Phenotypic deconstruction reveals involvement of manganese transporter malvolio in honey bee division of labor
1 Department of Entomology, University of Illinois at Urbana-Champaign, 320
Morrill Hall, 505 S. Goodwin Avenue, Urbana, IL 61801, USA
2 Neuroscience Program, University of Illinois at Urbana-Champaign, 320
Morrill Hall, 505 S. Goodwin Avenue, Urbana, IL 61801, USA
* Author for correspondence at present address: Howard Hughes Medical Institute, 500 EMRB, University of Iowa College of Medicine, Iowa City, IA 52242, USA (e-mail: yehuda-ben-shahar{at}uiowa.edu)
Accepted 21 June 2004
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Summary |
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Key words: mvl, Apis mellifera, foraging, sucrose response threshold, Hymenoptera.
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Introduction |
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One way to facilitate the molecular analysis of a complex behavioral
phenotype such as honey bee division of labor is to dissect it into simpler
behavioral components. For example, understanding the molecular mechanisms
underlying schizophrenia and other mental illnesses is simplified by
attempting to identify symptoms that are thought to represent components of
the disease that can each be studied independently; each of these symptoms is
an `endophenotype' of the whole disease
(Leboyer et al., 1998). Using
such an approach, we report on experiments that implicate increased manganese
transport by malvolio (mvl) into brain cells as one factor
influencing division of labor in honey bee colonies. mvl was
discovered in Drosophila melanogaster when screening for genes that
affect responsiveness to sucrose
(Rodrigues et al., 1995
); it
was later found to encode a manganese transmembrane transporter
(Orgad et al., 1998
;
Supek et al., 1996
). Flies
with mutations at the mvl locus showed reduced responsiveness to
sucrose, a deficit that was fully rescued by oral treatment with manganese
(Orgad et al., 1998
).
Manganese toxicity is known to negatively affect neural function, but the role
of manganese in natural neural and behavioral plasticity is poorly understood
(Takeda, 2003
; Takeda et al.,
2002
,
2003
;
Verity, 1999
).
We studied mvl and manganese transport for two reasons. First,
mvl and manganese influence responsiveness to sucrose in
Drosophila, and changes in sucrose responsiveness appear to be a
component of honey bee division of labor. In honey bees, there are genotypic
and phenotypic correlations between variation in sucrose responsiveness and
two aspects of division of labor, the age at onset of foraging and the
tendency to forage for either pollen or nectar
(Pankiw and Page, 1999).
Responsiveness to sucrose increases with age and is highest in foragers. In
addition, foragers that specialize on collecting pollen show higher sucrose
responsiveness than do nectar foragers. Bees selected for increased pollen
collection also show increased sucrose responsiveness and an earlier age at
onset of foraging (Page et al.,
1998
). African honey bees (Apis mellifera scutellata L.)
also show increased pollen collection, increased responsiveness to sucrose and
an earlier age at onset of foraging relative to bees derived from a mixture of
subspecies that originated in Europe
(Pankiw, 2003
). The causal
relationships between sucrose responsiveness and these two aspects of division
of labor are not understood, but the evidence for their association is
extensive.
The second reason we studied mvl is because an ortholog of another
gene involved in Drosophila feeding behavior, foraging
(Amfor), is also involved in honey bee behavioral maturation
(Ben-Shahar et al.,
2002a,b
).
Although the two genes have no known functional relationship, we wished to
explore further the idea (Ben-Shahar et
al., 2002b
) that some feeding-related genes in Drosophila
have been used in social evolution to regulate division of labor. Our goal was
to determine whether another gene that influences Drosophila feeding
behavior is also involved in honey bee division of labor, albeit in
association with a different behavioral component, sucrose responsiveness.
mvl has no known genetic or molecular connection to PKG
(cGMP-dependent protein kinase)-related pathways.
We tested the hypothesis that honey bee behavioral maturation is associated
with an increase in brain mvl expression, with foragers having higher
levels of mvl mRNA and manganese than nurses. We used manganese
treatment experiments to gain insight into whether mvl activity can
result in increased sucrose responsiveness, precocious foraging and increased
pollen foraging. We also performed similar experiments with cGMP (cyclic
guanosine monophosphate), a previously identified activator of foraging
behavior (Ben-Shahar et al.,
2002b), because it was not known whether this treatment also
affects sucrose responsiveness and pollen foraging. A fly genotype with higher
PKG activity shows increased sucrose responsiveness
(Scheiner et al., 2004
), but
treatment experiments with cGMP have not been performed.
