Primer effect of queen pheromone on juvenile hormone biosynthesis in adult worker honey bees

Juvenile hormone synthesis in adult worker honey bees was measured by an in vitro corpora allata bioassay. Adult queenless workers exhibit higher rates of juvenile hormone biosynthesis than queenright workers. Hormone synthesis is not correlated with the volume of the glands. Extract of queen mandibular glands, applied to a dummy, reduces juvenile hormone biosynthesis in caged queenless workers to the level of queenright workers. The same result was obtained with synthetic (E)-9-oxo-2-decenoic acid, the principal component of the queen mandibular gland secretion. This pheromonal primer effect may function as a key regulating element in maintaining eusocial colony homeostasis. The presence of brood does not affect the hormone production of the corpora allata.Abbreviations BSA bovine serum albumin - CA Corpora allata - JH juvenile hormone - 9-ODA (E)-9-oxo-2-decnoic acid     

The mode of regulation of the corpus allatum activity during starvation in adult females of the Colorado potato beetle, Leptinotarsa decemlineata (Say)

When two-day-old female Leptinotarsa decemlineata were starved, their corpus allatum activity, as measured by the radiochemical in vitro assay, was significantly reduced after 24 hr. Such a reduction was not observed when the nerve connections between the central nervous system and the retrocerebral complex were severed and the beetles starved up to 5 days. In some experiments, the rate of juvenile hormone biosynthesis in vitro, was substantiated by measurement of the juvenile hormone titre in the haemolymph by physico-chemical methods. It is concluded that intact nervous connections between the central nervous system and the corpora allata are essential for restraining the juvenile hormone biosynthesis during the initial stages of starvation.Corpora allata from 1-day starved insects were considerably stimulated in vitro by farnesenic acid indicating that juvenile hormone synthesis is controlled enzymatically at a stage prior to the final two steps in the pathway. However, on day 5 of starvation, rate-limitation may occur after formation of this intermediate, since farnesenic acid stimulation was much less at this time.Corpora allata of adult females newly emerged from the soil were activated within 4 hr regardless of feeding.     

Responsiveness of the adult cricket (Gryllus bimaculatus andAcheta domesticus) retrocerebral complex to allatostatin-1 from a cockroach,Diploptera punctata

Brain-retrocerebral complexes of female crickets,Gryllus bimaculatus andAcheta domesticus, treated with antibody to allatostatin-1 from a cockroach,Diploptera punctata, show extensive immunoreactivity. The results suggest that allatostatins or allatostatin-like molecules are produced in neurosecretory cells of the brain and are delivered to the corpora allata through nervous connections and/or via haemolymph. Radiochemical measurements of juvenile hormone III biosynthesis by isolated corpora cardiaca-corpora allata complexes from adultG. bimaculatus have been used to demonstrate an in vitro sensitivity of these glands to allatostatin-1 fromD. punctata. Allatostatin-1 is a relatively potent inhibitor of juvenile hormone III biosynthesis in corpora allata of both young adult females and males. In glands taken from 3-day virgin females, 50% inhibition of hormone biosynthesis is reached at ca. 3 nmol·l^-1 allatostatin-1. The inhibitory action of allatostatin-1 is rapid, dose-dependent and reversible. Addition of 200 mol·l^-1 farnesol to the incubation medium prevents inhibition of juvenile hormone III biosynthesis by allatostatin-1. Juvenile hormone III biosynthesis by isolated corpora allata of 3-day female house crickets,A. domesticus, is also susceptible to inhibition by 1 mol·l^-1 allatostatin-1.Abbreviations ASB2 Diploptera punctata allatostatin-5 - CA corpora allata - CC corpora cardiaca - Dip A-1 Diploptera punctata allatostatin-1 - HEPES 4-(2-hydroxyethyl)piperazine-1-ethanesulphonic acid - JH juvenile hormone(s) - Mas-AS Manduca sexta allatostatin - MF methyl farnesoate - NCA nervus corporis allati - NCC nervus corporis cardiaci - SEM standard error of mean - TRIS Tris(hydroxymethyl)aminomethane     

In vitro biosynthesis of juvenile hormone in larval honey bees: Comparison of six media

