Some problems of homeostasis of haemolymph metabolites in the locust

Sectioning of the afferent nerves (NCCI and II) to the locust corpora cardiaca, glandular lobe removal, cardiacectomy, or removal of the median neurosecretory cells of the brain, have no long-term effect on blood lipid concentration. After removal of the glandular lobe, haemolymph carbohydrate concentration is lowered and remains significantly so from the second to the sixth day after the operation but returns to normal within 10 to 15 days. Severance of the afferent nerves to the corpora cardiaca does not, however, affect blood carbohydrate concentration. The injection of a concentrated extract of glandular lobes into locusts deprived of their glandular lobes does not elicit a hyperglycaemic effect even when blood carbohydrate levels are low. Cauterization of the median neurosecretory cells of the brain, sectioning of the NCCI and II, or removal of the glandular lobes of the corpora cardiaca have no effect on haemolymph protein.After dilution of haemolymph constituents by the injection of water, carbohydrate and protein concentrations are not rapidly restored to their initial values. The lipid concentration, however, rapidly returns to its pre-injection level due to the mobilization of 16:16, 16:18, and 18:18 diglycerides. This occurs even in glandular lobe deprived, median neurosecretory cell cauterized, or headless locusts. These diglycerides are mobilized following the injection of solutions containing lipid, carbohydrate, and/or protein, and are the same diglycerides that are released from the fat body in response to adipokinetic hormone. It is concluded that the injection of large volumes of fluid causes lipid mobilization but adipokinetic hormone does not apear to be involved, and the mechanism of blood lipid homeostasis in the resting locust is not clear.     

The role of the glandular lobes of the corpora cardiaca during flight in Locusta

ABSTRACT. Flight performance in Locusta is reduced following severance of the major afferent nerves to the corpora cardiaca or removal of the glandular lobes of the corpora cardiaca. These operations prevent the release of adipokinetic hormone and the consequent mobilization of stored lipid. However, locusts deprived of about 90% of their glandular lobe tissue, while flying poorly, did mobilize lipid. It is suggested that the remaining glandular parenchyma cells are capable of secreting enough hormone to stimulate lipid mobilization, but that the concentration may be inadequate to encourage lipid utilization. After removal of all the glandular lobe parenchyma, the blood carbohydrate concentration was temporarily depressed. Nevertheless flight performance was equally poor, both when haemolymph carbohydrate levels were low and when they had returned to normal. After the injection of trehalose into operated control locusts and locusts deprived of their glandular lobes, flight was still markedly poorer in the operated insects, even though the injection of trehalose prevented adipokinetic hormone release in the intact locust. It seems that the poor flight performance of locusts deprived of their glandular lobes cannot be fully explained by the simple absence of adipokinetic hormone.     

Number and size of intracisternal granules in the adipokinetic hormone-secreting cells of locusts: a phase-dependent characteristic

Abstract.The intracisternal (= ergastoplasmic) granules in the adipokinetic hormone-secreting cells of the glandular lobes of the corpora cardiaca in Locusta migratoria migratorioides represent accumulations of adipokinetic prohormones within cisternae of the rough endoplasmic reticulum. Solitary locusts have more and larger intracisternal granules than gregarious locusts. This coincides with the general locomotor activity and thus the energy metabolism in solitary locusts being quite different from that of gregarious locusts, which apparently has consequences for the amounts of adipokinetic hormones synthesized and/or released and, consequently, for the storage of these hormones in the intracisternal granules. These granules apparently function as supplementary stores of secretory material.     

A possible site of action for adipokinetic hormone on the flight muscle of locusts

In mitochondrial preparations the oxidation of palmitate and of palmitoyl carnitine is stimulated by dilute extracts of the glandular lobes of the corpora cardiaca. Extracts of the storage lobes of the corpora cardiaca or of corpora allata are without effect. In a working single muscle preparation the entry of diglyceride into the muscle is also stimulated by tissue extracts containing adipokinetic hormone. This stimulation is, however, prevented by the addition of 2-bromostearic acid to the perfusion fluid. It is suggested that adipokinetic hormone may have a single site of action in the flight muscle and this may be in stimulating the entry of acyl groups into the mitochondria; perhaps by acting on the inner acyl carnitine transferase.     

