Changes in biosynthesis and degradation of juvenile hormone during breeding by burying beetles: a reproductive or social role?

Burying beetles, Nicrophorus orbicollis, depend on the location of an unpredictable resource, a small vertebrate carcass, for reproduction. When they discover a carcass, they undergo a correlated rapid rise in titers of juvenile hormone (JH) in the hemolymph and ovarian development. This study investigates the regulation of the changes in JH during breeding in both male and female burying beetles and the role of JH in ovarian development. JH biosynthesis by the corpora allata (CA), measured in vitro, increased in females within an hour of their discovery of a carcass and increased later in males. After returning to low rates as oviposition began, JH biosynthesis rose again 3 days later in females but not in males. Neither the ovaries nor testes synthesized JH. There was a concomitant fall in JH esterase activity within 12 h of discovery of the carcass in both males and females. Although the rise in JH titers and biosynthesis and the fall in JH esterase is correlated with ovarian development, application of methoprene or JH III in the absence of a carcass did not result in vitellogenin uptake by the oocytes. Therefore, we conclude that, in spite of the rapid rise in JH before oviposition, it is not sufficient to regulate vitellogenin synthesis and/or its uptake by the ovaries. We suggest that its role has been preempted to organize social behavior and coordinate parental behavior between mates.     

Plasticity in juvenile hormone in male burying beetles during breeding: physiological consequences of the loss of a mate

Burying beetles, Nicrophorus orbicollis, have facultative biparental care. They bury and prepare small vertebrate carcasses that provide food for their young. Here we establish the juvenile hormone (JH) profiles of paired females, paired males and single males and investigate some of the environmental and social factors that may affect these profiles. Before larvae hatch JH profiles of paired males and females were similar. However, after larvae hatch and during brood care, JH titers of females were very high and those of single males were significantly higher than those of paired males. We tested the hypothesis that higher JH was a response to the need for increased parental care by manipulating brood size. Although JH titers of single males caring for small versus large broods were not significantly different, when comparing JH titers and larval growth (a measure of parental effort), a significant positive correlation emerged. In contrast, we found that food quality had no effect on JH levels suggesting that increased feeding by males and females after carcass discovery cannot explain the elevation of JH. The regulation of JH in male burying beetles appears thus to be dependent on the presence of a mate and on critical stimuli from young.     

Social and nonsocial stimuli and juvenile hormone titer in a male burying beetle, Nicrophorus orbicollis

We investigated the interaction of social and nonsocial stimuli on juvenile hormone (JH) titer in male burying beetles (Nicrophorus orbicollis). The initial JH response to discovery of a carcass was substantial (10-15-fold increase over controls) and rapid (<1h), and occurred whether or not a female was present. By 3h after discovery, JH titers were declining, the decline being more pronounced when a female was not present. We also tested the effect of larval stimulation on JH titer in care-giving males by removing a male's brood and replacing it with a brood of first or third instar larvae. Males initially providing care for begging first instar larvae continued to maintain high titers of JH when the replacement broods were first but not third instars. Males caring for third instar larvae (normally low JH titers) maintained low levels of JH regardless of the developmental stage of the replacement brood. This suggests that once males begin to care for nutritionally independent third instar larvae, JH titers remain low regardless of subsequent larval stimulation. Burying beetles are socially and hormonally complex organisms in which stimuli from a breeding resource, mating partners, rivals and young interact to alter the JH profile of breeding adults.     

Quantification of vitellogenin-mRNA during maturation and breeding of a burying beetle

Burying beetles (Nicrophorus orbicollis) are unusual in that to breed they require an unpredictable and valuable resource, a small carcass. Thus the timing of reproduction is unpredictable and beetles' physiological response must be fast. We hypothesized that their pattern of vitellogenin (Vg) synthesis might reflect these requirements. We examined the expression of two Vg genes (sequenced for this study) during sexual maturation and through a reproductive bout. Vg-mRNA, juvenile hormone (JH) titers, ovarian development, and hemolymph concentrations of Vg were quantified in the same individuals. All four variables gradually increased during maturation to peak 15-20 days after eclosion. Twelve hours after the discovery of a carcass, a few hours before oviposition, mRNA was high, hemolymph Vg had decreased, JH and ovarian weight had increased. After oviposition, mRNA was low, hemolymph Vg concentrations and JH were high. This is consistent with our hypothesis that beetles produce and store Vg in the hemolymph prior to the discovery of a breeding resource and replace it quickly. Partial regression of these variables (with the effect of time removed) indicated that JH was not correlated with mRNA, hemolymph Vg, or ovarian weight at any time. Thus the role of JH as a gonadotropin remains unclear.     

