuclear single quantum correlation spectra were obtained. The spectra revealed excellent chemical shift dispersion in both the 1H and 15N dimensions with a well defined resonance pattern. Studies with 15N-leucine specifically labeled apoLp-III in the presence and absence of the micelle forming lipid, dodecylphosphocholine, provided evidence for a significant conformational change upon lipid association.

Adipokinetic hormones of insect: release, signal transduction, and responses.

Flight activity of insects provides an attractive yet relatively simple model system for regulation of processes involved in energy metabolism. This is particularly highlighted during long-distance flight, for which the locust constitutes a well-accepted model insect. Peptide adipokinetic hormones (AKHs) are synthesized and stored by neurosecretory cells of the corpus cardiacum, a neuroendocrine gland connected with the insect brain. The actions of these hormones on their fat body target cells trigger a number of coordinated signal transduction processes which culminate in the mobilization of both carbohydrate (trehalose) and lipid (diacylglycerol). These substrates fulfill differential roles in energy metabolism of the contracting flight muscles. The molecular mechanism of diacylglycerol transport in insect blood involving a reversible conversion of lipoproteins (lipophorins) has revealed a novel concept for lipid transport in the circulatory system. In an integrative approach, recent advances are reviewed on the consecutive topics of biosynthesis, storage, and release of insect AKHs, AKH signal transduction mechanisms and metabolic responses in fat body cells, and the dynamics of reversible lipophorin conversions in the insect blood.

Adipokinetic hormone-induced influx of extracellular calcium into insect fat body cells is mediated through depletion of intracellular calcium stores.

Adipokinetic hormone I (AKH I) needs extracellular Ca2+ for its activating action on glycogen phosphorylase in locust fat body in vitro. TMB-8 reduces this AKH effect significantly, indicating that for a major part, hormone action also requires the mobilization of Ca2+ from intracellular stores. Using 45Ca2+, AKH was shown to stimulate both the influx and the efflux of Ca2+. Thapsigargin also enhances the influx of extracellular Ca2+ into the fat body cells, indicating that the stimulating effect of AKH on Ca2+ influx may be mediated through depletion of intracellular Ca2+ stores as well. AKH is known to enhance cAMP levels in locust fat body. We show that elevation of cAMP with forskolin or theophylline leads to activation of glycogen phosphorylase, both in the presence and in the absence of extracellular Ca2+. The present data are discussed in an attempt to elucidate further the mechanism underlying transduction of the hormonal signal in locust fat body.

Signal transduction of adipokinetic hormones involves Ca2+ fluxes and depends on extracellular Ca2+ to potentiate cAMP-induced activation of glycogen phosphorylase.

Adipokinetic hormone (AKH)-induced mobilization of insect fat body glycogen occurs through activation of glycogen phosphorylase. In the migratory locust, signal transduction of AKH-I, -II and -III has been shown to involve the formation of cAMP. In the present study, we show that both the elevation of fat body cAMP levels and the activation of phosphorylase by the three AKHs in vitro depend on the presence of extracellular Ca2+; in the absence of Ca2+ in the medium, no phosphorylase activation occurs, whereas a concentration of at least 1.5 mM Ca2+ in the medium is required for maximal activation by each of the hormones. Furthermore, we show that AKH-I, -II and -III increase the influx of extracellular calcium into the fat body, as well as the efflux of cytosolic calcium from the fat body into the medium within 1 min of incubation. Although the time courses of their effects and the maximal responses to massive doses (40 nM) of the three hormones do not differ, AKH-III induces the highest increase in Ca2+ efflux when applied in a physiological dose (4 nM). No difference in the levels of Ca2+ influx induced by 4 nM of the hormones was observed. Quantitative analysis of the data suggests that the AKH-induced influx is larger than the efflux, implying a net rise in the fat body Ca2+ concentration.

The adipokinetic cells in the corpus cardiacum of Locusta migratoria preferentially release young secretory granules.

