Larval-pupal Apolysis Research Articles

High-Precision Calorimetry-Based Analysis of Pupal-Pharate Adult Development in Drosophila melanogaster.

In holometabolous insects, the larval body is almost completely decomposed and reconstructed into the adult body during the pupal-pharate adult stages. Therefore, the total energetic cost of this process is a key thermodynamic quantity necessary for evaluating the benefit of their life history. Here, we measured whole-body thermal dissipation of single pupae of the fruit fly, Drosophila melanogaster, during the period from puparium formation to adult eclosion as a function of age, using a high-precision isothermal calorimeter at T = 298 K. The mass-specific energy consumption during the period from the onset of larval-pupal apolysis to adult eclosion was determined to be 2.3 kJ/g for an individual of mass (adult) = 1.0 mg, while it was observed to follow Kleiber's law for individuals smaller than mass (adult) = 1.0 mg. During the pupal-pharate adult period, in addition to the U-shaped variation, several characteristic thermal dissipations related to various events, including somatic muscle contractions, ecdyses, pulsatile hormone secretion in a pharate adult, and vaporization of the exuvial fluid, were observed. The periodic bursts in the pharate adult stage grew exponentially, suggesting that the positive feedback in the metabolic system synchronized with the progression of development, making the energy consumption in this stage more efficient. The present study showed that high-precision calorimetry is a powerful and credible method for measuring not only the total energy spent during development but also the energy spent during every specific developmental event in an organism.

A Study of the Pupal Development of Five Forensically Important Flies (Diptera: Brachycera).

Holometabolous insects undergo complete metamorphosis, and hence, they have different phases of development (egg, larva, pupa, and adult), which occupy distinct ecological niches. The pupae of several fly species are surrounded by the puparium, which is a rigid structure, usually formed by the integument of the last larval instar. The puparium presents unique characteristics distinct from those of the larval and adult phases. During intrapuparial development, it is possible to distinguish at least four fundamental and continuous steps, namely: 1) larval-pupal apolysis, 2) cryptocephalic pupa, 3) phanerocephalic pupa, and 4) pharate adult. The objective of this work was to describe the external morphology of the distinct phase of development for five species that were collected, identified, and raised in the laboratory; intrapuparial development was studied by fixing immature specimens at regular intervals; the morphological analyses were performed with the aid of both light and scanning electron microscopy. Under the conditions established (27 ± 1.0 or 23 ± 1.0°C, 60 ± 10% relative humidity, 12 h of photoperiod), the minimum time for intrapuparial development was: 252 h for Megaselia scalaris (Loew 1966) (Phoridae), 192 h for Piophila casei (Linnaeus 1758) (Piophilidae), Fannia pusio (Wiedemann 1830) (Fanniidae), and Musca domestica (Linnaeus 1758) (Muscidae), and 96 h for Chrysomya megacephala (Fabricius 1794) (Calliphoridae). Intrapuparial development has defined steps, and distinct species responded differently to the same environmental conditions. In addition, it is possible to establish a sequential rule without ignoring the specific characteristics of each taxon.

First Insights into the Intrapuparial Development of Bactrocera dorsalis (Hendel): Application in Predicting Emergence Time for Tephritid Fly Control.

Intrapuparial development is a special pattern of metamorphosis in cyclorrhaphous flies, in which the pupa forms in an opaque, barrel-like puparium. This has been well studied in forensic insects for age estimations. In this study, the intrapuparial development of a quarantine agricultural pest, Bactrocera dorsalis (Hendel), was studied under a constant temperature of 27 ± 1 °C and 70 ± 5% relative humidity. Results showed that intrapuparial development could be divided into five stages: Larval-pupal apolysis, cryptocephalic pupa, phanerocephalic pupa, pharate adult, and emergent adult. It lays a morphology-based foundation for molecular mechanism studies and enhances the understanding of the physiological basis for changes in intrapuparial development. More importantly, the chronology of intrapuparial development can be used to predict the emergence time of tephritid flies, indicating when to spray insecticides to control these phytophagous agricultural pests. This may be an effective approach to reduce the use of insecticides and slow down the evolution of insecticidal resistance.

Chronology of the Intrapuparial Development of the Blowfly Chrysomya albiceps (Diptera: Calliphoridae): Application in Forensic Entomology.

