Developmental study of afferented and deafferented bee antennal lobes

The role of antennal sensory projections on the ontogeny of the bee antennal lobe was analyzed using both light and transmission electron microscopy. Normal and deafferented developing antennal lobes were examined. The results obtained show that (1) initiation of synaptogenesis in the antennal lobe is independent of the arrival of sensory inputs; (2) sensory inputs are necessary for setting up the glomerular antennal lobe organization; (3) regressive events, such as the reduction of synapse density, occur during the development of the antennal lobe; and (4) glomeruli formation appears as related to glia development.     

Comparative study of the antennal lobes and their afferent pathway in the worker bee and the drone (Apis mellifera)

Summary The topography of the antennal afferent path-ways was studied comparatively in the worker bee and the drone of Apis mellifera L. by cellular marking, following localized application of cobalt chloride to the cut end of one antenna. This study dealt principally with the first relay of the afferent pathway in the glomeruli of the antennal lobe. The counting and measurement of all the glomeruli were performed on 5 worker bees and 5 drones. An important sexual dimorphism, represented by 4 large and easily identifiable glomerular complexes, was demonstrated in the drone. In both worker bee and drone, four main regions of the glomerular neuropil were distinguished according to corresponding afferent bundles. The worker possessed 166 glomeruli and the drone 103. The number, position and dimensions of the glomeruli indicated that the glomerular organization was unvarying in worker bees and in drones. Concerning the internal structure of the glomeruli, two types were distinguished: the great majority (95%) exhibited a cortical layer, whereas in the 7 posterior glomeruli the synaptic fields of association seemed to be scattered throughout the whole volume. The main results of this work (glomerular invariance, sexual dimorphism) support the hypothesis of the functional unit of the glomeruli.Unité de Recherche Associée au CNRS, UA 483     

用激光共聚焦扫描技术研究昆虫触角叶的雌雄异构性

【目的】本研究探讨用激光共聚焦扫描显微镜对昆虫触角叶内结构的扫描技术。【方法】选取鳞翅目斜纹夜蛾Spodoptera litura, 蜚蠊目美洲大蠊Periplaneta americana和鞘翅目松墨天牛Monochamus alternatus, 仔细解剖得到昆虫完整脑组织, 经过Lucifer yellow染色、戊二醛固定、梯度酒精脱水和透明等一系列处理后, 用激光共聚焦扫描显微镜对昆虫触角叶结构进行分层扫描。【结果】结果显示: 经该方法处理后在激发光488 nm下能清晰扫描出昆虫触角内典型结构神经纤维球, 并且可清晰看到这3种昆虫雄性触角叶结构内的扩大型神经纤维球复合体(macroglomerular complex, MGC), 而在相应雌性昆虫体内都没有此复合体。另外通过5 μm分层扫描得到斜纹夜蛾、美洲大蠊和松墨天牛的触角叶平均厚度分别为130, 235和115 μm, 神经纤维球数量分别为35, 59和39个。【结论】激光共聚焦扫描技术是获得昆虫触角叶内部结构的一个可行方法。     

Postembryonic development of the antennal lobes in Periplaneta americana L.

Summary The postembryonic development of the antennal lobes of Periplaneta americana L. was examined with light- and electron-microscopical methods. There is no difference in the number of glomeruli and neurons in the antennal lobes of larval and adult animals. At hatching, the first larva already possesses the adult number of approximately 125 glomeruli and 500 to 560 deutocerebral neurons in the dorsolateral cell group of each antennal lobe. During postembryonic development the volume of the deutocerebral neurons increases three- to fourfold. The glomeruli of the first larva have about 7 % of the volume of the corresponding adult glomeruli. Since number, pattern, and size ratio of glomeruli (with the exception of the macroglomerulus) are constant in all larval stages and adult animals, it is possible to identify individual glomeruli. During the whole postembryonic development the ordinary glomeruli show a continuous volume increase, which parallels the increase in antennal sensory input. The macroglomerulus develops by way of special growth of two to four neuropil units, but not before the last three to four larval stages and only in males. Its growth precedes the formation of antennal pheromone receptors during the final molt; these receptors are known to project into the macroglomerulus. The development of the macroglomerulus in the last larval stages of the male may be caused by a genetically fixed growth program of specific deutocerebral neurons.Supported by the Deutsche Forschungsgemeinschaft (Scha 291/1)     

Development of neuronal connectivity in Drosophila antennal lobes and mushroom bodies

Recent advances in the study of the connectivity of Drosophila olfactory system include the demonstration that olfactory receptor neurons project to specific glomeruli according to the receptor type they express, and that their projection neuron partners are prespecified to innervate particular glomeruli by birth order or time. This same theme of sequential generation has been observed in the generation of the three major types of mushroom body neurons.     

