Disinhibition of oocyte growth in adult,virgin Periplaneta americana by corpus allatum denervation: Age dependency and relatedness to mating

Unilateral section of the nervi corporis allati I (NCA-1) of isolated, starved, adult, virgin Periplaneta americana disinhibited oocyte growth during a specific period following their adult emergence. The effect required that the corpus allatum (CA) be free of NCA-1 innervation for 4 days beyond the time the females were 7–8 days old. The onset of this sensitive period corresponds to when most isolated, starved virgins become sexually receptive. The results suggest that NCA-1 inhibition of CA activity, initiated about 7 days, is relieved by mating. When done on sexually receptive, starved virgins, unilateral NCA-1 section was as effective as insemination for stimulating growth and chorionation of the first generation of oocytes. Neural inhibition of juvenile hormone (JH) secretion by the CA may also explain diminished production of oocytes by isolated, fed virgins, for during 30 days following unilateral NCA-1 section they produced 2.6 to 5 times more oothecae than did controls with a single CA removed or after the sham operation. The number of oothecae deposited by fed virgins was similarly increased after bilateral NCA-1 section, but to a lesser extent than when the operation was done on fed, inseminated females of the same age. Specificity of the response of the CA to denervation was substantiated by experiments in which the CA were extirpated and reimplanted, by topically applying C₁₆JH, and by experiments in which the nervus corporis cardiaci 1 and 2 on the right or left side were severed.     

A stereoisomer of JHSB3 with 10S-configuration, 10S-JHSB3, biosynthesized by the corpus allatum of the brown marmorated stink bug,Halyomorpha halys (Hemiptera: Pentatomidae)

The products released by the corpus allatum (CA), an endocrine gland producing juvenile hormone (JH), were analyzed in three species of stink bugs. Liquid chromatography–mass spectrometry (LC–MS) analysis of the CA products from female adults of Halyomorpha halys (Stål), Nezara viridula (Linnaeus), and Nezara antennata Scott (Hemiptera: Pentatomidae) revealed that the CA biosynthesized JH III skipped bisepoxyside (JHSB₃) in all three species. In H. halys, in addition to JHSB₃, the CA also produced its stereoisomer, 10S-JHSB₃. A bioassay focusing on the number of antennal segments was adopted to examine biological activity of these products. When last instar nymphs were treated with either of the two JHs, they emerged as adults bearing nymphal type antennae with four segments in a dose-dependent fashion, indicating that both JHSB₃ and 10S-JHSB₃ had the JH activity in H. halys. Therefore, for the first time, 10S-JHSB₃ in H. halys was found to be a naturally occurring JH molecule with 10S-configuration.

     

Evidence of controlled corpus allatum activity in the adult Colorado potato beetle

The in vivo control of corpus allatum (CA) activity in females of Leptinotarsa decemlineata was investigated. Evidence was obtained that CA activity is adjusted by negative feedback when juvenile hormone (JH) titres are changed experimentally. Conclusions are based on determination of the rate of in vitro JH synthesis by the CA, on changes in CA volume, and on JH titres in the haemolymph. These assay methods are used alternatively in some of the experiments.After unilateral allatectomy, the remaining CA had doubled its activity 7 days later. On the other hand, the activity of CA in young adults was suppressed after the JH titre was elevated by the implantations of 2 CA taken from active females. Similarly, in beetles treated topically with exogenous JH the CA atrophied and showed a much reduced activity after 5 days. Denervation of CA in 0-day-old long-day and 7-day-old short-day females did not change CA activity when measured 1 day later.     

Effect of the brain and corpus cardiacum on egg production in Rhodnius prolixus

Using total egg production corrected for size of blood meal as an index of the activity of the corpus allatum (CA), the effects of various surgical manipulations of the neuroendocrine system have been examined. Isolation of the CA from its nervous connections increases egg production well beyond that of a normal insect, thus confirming that the CA is at least partly controlled by inhibitory nerves from the brain. Removal of the corpora cardiaca (CC) reduces the level of this increased egg production, and decapitation anterior to the CC results in a level of egg production that is greater than that found in females decapitated between the CA and CC. Implanting a CC together with a CA into a decapitated female results in a higher egg production than implanting a CA alone. These results demonstrate that an allatotropic influence is exerted by the CC. Experiments designed to examine the role of the brain were inconclusive and did not eliminate the possibility that the allatotropin from the CC originated in the brain.     

