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.

Locust adipokinetic hormones: carrier-independent transport and differential inactivation at physiological concentrations during rest and flight.

Since concomitant release of structurally related peptide hormones with apparently similar functions seems to be a general concept in endocrinology, we have studied the dynamics of the lifetime of the three known adipokinetic hormones (AKHs) of the migratory locust, which control flight-directed mobilization of carbohydrate and lipid from fat body stores. Although the structure of the first member of the AKHs has been known for 20 years, until now, reliable data on their inactivation and removal from the hemolymph are lacking, because measurement requires AKHs with high specific radioactivity. Employing tritiated AKHs with high specific radioactivity, obtained by catalytic reduction with tritium gas of the dehydroLeu2 analogues of the AKHs synthesized by the solid-phase procedure, studies with physiological doses of as low as 1.0 pmol per locust could be conducted. The AKHs appear to be transported in the hemolymph in their free forms and not associated with a carrier protein, despite their strong hydrophobicity. Application of AKHs in their free form in in vivo and in vitro studies therefore now has been justified. We have studied the degradation of the three AKHs during rest and flight. The first cleavage step by an endopeptidase is crucial, since the resulting degradation products lack any adipokinetic activity. Half-lives for AKH-I, -II and -III were 51, 40, and 5 min, respectively, for rest conditions and 35, 37, and 3 min, respectively, during flight. The rapid and differential degradation of structurally related hormones leads to changes in the ratio in which they are released and therefore will have important consequences for concerted hormone action at the level of the target organ or organs, suggesting that each of the known AKHs may play its own biological role in the overall syndrome of insect flight.

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.

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).