The influence of flight activity on the release of secretory granules from the adipokinetic cells in the corpus cardiacum of Locusta migratoria was studied. Two labeling methods, an enzymatical and a radioactive one, were used to label young, newly synthesized secretory granules and so distinguish them from older, preexisting granules. Both methods demonstrated that the ratio between the numbers of labeled and unlabeled secretory granules was lower in flight-stimulated adipokinetic cells than in unstimulated cells. This ratio was lower in both the cell bodies and the cell processes of flight-stimulated cells. After flight there was no detectable change in the total number of secretory granules, which indicates that the synthesis of new secretory granules is not inhibited by flight activity. Rather, the tendency of flight-stimulated cells to have more trans-Golgi networks labeled with wheat-germ agglutinin-conjugated horseradish peroxidase suggests that the synthesis of new secretory granules was enhanced by flight. The results led to the conclusion that young secretory granules were preferentially released over older secretory granules.

Preferential release of newly synthesized, exportable neuropeptides by insect neuroendocrine cells and the effect of ageing of secretory granules.

The release of newly synthesized neuropeptides was studied in an in vitro system using the adipokinetic hormone (AKH)-producing cells of an insect (Locusta migratoria) as a model system. Tritiated phenylalanine incorporated into three hormonal neuropeptides, AKH I, II and III, was used to distinguish newly synthesized hormones from older, preexisting ones. After pulse-chase labeling experiments of varying duration, the secretion of AKHs by the AKH cells was stimulated. Both hormones released into the incubation medium after stimulation and non-released hormones extracted from the tissue were separated by reversed-phase high performance liquid chromatography. Their radioactivity was measured by scintillation counting of the column eluate. The ratio between the specific radioactivities of the released and the non-released neuropeptides was always greater than 1.0, which indicates that the newly synthesized peptides are preferentially released. The percentages of newly synthesized (radioactive) AKHs which are released, increased until 8 1/4 h and decreased thereafter. The results indicate that after the packaging of the prohormones into secretory granules and their processing to bioactive AKHs, some further maturation of the secretory granules is required before they can release their content. After an 8 1/4 h incubation, secretory granules with radioactive AHKs enter a non-releasable pool consisting of older secretory granules.

Intracisternal granules in the adipokinetic cells of locusts are not degraded and apparently function as supplementary stores of secretory material.

The intracisternal granules in locust adipokinetic cells appear to represent accumulations of secretory material within cisternae of the rough endoplasmic reticulum. An important question is whether these granules are destined for degradation or represent stores of (pro)hormones. Two strategies were used to answer this question. First, cytochemistry was applied to elucidate the properties of intracisternal granules. The endocytic tracers horseradish peroxidase and wheat-germ agglutinin-conjugated horseradish peroxidase were used to facilitate the identification of endocytic, autophagic, and lysosomal organelles, which may be involved in the degradation of intracisternal granules. No intracisternal granules could be found within autophagosomes, and granules fused with endocytic and lysosomal organelles were not observed, nor could tracer be found within the granules. The lysosomal enzyme acid phosphatase was absent from the granules. Second, biochemical analysis of the content of intracisternal granules revealed that these granules contain prohormones as well as hormones. Prohormones were present in relatively higher amounts compared with ordinary secretory granules. Since the intracisternal granules in locust adipokinetic cells are not degraded and contain intact (pro)hormones it is concluded that they function as supplementary stores of secretory material.

Absence of coupling between release and biosynthesis of peptide hormones in insect neuroendocrine cells.

Adipokinetic hormone (AKH)-producing cells in the corpus cardiacum of the insect Locusta migratoria represent a neuroendocrine system containing large quantities of stored secretory peptides. In the present study we address the question whether the release of AKHs from these cells induces a concomitant enhancement of their biosynthesis. The effects of hormone release in vivo (by flight activity) and in vitro (using crustacean cardioactive peptide, locustamyoinhibiting peptide, and activation of protein kinase A and C) on the biosynthetic activity for AKHs were measured. The intracellular levels of prepro-AKH mRNAs, the intracellular levels of pro-AKHs, and the rate of synthesis of (pro-)AKHs were used as parameters for biosynthetic activity. The effectiveness of in vitro treatment was assessed from the amounts of AKHs released. Neither flight activity as the natural stimulus for AKH release, nor in vitro treatment with the regulatory peptides or signal transduction activators appeared to affect the biosynthetic activity for AKHs. This points to an absence of coupling between release and biosynthesis of AKHs. The strategy of the AKH-producing cells to cope with variations in secretory stimulation seems to rely on a pool of secretory material that is readily releasable and continuously replenished by a process of steady biosynthesis.