The effect of juvenile hormone on protein synthesis in the transparent accessory gland of male Rhodnius prolixus

Feeding stimulates the accumulation of protein by the transparent accessory reproductive glands (TARG), allatectomy abolishes the increase, and severing the nervous connections between the corpus allatum (CA) and the brain raises the level of accumulation after feeding. TARG removed from allatectomised females exhibit a markedly decreased level of incorporation of [³H]-leucine into protein in vitro, and topical application of juvenile hormone (JH) to allatectomised males restores the ability of the glands to incorporate [³H]-leucine in vitro. Severing the nerves between the CA and the brain increases the rate of incorporation in vitro, but in both operated and normal males, the rate of incorporation decreases with time after feeding. JH applied topically to intact animals during the falling phase of incorporation, or applied directly to TARG in vitro from such animals, stimulates incorporation. These data indicate that JH from the CA controls protein synthesis in the TARG and that the CA in the male of Rhodnius may be subject to different controls than in the female.     

The roles of Dippu-allatostatin in the modulation of hormone release in Locusta migratoria

Dippu-allatostatins (ASTs) have pleiotropic effects in Locusta migratoria. Dippu-ASTs act as releasing factors for adipokinetic hormone I (AKH I) from the corpus cardiacum (CC) and also alter juvenile hormone (JH) biosynthesis and release from the corpus allatum (CA). Dippu-AST-like immunoreactivity is found within lateral neurosecretory cells (LNCs) of the brain and axons within the paired nervi corporis cardiaci II (NCC II) to the CC and the CA, where there are extensive processes and nerve endings over both of these neuroendocrine organs. There was co-localization of Dippu-AST-like and proctolin-like immunoreactivity within these regions. Dippu-ASTs increase the release of AKH I in a dose-dependent manner, with thresholds below 10(-11)M (Dippu-AST 7) and between 10(-13) and 10(-12)M (Dippu-AST 2). Both proctolin and Dippu-AST 2 caused an increase in the cAMP content of the glandular lobe of the CC. Dippu-AST 2 also altered the release of JH from the locust CA, but this effect depended on the concentration of peptide and the basal release rates of the CA. These physiological effects for Dippu-ASTs in Locusta have not been shown previously.     

Function of the neuroendocrine complex in diapausing Pyrrhocoris apterus females

The role of the brain in inhibiting the action of corpora allata in diapausing short-day females was investigated by transplantation experiments. The function of the transplanted glands was evaluated by oviposition. Active glands from long-day females remained active for a long period of time after transplantation into short-day females, although in situ corpora allata were inhibited shortly after the transfer of females from long to short day. Moreover, inactive glands from short-day females became active after transplantation into other short-day females. In contrast, corpora allata remained inhibited when transplanted together with the brain in the neuroendocrine complex (brain-corpora cardiaca-corpus allatum) where the nervous connections between the brain and corpus allatum remained intact. It is therefore suggested that short-day conditions inhibit corpora allata via nervous connections with the brain.     

Neuropeptide regulators of the juvenile hormone biosynthesis (in vitro) in the beetle,Tenebrio molitor (Coleoptera,Tenebrionidae)

The genome of Tribolium castaneum encodes two allatostatin [AS type B; W(X)₆Wamide and AS type C; PISCF‐OH] and one allatotropin (AT) precursor, but no AS type A (FGLamide) (Tribolium Genome Sequencing Consortium, 2008: Nature 452:949–955). Here we studied the activity (in vitro) of peptides derived from these precursors on the synthesis/release of juvenile hormone (JH) III. The corpora cardiaca‐corpora allata (CC‐CA) complexes of adult females of another tenebrionid beetle, the mealworm Tenebrio molitor, were used. Incubating the gland complexes in a medium containing Trica‐AS B3 peptide, we showed that the peptide has allatostatic function in T. molitor. The activity of the type C AS depended on the age of the test animals and their intrinsic rate of JH III biosynthesis. The Trica‐AS C peptide inhibited the JH release from CA of 3‐day‐old females with a high intrinsic rate of JH synthesis, but activated JH release from the CA of 7‐day‐old females with a lower intrinsic rate of JH production. The allatotropin peptide (Trica‐AT) also activated the JH release from the CA of 7‐day‐old females in a dose‐dependent and reversible manner. Unexpectedly, a type A AS derived from the precursor of the American cockroach Periplaneta americana (Peram‐AS A2b) inhibited the JH release from the CA of younger and older females in the concentration range of 10^?8 to 10^?4 M, and the effects were fully reversible in the absence of peptide. These data suggest a complex role of allatoactive neuropeptides in the regulation of JH III biosynthesis in beetles. © 2010 Wiley Periodicals, Inc.     