Insects display different patterns of development, and blow flies have one of the most specialized patterns of intrapuparial development of all. In forensic entomology, pupae can be used as a tool to estimate the minimum postmortem time interval (minPMI). We analyzed the intrapuparial development of Chrysomya albiceps (Diptera: Caloricidade), whose larvae had been fed pig lungs and reared in a climate-controlled room at 28°C day/26°C night, 70 ± 10% RH, and 12 h of photophase and monitored daily. After the third-instar larvae abandoned their diet, the process of pupariation and pupation was monitored. At pre-established times, five pupae were collected, euthanized, and fixed in 5% formaldehyde, inside polypropylene test tubes with caps. Since they were the first, they were classified as 0 h pupae. Twelve collections occurred until the emergence of the adults, at 0, 2, 4, 6, 8, 10, 24, 30, 48, 54, 72, 78, 96, and 99 h (n = 84). The fixed pupae were dissected under the microscope, with the aid of anatomical tweezers and hypodermic needles, and photographed. The stages of metamorphosis and the morphological alterations occurring during the process were identified, described, and recorded before and after pupation. These phases were: pupation, larval pupal apolysis, cryptocephalic, phanerocephalic, pharate adult, emergence, and adult. The cryptophalic phase occurred between 4 and 6 h after pupation; the phanerocephalic phase between 6 and 10 h after; the pharate adult phase between 24 and 96 h after; and the imago/emergence phase 99 h after pupation.

Testicular maturation in the sheep bot fly Oestrus ovis.

The process of testicular maturation in relation to intrapuparial development was studied in the sheep nasal bot fly, Oestrus ovis L. (Diptera: Oestridae). After formation of the puparium during larval-pupal apolysis and the cryptocephalic pupal stage (approximately 24-72 h), spermatogonia had undergone mitotic divisions and sperm cysts had been formed. Five days after pupariation, spermatogonia transformed into primary spermatocytes during the phanerocephalic pupal stage, and secondary spermatocytes first appeared during the pupal-adult apolysis. Secondary spermatocytes began undergoing the second meiotic division by day 8 (transparent-eye pharate adult stage). By days 9 and 10, round spermatids were present and began to elongate by day 11. By day 12, the first bundles of tailed spermatozoa had appeared. By day 15 (the yellow-orange eye pharate adult stage), round, elongated, tailed and bundled spermatids were predominant and by day 17 differentiating spermatids occupied nearly 35% of the testicular cavity, and 60% was occupied by free sperm. By day 21 (the red-brown eye pharate adult stage), spermatozoa colonized the seminal vesicle. At emergence (approximately day 22), a complement of free sperm occupied the testis and the seminal vesicle, but groups of developing cells frequently remained in certain zones. Spermatogenesis was carried out after pupariation and spermiogenesis occurred during the pharate adult stage. After emergence, males possessed fully formed spermatozoa ready for ejaculation.

Intra-puparial development in Oestrus ovis (Diptera: Oestridae).

The phenology of intrapuparial development in Oestrus ovis L. is described, based on 302 specimens collected from the head cavities of goats and reared in the laboratory at a photoperiod of 12:12 (L:D) h and 32 and 16 degrees C. Dissection and histology of puparia at pupariation and at 3, 6, 9, 12, 15, 18, 21, 24, 30, 36, 42, 48, 66, and 72 h after pupariation and every day of the intra-puparial period showed that pupariation was achieved in approximately 12 h in heavily pigmented larvae (range, 2-46 h in postfeeding period). Larval-pupal apolysis began immediately after pupariation and was completed by 18-36 h after pupariation (prepupal period). The cryptocephalic pupa was found from this time to the 5th d, when head eversion occurred. Pupal-adult apolysis was initiated before head eversion and completed by day 7. The pharate adult presented progressive coloration in compound eyes (transparent, white, yellow, orange, red, brown, silver) while integumental pigmentation and sclerotization were in progress. Adult emergence occurred at 22 and 23 d in males and females, respectively. Changes in the weekly puparial weight of specimens reared under both field and laboratory conditions was described. It was concluded that although the intra-puparial development of O. ovis displayed some unique characteristics, it was essentially similar to other cyclorraphous flies. The actual pupal period of O. ovis lasted from the 2-7 d post-pupariation, whereas approximately two-thirds of the intra-puparial period was used for the maturation of the pharate adult.

