Brain cyclic AMP levels and the initiation of adult development in the Cecropia silkmoth.

A six-fold increase in the level of brain cyclic AMP is observed in chilled Cecropia pharate adults within 24 hr of transfer from the cold to room temperature. This increase is not observed in pupae chilled for a period insufficient to allow initiation of adult development, nor after injury to diapausing pupae. Other tissues show a variable and minor response during initiation. Injected dibutyryl cAMP will cause initiation in insufficiently chilled pupae, but not in dauer pupae. The possible relationship of this rise in cAMP to the process of initiation is discussed.     

Haemolymph ecdysone concentrations in Hyalophora cecropia pupae,dauer pupae and adults

Haemolymph ecdysone concentrations were determined by radioimmunoassay in diapausing pupae, pharate adults, adults, and chilled dauer pupae. The concentration in diapausing pupae after 6 months chilling (5.35 pg/μl) increased dramatically after 3 days at 27°C (>200 pg/μl) and then decreased to low levels in adult females (1.63 pg/μl). In adult males ecdysone was undetectable in all except one animal. Dauer pupae showed a decrease from 6.1 to 1.7 pg/μl 1 day after being transferred from 6 to 27°C. Over a 3-day period the value increased to 3.19 pg/μl and remained constant for more than a year. These results suggest that diapausing pupae with and without brain neurosecretory cells maintain a low concentration of ecdysone in the haemolymph.     

Effects of caffeine and aminophylline on adult development of the Cecropia silkmoth

Caffeine and aminophylline were potent inhibitors of adult development in the moth, Hyalophora cecropia, when administered prior to initiation of pharate adult development. A dose of 1 mg/g live weight was 100 per cent effective in bringing about this arrest. Cyasterone and α-ecdysone induced development in arrested animals and arrested dauer pupae. Brain transplant and perfusion experments indicated that synthesis or release of brain hormone was disrupted by these drugs. In addition, the effects of ecdysone and perfusion on arrested animals suggested that methylxanthines may also act at other sites in the sequence of events leading to adult development.     

Induction of dauer pupae by fenoxycarb in the silkworm,Bombyx mori

Topical application of fenoxycarb (1 μg per animal) at 129 or 132 h of the fifth instar larvae of the silkworm, Bombyx mori, did not induce morphological abnormalities in the pupal stage, but these animals became dauer (permanent) pupae. This condition of B. mori and the endocrine events leading to permanent pupae are discussed in this work. Application of fenoxycarb at 132 h of the fifth instar elicited a high ecdysteroid titre in the pharate pupal stage and a steadily high ecdysteroid titre in the pupal stage. The fenoxycarb-induced permanent pupae had non-degenerating prothoracic glands that secreted low amounts of ecdysteroid and did not respond to recombinant prothoracicotropic hormone (rPTTH) late in the pupal stage. The Bombyx PTTH titre in the haemolymph, determined by a time-resolved fluoroimmunoassay, was lower than that of controls at the time of pupal ecdysis, but higher than controls later in the pupal stage in fenoxycarb-treated animals. After application of fenoxycarb, its haemolymph level, measured by ELISA, reached a peak at pupal ecdysis, then remained low. These results suggest that the fenoxycarb-mediated induction of permanent pupae is only partially a brain-centred phenomenon. It also involves alterations in the hormonal interplay that govern both the initiation of pupal-adult differentiation and changes in the steroidogenic pathway of the prothoracic glands of B. mori.     

Relationship between a metabolic rhythm and emergence rhythm in Drosophila melanogaster

Oxygen consumption and lactic acid dehydrogenase (LDH) activity were determined for Drosophila melanogaster pupae and pharate adults exposed to 12 : 12 or 1 : 23 light-dark (LD) regime. Bimodal circadian fluctuations of oxygen consumption were found in pupae and pharate adults exposed to either LD regime and organisms appeared to demonstrate an anticipatory change in oxygen consumption associated with change in illumination. The oxygen-consumption trend for the entire period spent in the puparium showed a high at the time of emergence, but the diurnal rhythm showed a low at the time of emergence suggesting that emergence occurs at a low in the diurnal cycle. Emergence maximum showed a 3 hr lead over the oxygen-consumption maximum. Changing the LD regime produced similar changes in the phasing of both oxygen consumption and emergence rhythms. LDH activity did not demonstrate a detectable circadian rhythm but did show a steady decrease during pupal and pharate adult development.     