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Materials and methods |
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Bees used to measure brain mRNA and head manganese levels were collected
from either triple-cohort colonies
(Ben-Shahar and Robinson, 2001)
or single-cohort colonies. Triple-cohort colonies were used to study typical
patterns of behavioral development, with foragers older than nurse bees. They
were established by sequentially introducing three cohorts of 800-1000
one-day-old bees to a small hive at one-week intervals. Each colony was also
given two frames of honeycomb for food storage and brood rearing and an
unrelated, naturally mated, queen. Nurse bees were identified as one-week-old
bees that inserted their heads into honeycomb cells containing larvae, and
foragers as bees older than three weeks of age returning to the hive with
either clearly visible pollen loads on their hind legs or distended abdomens
(bearing either nectar or water).
Single-cohort colonies were used to uncouple behavioral status and chronological age. They were established with one cohort of 800-1000 one-day-old bees; because these colonies initially contain no old bees, some colony members initiate foraging as much as two weeks earlier than usual, enabling us to sample precocious foragers and normal age nurses, all 5-9 days of age.
Treatments
Groups of 50 one-day-old bees were placed in a 6x12x18 cm
wooden cage placed in an incubator (33°C, 95% relative humidity) for 4
days. Bees were treated orally with a 50% sucrose solution containing either
20 mmol l-1 MnCl2, 100 mmol l-1
ZnCl2, 20 mmol l-1 MnCl2 + 100 mmol
l-1 ZnCl2, 500 mmol l-1 8-Br-cGMP or sucrose
alone as a control (all compounds from Sigma, St Louis, MO, USA). Zinc
treatments were used because zinc is a known antagonist of malvolio
and antagonizes the behavioral effects of manganese in Drosophila,
possibly by inhibiting its uptake by malvolio
(Orgad et al., 1998).
8-Br-cGMP was previously shown (Ben-Shahar
et al., 2002b
) to cause precocious foraging, but effects on
sucrose responsiveness were not examined. Oral treatment was used for two
reasons. First, this was the method used to rescue the mvl mutant
effect in Drosophila (Orgad et
al., 1998
). Second, this non-invasive method works well for
treating bees that are placed in colonies in the field to determine effects on
age at onset of foraging (Ben-Shahar et
al., 2002b
). Solutions were made fresh daily.
Sucrose responsiveness
After 4 days of treatment, caged bees from each treatment group were
cold-anesthetized and placed in individual restrainers for use in a sucrose
response assay (Ben-Shahar and Robinson,
2001). Bees were tested in a sequential series of increasing
sucrose concentration: 0, 0.1, 0.3, 3, 10 and 30% (w/v). A bee extends its
proboscis reflexively when the antenna is stimulated with sucrose
(Page et al., 1998
). We
recorded the number of times each bee extended its proboscis (0-6); greater
sucrose responsiveness is reflected by higher numbers of extensions. We
performed four independent trials of this experiment, each with bees derived
from several naturally mated queens. Bees from the different colony sources
were mixed and used randomly for each treatment. Data were analyzed with a
general linear model (GLM) using both treatment and trial as factors (SAS
Institute, Cary, NC, USA).
Age at onset of foraging
After 4 days of treatment in a cage in the laboratory (see above), all
surviving bees from each cage were counted (80-100% survival) and placed into
a single-cohort colony, made with 1000 one-day-old (untreated) bees and a
queen. Observations at the hive entrance were made as previously described
(Ben-Shahar et al., 2002b
) to
ensure that we observed the onset of foraging in each colony; observations
then occurred for 4 h day-1, 2 h in the morning and 2 h in late
afternoon, times of peak foraging activity for these colonies. All bees
initiating foraging during the first 7 days of observations were marked with a
second spot of paint on their abdomens (so they were counted just once), and
the cumulative percentage of bees that foraged (precociously) was calculated
for each group. We performed six independent trials of this experiment, each
with bees derived from several naturally mated queens. Differences in the
proportion of bees starting to forage from each treatment group were evaluated
with multiple factor survival analysis with Cox proportional hazards
estimation (Ben-Shahar et al.,
2002b
). After concluding behavioral observations, each colony was
killed (liquid nitrogen) to census the number of bees from each treatment
group present. Proportions of foragers were calculated on the basis of these
censuses.
