Synaptic organization in the adult honey bee brain is influenced by brood-temperature control during pupal development

Recent studies have shown that the behavioral performance of adult honey bees is influenced by the temperature experienced during pupal development. Here we explore whether there are temperature-mediated effects on the brain. We raised pupae at different constant temperatures between 29 and 37 degrees C and performed neuroanatomical analyses of the adult brains. Analyses focused on sensory-input regions in the mushroom bodies, brain areas associated with higher-order processing such as learning and memory. Distinct synaptic complexes [microglomeruli (MG)] within the mushroom body calyces were visualized by using fluorophore-conjugated phalloidin and an antibody to synapsin. The numbers of MG were different in bees that had been raised at different temperatures, and these differences persisted after the first week of adult life. In the olfactory-input region (lip), MG numbers were highest in bees raised at the temperature normally maintained in brood cells (34.5 degrees C) and significantly decreased in bees raised at 1 degrees C below and above this norm. Interestingly, in the neighboring visual-input region (collar), MG numbers were less affected by temperature. We conclude that thermoregulatory control of brood rearing can generate area- and modality-specific effects on synaptic neuropils in the adult brain. We propose that resulting differences in the synaptic circuitry may affect neuronal plasticity and may underlie temperature-mediated effects on multimodal communication and learning.     

Regulation of behavioral maturation by a primer pheromone produced by adult worker honey bees

Previous research showed that the presence of older workers causes a delayed onset of foraging in younger individuals in honey bee colonies, but a specific worker inhibitory factor had not yet been identified. Here, we report on the identification of a substance produced by adult forager honey bees, ethyl oleate, that acts as a chemical inhibitory factor to delay age at onset of foraging. Ethyl oleate is synthesized de novo and is present in highest concentrations in the bee's crop. These results suggest that worker behavioral maturation is modulated via trophallaxis, a form of food exchange that also serves as a prominent communication channel in insect societies. Our findings provide critical validation for a model of self-organization explaining how bees are able to respond to fragmentary information with actions that are appropriate to the state of the whole colony.     

Visual working memory in decision making by honey bees

The robustness and plasticity of working memory were investigated in honey bees by using a delayed matching-to-sample (DMTS) paradigm. The findings are summarized as follows: first, performance in the DMTS task decreases as the duration between the presentation of the sample stimulus and the presentation of the comparison stimuli is increased. This decrease is well approximated by an exponential decay function. Performance is significantly better than random-choice level even at delays as long as 5 sec and is reduced to random-choice levels at an average delay time of 8.68 +/- 0.06 sec. Second, when the DMTS task involves two samples (one relevant, the other irrelevant), bees can be trained to learn to use the relevant sample to perform the task if (i) the relevant sample is always at a fixed position, or (ii) the relevant sample always has the same place in the sequence of presentation (always first or always second). Bees that have learned to use the relevant sample and to ignore the irrelevant sample can generalize this learning, and apply it to novel sets of sample and comparison stimuli that they have never previously encountered. The findings point to a remarkably robust, and yet plastic, working memory in the honey bee.     

Kin discrimination by worker honey bees in genetically mixed groups

We tested the hypothesis that in a genetically mixed assemblage of worker honey bees, individual workers would behave differently toward unfamiliar sisters than toward unfamiliar nonsisters. Groups of worker honey bees of mixed genetic composition were assembled by collecting pupae from separate colonies and placing the worker bees together on eclosion. A total of 10 workers, 5 from each of two kin groups, were used to form each group. When the workers were 5 days old, a worker of one of the two kin groups was introduced into the mixed group. This worker had previously been held in a group of its sisters, without contact with queen or nonsister bees. The interactions with the introduced bee indicate that in a mixed kin group, individual workers learn the composite identity of the group and do not attack unfamiliar bees differentially on the basis of kinship. However, kinship does influence the total number of interactions in which an introduced bee engages when placed in a genetically mixed group; bees interacted significantly more often with sisters than with nonsisters. There was a trend for bees to be involved in more feeding interactions with sisters. This finding indicates an ability of a bee to learn and use its own cues. In mixed groups, each bee maintains its genotypically correlated identity; the bees' odors do not comingle into a “group” or “gestalt” odor. The significance of these results is discussed in light of the genetic structure of natural colonies of honey bees.     