Glomerular interactions in olfactory processing channels of the antennal lobes

An open question in olfactory coding is the extent of interglomerular connectivity: do olfactory glomeruli and their neurons regulate the odorant responses of neurons innervating other glomeruli? In the olfactory system of the moth Manduca sexta, the response properties of different types of antennal olfactory receptor cells are known. Likewise, a subset of antennal lobe glomeruli has been functionally characterized and the olfactory tuning of their innervating neurons identified. This provides a unique opportunity to determine functional interactions between glomeruli of known input, specifically, (1) glomeruli processing plant odors and (2) glomeruli activated by antennal stimulation with pheromone components of conspecific females. Several studies describe reciprocal inhibitory effects between different types of pheromone-responsive projection neurons suggesting lateral inhibitory interactions between pheromone component-selective glomerular neural circuits. Furthermore, antennal lobe projection neurons that respond to host plant volatiles and innervate single, ordinary glomeruli are inhibited during antennal stimulation with the female’s sex pheromone. The studies demonstrate the existence of lateral inhibitory effects in response to behaviorally significant odorant stimuli and irrespective of glomerular location in the antennal lobe. Inhibitory interactions are present within and between olfactory subsystems (pheromonal and non-pheromonal subsystems), potentially to enhance contrast and strengthen odorant discrimination.     

Serotonin-immunoreactive neurons in the antennal lobes and suboesophageal ganglion of the honeybee

Summary We have used immunohistochemical methods to investigate the morphology of identified, presumptive serotonergic neurons in the antennal lobes and suboesophageal ganglion of the worker honeybee. A large interneuron (deutocerebral giant, DCG) is described that interconnects the deutocerebral antennal and dorsal lobes with the suboesophageal ganglion and descends into the ventral nerve chord. This neuron is accompanied by a second serotonin-immunoreactive interneuron with projections into the protocerebrum. Two pairs of bilateral immunoreactive serial homologues were identified in each of the three suboesophageal neuromeres and were also found in the thoracic ganglia. With the exception of the frontal commissure, no immunoreactive processes could be found in the peripheral nerves of the brain and the suboesophageal ganglion. The morphological studies on the serial homologues were extended by intracellular injections of Lucifer Yellow combined with immunofluorescence.     

Distribution of neuropeptides in the antennal lobes of male Spodoptera littoralis

Olfaction is an important sensory modality that regulates a plethora of behavioural expressions in insects. Processing of olfactory information takes place in the primary olfactory centres of the brain, namely the antennal lobes (ALs). Neuropeptides have been shown to be present in the olfactory system of various insect species. In the present study, we analyse the distribution of tachykinin, FMRFamide-related peptides, allatotropin, allatostatin, myoinhibitory peptides and SIFamide in the AL of the male Egyptian cotton leafworm, Spodoptera littoralis. Immunocytochemical analyses revealed that most neuropeptides were expressed in different subpopulations of AL neurons. Their arborisation patterns within the AL suggest a significant role of neuropeptide signalling in the modulation of AL processing. In addition to local interneurons, our analysis also revealed a diversity of extrinsic peptidergic neurons that connected the antennal lobe with other brain centres. Their distributions suggest that extrinsic neurons perform various types of context-related modulation.     

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

Levels of the biogenic amines dopamine, serotonin, and octopamine were measured in different brain regions of adult worker honey bees as a function of age-related division of labor, using social manipulations to unlink age and behavioral state. In the antennal lobes, foragers had higher levels of all three amines than nurses, regardless of age. Differences were larger for octopamine than serotonin or dopamine. In the mushroom bodies, older bees had higher levels of all three amines than younger bees, regardless of behavioral state. These correlative results suggest that increases in octopamine in the antennal lobes may be particularly important in the control of age-related division of labor in honey bees. Accepted: 10 February 1999     

Pheromone-evoked potentials and oscillations in the antennal lobes of the sphinx moth Manduca sexta