Tendency to forage for nectar or pollen
In addition to the observations described in the previous paragraph, we
also recorded whether each forager returned with either pollen or nectar. To
analyze the effects of treatment on foraging behavior, the proportions of
foragers returning with either nectar or pollen were analyzed with PROC GENMOD
(SAS Institute), with colony and treatment as factors. Since manganese was the
only treatment that affected response threshold to sugar we also used the
Contrast function under PROC GENMOD to test the more specific hypothesis that
the effect of manganese on pollen foraging is different from all other
treatments.
Cloning of Amvl
We identified two ESTs (expressed sequence tags) from a honey bee brain EST
project
(http://titan.biotec.uiuc.edu/bee/honeybee_project.htm)
annotated as orthologs of Drosophila mvl (GenBank accession no.
AY526611). After additional sequencing of these cDNAs, the putative protein
sequence of the honey bee mvl ortholog (Amvl) showed more
than 80% similarity to Drosophila mvl
(Fig. 1).
|
Real-time quantitative RT-PCR
Procedures and statistical analysis were as previously described
(Ben-Shahar et al., 2002a).
Sequences for mvl-specific primers and TaqMan® probe are given in
Table 1. In all experiments, we
collected bees from the different behavioral groups - nurses, pollen foragers
and nectar foragers - according to established methods
(Ben-Shahar et al., 2002a
).
Amvl expression was normalized to an RNA loading control
`housekeeping' gene, the honey bee rp49 gene, as previously described
(Ben-Shahar et al., 2002a
).
Data were analyzed by two-way analysis of variance (ANOVA) with behavior and
trial as factors. Data were also analyzed with pair-wise post hoc
tests using a Bonferroni adjustment. Bees in each trial were collected from
independent colonies that were either single-cohort or triple-cohort colonies
(Ben-Shahar et al., 2002a
),
which were each established with bees of mixed genetic backgrounds from
different source colonies. We used a relative measure of mRNA fold
differences, with each colony analyzed as an independent experiment. Hence, it
was impossible to compare expression levels between colonies (or colony types)
on an absolute basis.
|
Manganese quantification
Manganese concentrations in bee heads were measured using non-destructive
neutron activation analysis (Landsberger,
1994). Single bee heads, weighing 2-3 mg, were placed individually
in polyethylene vials and irradiated in the TRIGA® (Training, Research,
Isotopes, General Atomics) research reactor at a thermal neutron flux of
4x1012 neutrons cm-2 s-1 for 10 min at
a power level of 950 kW. The neutron reaction
55Mn(n,
)56Mn was used for the analysis employing
the 846.7 keV gamma-ray with its 2.56 h half-life. To avoid any spectral
interference from the 843.3 keV gamma ray belonging to 28Mg and its
9.45 min half-life, and to increase sensitivity by allowing other short-lived
isotopes to die away, a decay time of 2-4 h was used. To avoid any possible
manganese contamination from the original irradiated vial, all samples were
transferred into inert vials after irradiation. Calibration was done using a
National Institute of Standards and Technology certified biological reference
material, NIST 1575 Tomato Leaves with a manganese value of 675±15
p.p.m. For quality control, NIST 1575 Pine Needles were analyzed for
manganese. Our results of 203±5 p.p.m., 236±5 p.p.m. and
230±5 p.p.m. are in agreement with the certified value of 238±7
p.p.m. Differences in manganese levels were evaluated with ANOVA with a
post hoc test and a Bonferroni adjustment.
In-situ hybridization
Procedures and conditions were as previously described
(Ben-Shahar et al., 2002b).