Molecular characterization and gene expression in the eye of the apolipophorin II/I precursor from Locusta migratoria.

The transport of lipids via the circulatory system of animals constitutes a vital function that uses highly specialized lipoprotein complexes. In insects, a single lipoprotein, lipophorin, serves as a reusable shuttle for the transport of lipids between tissues. We have found that the two nonexchangeable apolipoproteins of lipophorin arise from a common precursor protein, apolipophorin II/I (apoLp-II/I). To examine the mechanisms of transport of lipids and liposoluble substances inside the central nervous system, this report provides the molecular cloning of a cDNA encoding the locust apoLp-II/I. We have recently shown that this precursor protein belongs to a superfamily of large lipid transfer proteins (Babin et al. [1999] J. Mol. Evol. 49:150-160). We determined that, in addition to its expression in the fat body, the locust apoLp-II/I is also expressed in the brain. Part of the signal resulted from fat body tissue associated with the brain; however, apoLp-II/I was strongly expressed and the corresponding protein detected, in pigmented glial cells of the lamina underlying the locust retina and in cells or cellular processes interspersed in the basement membrane. The latter finding strongly suggests an implication of apolipophorins in the transport of retinoids and/or fatty acids to the insect retina.

Electron microscopic visualization of receptor-mediated endocytosis of Dil-labeled lipoproteins by diaminobenzidine photoconversion.

We present a modified diaminobenzidine (DAB) photoconversion method that enables staining of internalized Dil-labeled lipoproteins without the apparent punctate background staining that was observed with the original DAB photoconversion method. This is illustrated by the localization of Dil-labeled insect lipoproteins in natural recipient cells that internalize these lipoproteins by receptor-mediated endocytosis. Exposure to Dil-excitation light of cells that had been incubated with Dil-labeled lipoproteins yielded a light- and electron-dense DAB reaction product. In addition to the expected staining, an apparent punctate background staining of vesicular structures hindered proper identification of Dil-containing vesicles because these background-stained vesicles were indistinguishable from putative late endosomal and lysosomal structures at the electron microscopic level. This background staining was completely abrogated by inhibition of peroxisomal catalase with aminotriazole. The conversion of DAB by the emitted light of Dil was not affected by aminotriazole. We conclude that specific staining of Dil-labeled intracellular structures can be achieved with the modified DAB photoconversion method reported here.

Trehalose inhibits the release of adipokinetic hormones from the corpus cardiacum in the African migratory locust, Locusta migratoria, at the level of the adipokinetic cells.

The effect of trehalose at various concentrations on the release of adipokinetic hormones (AKHs) from the adipokinetic cells in the glandular part of the corpus cardiacum of Locusta migratoria was studied in vitro. Pools of five corpora cardiaca or pools of five glandular parts of corpora cardiaca were incubated in a medium containing different concentrations of trehalose in the absence or presence of AKH-release-inducing agents. It was demonstrated that trehalose inhibits spontaneous release of AKH I in a dose-dependent manner. At a concentration of 80 mM, which is the concentration found in the hemolymph at rest, trehalose significantly decreased the release of AKH I induced by 100 microM locustatachykinin 1, 10 microM 3-isobutyl-1-methylxanthine (IBMX) or high potassium concentrations. The specificity of the effect of trehalose was studied by incubating pools of corpora cardiaca with the non-hydrolyzable disaccharide sucrose or with glucose, the degradation product of trehalose, both in the presence and absence of 10 microM IBMX. Sucrose had no effect at all on the release of AKH I, whereas glucose strongly inhibited its release. The results point to the inhibitory effect of trehalose on the release of AKH I being exerted, at least partly, at the level of the adipokinetic cells, possibly after its conversion into glucose. The data presented in this study support the hypothesis that in vivo the relatively high concentration of trehalose (80 mM) at rest strongly inhibits the release of AKHs. At the onset of flight, the demand for energy substrates exceeds the amount of trehalose that can be mobilized from the fat body and consequently the trehalose concentration in the hemolymph decreases. This relieves the inhibitory effect of trehalose on the release of AKHs, which in turn mobilize lipids from the fat body.