Social stimuli affect juvenile hormone during breeding in biparental burying beetles (Silphidae: Nicrophorus)

Extended biparental care is rare in insects but provides an excellent opportunity to investigate the interaction between the endocrine system and the physical and social environment in the regulation of this behavior. Burying beetles (Nicrophorus spp.) have facultative biparental care and depend on locating a small vertebrate carcass that they bury and prepare as food for their young. Commonly, both male and female Nicrophorus orbicollis remain in the burial chamber after eggs hatch to feed and guard the larvae. In both sexes, juvenile hormone (JH) rises rapidly in response to the discovery and assessment of the carcass; it returns to near baseline in 24 h; then in females it reaches very high titers at the onset of maternal care. In this paper, we investigate some social (presence of a mate, mating history, larval age) and environmental (carcass size) factors that may affect this endocrine profile. For females, neither the presence of a mate nor mating status (i.e., virginity) affected the initial rise of JH. However, the absence of a mate significantly depressed the JH rise in males. Eighty-seven percent of the single males buried the carcass like paired males but 87% also released pheromones to attract a mate. JH hemolymph titers in females whose broods were replaced every 24 h with newly hatch larvae were significantly higher than those of females rearing aging broods. Lastly, even though larger carcasses took longer to bury and prepare and oviposition was delayed, neither JH titers nor speed of ovarian development was affected by carcass size.     

Significance of haemolymph juvenile hormone titer changes in timing of migration and reproduction in adult Oncopeltus fasciatus

Previous studies had shown that both migratory flight and ovarian maturation in Oncopeltus fasciatus were stimulated by juvenile hormone (JH), yet the two behaviors were mutually exclusive. To understand the relationship of this hormone to these behaviors, haemolymph juvenile hormone titers were determined in both sexes of Oncopeltus throughout the adult stage by the Manduca pigmentation bioassay. Animals reared under long day conditions (17L:7D, 24°C) showed an immediate rise in haemolymph titers of JH after adult emergence whereas those reared in a short day photoperiod (12L:12D, 24°C) had a more gradual increase in hormone titers. Migratory flight behavior occurred during periods of intermediate hormone titers while oviposition did not begin until JH titers had reached their peak. It was concluded that lower JH titers normally stimulate flight in the prereproductive adult whereas higher titers are required for complete ovarian development. The corpus allatum in Oncopeltus thus coordinates migration and reproduction in response to the environmental cues of photoperiod, temperature, and food quality.     

Differential visual ornamentation between brood parasitic and parental cuckoos

The evolution of brood parasitism should affect adult phenotypic traits due to sexual selection as well as the parasite–host interactions, although it is rarely focused on. Sexual selection theory predicts extravagant secondary sexual characteristics in brood parasites whereas immature‐like modest sexual characteristics in parental species. This is because juvenile‐like immature traits can attract mates by exploiting parental care for young (i.e. attraction to young), and because the good parent process, which favours traits that signal parental care ability, would constrain the evolution of costly secondary sexual characteristics due to evolutionary trade‐offs between parental investment and sexually selected traits. Using a phylogenetic comparative approach, we studied plumage and bare‐part characteristics of adults in relation to brood parasitism in cuckoos (family Cuculidae), in which brood parasitism together with loss of parental care has evolved three times. As predicted, we found that nonparasitic cuckoos had plumage more similar to the juveniles than did brood parasitic cuckoos. Furthermore, nonparasitic cuckoos had a higher probability of having additional bare skin, that is a seemingly less costly, hatchling‐like trait, than did brood parasitic cuckoos. This finding further supports the link between parental care and sexual selection, although the influence of a parasite–host interaction cannot be excluded. The analysis of evolutionary pathways suggested interdependent evolution of additional bare skin and brood parasitism. Brood parasitism together with the loss of parental care may prevent the maintenance of a modest phenotype similar to the young, and vice versa in some cases.     