Summary We have formulated a tissue culture medium based on the components of larval honey bee hemolymph. Using an in vitro radiochemical assay to measure juvenile hormone biosynthesis, we compared our larval-based medium to four commercially available media (Grace’s, Medium-199; Shields and Sang M3, and Minimum Essential Medium), and a medium based on adult honey bee hemolymph. All media were formulated without methionine. There was no significant difference in the amounts of juvenile hormone produced by the larval medium and Grace’s; both of these media, however, were more suitable than the remaining four. Our larval-based tissue culture medium should prove useful in studies aimed at elucidating the underlying hormonal mechanism(s) of caste development in honey bees.     

Sex-specific developmental profiles of juvenile hormone synthesis in honey bee larvae

Summary Juvenile hormone synthesis in drone larvae of the honey bee was measured by an in vitro radiochemical assay. The developmental profile of corpora allata activity in male larvae showed considerable differences from queen larvae, the presumptive reproductive females, and was comparable to workers, the sterile female morph. Drone and worker larvae, however, differed drastically in the regulation of juvenile hormone biosynthesis, as revealed by the addition of farnesoic acid to the culture medium. This precursor stimulated juvenile hormone synthesis of drone glands nearly eightfold, whereas in worker larvae it is known to lead to an accumulation of methyl farnesoate. The sex-specific differences in endocrine activity indicate a role for juvenile hormone in the expression of genetically determined sexually dimorphic characters during metamorphosis, a role not currently accounted for in models describing endocrine regulation of insect development. Correspondence to: K. Hartfelder     

Identification and characterization of the diuretic hormone 31 receptor in the silkworm Bombyx mori

The receptor for diuretic hormone 31 (DH31R) was identified in the silkworm Bombyx mori. A heterologous expression system revealed that an orphan G-protein coupled receptor, BNGR-B1, responded to DH31 and upregulated the intracellular cAMP level. DH31R (BNGR-B1) was predominantly expressed in the anterior silk gland, midgut, and ovary, whereas DH31 was predominantly expressed in the central nervous system and midgut.     

Distribution of allatostatin in the adult cockroach,Diploptera punctata and effects on corpora allata in vitro

Direct radiochemical measurements of juvenile hormone synthesis showed that corpora allata from adult female Diploptera punctata can be inhibited in vitro by neuropeptides extracted from several ganglia of the central nervous system of females at many stages of the reproductive cycle. Extracts of protocerebra, corpora cardiaca, suboesophageal, thoracic and ventral ganglia all elicited dose-depedent reductions in juvenile hormone synthesis. On a ‘per organ’ basis, the protocerebrum contains the most extractable material. Inhibitory activity of extracts of suboesophageal, thoracic and 6th abdominal ganglia, like that of protocerebra (Rankin et al., 1986) was trypsin sensitive.Glands of high activity were less sensitive to protocerebral extract than those of low activity. The inhibitory effect on glands of low activity was maximal within 1 h, persisted in the presence of protocerebral extract for at least 46 h, and was abolished within 1 h after corpora allata were placed in normal medium. The inhibitory effect of protocerebral extract was not altered by the addition of magnesium to the medium. The extract had a specific effect on synthetic step(s) prior to methylation and epoxidation as demonstrated by enhanced juvenile hormone synthesis in the presence of inhibitory factor and the juvenile hormone precursor, farnesoic acid.     

Neuromodulation for behavior in the locust frontal ganglion

Neuromodulators orchestrate complex behavioral routines by their multiple and combined effects on the nervous system. In the desert locust, Schistocerca gregaria, frontal ganglion neurons innervate foregut dilator muscles and play a key role in the control of foregut motor patterns. To further investigate the role of the frontal ganglion in locust behavior, we currently focus on the frontal ganglion central pattern generator as a target for neuromodulation. Application of octopamine, a well-studied insect neuromodulator, generated reversible disruption of frontal ganglion rhythmic activity. The threshold for the modulatory effects of octopamine was 10^–6 mol l^–1, and 10^–4 mol l^–1 always abolished the ongoing rhythm. In contrast to this straightforward modulation, allatostatin, previously reported to be a myoinhibitor of insect gut muscles, showed complex, tri-modal, dose-dependent effects on frontal ganglion rhythmic pattern. Using a novel cross-correlation analysis technique, we show that different allatostatin concentrations have very different effects not only on cycle period but also on temporal characteristics of the rhythmic bursts of action potentials. Allatostatin also altered the frontal ganglion rhythm in vivo. The analysis technique we introduce may be instrumental in the study of not fully characterized neural circuits and their modulation. The physiological significance of our results and the role of the modulators in locust behavior are discussed.Abbreviation CPG central pattern generator - FG frontal ganglion - JH juvenile hormone - STNS stomatogastric nervous system     