Stimulation of glycogenolysis by three locust adipokinetic hormones involves Gs and cAMP.

Insect adipokinetic hormones (AKHs) have been shown to mobilize fat body carbohydrate by glycogen phosphorylase activation. In this study, the signal transduction pathways of AKH-I, -II and -III from the migratory locust are further elucidated. We show that the AKHs enhance fat body cAMP levels in vitro. For all hormones, maximal levels are reached after 1 min and correspond to a 200% increase compared to resting levels. Although cAMP levels induced by massive doses of AKH-I, -II and -III are equal, AKH-III is the most potent when applied in a physiological dose. This difference in potency also applies to glycogen phosphorylase activation. Cholera toxin (CTX) likewise ennhaces cAMP levels and phosphorylase activity, however pertussis toxin (PTX) has no effect. Increases induced by CTX and AKH are not additive, suggesting that they share the same pathway. Phosphorylase activation by the AKHs is strongly attenuated by guanosine-5'-O-(2-thiodiphosphate) (GDP beta S). These results demonstrate a role for cAMP in AKH signal transduction and indicate that the AKH receptor(s) are coupled to cAMP formation and glycogen phosphorylase activation via the stimulatory guanine nucleotide-binding protein (Gs).

New insights into adipokinetic hormone signaling.

Flight activity of insects comprises one of the most intense biochemical processes known in nature, and therefore provides an attractive model system to study the hormonal regulation of metabolism during physical exercise. In long-distance flying insects, such as the migratory locust, both carbohydrate and lipid reserves are utilized as fuels for sustained flight activity. The mobilization of these energy stores in Locusta migratoria is mediated by three structurally related adipokinetic hormones (AKHs), which are all capable of stimulating the release of both carbohydrates and lipids from the fat body. To exert their effects intracellularly, these hormones induce a variety of signal transduction events, involving the activation of AKH receptors, GTP-binding proteins, cyclic AMP, inositol phosphates and Ca2+. In this review, we discuss recent advances in the research into AKH signaling. This not only includes the effects of the three AKHs on each of the signaling molecules, but also crosstalk between signaling cascades and the degradation rates of the hormones in the hemolymph. On the basis of the observed differences between the three AKHs, we have tried to construct a physiological model for their action in locusts, in order to answer a fundamental question in endocrinology: why do several structurally and functionally related peptide hormones co-exist in locusts (and animals in general), when apparently one single hormone would be sufficient to exert the desired effects? We suggest that the success of the migratory locust in performing long-distance flights is in part based on this neuropeptide multiplicity, with AKH-I being the strongest lipid-mobilizing hormone, AKH-II the most powerful carbohydrate mobilizer and AKH-III, a modulatory entity that predominantly serves to provide the animal with energy at rest.

Differential induction of inositol phosphate metabolism by three adipokinetic hormones.

Many (in)vertebrates simultaneously release several structurally and functionally related hormones; however, the relevance of this phenomenon is poorly understood. In the locust e.g. each of three adipokinetic hormones (AKHs) is capable of controlling mobilization of carbohydrate and lipid from fat body stores, but it is unclear why three AKHs coexist. We now demonstrate disparities in the signal transduction of these hormones. Massive doses of the AKHs stimulated total inositol phosphate (InsPn) production in the fat body biphasicly, but time courses were different. Inhibition of phospholipase C (PLC) resulted in attenuation of both InsPn synthesis and glycogen phosphorylase activation. The AKHs evoked differential formation of individual [3H]InsPn isomers (InsP(1-6)), the effect being most pronounced for InsP3. 40 nM of AKH-I and -III induced a substantial rise in total InsPn and [3H]InsP3 at short incubations, whereas the AKH-II effect was negligible. At a more physiological dose of 4 nM, the AKHs equally enhanced Ins(1,4,5)P3 levels. The InsP3 effect was most prolonged for AKH-III. These subtle differences in InsPn metabolism, together with earlier findings on differences between the AKHs, support the hypothesis that each AKH exerts specific biological functions in the overall syndrome of energy mobilization during flight.