The role of juvenile hormone in competition and cooperation by burying beetles

Few studies have addressed the physiological mechanisms that modulate aggression in insects. In some social insects, there is a correlation of JH and aggression in colony defense and in the establishment of dominance, but only a few studies demonstrate a causal relationship. Burying beetles aggressively defend a breeding resource, a carcass, and juvenile hormone (JH) hemolymph titers increase rapidly upon the discovery of a carcass. In this study, I show that treatment with the JH analog, methoprene, in the absence of a carcass increases the probability of injuries from aggressive interactions, but treatment to one member of a pair of competing Nicrophorus orbicollis females does not increase the probability that she will win control of the resource. In addition, higher JH levels are not associated with greater competitive ability in communally breeding Nicrophorus tomentosus females. Treatment of one female N. tomentosus does not increase her share of the communal brood. Methoprene seems to make a less competitive female more persistent and less willing to concede, which, although maintaining her share of reproduction, results in her exclusion from the brood chamber.     

Identification of juvenile hormone III in the hemolymph of Blattella germanica adult females by gas chromatography-mass spectrometry

Juvenile hormone III was identified in purified hemolymph extracts of adult females of the cockroach Blatella germanica by gas chromatography-mass spectrometry with chemical ionization and selected ion monitoring. Under these conditions, juvenile hormones I and II were not detected within the sensitivity ranges of this analytical method. For quantification purposes a 5,5-bisdeuterated analog of juvenile hormone III was synthesized and used as an internal standard. In general, juvenile hormone III titers obtained correlated with data on oocyte growth and with hormone titers found from in vitro corpora allata incubations along the first gonotrophic cycle.     

Hormonal controls on reproduction in female heteroptera

A survey of the literature related to hormonal control of reproduction in female heteropterans reveals many common features. Juvenile hormone secreted by the corpus allatum is the principal hormone governing vitellogenesis. Feeding and nutrition affects egg development via juvenile hormone secreted by the corpus allatum, which is under inhibitory control from the brain. Information about feeding reaches the brain via a humoral route. Similarly, mating increases egg production by altering the degree of inhibition of the corpus allatum by the brain, although the nature of the mating stimulus and the route by which the stimulus reaches the brain remains uncertain. There may be one or more stimulatory factors acting on the corpus allatum. Progress in identifying controls on the corpus allatum is hindered by our ignorance of the identity of the juvenile hormone(s) acting in Heteroptera. Ovulation and oviposition are under the control of a myo-active peptide from the brain, and its release is governed by hormonal inputs from both mating and the ovary. The ovarian input appears to consist of ecdysteroid. A role for ovarian ecdysteroid in controlling the corpus allatum remains uncertain. Arch. Insect Biochem. Physiol. 35:443–453, 1997. © 1997 Wiley-Liss, Inc.     

Effects of experience and juvenile hormone on the organization of the mushroom bodies of honey bees

There is an age-related division of labor in the honey bee colony that is regulated by juvenile hormone. After completing metamorphosis, young workers have low titers of juvenile hormone and spend the first several weeks of their adult lives performing tasks within the hive. Older workers, approximately 3 weeks of age, have high titers of juvenile hormone and forage outside the hive for nectar and pollen. We have previously reported that changes in the volume of the mushroom bodies of the honey bee brain are temporally associated with the performance of foraging. The neuropil of the mushroom bodies is increased in volume, whereas the volume occupied by the somata of the Kenyon cells is significantly decreased in foragers relative to younger workers. To study the effect of flight experience and juvenile hormone on these changes within the mushroom bodies, young worker bees were treated with the juvenile hormone analog methoprene but a subset was prevented from foraging (big back bees). Stereological volume estimates revealed that, regardless of foraging experience, bees treated with methoprene had a significantly larger volume of neuropil in the mushroom bodies and a significantly smaller Kenyon cell somal region volume than did 1-day-old bees. The bees treated with methoprene did not differ on these volume estimates from untreated foragers (presumed to have high endogenous levels of juvenile hormone) of the same age sampled from the same colony. Bees prevented from flying and foraging nonetheless received visual stimulation as they gathered at the hive entrance. These results, coupled with a subregional analysis of the neuropil, suggest a potentially important role of visual stimulation, possibly interacting with juvenile hormone, as an organizer of the mushroom bodies. In an independent study, the brains of worker bees in which the transition to foraging was delayed (overaged nurse bees) were also studied. The mushroom bodies of overaged nurse bees had a Kenyon cell somal region volume typical of normal aged nurse bees. However, they displayed a significantly expanded neuropil relative to normal aged nurse bees. Analysis of the big back bees demonstrates that certain aspects of adult brain plasticity associated with foraging can be displayed by worker bees treated with methoprene independent of foraging experience. Analysis of the over-aged nurse bees suggests that the post-metamorphic expansion of the neuropil of the mushroom bodies of worker honey bees is not a result of foraging experience. © 1995 John Wiley & Sons, Inc.     