Cell differentiation during the ontogeny of larval salivary glands of the fly, Telmatoscopus albipunctatus

Using the larvae, pharate pupa, and pharate adults of the moth fly, Telmatoscopus albipunctatus, histological and ultrastructural features of the salivary glands were investigated. The gland lumen contains a milky secretion from the first instar. This secretion continues to ccur at all subsequent developmental stages; with the onset of the pharate pupal stage, however, the secretion becomes transparent and rather viscous. Histochemical tests revealed that it is mainly proteinaceous. Glands from the same developmental stage may respond differently to PAS-reaction.Various cell organelles were compared at consecutive stages of larval development and of secretory activity of the salivary glands. In first and second instar larvae autophagic vacuoles are virtually absent in the salivary gland cells. They were occasionally found in the third instar, when they appear to be engaged in the process of organelle turnover. Histolysis of the larval glands is initiated towards the close of the fourth instar when the number of autophagic vacuoles starts to increase. Simultaneously, the cytoplasm, previously full of ribosomes and endoplasmic reticulum, starts losing these structures. At the beginning of the pharate adult stage, the cytoplasm becomes practically devoid of all structures other than those engaged in autophagy.Polyteny of the chromosomes during ontogeny of the larval salivary glands is also discussed.     

Effect of whole-body irradiation of pharate adult Ceratitis capitata on the activity of acetylcholinesterase during final development

Acetylcholinesterase activity has been studied during the development of Ceratitis capitata and under several γ-irradiation conditions. A slight and gradual increase takes place until the 4 day pharate adult from which the activity rises more sharply reaching the maximum level at 3 to 4 days after emergence. Irradiation (8·4 krad) of 4 day pharate adults reduced emergence and the levels of enzymatic activity, but its effect on the non-emerged pharate adults is very similar to that on emerged adult insects. Doses of 18 krad completely prevented emergence.     

Ecdysteroids and diapause in pupae of Heliothis punctiger

Most pupae of H. punctiger enter diapause when reared at 19°C, 12L:12D. When pharate pupae were treated for only 12 hr at 28°C about 50% developed at 19°C. The proportion of non-diapausing pupae increased as the temperature at which the pharate pupal stage was spent increased.The quantity of injected 20-hydroxyecdysone necessary to promote development in diapausing pupae varied from about 1 μg g^?1 soon after pupation to about 4 μg g^?1 after 50 days. It fell somewhat after 150 days.Removing brains from non-diapausing pupae showed that the brain secreted its hormone at the time of pupation (or just before). However, if the pupae were kept at 19°C development did not occur unless the brain remained in situ for at least 20 hr at 28°C. Implanting brains from non-diapausing pupae into diapausing ones had no measurable effect.These results may be explained by postulating that the prothoracic gland is ‘activated’ by exposure to high temperature, but that it reverts to inactivity over a period at 19°C. The ‘active’ gland must then be stimulated by brain hormone for a long period to trigger secretion of its hormone, which results in development. Diapause is thus the result of the failure of the prothoracic gland to secrete.     

Radiation biology and inherited sterility of light brown apple moth (Lepidoptera: Tortricidae): developing a sterile insect release program

The radiation biology of two geographically isolated populations of the light brown apple moth [Epiphyas postvittana (Walker)] was studied in Australia and New Zealand as an initiation of a SIT/F1 sterility program. Pharate and < or = 2 d pre-emergence pupae were exposed to increasing radiation doses up to a maximum dose of 300 Gy. Fertility and other life history parameters were measured in emerging adults (parental) and their progeny (F1-F3 adults). Parental fecundity was significantly affected by increasing irradiation dose in pharate pupae only. For both populations, parental egg fertility declined with increasing radiation. This was most pronounced for the irradiated parental females whose fertility declined at a higher rate than of irradiated males. At 250 Gy, females < or = 2 d preemergence pupae produced few larvae and no adults at F1. No larvae hatched from 250 Gy-irradiated female pharate pupae. At 300 Gy, males still had residual fertility of 2-5.5%, with pharate pupae being the more radio-sensitive. Radiation-induced deleterious inherited effects in offspring from irradiated males were expressed as increased developmental time in F1 larvae, a reduction in percent F1 female survival, decreased adult emergence and increased cumulative mortality over subsequent generations. Males irradiated at > or = 150 Gy produced few but highly sterile offspring at F1 and mortality was > 99% by F2 egg.     