Using intra- and extracellular recording methods, we studied the activity of pheromone-responsive projection neurons in the antennal lobe of the moth Manduca sexta. Intracellularly recorded responses of neurons to antennal stimulation with the pheromone blend characteristically included both inhibitory and excitatory stages of various strengths. To observe the activity of larger groups of neurons, we recorded responses extracellularly in the macroglomerular complex of the antennal lobe. The macroglomerular complex is part of a specialized olfactory subsystem and the site of first-order central processing of sex-pheromonal information. Odors such as the pheromone blend and host-plant (tobacco) volatiles gave rise to evoked potentials that were reproducible upon repeated antennal stimulation. Evoked potentials showed overriding high-frequency oscillations when the antenna was stimulated with the pheromone blend or with either one of the two key pheromone components. The frequency of the oscillations was in the range of 30–50 Hz. Amplitude and frequency of the oscillations varied during the response to pheromonal stimulation. Recording intracellular and extracellular activity simultaneously revealed phase-locking of action potentials to potential oscillations. The results suggest that the activity of neurons of the macroglomerular complex was temporally synchronized, potentially to strengthen the pheromone signal and to improve olfactory perception. Accepted: 19 December 1997     

Immunocytochemistry of GABA in the antennal lobes of the sphinx moth Manduca sexta

Summary The ganglionated plexus of the trachea of mice was studied quantitatively with a histochemical method that stains electively the ganglion nerve cells in whole-mount preparations. The plexus lies exclusively over the muscular part of the trachea, dorsal to the muscle itself, and it varies considerably in pattern and extent between individual animals. In young adult mice the plexus contains on average 235 neurons, occurring singly or gathered in packed ganglia. The ganglion neurons are relatively small, the profile area of three quarters of them measuring between 150 and 275 m² with an average of 251 m². In ageing mice the average number of ganglion neurons is the same as in young animals; however, cell sizes are markedly increased, the average being 341 m². Among the ultrastructural features of the ganglia, is a capsule (perineurium) of very regular structure, the presence of collagen, capillaries and myelinated axons inside the ganglia, and the presence of only few and short dendrites, some of which are abutted by synapsing nerve endings.     

Synaptic connections of dopamine-immunoreactive neurons in the antennal lobes of Periplaneta americana. Colocalization with GABA-like immunoreactivity

Summary Dopamine-like immunoreactivity was demonstrated histochemically in about ten local interneurons in the antennal lobe of Periplaneta americana. The somata of these neurons are within the ventrolateral group of cell bodies. Additional immunohistochemical tests revealed that the same neurons also have a GABA-like immunoreactivity.Immunohistochemical dopamine staining (preembedding) of preparations in which the antennal receptor fibers had been caused to degenerate showed that in the glomerular neuropil these antennal fibers form output synapses on dopamine-immunoreactive neurons. The latter form output synapses on unstained neuron profiles.     

Synaptic connections of dopamine-immunoreactive neurons in the antennal lobes of Periplaneta americana. Colocalization with GABA-like immunoreactivity

Dopamine-like immunoreactivity was demonstrated histochemically in about ten local interneurons in the antennal lobe of Periplaneta americana. The somata of these neurons are within the ventrolateral group of cell bodies. Additional immunohistochemical tests revealed that the same neurons also have a GABA-like immunoreactivity. Immunohistochemical dopamine staining (preembedding) of preparations in which the antennal receptor fibers had been caused to degenerate showed that in the glomerular neuropil these antennal fibers form output synapses on dopamine-immunoreactive neurons. The latter form output synapses on unstained neuron profiles.     

Chromatic properties of interneurons in the optic lobes of the bee

Summary The chromatic properties of single units in the optic medulla and lobula of the worker bee were examined. This paper describes the spectral sensitivity, S () and the receptive fields of broad band units, ie. those neurons which receive qualitatively similar inputs from 2 or 3 colour receptor types.The simplest broad band unit responds with sustained excitation or inhibition to light of all colours. Intracellular staining has identified the sustained excitatory unit as the Y8 cell of the proximal medulla.More complex broad band units may receive a variety of colour inputs which sum with different weighting factors or the colour inputs may have different temporal patterning.Receptive fields tend to be large (diameter greater than 60°). The simplest broad band units show homogenous receptive fields which are uniform for all colours. More complex receptive fields contain different areas where different colours evoke an optimal response. No centre-sourround spatial antagonism was found.This work was supported by DFG grant no. Me 365/4 while J.K. was on an Alexander v. Humboldt Stipendium.We thank Joy Nelson for assistance with histology.     