Briefly, freshly dissected brains were immediately freeze-mounted on dry ice
with anterior side (identified by antennal lobes) up and transferred to the
cryostat (Bright Inst. Co., Huntingdon, UK; -20°C). Brains were sectioned
(12 µm) and dry-mounted on glass slides. Hybridization was performed in 50%
formamide buffer with a digoxygenin-labeled anti-sense RNA probe (Roche,
Basel, Switzerland) at 60°C. Sense probe was used as control. Expression
patterns were studied in brains from three nurses and two pollen and nectar
foragers of typical ages. The cloned Apis mvl was used as a template
for in vitro transcription of riboprobes (827 bp long).
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Results |
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Foragers had higher levels of manganese in their heads than did nurse bees (Fig. 2C). Bonferroni post hoc analysis (P<0.05) indicated that manganese levels in pollen foragers were higher than nurses, with nectar foragers exhibiting intermediate levels. Levels of manganese in bees from single-cohort colonies also varied significantly (P<0.02) with behavior. Bonferroni post hoc analysis indicated that manganese levels in pollen foragers were higher than in nurses, with no difference between nectar foragers and nurses.
In situ hybridization analysis revealed that Amvl is
widely expressed in the honey bee brain
(Fig. 3). High expression
levels were observed in the antennal lobes and the subesophageal ganglion. In
contrast to Amfor, a previously identified gene affecting foraging
behavior (Ben-Shahar et al.,
2002b), Amvl was not highly expressed in the mushroom
bodies. There were no obvious spatial differences between nurses and foragers
in expression patterns. It is thus likely that the foraging-related increase
detected with qRT-PCR was mainly the result of increased expression in the
same cells rather than additional neurons expressing this gene.
|
Effects of manganese, zinc and cGMP treatments on sucrose responsiveness
Treating bees with manganese caused a significant increase in
responsiveness to sucrose (Fig.
4A). This effect was not seen in bees treated with zinc, manganese
plus zinc, or cGMP (Fig. 4A).
Manganese-treated bees showed a significant increase in head manganese levels
(Fig. 4B), suggesting that the
treatments were effective in elevating manganese levels in the brain.
|
Effects of manganese, zinc and cGMP treatments on age at onset of foraging
Manganese treatments caused precocious foraging in honey bee colonies. An
even stronger effect was seen in bees treated with cGMP; Ben-Shahar et al.
(2002b) also reported a strong
effect of cGMP treatment. There was no effect of zinc or manganese plus zinc
on age at onset of foraging (Fig.
5A; multifactorial survival analysis; treatment,
P<0.001; colony, P<0.004). Although we started our
treatment experiments with equal amounts of treated bees, a final census
revealed varying amounts of bees from each group present in the colony. Bees
treated with 8-Br-cGMP tend to attempt to initiate flight almost immediately
upon being introduced to the colony, which may explain their somewhat smaller
numbers in the final census.
|
Effects of manganese, zinc and cGMP treatments on the tendency to forage for nectar or pollen
There was a marginal overall effect of treatment on the proportion of
pollen foragers in each colony (Fig.
5B; P=0.047). Manganese treatment was significantly
different from all other treatments (P<0.01; Contrast analysis;
PROC GENMOD), but there were strong colony differences (P<0.001)
and no consistent trends within each colony.
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Discussion |
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Manganese deficiency results in a variety of neural deficits, perhaps
mediated by AMPA and NMDA receptor functions
(Takeda, 2003;
Takeda et al., 2002
) or other
types of receptors or ion channels (Wang
et al., 2003
). Manganese may also function in brain metabolism as
a cofactor for enzymes such as superoxide dismutases
(Zelko et al., 2002
).
mvl-mediated transport is apparently not the only way manganese can
enter a cell because, in Drosophila, mutations of the mvl
locus are not lethal and behavioral defects are rescued by manganese treatment
(Orgad et al., 1998
;
Rodrigues et al., 1995
). Also,
some evidence suggests that manganese ions can also permeate the cell membrane
via voltage-gated calcium channels
(Nasu, 1995
). However, the
observation that zinc, a known antagonist of mvl
(Orgad et al., 1998
;
Supek et al., 1996
),
attenuated the behavioral effect of manganese suggests that manganese
transport via mvl is a primary route into brain cells, either
neurons, glia or both.