Immunological evidence for an allatostatin-like neuropeptide in the central nervous system of Schistocerca gregaria,Locusta migratoria and Neobellieria bullata

Methanolic brain extracts of Locusta migratoria inhibit in vitro juvenile hormone biosynthesis in both the locust L. migratoria and the cockroach Diploptera punctata. A polyclonal antibody against allatostatin-5 (AST-5) (dipstatin-2) of this cockroach was used to immunolocalize allatostatin-5-like peptides in the central nervous system of the locusts Schistocerca gregaria and L. migratoria and of the fleshfly Neobellieria bullata. In both locust species, immunoreactivity was found in many cells and axons of the brain-retrocerebral complex, the thoracic and the abdominal ganglia. Strongly immunoreactive cells were stained in the pars lateralis of the brain with axons (NCC II and NCA I) extending to and arborizing in the corpus cardiacum and the corpora allata. Although many neurosecretory cells of the pars intercerebralis project into the corpus cardiacum, only 12 of them were immunoreactive and the nervi corporis cardiaci I (NCC I) and fibers in the nervi corporis allati II (NCA II) connecting the corpora allata to the suboesophageal ganglion remained unstained. S. gregaria and L. migratoria seem to have an allatostatin-like neuropeptide present in axons of the NCC II and the NCA I leading to the corpus cardiacum and the corpora allata. All these data suggest that in locusts allatostatin-like neuropeptides might be involved in controlling the production of juvenile hormone by the corpora allata and, perhaps, some aspects of the functioning of the corpus cardiacum as well. However, when tested in a L. migratoria in-vitro juvenile hormone-biosynthesis assay, allatostatin-5 did not yield an inhibitory or stimulatory effect. There is abundant AST-5 immunoreactivity in cell bodies of the fleshfly N. bullata, but none in the CA-CC complexes. Apparently, factors that are immunologically related to AST-5 do occur in locusts and fleshflies but, the active protion of the peptide required to inhibit JH biosynthesis in locusts is probably different from that of AST-5.     

Physiological correlates of division of labor among similarly aged honey bees

Hormone analyses and exocrine gland measurements were made to probe for physiological correlates of division of labor among similarly aged adult worker honey bees (Apis mellifera L.). Middle-age bees (ca. 2 weeks old) performing different tasks showed significant differences in both juvenile hormone (JH) biosynthesis rates and hemolymph titers; guards and undertakers had high JH, and wax producers and food storers, low JH. Guards and undertakers had similar hormone levels to foragers, even though they were 10 days younger than foragers. No differences in JH were detected among young bees (1-week-old queen attendants and nurses) or older bees (3–4 week-old pollen foragers, non-pollen foragers, and soldiers). Hypopharyngeal gland size was inversely correlated with worker age and rate of JH biosynthesis, but soldiers had significantly larger hypopharyngeal glands than did foragers, despite their similar age and JH level. Results from soldiers indicate that exocrine gland development is not always linked with age-related behavior and endocrine development; they also support the recent claim that soldiers constitute a group of older bees that are distinct from foragers. Hormonal analyses indicate that the current model of JH's role in honey bee division of labor needs to be expanded because high levels of JH are associated with several other tasks besides foraging. JH may be involved in the regulation of division of labor among similarly aged workers in addition to its role in age-related division of labor.Abbreviations JH Juvenile hormone - RIA radioimmunoassay - CA corpora allata - HPLC high performance liquid chromatography - TLC thin layer chromatography     