In vivo effects of allatostatins in crickets,Gryllus bimaculatus (Ensifera: Gryllidae)

Two types of cricket allatostatins, Grb-AST A1 and Grb-AST B1, were injected into adult female crickets three times each day on days 0–3 and once on day 4 after adult emergence to test their activity in vivo. On day 4, body weight, ovary weight, number of eggs per ovary, length of the terminal oocytes, ovarian ecdysteroid biosynthesis, and hemolymph titers of ecdysteroids were lower in the allatostatin-injected animals compared with untreated and Ringer-injected controls. Effects of the injected allatostatins on hemolymph juvenile hormone titers were inhomogeneous, and no differences were found in the capacity of the corpora allata to produce juvenile hormone ex vivo. The hemolymph titers of yolk proteins (vitellogenins) were almost twice as high in the allatostatin-injected animals as in the control animals. The effects of the injected allatostatins and their interactions with the endocrine system of the animal are discussed. Arch. Insect Biochem. Physiol. 38:32– 43, 1998. © 1998 Wiley-Liss, Inc.     

Interspecific interactions change the outcome of sexual conflict over prehatching parental investment in the burying beetle Nicrophorus vespilloides

Sexual conflict arises when the optimal reproductive strategy differs for males and females. It is associated with every reproductive stage, yet few studies have considered how the outcome may be changed by interactions with other species. Here, we show that phoretic mites Poecilochirus carabi change the outcome of sexual conflict over the supply of prehatching parental investment in the burying beetle Nicrophorus vespilloides. Burying beetles require a small dead vertebrate for reproduction, which they prepare by shaving it, rolling up the flesh, and burying it. When pairs were given a medium‐sized mouse to prepare (13–16 g), mites changed how the costs of reproduction were divided between the sexes, with males then sustaining greater costs than females. We found no equivalent difference when pairs prepared larger or smaller carcasses. Thus, our experiment shows that the outcome of sexual conflict over prehatching parental investment is changed by interactions with other species during reproduction.     

Seasonal changes in juvenile hormone titers and rates of biosynthesis in honey bees

Honey bee colonies can respond to changing environmental conditions by showing plasticity in age related division of labor, and these responses are associated with changes in juvenile hormone. The shift from nest taks to foraging has been especially well characterized; foraging is associated with high juvenile hormone titers and high rates of juvenile hormone biosynthesis, and can be induced prematurely in young bees by juvenile hormone treatment or by a shortage of foragers. However, very few studies have been conducted that study plasticity in division of labor under naturally occurring changes in the environment. To gain further insight into how the environment and juvenile hormone influence foraging behavior, we measured juvenile hormone titers and rates of biosynthesis in workers during times of the year when colony activity in temperate climates is reduced: late fall, winter, and early spring. Juvenile hormone titers and rates of biosynthesis decreased in foragers in the fall as foraging diminished and bees became less active. This demonstration of a natural drop in juvenile hormone confirms and extends previous findings when bees were experimentally induced to revert from foraging to within-hive tasks. In addition, endocrine changes in foragers in the fall are part of a larger seasonally related phenomenon in which juvenile hormone levels in younger, pre-foraging bees also decline in the fall and then increase the following spring as colony activity increases. The seasonal decline in juvenile hormone in foragers was mimicked in summer by placing a honey bee colony in a cold room for 8 days. This suggests that seasonal changes in juvenile hormone are not related to photoperiod changes, but rather to changes in temperature and/or colony social structure that in turn influence endocrine and behavioral development. We also found that active foragers in the late winter and early spring had lower juvenile hormone levels than active foragers in late spring. In light of recent findings of a possible link between juvenile hormone and neuroanatomical plasticity in the bee brain, these results suggest that bees can forage with low juvenile hormone, after previous exposure to some threshold level of juvenile hormone leads to changes in brain structure.     