Proctolin: A possible releasing factor in the corpus cardiacum/corpus allatum of the locust

The corpus cardiacum (CC) and corpus allatum (CA) of the locust, Locusta migratoria, contain intense proctolin-like immunoreactivity (PLI) within processes and varicosities. In contrast, in the cockroach, Diploptera punctata, although a similar staining pattern occurs within the CC, PLI appears absent within the CA. The possible role of proctolin as a releasing factor for adipokinetic hormone (AKH) and juvenile hormone (JH) was investigated in the locust. Proctolin caused a dose-dependent increase in AKH I release (determined by RP-HPLC) from the locust CC over a range of doses with threshold above 10(-8)M and maximal release at about 10(-7)M proctolin. Isolated glandular lobes of the CC released greater amounts of AKH I following treatment with proctolin and in these studies AKH II was also released. Confirmation of AKH I release was obtained by injecting perfusate from incubated CCs into locusts and measuring hemolymph lipid concentration. Perfusate from CC incubated in proctolin contained material with similar biological activity to AKH. Proctolin was also found to significantly increase the synthesis and release of JH from locust CA, with the increase being greatest from CAs that had a relatively low basal rate of JH biosynthesis (<35 pmol h(-1) per CA). In contrast, proctolin did not alter the synthesis and release of JH from the cockroach CA. These results suggest that proctolin may act as a releasing factor for AKHs and JH in the locust but does not act as a releasing factor for JH in the cockroach.     

A factor causing stable stimulation of juvenile hormone synthesis by Diploptera punctata corpora allata in vitro

Co-incubation of corpora allata (CA) from the cockroach, Diploptera punctata, with ovaries, fat body or muscle but not brain or testis, leads to a substantial increase in juvenile hormone synthesis. Incubation of the glands in medium pre-conditioned with ovaries also stimulates JH synthesis. The ovary was used as a convenient source of stimulatory factor for a detailed analysis of its physiological effects on the CA. The increase in JH synthesis is stable, maintained over 24h after exposure to the stimulatory factor. Stimulation is dose-dependent, and the corpora allata show an exquisite relationship between sensitivity to this factor and developmental stage. Day 0 and day 1 glands, as well as glands from post-vitellogenic females, are sensitive to stimulation, whereas glands from vitellogenic females are not sensitive. Corpora allata attached to the brain do not respond to the stimulatory factor, and denervation in vivo leads to an increase in JH synthesis by the glands and a loss in sensitivity to the factor. These data suggest that glands from pre- and post-vitellogenic females are inhibited by their nervous connection to the brain. In contrast, glands from vitellogenic females are normally responding to the endogenous stimulatory factor and are thus no longer stimulated in vitro. Co-incubation of CA with allatostatin and conditioned medium still leads to a stimulation of JH synthesis, suggesting that the restraining effect of the nervous connections to the brain is not caused by allatostatin. The CA cell number increases between emergence and day 2, then remains stable until after oviposition. The stimulatory factor accelerates the increase in cell number in young adult females. The results are interpreted as providing evidence for a constitutive change in CA activity caused by a humoral factor produced by various tissues including the ovary, and modulated by nervous connections to the brain.     