In vivo studies on the release of hormones from the corpora cardiaca of locusts

Summary Sectioning of the afferent nerves (NCCl and NCCll) to the locust corpus cardiacum prevents thein vivo release of adipokinetic hormone from the glandular lobes. This failure to release the hormone during flight and the consequent lack of lipid mobilisation brings about an impairment of flight performance which can be corrected by injections of corpus cardiacum extracts. Sectioning of the NCCl and NCCll reduces markedly the activity of the corpora allata. However, the poor flight performance of allatectomised locusts is not related to an inability to mobilise lipid since injections of corpus cardiacum extract which will mobilise fat body lipid in these locusts have no effect on flight performance. The results of individual sectioning of the NCCl and NCCll suggest that a double innervation of the glandular lobes functionsin vivo to control adipokinetic hormone release but that the NCCl alone may control the release of the diuretic hormone.     

Comparative immunocytochemical study of release sites of insulin,glucagon and AKH-like products in Locusta migratoria,Periplaneta americana,and Carausius morosus

Summary Insulin, glucagon and adipokinetic hormone antisera were applied to the corpora cardiaca, perisympathetic organs, neurohemal areas and peripheral neurosecretory cells of three insect species, the locust Locusta migratoria, the cockroach Periplaneta americana, and the stick insect Carausius morosus. The neurohemal part of the corpora cardiaca was shown to be immunoreactive to both insulin and glucagon antisera while the glandular cells reacted to adipokinetic hormone antisera. The perisympathetic organs seem to be devoid of these three substances, but certain peripheral neurohemal areas contained AKH and glucagon immunoreactive products. The latter were found to originate in the peripheral neurosecretory cells.     

Adipokinetic hormone content of the corpora cardiaca in gregarious and solitary migratory locusts

Abstract. The adipokinetic hormone (AKH-I and AKH-II) content of the corpora cardiaca from adult males of crowded (gregarious) and isolated (solitary) Locusta migratoria migratorioides (Reiche & Fairmaire) was quantified by reverse-phase high-performance liquid chromatography.Significantly less total hormone was found in the corpora cardiaca of crowded locusts than in those glands of isolated locusts at the age of 12–19 days after fledging.The ratio of AKH-I/AKH-II was higher in crowded than in isolated locusts at this age.From the age of 12–19 days to that of 25–30 days, AKH content increased significantly in the corpora cardiaca of crowded locusts, but no such increase was found in the glands of isolated locusts, and at 25–30 days there were no significant differences in the AKH content of the glands from crowded and isolated locusts.     

Adipokinetic and hyperglycaemic factor(s) in the corpora cardiaca/corpora allata complex of the stick insect, Carausius morosus.

ABSTRACT. The neurosecretory corpora cardiaca/corpora allata complex of Carausius morosus , a wingless insect of the superorder Orthopteroidea, contains adipokinetic and hyperglycaemic factor(s) capable of elevating lipids in locusts, and carbohydrates in cockroaches. Neither activity can be demonstrated in the stick insect itself, however. In addition, in locusts the Carausius gland extract is able to elevate levels of cyclic AMP in the fat body.     

Adipokinetic Hormone and Flight Fuel Related Characteristics of Density-Dependent Locust Phase Polymorphism: A Review

Recent findings on differences between the gregarious and solitary phases of locusts are reviewed in relation to flight fuel utilization, adipokinetic responses, and adipokinetic hormones. Laboratory results obtained with Locusta migratoria migratorioides show that the amount of lipid reserves, resting levels of haemolymph lipids, and hyperlipaemic responses to flight and to injection of corpus cardiacum extract or of synthetic adipokinetic hormones, are higher in crowded than in isolated locusts. No major phase-dependent differences seem to exist in flight-related carbohydrate metabolism. The adipokinetic hormone content of the corpora cardiaca is higher in younger isolated locusts than in crowded ones. Adipokinetic hormone precursor-related peptide content of the corpora cardiaca is also higher in isolated than in crowded locusts. Crowded locusts have higher lipid reserves and higher hyperlipaemic responses to flight than isolated locusts also in Schistocerca gregaria and, following injection of synthetic adipokinetic hormone, the formation of low density lipophorin is higher in crowded than in isolated locusts of this species. The laboratory results obtained with isolated and crowded locusts are extrapolated to understand the ecophysiology of the migrations of solitary and gregarious field populations of L.m. migratorioides according to available information on the differences in the migration of the two phases. It is inferred that in this species solitary locusts have a rather coarse adipokinetic strategy focused on a single prereproductive long-distance migratory flight, whereas gregarious locusts possess a fine adipokinetic balance for reiterative, sometimes unpredictably long-distance, migrations in the prereproductive, as well as reproductive, periods. The differences between the adipokinetic strategies of solitary and gregarious S. gregaria seem to be less dramatic, nevertheless, they indicate a better adaptation of the gregarious phase to prolonged flights.     