Effects of social environment and worker mandibular glands on endocrine-mediated behavioral development in honey bees

Previous studies suggest that older honey bee workers possess an inhibitory signal that regulates behavioral development in younger bees. To study how this inhibitor is transmitted, bees were reared for 7 days in double-screen cages, single-screen cages, or unrestricted in a typical colony (control bees). Double-screen cages prevented physical contact with colony members while single-screen cages allowed only antennation and food exchange. Bees reared in double-screen cages showed accelerated endocrine and behavioral development; they had significantly higher rates of juvenile hormone biosynthesis and juvenile hormone titers than did control bees and also were more likely to become precocious foragers. Relative to the other two groups, bees reared in single-screen cages showed intermediate juvenile hormone biosynthesis rates and titers, and intermediate rates of behavioral development. These results indicate that physical contact is required for total inhibition. We also began to test the hypothesis that worker mandibular glands are the sources of an inhibitory signal. Old bees with mandibular glands removed were significantly less inhibitory towards young bees than were sham-operated and unoperated bees. These results suggest that an inhibitor is produced by the worker mandibular glands. Accepted: 29 January 1998     

Shared or Unshared Parental Care in Overwintering Brent Geese (Branta bernicla hrota)

We examine brood size effects on the behaviour of wintering parent and juvenile brent geese (Branta bernicla hrota) to test predictions of shared and unshared parental care models. The behaviour of both parents and offspring appear to be influenced by declining food availability over the winter. Parental vigilance increased with brood size and may be explained by vigilance having functions in addition to antipredator behaviour where the benefits are shared among the brood. There was no increase in parental aggression with brood size and this does not fit the prediction of shared care. Nevertheless, large families are able to monopolize better feeding areas compared with smaller families and large families static feed more but walk feed less than do small families, the former apparently being the preferred mode. The presence of additional young, rather than increasing the amount of parental aggression, seems to enhance the family's competitive ability. Because parents with large broods benefit from enhanced access to resources there is likely to be no additional significant cost in the parental care of larger broods (sensu Trivers 1972 ).     

Resource defense and juvenile hormone: the "challenge hypothesis" extended to insects

As predicted for vertebrates by the challenge hypothesis, the endocrine system of invertebrates can respond to social stimuli to modulate aggression. Testosterone (T) is generally considered to moderate aggression associated with reproduction, i.e. the establishment of breeding territories, mate guarding and offspring defense; juvenile hormone (JH) serves an analogous function in burying beetles. Hemolymph titers of JH increase significantly in Nicrophorus orbicollis, a species with facultative biparental care, when challenged by an intruder to defend their resource. During the first 12 h after the discovery of a carcass, the necessary breeding resource, competition is intrasexual, and JH of males responds only to a challenge by males, and JH of females responds only to a challenge by female intruders. After this period, competition is intersexual, and JH increases significantly in both males and females challenged by an intruder of either sex. In contrast, JH titers in a nonparental species are much higher throughout the breeding season, and neither males nor females respond hormonally to an intruder. These findings support the challenge hypothesis and suggest that mating systems and breeding strategies can promote plastic responses in insect, as well as vertebrate, endocrine systems.     

Difference in parenting in two species of burying beetle, <Emphasis Type="Italic">Nicrophorus orbicollis</Emphasis> and <Emphasis Type="Italic">Nicrophorus vespilloides</Emphasis>

Burying beetles (Nicrophorus) are model parents among insects, with all studied species known to regurgitate flesh from vertebrate carcasses to their offspring. However, most studies focus on a very few species, yet the interpretation of the function and importance of care is typically generalized to all burying beetles. Here we characterize subtle variation within and between individuals and sexes, and how this variation differs between two species of burying beetle. We find that Nicrophorus orbicollis exhibits low variance, with a normal distribution of parental care provided during peak care periods. In N. vespilloides, however, the distribution is more uniform as values of care are spread across the possible phenotypic spectrum. This suggests that there is stabilizing selection on care in N. orbicollis, but relaxed or disruptive selection in N. vespilloides. Although repeatability was similar between both species, transitions from other care behaviors into feeding were more common in N. orbicollis than N. vespilloides. Thus, while parenting is coarsely similar across the genus, variation in its expression should not be extrapolated to all Nicrophorus. We suggest that subtle variation both within and among species merits greater attention, and could inform us about the factors that lead to different distributions of care.