Thomas J. Kelly

The effects of nutrition and methoprene treatment on ovarian ecdysteroid synthesis in Drosophila melanogaster

Radioimmunoassay of in vitro culture medium from ovaries of Drosophila melanogaster indicates that detectable ovarian ecdysteroid synthesis begins between 6 and 12 h after eclosion and reaches a peak between 24 and 30 h, when animals are reared at 25°C, 12 h photophase. Analysis of 24 and 72 h medium by a combination of high-performance liquid chromatography and radioimmunoassay demonstrates three ecdysteroid regions, two comigrating with known standards of ecdysone and 20-hydroxyecdysone and a third highly polar region containing one or more unidentified radioimmunoassay-active ecdysteroids. In 72 h medium the polar region comprises the majority of radioimmunoassay-active material while in 24 h medium the majority is in the ecdysone region. Provision of a nutritionally deficient diet to females at adult eclosion prevents the normal increase in vitellogenic-stage follicles and ovarian ecdysteroid synthesis. Methoprene treatment of such females stimulates a transient burst of ovarian ecdysteroid synthesis and the production of near normal numbers of vitellogenic oocytes by 24 h, although by 48 h the number of vitellogenic oocytes is less than normal.

Ecdysteroid titers and molting aberrations in last-stage Oncopeltus nymphs treated with insect growth regulators

Abstract The ecdysteroid titers of insect growth regulator (IGR)-treated and untreated large milkweed bug nymphs, Oncopeltus fasciatus (Dallas) (Heteroptera: Lygaeidae), were determined by radioimmunoassay. All of the IGRs tested affected the ecdysteroid levels and the ecdysis of both sexes in some way. AI3-63604, a juvenile hormone mimic, accelerated ecdysteroid production and its subsequent decline, shortened the stadium, and induced a supernumerary nymphal molt. Diflubenzuron had no effect on the onset of ecdysteroid production but slightly retarded the decline in ecdysteroid titers before adult ecdysis, and the ecdysis was incomplete. Ecdysteroid production was delayed and erratic in nymphs treated with azadirachtin and AI3-63967, but the effect on ecdysis was different for each compound. Azadirachtin caused incomplete adult ecdysis, whereas AI3-63967 completely prevented any attempts at ecdysis. The large peak of exdysteroid activity associated with apolysis was absent in starved insects, and they made no attempt to ecdyse.

Juvenile hormone and ovarian maturation in the diptera: A review of recent results

Abstract The roles of the JHs and ecdysteroids in ovarian maturation of adult Aedes aegypti, A. atropalpus, Musca domestica and Drosophila melanogaster have been characterized by comparing the effects of surgical (allatectomy, ovariectomy, decapitation, abdominal ligation), genetic, and nutritional (sugar feeding) manipulations. The results show that in all species JHs, or their mimics, and 20-hydroxyecdysone act in combination to stimulate ovarian maturation and vitellogenin (Vg) synthesis and that high doses of exogenous JHs, or their mimics, stimulate ovarian ecdysteroid synthesis, at least in A. aegypti, A. atropalpus, and D. melanogaster. Thus the gonadotropic regulatory mechanisms that exist in various dipteran species are more similar than originally suspected. In M. domestica, and possibly D. melanogaster, 20-hydroxyecdysone is present in the hemolymph at vitellogenic levels in newly emerged females and may persist in ovariectomized adults. If true for D. melanogaster, this could explain why topical application of JH, or its mimics, to ovariectomized, isolated abdomens is effective in stimulating Vg synthesis in the absence of the ovaries.

Hormonal regulation of ovarian ecdysteroid production in the autogenous mosquito, Aedes atropalpus

The effects of juvenile hormone (JH) and egg development neurosecretory hormone (EDNH) on ovarian ecdysteroid production during vitellogenesis in the autogenous mosquito, Aedes atropalpus, were investigated using in vitro techniques coupled with radioimmunoassay (RIA) and high-pressure liquid chromatography (HPLC). Normal females were characterized by quantitative, qualitative, and temporal patterns of in vitro ovarian ecdysteroid production. Females decapitated at emergence showed little ovarian ecdysteroid production and did not undergo vitellogenesis. A 500-ng dose of JH-I applied topically to decapitated females restored normal patterns of ecdysteroid production. In both normal and experimental females, ecdysone constituted the major portion of the ecdysteroids secreted by the ovaries in vitro. However, significant amounts of other RIA-active materials were detected, one of which was probably 20-OH-ecdysone. Fat body incubations indicated that these tissues produce little RIA-active material during the peak of vitellogenesis. During that period, the ovaries were the major source of ecdysteroid. Various doses of JH-I, applied to abdomens isolated at emergence, enhanced ovarian responsiveness to subsequent applications of head extracts containing EDNH both in vivo and in vitro. A 500-ng dose of JH-I, applied topically to blood-fed, decapitated Aedes aegypti, stimulated a significant increase in in vitro ovarian ecdysteroid production. Similarities between these data and those demonstrating prothoracicotropic effects of JH in the Lepidoptera are discussed.

Makisterone A: its distribution and physiological role as the molting hormone of true bugs

Makisterone A, a 28-carbon (C-24 alkyl) hexahydroxy steroid, has been identified by mass spectrometry as the major ecdysteroid in last-stage larvae of the large milkweed bug,Oncopeltus fasciatus, a phytophagous hemipteran. Similarly, it is a major molting hormone in 2 phytophagous and 1 predacious species of Hemiptera belonging to the group, Pentatomomorpha. It is not, however, a major ecdysteroid in another group of Hemiptera, the Cimicomorpha, where 1 predacious and 2 hematophagous species contain ecdysone and 20-hydroxyecdysone as their major molting hormones.