Evidence for the presence of thyroid stimulating hormone, thyroglobulin and their receptors in Eisenia fetida: a multilevel hormonal interface between the nervous system and the peripheral tissues

The present study describes the localization and distribution of thyroid-stimulating hormone (TSH), thyroglobulin (TGB) and their receptors in Eisenia fetida (Annelida, Oligochaeta) as revealed by immunohistological methods. Immunopositive neuronal and non-neuronal cells are present in both the central nervous system and some peripheral organs (e.g. foregut and coelomocytes). TSH- and TGB-immunopositive neurons in the various ganglia of the central nervous system are differentailly distributed. Most of the immunoreactive cells are found in the suboesophageal ganglion. The stained cells also differ in their shapes (round, oval, pear-shaped) and sizes (small, 12–25 μm; medium, 20–35 μm; large, 30–50 μm). In all ganglia of the central nervous system, TSH-positive neurons additionally show gamma aminobutyric acid (GABA) immunopositivity. Non-neuronal cells also take part in hormone secretion and transport. Elongated TSH-positive cells have been detected in the capsule of the central ganglia and bear granules or vacuoles in areas lacking neurons. Many of capillaries show immunoreactivity for all four tested antibodies in the entire central nervous system and foregut. Among the coelomocytes, granulocytes and eleocytes stain for TSH and its receptor and for TGB but not for thyroid hormone receptor. Most of the granulocytes are large (25–50 μm) but a population of small cells (10–25 μm) are also immunoreactive. None of the coelomocytes stain for GABA. We therefore suggest that the members of this hormone system can modify both metabolism and immune functions in Eisenia. Coelomocytes might be able to secrete, transport and eliminate hormones in this system.This work was supported by the MTA-PTE Adaptation Biology Research Group and National Research and Developmental Fund (NKP 1/048/2001). M.W. is in receipt of a János Bolyai Scholarship.     

Colony Integration in Honey Bees: Mechanisms of Behavioral Reversion

After confirming that worker honey bees (Apis mellifera) can revert from foraging to brood care, we determined whether juvenile hormone (JH) mediates this form of plasticity in behavioral development and whether worker age and genotype influence the probability of its expression. Measurements of JH titers support the hypothesis that plasticity in honey bee behavioral development is a consequence of modulation of JH by extrinsic factors. Observations of individually marked bees in a colony composed of two phenotypically distinguishable subfamilies revealed that the likelihood of undergoing behavioral reversion was influenced by worker age but not by worker genotype. The effect of worker age on reversion is consistent with a previously formulated model for the regulation of age polyethism in honey bees that predicts that workers of different ages have different response thresholds for task-associated stimuli. The lack of a genotypic effect on reversion is in contrast to results for other forms of behavioral plasticity.     

Endocrine modulation of a pheromone-responsive gene in the honey bee brain

Pheromones cause dramatic changes in behavior and physiology, and are critical for honey bee colony organization. Queen mandibular pheromone (QMP) regulates multiple behaviors in worker bees (Slessor et al. in J Chem Ecol 31(11):2731–2745, 2005). We also identified genes whose brain expression levels were altered by exposure to QMP (Grozinger et al. in Proc Natl Acad Sci USA 100(Suppl 2):14519–14525, 2003). Krüppel-homolog 1 (Kr-h1) RNA levels were significantly downregulated by QMP, and were higher in foragers than in nurses (Whitfield et al. in Science 302(5643):296–299, 2003). Here we report on results of behavioral and pharmacological experiments that characterize factors regulating expression of Kr-h1. Foragers have higher brain levels of Kr-h1 than in-hive bees, regardless of age and pheromone exposure. Furthermore, forager Kr-h1 levels were not affected by QMP. Since the onset of foraging is caused, in part, by increasing juvenile hormone blood titers and brain octopamine levels, we investigated the effects of octopamine and methoprene (a juvenile hormone analog) on Kr-h1 expression. Methoprene produced a marginal (not significant) increase in Kr-h1 expression, but Kr-h1 brain levels in methoprene-treated bees were no longer downregulated by QMP. Octopamine did not modulate Kr-h1 expression. Our results demonstrate that the gene expression response to QMP is not hard-wired in the brain but is instead dependent on worker behavioral state.     