Tolerance of pupae and pharate adults of Callosobruchus subinnotatus Pic (Coleoptera: Bruchidae) to modified atmospheres: a function of metabolic rate

Three developmental stages (pupae, early pharate and late pharate adults) of Callosobruchus subinnotatus (Pic.) were investigated for their tolerance or susceptibility to four modified atmospheres. Two of these atmospheres were hypercarbic and two were hypoxic. The hypercarbic atmospheres were found to cause mortality earlier than hypoxic atmospheres. Late pharate adults died earlier than pupae or early pharate adults. Late pharate adults that survived the exposure took a longer time to eclose than the pupae or early pharate adult.Using high resolution microrespirometric techniques, it was possible to record the oxygen consumption rate and CO(2) output of different developmental stages in air. The metabolic rate was determined manometrically as the oxygen uptake rate at an ambient temperature of 25 degrees C. The oxygen uptake rate differed significantly between groups of 20 individuals of different stages (p<0.01; t-test). The lowest rate of oxygen uptake (510.6+/-52.2 &mgr;l g(-1) h(-1)) was recorded in pupae. Higher oxygen uptake rates were found in early pharate adults (668.4+/-45.6 &mgr;l g(-1) h(-1)) and late pharate adults (1171.2+/-45.0 &mgr;l g(-1) h(-1)), and adult beetles (1310.4+/-53.4 &mgr;l g(-1) h(-1)). The patterns of CO(2) release were similar to those of oxygen uptake. CO(2) release was highest in eclosed adults and late pharate adults followed by early pharate adults, and lowest in pupae. The mode of CO(2) release ranged from continuous CO(2) release in pupae to discontinuous CO(2) release in late pharate and eclosed adults. Thus, high metabolic rates, and perhaps, in conjunction with discontinuous CO(2) of late pharate adults are responsible for their higher susceptibility to modified atmospheres than pupae and early pharate adults.     

The clearance of purines from the haemolymph of the cecropia silkmoth

The clearance of adenine, caffeine, guanine, theophylline, and xanthine from the haemolymph of diapausing pupae and pharate adults of Hyalophora cecropia was studied. In all animals caffeine and theophylline persisted, while the other purine bases were cleared within a few days.     

Morphogenetic effects of juvenile hormone and juvenile hormone mimics on adult development of Drosophila

The morphogenetic effects of t,t-farnesol, Law-Williams juvenile hormone analogue, dichlorofarnesenic acid ethyl ester (DFAEE), and a syntetic racemic or isomeric mixture of C₁₈ juvenile hormone (JH), when applied topically to pharate pupae and adults of D. melanogaster have been studied. Of these various agents tested, only DFAEE and JH affected adult development and eclosion and the pharate pupae were the most sensitive to these agents. The racemic mixture of JH induced the secretion, in the abdomen, of a supernumerary cuticle indistinguishable from that of the pupa; it, in addition, retarded the synthesis of brown eye pigments, general body pigmentation, and affected the differentiation of various internal organs and cuticular structures of the abdomen. By comparing the effects of JH with those of Minute (M) and bobbed (bb) mutations on the adult development, it is suggested that JH, by retarding genetic translation mimics M or bb.     

Disk electrophoresis fractionation of hemolymph proteins during the metamorphosis of different castes and sexes of Formica pratensis