Larval moulting hormone of trichophoran Hemiptera-Heteroptera: Makisterone A, not 20-hydroxyecdysone

Abstract The haemolymph ecdysteroids were examined in fifth-stage larvae of Nezara viridula, Podisus maculiventris and Dysdercus cingulatus (Hemiptera-Heteroptera) using high-pressure liquid chromatography to separate the ecdysteroids and a radioimmunoassay to detect the fractionated ecdysteroids. The length of the fifth stage ranged from 5 to 8 days, and a peak in ecdysteroid titre (1700–2650 ng/ml) occurred 2–3 days prior to ecdysis to the adult. An ecdysteroid matching the retention time of makisterone A (24-methyl-20-hydroxyecdysone) was clearly present in haemolymph taken at the time of peak titre in all 3 of these true bugs, whereas little, if any, ecdysone or 20-hydroxyecdysone was detected. These data, along with previously reported data for the milkweed bug Oncopeltus fasciatus, are persuasive evidence that makisterone A is the larval moulting hormone of a group of closely related Heteroptera called the Trichophora (Lygaeoida, Pentatomoidea, Pyrrhocoroidea and Coreoidea).

Ecdysteroid fluctuations in adult Drosophila melanogaster caused by elimination of pupal reserves and synthesis by early vitellogenic ovarian follicles

Abstract Radioimmunoassay (RIA) of whole body extracts of Drosophila melanogaster males and females demonstrates that at eclosion all individuals contain high levels of ecdysteroid. Highly polar ecdysteroids (presumably metabolites) in the meconium represent approximately half of the total ecdysteroid RIA-activity present at this time and are subsequently eliminated. Ecdysteroids remaining after the elimination of the meconium are also highly polar as shown by reverse-phase high pressure liquid chromatography (RP-HPLC). The amount of ecdysteroid RIA-activity found in whole body extracts declines in both sexes until 18 h post-eclosion when levels begin to increase in the female and drop to undetectable levels in the male. In the female the ovaries are the major source of ecdysteroid. The increase in whole body ecdysteroid in the female coincides with the initiation of ovarian ecdysteroid production and accumulation. Topical application of methoprene, a juvenile hormone (JH) analog, stimulates ovarian ecdysteroid synthesis in apterous-ts 78j (ap ts 78j ) , a temperature-sensitive juvenile hormone-deficient mutant, corroborating previous results suggesting a role of juvenile hormone in ovarian ecdysteroid production. Stage 8–9 follicles, whose development is juvenile hormone dependent, are shown to be the most active in ecdysteroid production. The regulatory potential of these stages is discussed.

Inhibitory effects of oostatic hormone on ovarian maturation and ecdysteroid production in diptera

Extracts from post-vitellogenic ovaries of Musca domestica, Aedes atropalpus and Drosphila melanogaster were examined for the ability to inhibit the vitellogenic phase of growth in newly emerged A. atropalpus and D. melanogaster adult females. All extracts were inhibitory in A. atropalpus. D. melanogaster females were less sensitive than A. atropalpus females, only showing inhibition at a two-fold greater concentration of M. domestica extract. The titre of oostatic hormone in A. atropalpus ovaries correlated with the decline in ecdysteroid synthesis by these ovaries. A. atropalpus ovaries containing post-vitellogenic follicles were shown to inhibit ovarian ecdysteroid synthesis in vitro by vitellogenic ovaries of both A. atropalpus and D. melanogaster. D. melanogaster ovaries were not capable of eliciting such an inhibition in vitro, at least with the numbers of ovaries utilized in our experiments.

Relationship between the corpus cardiacum-allatum complex and ovaries with the haemolymph ecdysteroid profile in the housefly, Musca domestica☆

Abstract Haemolymph from female houseflies at 4, 48 and 96 h after emergence contained approximately equivalent amounts of 20-hydroxyecdysone that varied from 2.9 to 4.1 pg/μl. Polar ecdysteroids decreased with the insect age from 12.9 pg/μl at 4 h to 5.1 pg/μl at 96 h. At 48 h the haemolymph contained ecdysone and 20,26-dihydroxyecdysone in addition to 20-hydroxyecdysone and the polar materials. When ovaries were culture in vitro , the culture medium contained ecdysone as the major ecdysteroid (70%) with lesser concentrations of 26-hydroxyecdysone, 20-hydroxyecdysone, and 20,26-dihydroxyecdysone. Haemolymph from ovariectomized flies contained 20-hydroxyecdysone, some polar materials, and very little ecdysone. Thus, it appears that the ovaries are the source of ecdysone. Since all samples contained 20-hydroxyecdysone, its source is not known. Removal of the corpus allatum-cardiacum complex decreased the amounts of all ecdysteroids in the haemolymph sample. Ecdysone and 20-hydroxyecdysone were both present in amounts of 1 pg/μl and 20,26-dihydroxyecdysone was barely detectable.

Prothoracicotropic Hormone Stimulation of Ecdysone Synthesis by the Prothoracic Glands of the Gypsy Moth, Lymantria Dispar

Kopec (1922), using Lymantria, first demonstrated that a factor(s) from the head was necessary for insect metamorphosis. This factor was later termed prothoracicotropic hormone (PTTH) (Kobayashi et al., 1965). PTTH stimulates ecdysone synthesis by the paired prothoracic glands (PGs) in lepidopterans, and ecdysone is subsequently converted by other tissues to the active hormone, 20-hydroxyecdysone (see Gilbert et al., 1980). In vitro bioassays for PTTH have been established in both Bombyx mori (Nagasawa et al., 1984; Okuda et al., 1985) and Manduca sexta (Bollenbacher et al., 1979) and have been utilized to quantitate PTTH activity in various tissues and fractions (see Ishizaki and Suzuki, 1984; Bollenbacher and Granger, 1985).