Adipokinetic hormone and flight metabolism in the locust

In adult male Schistocerca 20 min of tethered flight causes a halving of the haemolymph carbohydrate concentration. Injection of a proteinaceous emulsion of diglyceride 30 min before flight reduces both flight speed and carbohydrate utilisation. This effect can be overcome by the injection of trehalose immediately before the flight. If, in addition to the diglyceride, a dilute extract of the glandular lobes of the corpora cardiaca is injected immediately before flight, either with or without additional trehalose, carbohydrate utilisation is drastically reduced whereas flight speed is unaffected. It is argued that diglyceride competes with trehalose as a substrate for the flight muscles and that adipokinetic hormone from the glandular lobes of the corpora cardiaca stimulates the oxidation of diglyceride in these muscles during flight. This brings about a more complete (non-competitive) inhibition of trehalose utilisation by the flight muscles.     

A possible role of Schisto FLRFamide in inhibition of adipokinetic hormone release from locust corpora cardiaca

The distribution and actions of FMRFamide-related peptides (FaRPs) in the corpora cardiaca of the locust Locusta migratoria were studied. Antisera to FMRFamide and SchistoFLRFamide (PDVDHVFLRFamide) label neuronal processes that impinge on glandular cells in the glandular lobe of the corpora cardiaca known to produce adipokinetic hormones. Electron microscopic immunocytochemistry revealed that these FaRP-containing processes form synaptoid contacts with the glandular cells. Approximately 12% of the axon profiles present in the glandular part of the corpus cardiacum contained SchistoFLRFamide-immunoreactive material. Retrograde tracing of the axons in the nervus corporis cardiaci II with Lucifer yellow revealed 25–30 labelled neuronal cell bodies in each lateral part of the protocerebrum. About five of these in each hemisphere reacted with the SchistoFLRFamide-antiserum. Double-labelling immunocytochemistry showed that the FaRP-containing processes in the glandular lobe of the corpora cardiaca are distinct from neuronal processes, reacting with an antiserum to the neuropeptide locustatachykinin. The effect of the decapeptide SchistoFLRFamide and the tetrapeptide FMRFamide on the release of adipokinetic hormone I (AKH I) from the cells in the glandular part of the corpus cardiacum was studied in vitro. Neither the deca- nor the tetrapeptide had any effect on the spontaneous release of AKH I. Release of AKH I induced by the phosphodiesterase inhibitor IBMX, however, was reduced significantly by both peptides. These results point to an involvement of FaRPs as inhibitory modulators in the regulation of the release of adipokinetic hormone from the glandular cells.     

Fuels for flight in Locusta

In Locusta migratoria, carbohydrate is the predominant energy source for the first 20 to 30 min of flight. During this time carbohydrate is utilized at the rate of 120 μg/min, but thereafter it is used at only 8 to 9 per cent of the initial rate. Initially carbohydrate is not mobilized to any extent from body stores. Lipid, however, is mobilized (from stores in the fat body) in response to adipokinetic hormone released from the glandular lobes of the corpora cardiaca. The mobilized lipid is diglyceride, and 16 : 18 and 18 : 18 diglycerides predominate. The rate of diglyceride mobilization is rapid, both during flight and in response to extracts of corpora cardiaca. The release of adipokinetic hormone during flight has also been estimated and is found to be more rapid than previously suggested. It is calculated that the rate of lipid utilization increases during flight from virtually zero during the first few minutes to 35 μg/min (15 to 30 min) and finally reaches about 85 μg/min during the second 30 min of flight.     