Insect adipokinetic hormone stimulates inositol phosphate metabolism: roles for both Ins(1,4,5)P3 and Ins(1,3,4,5)P4 in signal transduction?

Adipokinetic hormones (AKHs) control the mobilization of energy reserves from the insect fat body as fuels for flight activity. As a part of our investigations on AKH signal transduction, we demonstrate in this study that the inositol lipid cycle may be involved in the action of AKH-I on fat body of the migratory locust. We show that [3H]inositol is incorporated into fat body phosphoinositides in vitro, whose hydrolysis leads to the formation of the following inositol phosphates (InsPs): Ins(1 and/or 3)P, Ins(4)P, Ins(1,3)P2, Ins(1,4)P2, Ins(3,4)P3, Ins(1,3,4)P3, Ins(1,4,5)P3 and Ins(1,3,4,5)P4. AKH stimulates the formation of these isomers, eliciting an increase in radioactivity of total InsPs already after 1 min. Mass measurements show that Ins(1,4,5)P3 levels are substantially enhanced by AKH, which is indicative of hormonal activation of phospholipase C. In cell-free tissue preparations, Ins(1,4,5)P3 is metabolized through dephosphorylation as well as further phosphorylation. Ins(1,3,4,5)P4 is dephosphorylated primarily to Ins(1,3,4)P3, although the ability for its reconversion to Ins(1,4,5)P3 suggests that in vivo Ins(1,3,4,5)P4 may function as a rapidly mobilizable pool for Ins(1,4,5)P3 generation. Metabolic pathways for the conversion of InsPs to inositol in the locust fat body are proposed.

Adipokinetic hormones. Coupling between biosynthesis and release.

During long-distance flight of migratory locusts, the dramatic energy demand of the flight muscles is controlled by three adipokinetic hormones (AKHs). These peptide hormones regulate the mobilization of lipid and carbohydrate stored in the fat body to serve as energy substrates for the flight muscles. Despite the relatively huge quantities of the three AKHs that are stored in the corpora cardiaca, flight induces a differential 2-4-fold increase in the mRNAs for the three hormones. Moreover, newly synthesized AKHs can be released only during a restricted period of time, suggesting that by far most of the stored hormones are physiologically inactive. This raises the question of how the biosynthetic activity in the AKH-producing cells is coupled to their secretory activity. The present review discusses the potential mechanisms by which generation and release of mixtures of bioactive neurohormones are controlled and how peptidergic neuroendocrine cells cope with variations in physiological stimulation, with the AKH-producing cells serving as a model system.

Multifactorial control of the release of hormones from the locust retrocerebral complex.

The retrocerebral complex of locusts consists of the corpus cardiacum, the corpora allata, and the nerves that connect these glands with the central nervous system. Both corpus cardiacum and corpora allata are neuroendocrine organs and consist of a glandular part, which synthesizes adipokinetic hormones and juvenile hormone, respectively, and of a neurohemal part. The glandular adipokinetic cells in the corpus cardiacum appear to be subjected to a multitude of regulatory stimulating, inhibiting, and modulating substances. Neural influence comes from secretomotor cells in the lateral part of the protocerebrum. Up to now, only peptidergic factors have been established to be present in the neural fibres that make synaptic contact with the adipokinetic cells. Humoral factors that act on the adipokinetic cells via the hemolymph are of peptidergic and aminergic nature. In addition, high concentrations of trehalose inhibit the release of adipokinetic hormones. Although there is evidence that neurosecretory cells in the protocerebrum are involved in the control of JH biosynthesis, the nature of the factors involved remains to be resolved.