Our results suggest that mvl-mediated manganese transport is
involved in the response to a rewarding stimulus such as sucrose. Manganese is
thought to function in the mammalian dopaminergic system, which plays a
central role in regulating the response to various types of pleasurable
stimuli (Salamone et al.,
2003). Similar to the role of dopamine in mammals, octopamine is
associated with the sucrose reward system in honey bees
(Hammer and Menzel, 1998
;
Menzel et al., 1999
), and this
neurochemical has been implicated in both honey bee responsiveness to sucrose
(Page et al., 1998
;
Pankiw and Page, 2003
) and
division of labor. Octopamine levels are higher in forager honey bees,
especially in the antennal lobes (Schulz
and Robinson, 1999
), and octopamine treatment causes precocious
foraging (Schulz and Robinson,
2001
). In addition, we found mvl expressed in the
subesophageal ganglion, and cells in this neuropil have been shown to be both
octopaminergic and responsive to sucrose reward
(Hammer and Menzel, 1998
;
Schroter and Menzel, 2003
).
Perhaps manganese effects in flies and bees are mediated by this
neuromodulatory system.
Manganese treatment showed a strong association between responsiveness to
sucrose and age at onset of foraging, but the association with foraging
specialization was weaker. Our results with different colony types suggest the
possibility of an interaction between bee age and foraging specialization;
however, such an age effect has not yet been detected in other studies (see
Pankiw and Page, 1999).
Perhaps we failed to detect a stronger effect of manganese treatment on pollen
foraging in typical-age foragers because we sampled too coarsely (only on the
bees' first foraging trips) or because the tendency to collect nectar or
pollen is influenced by a variety of colony and environmental factors
(Seeley, 1995
) that we could
not control in this experiment. Tests under more artificial conditions might
be more appropriate. Another possibility is that a more chronic treatment,
extending throughout the bees' foraging career, might have better revealed
effects of manganese on pollen foraging; in this study, the bees were treated
for only the first four days of adulthood, prior to the initiation of their
foraging career.
Our findings illustrate that molecular analysis of a complex behavioral
phenotype such as honey bee division of labor is facilitated by dissecting it
into simpler behavioral components. The foraging gene influences
honey bee behavioral maturation at least in part via effects on
phototaxis (Ben-Shahar et al.,
2002a,b
),
and mvl appears to influence behavioral maturation at least in part
via effects on responsiveness to sucrose. The results of manganese
treatment support the notion that responsiveness to sucrose is related in some
way to behavioral maturation in honey bees. However, the causal relationships
between them are not understood. Bees that specialize in collecting pollen
show increased responsiveness to sucrose in the laboratory relative to bees
that specialize in collecting nectar
(Pankiw and Page, 1999
); it is
not clear how increased responsiveness to sucrose causes pollen foraging.
Either an increase in responsiveness in some way facilitates the ability of
bees to leave the hive and collect food, especially pollen, or the increase in
responsiveness is itself associated with another behavioral change that is
more causally related to the transition to foraging behavior.
Previous findings have shown genotypic and phenotypic correlations between
variation in responsiveness to sucrose and two aspects of division of labor in
honey bees: the age at onset of foraging and the tendency to forage for either
pollen or nectar (Pankiw and Page,
1999). Our results demonstrate that, while there are molecular
pathways common to sucrose responsiveness and division of labor, linkages
between them can be dissociated. cGMP treatment affected age at onset of
foraging and phototaxis (Ben-Shahar et al.,
2002b
) but did not affect responsiveness to sucrose in the current
study. We also showed that manganese treatment affected both responsiveness to
sucrose and age at onset of foraging, but the association with foraging
specialization was weaker. Other studies have shown that cAMP treatments
increase sucrose responsiveness but do not affect age at onset of foraging
(Ben-Shahar et al., 2002b
;
Scheiner et al., 2003
), while
juvenile hormone affects both (Pankiw and
Page, 2003
; Schulz et al.,
2002
). It is not known how many independent molecular pathways in
the brain are involved in the regulation of honey bee behavioral maturation.
The results presented here and elsewhere
(Ben-Shahar et al., 2002a
;
Whitfield et al., 2003
)
suggest that there are multiple independent pathways. This is consistent with
the fact that honey bee behavioral maturation involves changes in
responsiveness to stimuli in various modalities, in addition to changes in
other neural and physiological processes
(Robinson, 2002
).