Plasma cortisol response to stress in juvenile rainbow trout is influenced by their life history during early development and by egg cortisol content

In this study, the consequences of the exposure of rainbow trout (Oncorhynchus mykiss) to a brief stress during early development were explored on the later response of fingerlings to stress. Firstly, we analyzed the ontogeny of cortisol production and that of the initial cortisol response to stress in developing fish. It is only at the eyed stage that the embryos started to produce some basal cortisol. The HPI (hypothalamic-pituitary-interrenal axis) was however not functional before hatching, as exposure of the embryos to a stress did not trigger any cortisol response. A cortisol response to an acute stress was detected 9 days after hatching. In a second set of experiments, we showed that a very brief stress applied at 3 different early stages (eyed, hatching, and yolk resorption) reduced the later cortisol response to stress of 5-month-old fingerlings. This reduction is not likely to be due to alterations in the fish interrenal sensitivity because the 5-month-old fingerlings responded to ACTH treatment (only one dose tested) within the same magnitude as the fish that were not stressed during early development. An experimentally induced increase in egg cortisol just after fertilization also induced a reduction in stress sensitivity of 5 month old fingerlings, which was dose-dependent This study shows for the first time that the responsiveness of the corticotrope axis in 5-6 months old rainbow trout was influenced both by early stress exposure and by initial egg cortisol levels. Whether the HPI was functional or not at the time the initial stress was applied only had a small influence on the later unambiguous effect of early stress exposure.     

Galanin-like immunoreactivity is influenced by estrogen in peripubertal and adult rats.

Galanin gene expression in the anterior pituitary is potently stimulated by estrogen in adult rats. To evaluate the influence of estrogen on galanin during the peripubertal period 30- to 32-day-old female rats were treated with pregnant mare serum gonadotropin (PMSG, 10 IU s.c., 10.00 h). Galanin-like immunoreactivity (galanin-LI) in hypothalamic and pituitary tissues was evaluated 1, 2 or 3 days after PMSG treatment between 17.00 and 19.00 h. The PMSG treatment stimulated 17 beta-estradiol secretion, which induced a midafternoon LH surge 2 days after the PMSG treatment. Concentrations of galanin-LI at the time of this LH surge were elevated 82% in the anterior pituitary and 58% in the hypothalamus (without the median eminence) when compared to saline-treated female rats. On the 3rd day after the PMSG injection, galanin-LI was increased 236% in the anterior pituitary, 88% in the neurointermediate lobe and 39% in the median eminence compared to saline-treated female rats. These changes in galanin-LI were not observed in similarly aged male rats or ovariectomized rats treated with PMSG. In adult male rats, daily injections with 17 beta-estradiol valerate (10 micrograms/daily s.c.) for 1 week increased galanin-LI in the median eminence and neurointermediate lobe to an extent similar to that seen in juvenile female rats following PMSG treatment. In contrast, the high serum levels of 17 beta-estradiol achieved after 17 beta-estradiol valerate treatment increased galanin-LI in the anterior pituitary 65-fold. These studies indicate that galanin-LI is influenced by estrogen in peripubertal and adult rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cognitive aging is linked to social role in honey bees (Apis mellifera)