Biosynthesis and regulation of juvenile hormone III,juvenile hormone III bisepoxide,and methyl farnesoate during the reproductive cycle of the grey fleshfly,Neobellieria (Sarcophaga) bullata

To study the effect of brain signals on the biosynthesis of juvenile hormone by the corpora allata of the grey fleshfly Neobellieria bullata, exposed corpora allata connected to the brain were surgically removed from sugar-fed flies and incubated in vitro with L -[³H-methyl]methionine. After incubation, the media together with the tissues were analyzed by HPLC. [³H]Juvenile hormone III (JH III), [³H]JH III bisepoxide (BE), [³H]methyl farnesoate (MF) and an unknown [³H]labeled metabolite (Un) were identified as the primary products. The rate of synthesis of [³H]JH III bisepoxide was higher than that of [³H]JH III, [³H]MF and [³H]Un. Two days after a liver meal, female flies synthesized more JH III, MF, BE, and the Un than did males. Synthesis of JH III, BE, and MF in females was lower during the previtellogenic, sugar-feeding period than during the vitellogenic liver-feeding period. Isolated corpus cardiacum–corpus allatum (CC-CA) complexes that were incubated in vitro synthesized less JH III, MF, and BE, as compared to complexes that were attached to the brain, indicating that the brain probably modulates the biosynthesis of JH III, MF, and BE in the corpora allata. Upon incubation of brain–CC–CA complexes with Neb-TMOF (10^–8 M), Neb-colloostatin (10^–8 M), ovarian, or brain extracts resulted in significant inhibition of JH III and BE biosynthesis in the presence of ovarian extracts. These results indicate that allatostatin-like factors are present in the ovary of the flesh fly. Arch. Insect Biochem. Physiol. 37:248–256, 1998. © 1998 Wiley–Liss, Inc.     

Allatostatins and allatotropin in the corpus cardiacum/corpus allatum complex of larval and adult lepidopterans studied by confocal laser scanning microscopy: correlation to juvenile hormone biosynthesis

Peptidergic innervation of the corpus cardiacum/corpus allatum (CC/CA) retrocerebral complex, and neurosecretory areas of the brain of the lepidopterans Lacanobia oleracea, Heliothis virescens and Manduca sexta was studied by immunocytochemistry linked to confocal laser scanning microscopy. The patterns of immunostaining resulting from the simultaneous application of fluorochrome-conjugated antibodies against Manduca sexta allatostatin (Mas-AS), M. sexta allatotropin (Mas-AT), and a representative of the –Y/FXFGL-NH₂ superfamily of allatostatins was correlated with the physiological effects of these putative allatoregulatory peptides on juvenile hormone (JH) biosynthesis by the corpora allata. Whereas the two types of allatostatin immunoreactivity are present in both larval and adult CA of the three species, allatotropin immunoreactivity occurs only in the adult gland. The conclusion that withdrawal of the stimulatory effect of allatotropin is unlikely to be involved in the downregulation of CA activity prior to the onset of metamorphosis, but that an inhibitory influence of at least Mas-AS is important, is borne out in physiological experiments on JH biosynthesis in M. sexta larvae (Mas-AS inhibitory, Mas-AT without effect). Immunoreactivity to the Y/FXFGL-NH₂ allatostatins is present in both larval and adult CA and CC, frequently co-localised with Mas-AS. The function of this peptide family in the retrocerebral complex remains enigmatic since experiments on JH biosynthesis, either when the peptide is administered alone, or together with Mas-AS, show no effect on JH biosynthesis.Financial support was provided by The Wellcome Trust (063367/Z/00) (to A.T.) and by the Pesticide Safety Directorate of the Department for Environment, Food and Rural Affairs (to N.A. and R.J.W.)     

The effect of severing the dorsal vessel on egg production in Rhodnius prolixus

Severing the dorsal vessel (DV) behind the corpus allatum (CA), or in the anterior part of the abdomen of Rhodnius prolixus, greatly reduces egg production, an effect which is abolished by the topical application of juvenile hormone l (JH l). Severing the DV in the posterior abdomen does not result in a marked reduction of egg production, although severing the alary muscles in segments V and VI has a similar effect to severing the DV in the anterior abdomen. Reduced egg production caused by severing the DV on day 8 postemergence does not occur if the nerves connecting the CA to the brain are severed on day 1 post emergence. However, egg production is reduced if the DV is severed on day 1 post emergence and the connections between the brain and the CA severed on day 8, suggesting that inhibition of the CA caused by severance of the DV requires innervation from the brain. An isolated CA implanted into an animal decapitated immediately after feeding escapes from the inhibition imposed by severance of the DV. Conversely, the CA in an insect, the head of which has been decapitated just anterior to the CA, remains inhibited. This result suggests that the head posterior to the brain must be present to maintain inhibition. It is concluded that DV severance acts on the brain via some humoral influence to impose inhibition on the CA, and that an endocrine center in the head is required in order to maintain the inhibition.     