Diglyceride-transporting lipoproteins inLocusta

Summary The majority of haemolymph proteins obtained from resting mature locusts are soluble in 50% saturated (NH₄)₂SO₄ solution. Chromatographically, these proteins may be resolved on Sepharose 6B into three partially included fractions. The leading high molecular weight fraction, A, is predominant and all the diglyceride in resting blood is associated with it. An additional higher molecular weight fraction, A⁺, is present in the haemolymph of locusts injected with extracts of the glandular lobes of the corpora cardiaca (elevated blood), and this fraction carries all the increased diglyceride. Fraction A⁺ is better resolved on Ultrogel ACA 22. A useful rapid technique for studying the formation of A⁺ lipoprotein during lipid mobilization has been developed by the use of lipoproteinpolyanion-metal ion complex formation. A⁺ is soluble after heparin treatment whereas the yellow leading edge of fraction A precipitates. Concomitant with the appearance of A⁺ there is a marked depletion of a lower molecular weight non lipid-carrying protein fraction, C. The ability to form fraction A⁺ after injections of glandular lobe extract cannot be demonstrated in fledglings (immature locusts within 2 or 3 days after the imaginal moult) where fraction C is virtually absent. It is suggested that during lipid mobilization, some lipoprotein from fraction A combines with protein from fraction C to form A⁺. A direct effect of adipokinetic hormone on the process of A⁺ formation cannot be excluded.     

The effects of corpora cardiaca on tethered flight in the Locust

Summary Removal of the cerebral neurosecretory cells tas no effect on the tethered flight pattern of adult maleLocusta. However, the flight pattern is markedly influenced by removal of the glandular lobes of the corpora cardiaca or by injection of 0.01 pair of corpora cardiaca. Changes in flight speed, brought about by such injections, may reflect differences in substrate utilisation in response to hormonal factor(s) from the corpora cardiaca. It is suggested that one effect of corpus cardiacum factor(s) may be the suppression of carbohydrate utilisation by the flight muscles.     

Diuretic hormone: another peptide with widespread distribution within the insect CNS

ABSTRACT. Diuretic hormone is a peptide neurohormone of as yet unknown chemical structure. Attempts to characterize the locust hormone have concentrated on the storage lobes of the corpora cardiaca as a source of material. Bioassay data now indicates that the occurrence of diuretic activity is more widespread in the locust CNS. Dose-response curves indicate that the levels of the diuretic factor(s) in the brain, sub-oesophageal and thoracic ganglia are remarkably similar to those found in the corpora cardiaca; activity was also detected in the abdominal ganglia (1–5) and terminal abdominal ganglion. Reverse-phase HPLC has confirmed that the diuretic material found in the various parts of the CNS is chemically similar to at least one of the diuretic factors separated from the corpora cardiaca. The significance of the widespread distribution of this peptide hormone is considered.
Recent evidence of other workers has indicated the possible structural similarity between vertebrate vasopressin and the diuretic factor in the locust sub-oesophageal ganglion. Chromatographic criteria have been used to demonstrate that the diuretic hormones isolated in our work are not vasopressin-like molecules, but are chemically distinct entities.     

Hormonal regulation of proline synthesis and glucose release in the fat body of the Colorado potato beetle, Leptinotarsa decemlineata

In the adult males of the Colorado potato beetle, Leptinotarsa decemlineata, corpora cardiaca extract injected in vivo gives rise to an increase in glucose and a decrease in alanine concentration of the haemolymph. The regulation of proline synthesis and glucose release in the fat body of the Colorado potato beetle was investigated further in vitro. Proline regulates its own synthesis by a feedback inhibition. Moreover, a factor present in extracts from the corpora cardiaca stimulates synthesis in the fat body in vitro. This effect was demonstrated with corpora cardiaca extracts from beetles, locusts and cokroaches. Also, synthetic adipokinetic hormone stimulates proline synthesis in the fat body of the Colorado beetle. In addition, a release of glucose from the fat body of the beetle could be evoked by the addition of locust and beetle corpora cardiaca extracts or synthetic adipokinetic hormone. The physiological significance of both effects is discussed.     