Changes of acid phosphatase content and activity in the fat body and the hemolymph of the flesh flyNeobellieria (sarcophaga) bullata during metamorphosis

Changes in the specific and total activity of the lysosomal marker enzyme acid phosphatase (Acph) and in the amount of enzyme protein were examined in the fat body and the hemolymph from the last larval molt to the larval-pupal apolysis. The specific activity showed minor changes during the last larval period. In contrast, the total activity of the enzyme was low during the feeding period and higher during the wandering stage and strikingly increased at the time of puparium formation. We purified a protein having para-nitrophenyl phosphate phosphatase (Acph) activity and raised antisera against it. The amount of Acph protein in the fat body and hemolymph was examined using an ELISA. The specific Acph content showed little variation, but the total amount of the enzyme protein showed a stepwise increase in both organs during last larval stage and was markedly elevated in the pupal stage in the fat body. In contrast, a considerable decrease in the amount of Acph protein was observed in the hemolymph during this period. These data were in agreement with immunohistochemical observations showing an accumulation of the enzyme protein in fat body cells during the prepupal stage with a concomitant disappearance of the enzyme from the hemolymph. Inhibition of ecdysteroid secretion by water stress prevented the changes both in total enzyme activity and in the amount of Acph protein. However, Acph protein content and enzyme activity could be restored when the water stress was followed by a 20-hydroxyecdysone (20-HE) treatment. Taken together, our data show that Acph is secreted by fat body cells into the hemolymph during the larval stage, where it is stored in an inactive form. Increase in the 20-HE titer at the end of last larval stage reverses this process, and the enzyme is taken up by the fat body cells, where it becomes activated and appears in auto- and heterophagic vacuoles. Arch. Insect Biochem. Physiol. 34:369–390, 1997. © 1997 Wiley-Liss, Inc.

Changes of acid phosphatase content and activity in the fat body and the hemolymph of the flesh fly Neobellieria sarcophaga bullata during metamorphosis

Changes in the specific and total activity of the lysosomal marker enzyme acid phosphatase (Acph) and in the amount of enzyme protein were examined in the fat body and the hemolymph from the last larval molt to the larval-pupal apolysis. The specific activity showed minor changes during the last larval period. In contrast, the total activity of the enzyme was low during the feeding period and higher during the wandering stage and strikingly increased at the time of puparium formation. We purified a protein having para-nitrophenyl phosphate phosphatase (Acph) activity and raised antisera against it. The amount of Acph protein in the fat body and hemolymph was examined using an ELISA. The specific Acph content showed little variation, but the total amount of the enzyme protein showed a stepwise increase in both organs during last larval stage and was markedly elevated in the pupal stage in the fat body. In contrast, a considerable decrease in the amount of Acph protein was observed in the hemolymph during this period. These data were in agreement with immunohistochemical observations showing an accumulation of the enzyme protein in fat body cells during the prepupal stage with a concomitant disappearance of the enzyme from the hemolymph. Inhibition of ecdysteroid secretion by water stress prevented the changes both in total enzyme activity and in the amount of Acph protein. However, Acph protein content and enzyme activity could be restored when the water stress was followed by a 20-hydroxyecdysone (20-HE) treatment. Taken together, our data show that Acph is secreted by fat body cells into the hemolymph during the larval stage, where it is stored in an inactive form. Increase in the 20-HE titer at the end of last larval stage reverses this process, and the enzyme is taken up by the fat body cells, where it becomes activated and appears in auto- and heterophagic vacuoles. Arch. Insect Biochem. Physiol. 34:369–390, 1997. © 1997 Wiley-Liss, Inc.

The onset of metamorphosis in Tenebrio molitor larvae (Insecta, Coleoptera) under grouped, isolated and starved conditions

The post-embryonic development of the beetle Tenebrio molitor presents a variable number of larval instars. Several parameters (instar length, time of apolysis and cuticle deposition) were compared during the larval-larval and larval-pupal cycles of mealworms over 50 mg, reared in grouped or isolated conditions. In grouped conditions comparable to mass breeding, larval-larval and larval-pupal apolyses were found to occur at the same time, but instar duration was longer in the case of prepupae. However, isolation was found to accelerate larval-pupal (but not larval-larval) apolyses and to reduce the number of larval instars, whereas starvation inhibited larval-larval (but not larval-pupal) development by delaying or preventing larval apolysis. Our observations confirm that the onset of metamorphosis is linked to a loss of sensitivity to starvation and to an increased requirement of isolation. In isolated conditions, larval-pupal apolysis was found to be preceeded by a characteristic behaviour (transitory period), homologous to the wandering stage of Lepidoptera. Ecdysteroid titres, measured with a radioimmunoassay, showed a very different pattern in a larval-larval cycle (a single high peak beginning at apolysis) compared to a larval-pupal one (several smaller peaks, the first one during the transitory period). The involvement of ecdysteroids and other hormones in the control of developmental variability is discussed.