There are now two genes involved in Drosophila feeding behavior
that have been implicated in controlling the age at onset of foraging in honey
bees: malvolio (present study) and foraging (Ben-Shahar et
al.,
2002a,b
).
These results support the idea that some genes that are involved in
feeding-related behaviors in Drosophila have been used in social
evolution to regulate honey bee division of labor. It is reasonable to assume
that the evolution of social behavior acted, in part, on conserved mechanisms
that control responses to other stimuli in the environment
(Robinson and Ben-Shahar,
2002
). Social cues, like other environmental cues, convey
information critical for animal survival and reproduction. Genes involved in
orchestrating the perception and processing of sensory information and the
responses that are then triggered
(Robinson and Ben-Shahar,
2002
) are thus likely to figure prominently in social evolution.
Further studies on mvl and other genes that are involved in feeding
behavior in Drosophila may provide important insights into the
regulation of division of labor, as well as to the neural mechanisms
underlying the regulation of food intake.
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Acknowledgments |
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Footnotes |
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References |
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Becker, J. B., Breedlove, S. M. and Crews, D. (1992). Behavioral Endocrinology. Cambridge, MA: MIT Press.
Ben-Shahar, Y. and Robinson, G. E. (2001). Satiation differentially affects performance in a learning assay by nurse and forager honey bees. J. Comp. Physiol. A 187,891 -899.[Medline]
Ben-Shahar, Y., Leung, H.-T., Pak, W. L., Sokolowski, M. B. and Robinson, G. E. (2002a). cGMP-dependent changes in phototaxis: a possible role for the foraging gene in honey bee division of labor. J. Exp. Biol. 206,2507 -2515.[CrossRef]
Ben-Shahar, Y., Robichon, A., Sokolowski, M. B. and Robinson, G.
E. (2002b). Influence of gene action across different time
scales on behavior. Science
296,741
-744.
Hammer, M. and Menzel, R. (1998). Multiple
sites of associative odor learning as revealed by local brain microinjections
of octopamine in honeybees. Learn. Mem.
5, 146-156.
Landsberger, S. (1994). Delayed instrumental neutron activation analysis. In Chemical Analysis by Nuclear Methods (ed. Z. B. Alfassi), pp.121 -142. New York: John Wiley & Sons.
Leboyer, M., Bellivier, F., Nosten-Bertrand, M., Jouvent, R., Pauls, D. and Mallet, J. (1998). Psychiatric genetics: search for phenotypes. Trends Neurosci. 21,102 -105.[CrossRef][Medline]
Menzel, R., Heyne, A., Kinzel, C., Gerber, B. and Fiala, A. (1999). Pharmacological dissociation between the reinforcing, sensitizing, and response-releasing functions of reward in honeybee classical conditioning. Behav. Neurosci. 113,744 -754.[CrossRef][Medline]
Nasu, T. (1995). Actions of manganese ions in contraction of smooth muscle. Gen. Pharmacol. Vasc. Syst. 26,945 -953.[CrossRef]
Orgad, S., Nelson, H., Segal, D. and Nelson, N.
(1998). Metal ions suppress the abnormal taste behavior of the
Drosophila mutant malvolio. J. Exp.
Biol. 201,115
-120.
Page, R. E., Jr, Erber, J. and Fondrk, M. K. (1998). The effect of genotype on response thresholds to sucrose and foraging behavior of honey bees (Apis mellifera L.). J. Comp. Physiol. A 182,489 -500.[CrossRef][Medline]
Pankiw, T. (2003). Directional change in a suite of foraging behaviors in tropical and temperate evolved honey bees (Apis mellifera L.). Behav. Ecol. Sociobiol. 54,458 -464.[CrossRef]
Pankiw, T. and Page, R. E., Jr (1999). The effect of genotype, age, sex, and caste on response thresholds to sucrose and foraging behavior of honey bees (Apis mellifera L.). J. Comp. Physiol. A 185,207 -213.[Medline]
Pankiw, T. and Page, R. E., Jr (2003). Effect of pheromones, hormones, and handling on sucrose response thresholds of honey bees (Apis mellifera L.). J. Comp. Physiol. A 189,675 -684.