Aging is associated with cognitive impairment in numerous animal species. Across taxa, decline in learning performance is linked to chronological age. The honey bee (Apis mellifera), in contrast, offers an opportunity to study such aspects of aging largely independent of age per se. This is because foraging onset can be decoupled from chronological age, although workers typically first perform tasks inside the nest and later forage outside the hive. Further, early phases of foraging are characterized by growth of specific brain neuropiles, whereas late phases of the forager life-stage are accompanied by accelerated rates of physiological senescence. Yet, it is unclear if these patterns of senescence include cognitive function. The flexibility of worker ontogeny, however, suggests that the bee can become an attractive model for studies of plasticity in cognitive aging that ultimately may lead to insight into mechanisms that govern age-related cognitive decline. To address this potential, we studied effects of honey bee chronological age and of social role on sensory sensitivity and associative olfactory learning performance. Our results show a decline in olfactory acquisition performance that is linked to social role, but not to chronological age. This decline occurs only in foragers with long foraging duration, but at the same time the foragers show less generalization of odors, which is indicative of more precise learning. Foragers that are reversed from foraging to nest tasks, furthermore, do not show deficits in olfactory acquisition. These results point to complex effects of aging on associative learning in honey bees.     

Rate of coronary vascularization during embryonic chicken development is influenced by the rate of myocardial growth.

OBJECTIVE: We tested the hypothesis that the degree of coronary microvessel formation in the embryonic heart is regulated by the magnitude of myocardial growth. METHODS: The outflow tract of Hamburger-Hamilton stage 21 chicken hearts (prior to the onset of coronary vasculogenesis) was constricted in ovo with a loop of 10-0-nylon suture, and the hearts were studied at stages 29 and 36. RESULTS: At stage 29 ventricular mass was 64% greater in the pressure-overloaded than in the hearts of sham-operated controls, but vascular volume density and numerical density, determined by electron microscopic morphometry, were identical. As demonstrated by histological morphometric evaluation, the compact region of the left ventricle at stage 29 was 43% thicker than the shams. However, by stage 36 heart mass, thickness of the compact region, and overall wall thickness (demonstrated by scanning electron microscopy) were significantly less than in the sham group of this stage, but vascular volume density was virtually identical in the two groups. Formation of the two main coronary arteries was clearly impeded in the banded hearts, i.e., the coronaries were stunted in their development or failed to completely form coronary ostia. CONCLUSIONS: Vascular growth is proportional to myocardial growth in the embryonic, overloaded heart, but the persistence of the pressure overload results in a failure of or severe limitations in coronary artery development. These data support the hypothesis that vascular growth during this period of development is regulated, at least in part, by the rate and magnitude of myocardial growth.     

Development of sun compensation by honeybees: how partially experienced bees estimate the sun's course

Honeybees and some other insects, in learning the sun's course, behave as if they can estimate the sun's position at times of day when they have never seen it, but there are competing ideas about the computational mechanisms underlying this ability. In an approach to this problem, we provided incubator-reared bees with opportunities to fly and see the sun only during the late afternoon. Then, on a cloudy day, we allowed bees to fly for the first time during the morning and early afternoon, and we observed how they oriented their waggle dances to indicate their direction of flight relative to the sun's position. The clouds denied the bees a direct view of celestial orientation cues and thus forced them to estimate the sun's position on the basis of their experience on previous evenings. During the test days, experience-restricted bees behaved during the entire morning as if they expected the sun to be in an approximately stationary position about 180 degrees from the average solar azimuth that they had experienced on previous evenings; then from about local noon onward they used the evening azimuth. This pattern suggests that honeybees are innately informed of the general pattern of solar movement, such that they can generate an internal representation that incorporates spatial and temporal features of the sun's course that they have never directly seen.     

Comb wax mediates the acquisition of nest-mate recognition cues in honey bees

Honey bees, Apis mellifera, acquire nest-mate recognition cues from wax, the predominant material used in nest construction. Exposure of a newly emerged worker bee to wax-comb substrate significantly reduced the acceptability of that worker to sister bees. Cues acquired from the comb provided colony-specific information about the identity of worker bees; moreover, the effect of comb exposure has been previously shown to override individually produced cues. Food odors (anise oil), when dissolved in paraffin wax, affected worker-recognition characteristics but food odors did not affect these characteristics when fed to bees in sugar candy. Paraffin wax alone did not affect the recognition cues of bees, showing that the wax can be a neutral medium for the transmission of cues. The wax comb in the colony and the hydrocarbon outer layer of the bee cuticle may be a continuous medium for any hydrocarbon-soluble substances used by honey bees in nest-mate recognition; if so, a mechanism by which environmental cues are acquired by honey bees is provided.     