Ecdysteroid titres and integument changes in superparasitized puparia of Anastrepha suspensa (Diptera: Tephritidae)

Larval-pupal apolysis of Anastrepha suspensa is inhibited when superparasitized (up to 7 parasites/host) by the solitary endoparasite Biosteres longicaudatus. Ecdysone 20-monooxygenase activity was similar in superparasitized hosts and controls and could not explain the significantly ( P < 0.05) high ecdysteroid titres, especially in 36–48-h-old hosts. In the control, synthesis of the outer and inner epicuticules coincided with a significantly ( P < 0.01) low ecdysteroid titre. Presumably, an inhibitory effect of high ecdysteroid titres and the abnormal proliferation of microvilli at the cuticle-epidermis interface led to the inability of superparasitized hosts to synthesize new cuticle. Moreover, the migration and exocytosis of vesicles observed at the cell apices in the control did not occur in superparasitized individuals which did not apolyse. Viral particles, presumably of the rhabdovirus type (300 × 80 nM) proliferated in large vesicles bordering the basement membrane of epidermal cells in superparasitized puparia. These viruses may have affected the migration of vesicles to the cell apices and the activation of the moulting fluid. Pox virus-like particles (240 nM in diameter) were also observed in haemocytes adjacent to the epidermal cells. Superparasitized A. suspensa were unable to initiate larval-pupal apolysis due presumably, to: (a) their inability to secrete moulting gel and (b) high ecdysteroid titres that may also have inhibited the synthesis of a new cuticle.

The role of 20-hydroxyecdysone in the moulting of Biosteres longicaudatus, a parasite of the Caribbean fruit fly, Anastrepha suspensa

Ecdysteroid levels in whole body extracts of parasitized and unparasitized Anastrepha suspensa peaked 24 h after initial puparium formation. Six to eight-hour old first instars of the parasite, Biosteres longicaudatus, dissected from the host 16–18 h prior to its ecdysteroid peak never moulted in 20-hydroxyecdysone free media, while > 15% removed 6–12 h afterwards (i.e. at 24–36 h old) did. Isolation of 6–8-h old parasites within ligated abdomens of pre-wandering host larvae inhibited the parasites' moult. However, topical applications of 20-hydroxyecdysone (1.3 μg) to each host abdomen 24 and 48 h after ligation elicited host pupariation and larval-pupal apolysis as well as parasite moulting. Pre-treatment of host larvae with 2.7 μg/g hydroprene prior to parasitization and ligation prolonged host larval life. Subsequent 20-hydroxecdysone therapy of these hydroprene-treated, ligated abdomens increased parasite moult above that of abdomens receiving 20-hydroxyecdysone treatments alone. Six to eight-h old parasite first instars incubated for 4 days in 20-hydroxyecdysone-free medium increased in length by 78.3% and in head-capsule width by 23.8%. After a 24-h pulse of 10 −6 M 20-hydroxyecdysone only those that exceeded 1400 μ [X̄ = 14.1 ± 0.9 μ (SD)] in length and 250 μ in headcapsule width, moulted. A comparison of parasite growth in vivo and in vitro suggested that size may also affect moulting competence.

Ecdysteroid levels and integument changes in post-embryonic stages of Anastrepha suspensa

Ecdysteroid levels in larvae and pupae of Anastrepha suspensa (Diptera: Tephritidae) were measured by radioimmunoassay. These levels were correlated with histological changes throughout the development of the post-embryonic stages. Ecdysteroid levels increase rapidly throughout the last-larval instar and on the last day of this stage are 283 pg equivalents of 20-hydroxyecdysone per insect (14.5 ng/g) when wandering behaviour is initiated. At the end of this period the puparium is formed and within 24 h, the ecdysteroid rises to its highest peak (625 pg equivalents of 20-hydroxyecdysone/insect). Larval-pupal apolysis is initiated within 24 h later and the pupal cuticle is then secreted. Two days later, the ecdysteroids reach their lowest levels (75 pg equivalents of 20-hydroxyecdysone/insect or 0.6 ng/g) and most of the larval fat body and other tissues have been histolysed. In 5- to 10-day old pupae ecdysteroid levels again increase and remain relatively high throughout. During this period the larval epidermis is replaced by imaginal epidermis, imaginal discs begin to proliferate and the adult cuticle is secreted. Ecdysteroid levels finally fall 2 days prior to adult emergence. HPLC determinations indicate that 20-hydroxyecdysone is the predominant free ecdysteroid, and along with ecdysone, is readily detectable in all postembryonic stages of this species. An unusually high and unexplained peak of 20-hydroxyecdysone occurs in the pharate adult. This peak probably consists of ecdysone metabolites with retentions similar to that of 20-hydroxyecdysone and to which the antiserum is sensitive.