Effects of chilling stress on allatal growth and juvenile hormone synthesis in the cockroach, Diploptera punctata

During the ovarian cycle of the cockroach, Diploptera punctata, a mitotic wave occurs in the corpora allata before an increase in gland volume and juvenile hormone (JH) synthesis. Previous studies have demonstrated that the brain inhibits mitosis and JH synthesis in corpus allatum (CA) cells until adult females have mated. Herein, we report that chilling stress effectively suppresses mating induced proliferation of CA cells. In mated females, chilling on melting ice for 0.5-3 hours caused a strong, dose-dependent decrease in mitotic activity. In insects chilled for 3 hours, although the mitotic wave in the CA was practically abolished, CA volume and JH synthesis finally reached peak levels typical of unchilled insects, despite a 2-day delay. Consequently, oocyte maturation and oviposition were also delayed by 2 days, yet in both chilled and unchilled insects, peak values of basal oocyte length were the same. By allowing virgin females to mate on different days after chilling, we found that the chilling effect could be retained in the insect body for at least 2 days. During this period, signals from mating could not effectively remove inhibition of CA cell proliferation. Unilaterally disconnecting the CA from the brain revealed that chilling stress mediated CA cell proliferation via the brain, and did not directly affect the CA.     

Juvenile hormone biosynthesis in diapause and nondiapause females of the adult blow fly Protophormia terraenovae

In vitro synthetic activities of juvenile hormones (JH) were examined using a radiochemical assay in diapause females and reproductive females of the blow fly, Protophormia terraenovae. Thin layer chromatography showed that products of the corpus allatum (CA) comigrated with a synthetic sample of JH III bisepoxide but neither with JH III nor methylfarnesoate. JH synthetic activities increased in females reared under LD 18:6 at 25 degrees C, as the ovaries developed. The synthetic activities remained low in previtellogenic females reared under LD 12:12 at 20 degrees C. Removal of the pars intercerebralis completely prevented ovaries from development under reproductive conditions, and removal of the pars lateralis caused partial or full development of ovaries under diapause-inducing conditions. In these operated animals, the JH synthetic activities were not significantly different from those of the intact and sham-operated animals. The results indicate that the CA in P. terraenovae produces mainly JH III bisepoxide and a decrease in the JH production rate is a cause of diapause induction. PI neurons and PL neurons in the brain do not directly mediate changes in the JH production rate, but regulate ovarian development cooperatively with some unknown allatostatic and allatotropic factors.     

Inhibition of DD2R gene expression in the corpus allatum activates alkaline phosphatase in female Drosophila melanogaster

Tissue-specific inhibition of the expression of the D2-like dopamine receptor gene (DD2R) in the corpus allatum (CA), which is a gland that synthesizes the juvenile hormone (JH), was tested for effect on alkaline phosphatase (AP) activity and the intensity of the AP response to heat stress (stress reactivity) in female Drosophila melanogaster. AP activity and AP stress reactivity in transgenic females with lower DD2R expression in the CA were higher than in control flies. A pharmacological elevation in JH increased AP activity in females of the control strains. DD2R was assumed to mediate the inhibitory effect of dopamine of JH synthesis in the CA of D. melanogaster.     

Time-course haemolymph juvenile hormone titres in solitarious and gregarious adults of Schistocerca gregaria, and their relation to pheromone emission, CA volumetric changes and oocyte growth

The haemolymph JH III titres in solitarious and gregarious adult desert locusts, Schistocerca gregaria, were examined in relation to corpus allatum (CA) volumes, aggregation-maturation pheromone production in males and oocyte growth in females. The JH titres of gregarious females were generally higher than those of solitarious females at all ages studied. The titre patterns, however, were similar: relatively high on day 10, dropping to low levels between days 20 and 25, before rising again by day 25. In the solitarious males, the JH titre was very low on day 10 after fledging, but increased gradually and reached a maximal amount on day 30. The JH titre in gregarious males was low on day 10, elevated on day 15 coinciding with the start of the production of the pheromone, and dropped to a relatively low level on day 20 around the time of maximal pheromone production, then rising again by day 25. These results suggest that biosynthesis of the pheromone is associated with a high JH titre peak in the haemolymph. Although a clear relationship was found during the first gonadotropic cycle between JH titres, on one hand, and CA volume and oocyte growth, on the other, in both phases, no such correlation could be discerned in the second cycle.     