Reproduction in worker honey bees is associated with low juvenile hormone titers and rates of biosynthesis.

Three experiments were performed to determine the role of juvenile hormone (JH) in worker reproduction in queenless colonies of honey bees. In Experiment 1, egg-laying workers had low hemolymph titers of JH, as did bees engaged in brood care, while foragers had significantly higher titers. Experiment 2 confirmed these findings by demonstrating that laying workers have significantly lower rates of JH biosynthesis than foragers do. In Experiment 3, ovary development was inhibited slightly by application of the JH analog methoprene to 1-day-old bees, but was not affected by application to older bees, at least some already displaying egg-laying behavior. These results, which are consistent with earlier findings for queen honey bees, are contrary to a common model of insect reproduction, in which elevated JH titers trigger ovary development, which then leads to oviposition. Previous experiments have demonstrated that JH regulates nonreproductive behavior in workers that is associated with colony division of labor; perhaps this function is incompatible with a traditional role for JH in reproduction.     

Cellular degradation activity is maintained during aging in long-living queen bees

Queen honeybees (Apis mellifera) have a much longer lifespan than worker bees. Whether cellular degradation activity is involved in the longevity of queen bees is unknown. In the present study, cellular degradation activity was evaluated in the trophocytes and oenocytes of young and old queen bees. The results indicated that (i) 20S proteasome activity and the size of autophagic vacuoles decreased with aging, and (ii) there were no significant differences between young and old queen bees with regard to 20S proteasome expression or efficiency, polyubiquitin aggregate expression, microtubule-associated protein 1 light chain 3-II (LC3-II) expression, 70 kDa heat shock cognate protein (Hsc70) expression, the density of autophagic vacuoles, p62/SQSTM1 expression, the activity or density of lysosomes, or molecular target of rapamycin expression. These results indicate that cellular degradation activity maintains a youthful status in the trophocytes and oenocytes of queen bees during aging and that cellular degradation activity is involved in maintaining the longevity of queen bees.     

Plasma-sulfated C21-steroids increase during the periovulatory period in female common wolffish and are influenced by temperature during vitellogenesis

Plasma concentrations of steroids during the periovulatory period were measured in female common wolffish reared at three different temperatures. Steroids were quantified by radioimmunoassay (RIA). Two "broad-spectrum specificity" RIAs-one which detects C21-steroids with a 17,20beta-dihydroxyl configuration (17,20beta-steroids) and the other which detects C21-steroids with a 5beta-reduced, 3alpha-hydroxyl configuration (5beta,3alpha-steroids)-picked up very large amounts of cross-reacting material (1.7 microg ml(-1) in one fish) in the sulfate fraction of plasma from ovulating females. Reverse-phase high-performance liquid chromatography and thin-layer chromatography revealed two major steroids: 5beta-pregnane-3alpha,17,20beta-triol (80%) and 5beta-pregnane-3beta,17,20beta-triol (20%). The sulfated forms of these steroids were elevated 4 to 6 days before and during ovulation, compared with those of females in vitellogenic and postspawning condition, in which concentrations were below 2.0 ng ml(-1). In the three groups of fish held at 4, 8, and 12 degrees C during vitellogenesis, but returned to 4 degrees C just prior to the spawning season, the mean concentrations of sulfated 17,20beta-steroids in ovulating females were 530, 635, and 325 ng ml(-1), respectively. The corresponding concentrations of free 17,20beta-dihydroxy-4-pregnen-3-one (17,20beta-P; the maturation-inducing steroid in many teleosts) were 0.88, 0.86, and 0.57 ng ml(-1), respectively. Only minute amounts of 17,20beta,21-P and its sulfated derivatives were detected. Significantly lower steroid concentrations in the 12 degrees C group indicate that steroid synthesis and/or metabolism during the periovulatory period are influenced by the temperature experienced during vitellogenesis. In male fish, plasma concentrations of both sulfated 17,20beta-steroids and free 17,20beta-P were low (< 2.0 ng ml(-1)) at all times.     