Phylogenetic differentiation of glandular cells in corpora cardiaca as studied immunocytochemically with region-specific antisera to adipokinetic hormone

Immunocytochemical reactions with region-specific antibodies to adipokinetic hormone were used to study the stainability and structural organization of the intrinsic glandular cells of the corpora cardiaca of 19 insect species belonging to 10 orders. There was considerable variation in the nature of the glandular product as well as in morphology of the glandular cells, and in their association with the storage compartment of the corpus cardiacum. The apterygote Folsomia only contains substances reacting with the C-terminal specific antibody No. 241, whereas Thermobia has glandular cells which stain either with this serum or with the N-terminal specific antiserum No. 433. Glandular cells in Locusta may either react with only the N-terminal specific antiserum or with both types of antiserum. All other species studied contain only glandular cells reacting with the N-terminal specific antiserum 433, except for the dipteran species which were negative.It was attempted to relate the immunological staining with the chemical properties of the substances so far isolated from the corpora cardiaca. The glandular cells are variable in the number, shape, and the degree of clustering and association with the storage zones. In some species they are entirely separated, in others they are embedded in the storage zone. Cell processes may be very long or absent.Neurones in the brain were generally left unstained by serum 433, indicating that the substance revealed earlier by antiserum 241 differs from adipokinetic hormone.     

Neuroendocrine control of blood lipids in the locust, Locusta migratoria

In order to determine the best conditions, the influence of various parameters on the haemolymph lipid concentration were studied. These parameters are the age, the sex and the feeding of the animals, the time and the number of the haemolymph sample-taking and the temperature of the locust culture. A large in vivo increase in haemolymph lipid concentration was obtained in locusts which received extracts of the whole CC and of their glandular or neurohemal lobes. Reversely, a decrease in this concentration was obtained in locusts operated 7 days before (cardiacectomy or glandular lobe removal). Moreover the pars intercerebralis extracts increased the level of haemolymph lipids. We conclude that adipokinetic factors are present, both in the glandular lobes of the CC and in their neurohemal lobes. It is likely that the latter partly originate from the pars intercerebralis. Results of allatectomy and injections of corpora allata extracts led to the conclusion that corpora allata contain an adipokinetic factor, the juvenile hormone. and factors that inhibit the haemolymph lipid concentration. Finally, from different injections of neurotransmitters and drugs it is argued that it is mainly octopamine which is involved in the mechanism governing the increase of the level of haemolymph lipids.     

Locust corpora cardiaca contain an inactive adipokinetic hormone

A neuropeptide from the migratory locust, Locusta migratoria, has been identified as a novel member of the family of adipokinetic hormones (AKHs). The peptide is probably synthesised in the brain because it is the first AKH found in the storage lobe, whilst the three 'classic' Locusta AKHs are present in the glandular lobe of the corpora cardiaca. In locusts, the peptide has no biological activity usually associated with AKHs. There is only 36-56% sequence identity with the three Lom-AKHs, but 78% identity with the Drosophila melanogaster AKH, Drm-HrTH. The new peptide is active in the American cockroach, Periplaneta americana, and was provisionally named 'L. migratoria hypertrehalosaemic hormone', Lom-HrTH; its biological role in locusts remains to be established. The high degree of identity with Drm-HrTH suggests that Lom-HrTH is an ancient molecule.     

Synthesis of shrimp red pigment-concentrating hormone analogs and their biological activity in locusts

Two analogs of the red pigment-concentrating hormone (RPCH) have been synthesized by the solid-phase method: [Thr6]-RPCH (I) and [Tyr4, Thr6]-RCPH (II). Analog I has the same amino acid composition as the second adipokinetic hormone (AKH-II) isolated from locust corpora cardiaca. Bioassay for lipid-mobilizing activity in adult male locusts gave the following increases in hemolymph lipid content: AKH-I, 3.5; I, 2.4; II, 2.9. The biological response shown by I lends support to the conclusion that its sequence is that of the presumptive AKH-II. Replacement of Phe in position 4 by Tyr does not reduce the adipokinetic response.