Timing of the Action of Photoperiod and Temperature On Events Leading to Diapause and Development in Pupae of Heliothis Punctigera (Lepidoptera: Noctuidae)

ABSTRACT Diapause in H. punctigera occurs in the pupa at some stage between pupation and the pupal-adult apolysis. Its occurrence is induced by 12 L: 12 D photoperiod, which is specifically effective during the last half of larval life, and by low temperature (19 °C). Irrespective of the conditions during larval life, the insects do not diapause if subjected to high temperature (28 °C) after feeding ceases and before the pupal ecdysis. The insects are most sensitive to high temperature during the short perioid between the larval-pupal apolysis and pupal ecdysis. At both 28° and 19 °C the brain and thoracic glands secrete almost simultaneously to trigger the larval-pupal apolysis, which occurs almost immediately after the release of the hormones. High temperature during the prepupal period results in an increased rate of development from pupal ecdysis to emergence of the adult.

Cuticular cycle and molting hormone levels during the metamorphosis of Tenebrio molitor (Insecta Soleoptera)

The cuticular cycle of Tenebrio molitor (apolysis, synthesis of outer and inner epicuticle, fibrous cuticle deposition) was studied during the last larval and pupal stages by electron microscopy. Concurrently, molting hormone (MH) titers in the hemolymph were determined by a radioimmunoassay method. It appears, both in larvae and in pupae, that the MH peak coincides with the initiation of pre-ecdysial cuticle deposition (i.e., outer epicuticle synthesis). Thus MH is involved in the induction of cuticular synthesis; however, its role in inducing larval-pupal apolysis is questionable. We note that this peculiar apolysis occurs long before MH release.

Changes in ecdysone content during postembryonic development of the wax moth, Galleria mellonella: The role of the ovary

The ecdysone titers of representative developmental stages during the life cycle of Galleria mellonella were determined by radioimmunoassay. During the last larval instar two ecdysone peaks are present, the first peak occurring relatively late in the instar concurrent with the initiation of wandering and the second separated from the first by only 1.5 days. The second increase in ecdysone titer marks larval-pupal apolysis and is only slightly larger than the first peak in contrast to studies on other Lepidoptera. Two days after pupation a large ecdysone peak occurs denoting the beginning of pharate adult development and the titer declines thereafter. In female pharate adults, but not in males, the ecdysone titer rises to the maximum for the life cycle at about the time of adult eclosion and then declines slowly during the next 4 days. When larvae are gonadectomized, the resulting females no longer yield the ecdysone peak found under normal conditions at about the time of pupal-adult ecdysis. Extraction of ovaries from late pharate adult G. mellonella and analyses of ovarian in vitro secretory kinetics demonstrate unequivocally that the developing ovary synthesizes the ecdysones characterizing the female at the time of adult emergence. Further studies revealed that most of the ovarian ecdysones are associated with mature oocytes and are probably synthesized by the nurse and/or follicle cells. The ovarian ecdysoones are a mixture of α- and β-ecdysone and another molecular species tentatively identified as 2-deoxy-α-ecdysone.

Epidermal cell development and control of cuticle deposition during the last larval instar of Manduca sexta

The development of the epidermis and the control of cuticle deposition in last instar larvae and pharate pupae of Manduca sexta were investigated by light microscopy and in vitro techniques. On day 1 after ecdysis to the fifth instar the epidermis exhibits cuboidal morphology, but columnar morphology is soon manifested with the onset of cell growth. At the time of epidermal “reprogramming” from larval-pupal syntheses (day 3), intercellular spaces appear which persist throughout the remainder of the instar. Calculations of cell density and nuclear diameter suggest that a mitotic wave occurs between days 5 and 6; the remainder of the instar is marked by slow cellular growth. Secretion of larval cuticle continues until the time of larval-pupal apolysis, but the greatest rate of secretion occurs throughout days 2–4. In vivo and in vitro experiments indicate that the brain and fat body play a permissive role in the secretion of cuticle. Evidence is presented which suggests that the brain may elicit the release of cuticular precursors or regulatory agents from the fat body. These findings are compared to previous descriptions of epidermal development in other species and discussed in light of the “quantal mitosis” theory of cellular differentiation.