Biosynthetic maturation of the corpus allatum of the female adult medfly,Ceratitis capitata,and its putative control

The major juvenile hormone (JH) homolog synthesized in vitro by the adult female Medfly (Ceratitis capitata) corpus allatum (CA) is JHB(3), with JH-III the minor homolog. Methyl-incorporation in vitro in post-eclosion virgin females is age-dependent. Basal activity occurs during the first four days post-eclosion and increases significantly thereafter, peaking at five days. Biosynthetic maturation of the mated female CA is delayed by one day and reduced considerably. The delayed response may be due to direct cerebral or neural inhibition. Synthetic Drosophila melanogaster sex peptide depresses JH biosynthesis by the Medfly female CA in vitro. Male-derived accessory gland peptides of the Medfly are transferred to the female during mating and a Medfly SP-analog may be responsible for down-regulation of JH synthesis by the CA in mated Medfly females. Mevinolin, an inhibitor of the mevalonate pathway, significantly reduces the biosynthesis of JHB(3), while farnesoic acid, a proximate precursor of JHIII, significantly stimulates the biosynthesis of both JHB(3) and JHIII in vitro.     

A pharmacological and endocrinological study of female insemination inPhormia regina (Diptera: Calliphoridae)

Injections of octopamine, dopamine, and the octopaminergic agonists, clonidine and naphazoline, into the thoracic hemocoel enhanced female insemination in sugar-fed (sexually unreceptive)Phormia regina. Topical applications of the juvenile hormone (JH) analogue, methoprene, also enhanced female insemination in sugar-fed (sexually unreceptive)P. regina. Since JH plays a role in receptivity in protein-fed females, it was originally hypothesized that one agonist, clonidine, enhanced female insemination by acting on the corpus allatum (CA) to increase JH biosynthesis. Two or three doses of the antiallatal agent, precocene II, prior to administration of clonidine, did not inhibit clonidine-enhanced female insemination. Removal of the corpus allatum also did not have a significant effect on clonidine-enhanced female insemination. Measurement of juvenile hormone (JH) biosynthesis/release in corpora allata, which were removed at 1, 3, 5, and 7 h postinjection, revealed that clonidine does not affect JH biosynthesis/release. Our study reveals a possible role for the biogenic amines in female insemination in insects. We suggest that the octopaminergic agonist, clonidine, acts downstream from the corpus allatum on the regulatory mechanisms involved in female insemination.     

Juvenile hormone in earwigs: Roles in oogenesis,mating, and maternal behaviors

The role of juvenile hormone (JH) in courtship, mating, maternal behavior, and the ovarian cycle was studied in the ring-legged earwig, Euborellia annulipes (Lucas). The single, median corpus allatum makes and secretes JH III. JH III production was low in newly eclosed adult females, increasing as oocytes developed, maximal at about the time of oviposition, and low again in brooding females. Application of 35 or 122 μg JH III to newly eclosed females hastened the onset of courtship behavior, but had no effect on the age at which females first mated nor on the duration of mating, though the trend is toward advanced onset. Hormone treatment advanced the age of first oviposition and reduced clutch size and the proportion of eggs hatching but did not affect the interval from oviposition of the first clutch to oviposition of the second clutch, nor the size and proportion hatching of the second clutch. Acetone treatment and treatment with 6 μg JH III did not affect these parameters. Application of 50 μg JH III to females on the day of oviposition shortened the duration of maternal care and advanced the onset of the second gonadotropic cycle, compared with that of acetone-treated and precocene II-treated females. The duration of maternal care was positively correlated with the proportion of eggs hatching. JH titer analysis confirmed JH III to be the predominant hormone in this species and clearly demonstrated the absence of other homologues. This work also confirmed our hypothesis that intermediate to high levels of JH are associated with oocyte growth, mating, and cessation of maternal care; low levels of JH are associated with the period of maternal behavior and slow ovarian development. We are currently investigating factors which might regulate corpus allatum activity during the reproductive cycle and the subsequent period of maternal care. Arch. Insect Biochem. Physiol. 35:427–442, 1997. © 1997 Wiley-Liss, Inc.