Using polyacrylamide electrophoresis the proteins of the haemolymph of the different developmental stages can be separated into eight strong and nine weak coloured fractions during the cocoon period of Formica pratensis. The proteins were stained with aniline black and measured quantitatively by a Chromoscan densitometer. The values were compared with those maintained with bovine serum albumin.The total protein content of the haemolymph was calculated as the sum of the different fractions; at maximum it amounts to 2·1 per cent (w/v). The maximum is reached during the pharate pupal stage and during the pigmentation of the eyes; the minimum can be observed at the end of pupal ecdysis. At the beginning of body pigmentation in all the forms the protein content of the haemolymph was very much reduced, especially in workers and females.All fractions change independently resulting in a different composition of the haemolymph proteins in pharate pupae, eclosed pupae, and pharate adults. The slow-running fractions f1, f3, f5, and f6 and the mean bands f8 and f11 are reduced weakly until body pigmentation, and from the eleventh day strongly in both castes. All fractions are reduced during the cocoon period, but mostly the slow-running ones. Only the front band f14 increases to nearly twice that of the protein content. The importance of the changes in the protein fractions for development of different organs and for the synthesis of the haemolymph proteins and the influence of hormones are discussed.     

Larval dispersal and cannibalism in a natural population of Zophobas atratus (Coleoptera: Tenebrionidae)

A natural population, of the tenebrionid beetle Zophobas atratus was studied in an attic in El Zamorano, Honduras. Larvae were living in bat guano and were, densest in the deepest part of the guano. No pupae or pharate pupae were found in the guano: they were always located a small distance away from the guano pile. Adults occurred on the guano surface and the beams and tiles of the roof. When larvae of three different ages and weights were marked and released in the guano, only the older, larger larvae were recaptured away from the guano, indicating a correlation between larval dispersal and increased weight and/or age. When 50 pharate pupae were exposed to a natural population of active larvae, only 4 (8%) survived after 5 days. This intense cannibalism of pharate pupae is consistent with the hypothesis that inhibition of pupation by crowding and the associated pre-pupal dispersal evolved, at least partly, in response to cannibalism.     

Fatty acid profile of the cabbage looper, Trichoplusia ni, and the effect of diet and rearing conditions

The fatty acids, palmitic (16:0), palmitoleic (16:1), stearic (18:0), oleic (18:1), linoleic (18:2), and linolenic (18:3) were measured by GLC in Trichoplusia ni (Hübn.) from larvae reared on three different diets. The effect of rearing temperature and humidity on the levels of these fatty acids in pupae and pharate adults was determined. T. ni tended to accumulate 18:3 somewhat in proportion to its level in the diet. The respective levels (relative percentage) of this fatty acid in the diet and in the pupae were: bean leaves, 73·6 and 56·7; the diet based on lima beans, 10·2 and 3·5; and a meridic diet containing wheatgerm oil, 5·6 and 1·8. Pupae from larvae reared on a diet marginal in 18:3 content produced adults with deformed wings when reared at 30°C and normal wings when reared at 23 or 24°C, but there was no difference in their tissue level of 18:3. The phospholipids of last instar larvae, pharate pupae, pupae, pharate adults, and emerged adults are made up mostly of 18:0, 18:2, and 18:3 while the triglycerides of these stages contain relative large quantities of 16:0, 16:1, and 18:1. Pupal rearing temperature did not appear to influence the level of 18:3 in these two fractions enough to account for the degree of deficiency expressed as adult wing deformity. Both high (95%) and low (20%) relative humidity have an adverse effect on wing development but this effect can be overcome by 18:3 supplementation of the larval diet.     

Ecdysone titers and prothoracic gland activity during the larval-pupal development of Manduca sexta

The titer of ecdysone in whole animal extracts of Manduca sexta was determined by radioimmunoassay during the fifth (last) larval instar, pharate pupal development and pupation. A subtle peak in ecdysone concentration was noted at day 4 (just prior to the onset of the wandering stage) and a second and greater peak at day 8.5 (coincident with pharate pupal development). The titer fluctuations during development were a result of changes in tissue ecdysone and not of alterations in the ecdysone content of the gut. When prothoracic gland secretory activity was analyzed in vitro at the same stages, the most rapid rate of α-ecdysone secretion was shown to occur on day 7 (one day prior to the peak in whole-animal ecdysone concentration). An earlier peak in prothoracic gland activity may occur at day 4–5. Thin layer and gas-liquid chromatographic analyses revealed developmental changes in the ratio of β:α-ecdysone in hemolymph and whole-animal extracts. It is suggested that the steroid-hydroxylating capacity of the insect increases during the instar.     