The role of exogenous JH I,JH III and Anti-JH (precocene II) on queen induction of 4.5-day-old worker honey bee larvae

Worker larvae at an age of 4½ days were fed one of several mixtures of reconstituted royal jelly adjusted to a refractive index of 1.3825 and supplemented with JH I, JH III or Anti-JH (precocene II). In addition, juvenile hormone was topically applied to larvae of the same age. It was readily apparent that caste induction is concentration-dependent and that 4?-day-old worker larvae can still develop into queens under laboratory conditions, providing that they have not stopped feeding or can be induced to commence feeding again. These findings are contrary to the general belief that queen induction is not possible after a socalled sensitive period of 3–3½ days. Queens resulted only from honey bee larvae exposed to royal jelly containing 1 μg of JH I. In addition, oral application at this concentration resulted in the only case in which the normal mean weights of worker honey bees were exceeded. All other concentrations of juvenile hormone were not sufficient to initiate queen induction, although its lower concentration may have influenced the production of intercastes.Precocene II did not play a role in queen induction and it also did not interfere with the growth of developing larvae or adults. In addition, the lack of malformations in honey bees treated with precocene II indicates that the use of such a compound as a control agent in insect populations will probably not be detrimental to honey bee larvae that are at least 4½ days old. However, large doses of precocene will quickly kill most 3½-day-old honey bee larvae.The evidence presented here clearly indicates that caste determination is regulated by the endocrine system in honey bee larvae. Food intake in honey bee larvae may well be regulated by the endocrine system. Thus, an apparently inhibited corpus allatum (C.A.) could be reactivated by food intake coupled with juvenile hormone. The food intake restriction that worker larvae normally encounter in the hive probably results in a cessation of C.A. activity. The increase in food intake by queen larvae, on the other hand, carries an increase in growth and accompanying morphological changes necessary for queen development. This concept may also explain the development of intercastes encountered in in vitro studies. Only those larvae that follow a normal food intake sequence, i.e. moderate during the first 3–4 days or so, will develop into queens. Conversely, those larvae that take in too much food during the early portion of development may achieve incomplete development of the neurosecretory system and, thus, develop into intercastes.     

Afferent and efferent components of joint position sense; interpretation of kinaesthetic illusion

The present model of joint angle perception is based on the following hypotheses: the perception and control of joint angle are closely interrelated processes; central motor commands are adequately expressed by shifts of an equilibrium point resulting from the interaction of antagonistic muscles and a load; two fundamental commands-reciprocal (r) and coactivative (c) provide for changes in activity of the antagonistic muscle pair. The dependence of joint angle on static muscle torque and r and c commands is derived (Eq. 5). The following principles of joint position sense are formulated: 1) the r component of the efferent copy plays the role of a reference point which shifts during voluntary regulation of muscle state, but remains unchanged during any passive alterations of joint position; 2) muscle afferent signals deliver not absolute but relative information (i.e. measured relatively to the central reference point). These signals turn out to be related to active muscle torque; 3) the nervous system evaluates muscle afferent signals on the basis of a scale determined by the level of coactivation of the antagonistic muscles. Kinaesthetic illusions appear to be due to disruptions in perception of afferent and/or efferent components of position sense. The present model is consistent with all the variety of kinaesthetic illusions observed experimentally. A qualitative neurophysiological schema for joint angle perception is proposed involving efferent copy and information concerning muscle torque delivered by the tendon organ, muscle spindle, and perhaps, articular receptors. It is known that the cerebellum incorporates both afferent and efferent information concerning movement. One may presume that it plays an essential role in position sense.     