Desacyl ghrelin is influenced by changes in insulin concentration during an insulin tolerance test

ObjectiveGhrelin, a gut-brain peptide, regulates energy homeostasis and glucose metabolism and is present in acylated and nonacylated form in the circulation. Although desacyl ghrelin (DAG), the predominant form of ghrelin, is associated with insulin sensitivity and improved metabolic state, not much is known about its direct regulation by insulin. We aimed to assess changes in DAG in response to the rapid increase in insulin concentration during an insulin tolerance test (ITT) in normal weight and obese subjects.DesignWe performed an observational single center study. An ITT was assessed in eight subjects (four males), median age of 29.9 years (range 19.6–42.0). DAG concentrations were measured at 20, 40, 60 and 90 min after insulin infusion. Homeostatic Model Assessment (HOMA) was calculated from fasting insulin and glucose. Body mass index (BMI) and waist circumference were assessed.ResultsThree subjects were obese (BMI ≥ 30 kg/m²), one subject was overweight (BMI = 25–30 kg/m²) and four subjects had normal weight (BMI = 18.5–25 kg/m²). Median DAG decreased after insulin infusion (90 pg/mL, p = 0.028), especially in normal weight subjects. Baseline DAG was lower in subjects with higher BMI (ρ = ? 0.76, p = 0.028) and higher fasting insulin (ρ = ? 0.76, p = 0.030). DAG changes correlated with fasting insulin levels (ρ = ? 0.85, p = 0.007), HOMA (ρ = ? 0.86, p = 0.007), BMI (ρ = ? 0.83, p = 0.010) and waist circumference (ρ = ? 0.93, p < 0.001).ConclusionDAG levels rapidly decreased in response to insulin administration in normal subjects, but not in insulin-resistant obese who are in a state of relative DAG deficiency.     

Fluorescent pigment and pentane production by lipid peroxidation in honey bees, Apis Mellifera

The concentration of fluorescent pigment associated with the oxidation of polyunsaturated lipids is significantly higher in old worker bees than in newly emerged worker adults. Pentane and hexane are produced by bees, presumably by decomposition of lipid hydroperoxides formed by peroxidation of membrane lipids. Protection against lipid peroxidation by vitamin E and glutathione peroxidase appears to be minimal in bees.     

Common endocrine and genetic mechanisms of behavioral development in male and worker honey bees and the evolution of division of labor.

Temporal polyethism is a highly derived form of behavioral development displayed by social insects. Hormonal and genetic mechanisms regulating temporal polyethism in worker honey bees have been identified, but the evolution of these mechanisms is not well understood. We performed three experiments with male honey bees (drones) to investigate how mechanisms regulating temporal polyethism may have evolved because, relative to workers, drones display an intriguing combination of similarities and differences in behavioral development. We report that behavioral development in drones is regulated by mechanisms common to workers. In experiment 1, drones treated with the juvenile hormone (JH) analog methoprene started flying at significantly younger ages than did control drones, as is the case for workers. In experiment 2, there was an age-related increase in JH associated with the onset of drone flight, as in workers. In experiment 3, drones derived from workers with fast rates of behavioral development themselves started flying at younger ages than drones derived from workers with slower rates of behavioral development. These results suggest that endocrine and genetic mechanisms associated with temporal polyethism did not evolve strictly within the context of worker social behavior.