Regulatory mechanisms in phenotypic plasticity of diapause and nondiapause pupal colouration of the swallowtail butterfly Papilio machaon

Nondiapause pupae of Papilio machaon L. exhibit pupal colour diphenism comprising green–yellow and brown–white types. To understand the regulatory mechanism underlying the control of pupal colouration in P. machaon, the effect of environmental cues on diapause and nondiapause pupal colouration is investigated. When larvae reared under short‐day and long‐day conditions are allowed to pupate in sites with a smooth surface and a yellow background colour, all diapause pupae exhibit a brown–white type and 89.5% of nondiapause pupae exhibit a green–yellow type, respectively. With rough‐surface pupation sites, all diapause pupae exhibit brown–white and intermediate types, whereas a large proportion of nondiapause pupae exhibit brown–white and intermediate types, although some exhibit a green–yellow type. When extracts prepared from the head‐thoracic and thoracic‐abdominal regions of larval central nervous systems are injected into the ligated abdomens of P. machaon short‐day pharate pupae, all recipients exhibit a brown–white colouration. Furthermore, when each extract is injected into the ligated abdomen of Papilio xuthus L. short‐day pharate pupae with orange‐pupa‐inducing factor activity, recipients injected with the head‐thoracic extract exhibit the brown type, whereas those injected with the thoracic‐abdominal extract exhibit an orange colour. The results indicate that the response to the environmental cues of pupation site in P. machaon changes according to the photoperiodic conditions experienced during larval stages, and that at least two hormonal factors producing brown–white pupae are located in the larval central nervous system, with the secretion of these factors being regulated by the recognition of environmental cues in long‐day larvae.     

Response of storage protein levels to variation in dietary protein levels

Storage proteins have been found to play a major role in insect metamorphosis and egg production and are accumulated during the actively feeding larval stage. Yet few studies have focused on how nutrition affects storage protein levels. Three storage proteins were identified in male and female Heliothis virescens pupae, one arylphorin and two putative high-methionine hexamers. Storage proteins were quantified in early pupae and in pharate adults. Storage protein levels peaked in 48-h pupae and were more abundant in females across all stages. Both male and female pharate adults retained a portion of total storage protein levels and females retained greater levels overall. In females, post-eclosion protein reserves will likely be used toward egg manufacturing, while the role of protein reserves in males remains speculative. In our previous study of H. virescens larvae, we found that protein-derived growth in females progressively increased as dietary protein levels increased. Our present data show that levels of storage protein also increased progressively along with dietary protein levels. This suggests that females allocated protein, in excess of adult tissue formation needs, toward storage protein. Our study is the first to demonstrate how responsive storage protein levels can be in face of varying levels of dietary protein.     

Ecdysteroid synthesis by testes of 5th instars and pupae of the European corn borer,Ostrinia nubilalis (Hubner)

Summary

Testes from young fifth instar Ostrinia nubilalis produced very small amounts of ecdysteroids while those from larvae that had purged their gut produced considerably more immunodetectable ecdysteroid in vitro. Larval testes that had fused produced 2.2 times more ecdysteroid than those that remained separate. It was the sheath of the testes rather than the contents that was physiologically active. Synthesis was questionable in testes from day-1 pupae and was not observed in testes from pharate pupae, from day-2 pupae or from pharate adults. Thus, synthesis only occurred at specific times in the life cycle. Ecdysteroid profiles for testes from wandering larvae whose testes had fused showed a net increase in all normally observed ecdysteroids, with the greatest increase being in 20-hydroxyecdysone. For testes from day-1 pupae, the nature of the ecdysteroid profile changed after 24 h of incubation, with some ecdysteroids showing increases and other decreases. There appear to be considerable differences among species regarding the times of testis synthesis and the amounts and nature of the ecdysteroids synthesized.     

Juvenile hormone in Drosophila melanogaster: Identification and titer determination during development

Juvenile hormone (JH) III was identified in whole-body extracts of eggs, larvae, pupae, pharate adults and adults of Drosophila melanogaster. Titers of the hormone varied according to stage of development. Highest levels were found in post-feeding (wandering) larvae and adults; only low levels were found in feeding last-stadium larvae and in pupae. None of the other known JHs were detected (limit of detection ≈0.01 ng/g). These results are discussed in the light of the known physiological roles of JH in the development of Drosophila and other insects.