Biochemistry of development in insects. Incorporation of fatty acids into different lipid classes.

1. To study the different metabolic behaviour of various stages of development of the insect Ceratitis capitata, the incorporation of labelled decanoic, lauric, myristic, palmitic, stearic, oleic, and linoleic acids into triacylglycerols by insect homogenates was investigated. The time-course of incorporation of labelled fatty acids was firstly studied by using oleic acid; it showed that after 10 min of incubation the levels of radioactivity incorporated into triacylglycerols and those remaining in the free fatty acids were practically unchanged. 2. All labelled fatty acids were efficiently incorporated by larval homogenates; however, most of the radioactivity remained as free fatty acids in the presence of pharate adult homogenates, palmitic, and stearic acids being the most scarcely incorporated by this stage of development of the insect. 3. Plots of triacylglycerol and free fatty acid radioactivites versus the stage of development defined a crossing-zone in coincidence with the larval-pupal apolysis. This metabolic difference between larval and pharate adult homogenates could not be explained through differences in the acyl-CoA synthetase activity of the insect; this enzyme activity was notably higher in pharate adult homogenates than in the larval homogenates whatever would be the nature of the fatty acid. 4. [14C]Triolein was scarcely hydrolyzed by both larval and pharate adult homogenates. 5. Double-label experiments were carried out by incorporating either [3H]oleic acid or [3H]-palmitic acid and [14C]glycerol 3-phosphate by larval and pharate adult homogenates at different incubation intervals. Diacylglycerols, triacylglycerols, and phosphoglycerides were isolated and the 14C/3H molar ratio calculated. Results suggest the existence of a different acyltransferase activity in the different stages of development of the insect.

Effect of a juvenile hormone analogue on phosphatase activity in pupae of the stable fly, Stomoxys calcitrans

The alkaline phosphatase activity of stable fly pharate pupae treated with 10 ng of an insect juvenile hormone analogue (JHA), Stauffer R-20458, or untreated pupae was rhythmic and peaked 48 and 96 hr after larval-pupal apolysis. Those treated with 10 μg were not rhythmic and peaked at 48 and 120 hr. Acid phosphatase activity showed a general increase throughout pupal-adult transformation and was three- to four-fold higher than alkaline phosphatase activity. At 24 and 48 hr after larval-pupal apolysis, acid phosphatase was lower in the treated animals, but at 72 and 120 hr, it was higher. Neither inhibition nor enhancement of acid and alkaline phosphatase activity or by three other JHAs could be demonstrated in vitro.

Cyclic AMP levels in Drosophila during postembryonic development and in the adults

The physiological content of Drosophila melanogaster tissues in cyclic AMP has been determined and its variations studied during postembryonic development and in the adults. Marked variations were observed, especially during metamorphosis where the ratio between the lowest and highest values (0·35 to 17·25 pmoles/mg protein) was 1 44 . In larvae the variations of cyclic AMP level were not clearly related to the larval ecdyses, but the steps of metamorphosis, i.e. formation of the puparium, larval-pupal apolysis, and pupal-adult apolysis, were accompanied with rapid and drastic rises of cyclic AMP, up to the highest value mentioned. We therefore deduce that cyclic AMP is involved in the metamorphosis of D. melanogaster as a chemical signal. In adults, the cyclic AMP level was remarkably constant and was around 7 pmoles/mg protein.

Incorporation of [ 32P]phosphate, [ 14C]acetate, and [ 3H]glycerol into phospholipids of Ceratitis capitata larvae. Turnover of labelled phospholipids during metamorphosis

Biosynthesis of phospholipids during metamorphosis of Ceratitis capitata has been investigated using disodium [ 32P]orthophosphate, sodium [ 14C]acetate, and [ 3H]glycerol as labelled precursors. Specific activities were determined in phosphatidylcholine, phosphatidylethanolamine, their lysoderivatives, and phosphatidylserine, obtained from the developmental stages of larva, larval-pupal apolysis, pharate adult, and emerged adult. The isotope incorporation into phospholipids of larvae exhibits the highest specific activities in phosphatidylserine and phosphatidylcholine. However, a clear tendency to equalize the values of the three main phospholipids is shown during metamorphosis of the insect, which might suggest the existence of reactions of base conversions.