Robinson, G. E. (2002). Genomics and integrative analyses of division of labor in honeybee colonies. Am. Nat. 160,S160 -S172.[CrossRef]
Robinson, G. E. and Ben-Shahar, Y. (2002). Social behavior and comparative genomics: new genes or new gene regulation? Genes Brain Behav. 1,197 -203.[CrossRef][Medline]
Rodrigues, V., Cheah, P. Y., Ray, K. and Chia, W. (1995). malvolio, the Drosophila homologue of mouse NRAMP-1 (Bcg), is expressed in macrophages and in the nervous system and is required for normal taste behaviour. EMBO J. 14,3007 -3020.[Abstract]
Salamone, J. D., Correa, M., Mingote, S. and Weber, S. M.
(2003). Nucleus accumbens dopamine and the regulation of effort
in food-seeking behavior: implications for studies of natural motivation,
psychiatry, and drug abuse. J. Pharmacol. Exp. Ther.
305, 1-8.
Scheiner, R., Muller, U., Heimburger, S. and Erber, J. (2003). Activity of portein kinase A and gustatory responsiveness in the honey bee (Apis mellifera A.). J. Comp. Physiol. A 189,427 -434.
Scheiner, R., Sokolowski, M. B. and Erber, J.
(2004). Activity of cGMP-dependent protein kinase (PKG) affects
sucrose responsiveness and habituation in Drosophila melanogaster.
Learn. Mem. 11,303
-311.
Schroter, U. and Menzel, R. (2003). A new ascending sensory tract to the calyces of the honeybee mushroom body, the subesophageal-calycal tract. J. Comp. Neurol. 465,168 -178.[CrossRef][Medline]
Schulz, D. J. and Robinson, G. E. (1999). Biogenic amines and division of labor in honey bee colonies: behaviorally related changes in the antennal lobes and age-related changes in the mushroom bodies. J. Comp. Physiol. A 184,481 -488.[CrossRef][Medline]
Schulz, D. J. and Robinson, G. E. (2001). Octopamine influences division of labor in honey bee colonies. J. Comp. Physiol. A 187,53 -61.[Medline]
Schulz, D. J., Sullivan, J. P. and Robinson, G. E. (2002). Juvenile hormone and octopamine in the regulation of division of labor in honey bee colonies. Horm. Behav. 42,222 -231.[CrossRef][Medline]
Seeley, T. D. (1995). The Wisdom of the Hive: the Social Physiology of Honey Bee Colonies. Cambridge, MA: Harvard University Press.
Supek, F., Supekova, L., Nelson, H. and Nelson, N.
(1996). A yeast manganese transporter related to the macrophage
protein involved in conferring resistance to mycobacteria. Proc.
Natl. Acad. Sci. USA 93,5105
-5110.
Takeda, A. (2003). Manganese action in brain function. Brain Res. Rev. 41, 79-87.[Medline]
Takeda, A., Sotogaku, N. and Oku, N. (2002). Manganese influences the levels of neurotransmitters in synapses in rat brain. Neuroscience 114,669 -674.[CrossRef][Medline]
Takeda, A., Sotogaku, N. and Oku, N. (2003). Influence of manganese on the release of neurotransmitters in rat striatum. Brain Res. 965,279 -282.[CrossRef][Medline]
Verity, M. A. (1999). Manganese neurotoxicity: a mechanistic hypothesis. Neurotoxicology 20,489 -497.[Medline]
Wang, J., Luthey-Schulten, Z. A. and Suslick, K. S.
(2003). Is the olfactory receptor a metalloprotein?
Proc. Natl. Acad. Sci. USA
100,3035
-3039.
Whitfield, C. W., Cziko, A. M. and Robinson, G. E.
(2003). Gene expression profiles in the brain predict behavior in
individual honey bees. Science
302,296
-299.
Zelko, I. N., Mariani, T. J. and Folz, R. J. (2002). Superoxide dismutase multigene family: a comparison of the CuZn-SOD (SOD1), Mn-SOD (SOD2), and EC-SOD (SOD3) gene structures, evolution, and expression. Free Radic. Biol. Med. 33,337 -349.[CrossRef][Medline]