Juvenile hormone stimulation of ornithine decarboxylase activity during vitellogenesis in Drosophila melanogaster

Summary The activity of ornithine decarboxylase (ODC), the rate-limiting enzyme in polyamine biosynthesis, becomes elevated in intact female Drosophila melanogaster shortly after adult eclosion. This activity reaches a peak at 24 h following eclosion, and then drops to lower levels by 48 h. This pattern is not observed in males, consistent with the hypothesis that polyamine synthesis is involved in ovarian maturation in Drosophila. Abdomens isolated within 2 h of adult eclosion do not display elevated ODC activity or ovarian maturation. However, a 250-ng dose of the juvenile hormone analog methoprene (ZR-515) applied in acetone to these abdomens, recovers ovarian maturation and causes a 5–10 fold increase in enzyme activity over controls treated with acetone alone. The same dose of the inactive precursor methyl farnesoate caused no such increase, whereas a 500-ng dose of the newly discovered natural Drosophila JHB₃ stimulated a four-fold response. The response to methoprene was dose-dependent, showing stimulatory activity at a dose as low as 10 ng. This stimulation by JHA is rapid, occurring between 1 and 3 h following hormone treatment, reminiscent of JH induction of fat body vitellogenin synthesis in Drosophila. Elevated ODC activity appeared to be localized in the adult fat body. During embryogenesis, ODC activity remained undetectable until just prior to hatching, when a large increase was detected. We postulate that JH may, either directly or indirectly, regulate polyamine biosynthesis in vivo, and that this synthesis may be required for the production of macromolecules during Drosophila vitellogenesis or embryogenesis.Abbreviations JH juvenile hormone - JHA juvenile hormone analog - ODC ornithine decarboxylase - SAMDC S-adenosyl-methionine decarboxylase - JHB ₃ juvenile hormone III bisepoxide     

Neuronal input contributes to sequence of Aplysia egg laying behaviors

Egg laying in Aplysia is controlled by the bag cell neuroendocrine system, which releases multiple peptides during a long-lasting electrical discharge. Following the discharge, a fixed sequence of head and neck movements is performed in which two phases can be distinguished: an appetitive or preparatory phase, in which the substrate is prepared, and a consummatory phase, when the egg string is deposited. During egg laying, feeding responses are suppressed. In this study, Aplysia fasciata was used. When the movement of the egg string through the genital groove was prevented by ligation, lesions of the nerve innervating the genital pore completely abolished the consummatory egg-laying behaviors. This shows that a nervous connection between the genital pore area and the central nervous system is important for the consummatory egg-laying behaviors.We found that suppression of feeding responses to seaweed occurred only during the consummatory phase of egg laying in controls, but animals with ligated genital grooves continued to show normal responses to food. It is hypothesized that a neuronal feedback, possibly together with the bag cell peptides, is critical for the temporal organization of egg-laying behavior in Aplysia.Abbreviations CNS central nervous system - ELH egg laying hormone     

Effects of parasitization of Trichoplusia ni by Chelonus sp.

ABSTRACT. Parasitization of Trichoplusia ni (Huebner) (Lepidoptera: Noctuidae) by Chelonus sp. (Hymenoptera: Braconidae), an egg-larval parasitoid, leads to precocious cocoon spinning of the host in the fourth (penultimate) stadium followed by parasitoid emergence from the prepupa. We have investigated the mechanism by which Chelonus sp. disrupts host development. The developing larva and fluids injected by the adult female separately from the egg, are not the source of these effects, but it remains a possibility that the teratocytes, originating from the trophamnion of the parasitoid egg, are responsible. The titre of the juvenile hormone esterase activity in the haemolymph of the parasitized fourth instar host is similar to that in the initial period of the final instar of normal T. ni, but lacks the postwandering peak of activity. The increased JHE activity leads to a reduced JH titre early in the fourth stadia. This indicates that disruption of host development occurs within 12h after apolysis to the fourth stadium, if not before. Anti-juvenile hormone activity is not detected in extracts of parasitized T. ni. The morphological and behavioural changes associated with precocious development of the T. ni host are prevented by applications of juvenile hormone I, juvenile hormone II and the juvenoid, Ro 10–3108, but not juvenile hormone III and the juvenoid R 20458. However, these applications fail to prevent the onset of juvenile hormone esterase activity, another marker of precocious development. These observations indicate that simple anti-juvenile hormone activity may not be the mechanism of disruption of host development. Development of the parasitoid is disrupted by application of Ro 10–3108 and juvenile hormones I, II and III, but timing of eclosion is only affected by application of juvenile hormone I, juvenile hormone II and Ro 10–3108. This observation may indicate a discrimination by the parasitoid between its own juvenile hormone III and the host's juvenile hormone II.