Sexually dimorphic and isomorphic glomeruli in the antennal lobes of the sphinx moth Manduca sexta

Antennal lobes of adult male and female Manduca sexta were compared in order to investigate the nature and extent of sexual dimorphism of the primary olfactory center of this lepidopteran species. Complete identification of the glomeruli led to the conclusion that all female glomeruli have homologous male counterparts. Thus, there is no sex-specific glomerulus present in one sex and absent in the other. Sexual dimorphism (i.e. glomeruli present but morphologically different in males and females), however, does occur in the three glomeruli composing the male macroglomerular complex. The female homologs of this complex consist of two previously identified 'large female glomeruli' and one newly identified normal-sized glomerulus. The lateral and medial large female glomeruli are interpreted to be homologous to the first two macroglomerular-complex glomeruli-the cumulus and toroid 1. The third male component, the toroid 2, was tentatively identified with a normal-sized spheroidal glomerulus of the female, called here the 'small female glomerulus'. The 60 'ordinary' glomeruli that make up the rest of the glomerular neuropil were found to be homologous in males and females, with the exception of two anomalous (or uncertain) glomeruli. Some variations in relative position and size observed among those glomeruli suggest a diffuse, quantitative kind of sexual dimorphism.     

Foraging experience, glomerulus volume, and synapse number: A stereological study of the honey bee antennal lobe

The primary antennal sensory centers (antennal lobes) in the brain of the honeybee are highly compartmentalized into discrete spheres of synaptic neuropil called glomeruli. Many of the glomeruli can be identified according to their predictable size and location. This study examines T1-44, a prominent glomerulus on the dorsal surface of the antennal lobe. Previously, we have shown that the volume of T1-44 in 4-day-old workers performing tasks within the hive is significantly smaller than in foragers and that increases in volume are accompanied by an increase in total synapse number in this glomerulus. Here we examine whether foraging experience is essential for either changes in volume or for changes in synapse numbers in glomerulus T1-44. Five-day-old bees reared under normal colony conditions were compared with 5-day-old bees reared under isolated conditions, and also to 5-day-old bees that had been induced to forage precociously. A combination of light and electron microscopy was used to compare T1-44 volumes and synapse numbers in these three groups. Two groups of 11-day-old bees, precocious foragers and nonforagers, were also examined. The Cavalieri direct estimator of volume was applied to 1.5 microm sections of resin embedded brains. Selected sections were then re-embedded and prepared for transmission electron microscopy. Synapse densities were determined using the physical disector method on electron micrographs. Synapse density and glomerulus volume were combined to give an unbiased estimate of the total number of synapses. This study shows that while both volume and synapse numbers can be induced to increase prematurely in young (5-day-old) precocious foragers, foraging experience is not essential for these structural changes to occur in glomerulus T1-44.     

Male-specific,sex pheromone-selective projection neurons in the antennal lobes of the mothManduca sexta

A subset of olfactory projection neurons in the brain of male Manduca sexta is described, and their role in sex pheromone information processing is examined. These neurons have extensive arborizations in the macroglomerular complex (MGC), a distinctive and sexually dimorphic area of neuropil in the antennal lobe (AL), to which the axons of two known classes of antennal pheromone receptors project. Each projection neuron sends an axon from the AL into the protocerebrum. Forty-one projection neurons were characterized according to their responses to electrical stimulation of the antennal nerve as well as olfactory stimulation of antennal receptors. All neurons exhibited strong selectivity for female sex pheromones. Other behaviorally relevant odors, such as plant volatiles, had no obvious effect on the activity of these neurons. Two broad physiological categories were found: cells that were excited by stimulation of the ipsilateral antenna with pheromones (29 out of 41), and cells that received a mixed input (inhibition and excitation) from pheromone pathways (12 out of 41). Of the cells in the first category, 13 out of 29 were equally excited in response to stimulation of the antenna with either the principal natural pheromone (bombykal) or a mimic of a second unidentified pheromone ('C-15') and were similarly excited by the natural pheromone blend. The remaining 16 out of 29 cells responded selectively, and in some cases, in a dose-dependent manner, to stimulation of the antenna with bombykal or C-15, but not both. Some of these neurons had dendritic arborizations restricted to only a portion of the MGC neuropil, whereas most had arborizations throughout the MGC. Of the cells in the second category, 9 out of 12 were excited by bombykal, inhibited by C-15, and showed a mixed response to the natural pheromone blend. For the other 3 out of 12 cells, the response polarity was reversed for the two chemically-identified odors. Two additional neurons, which were not tested with olfactory stimuli, were tonically inhibited in response to electrical stimulation of the ipsilateral antennal nerve. These observations suggest that some of the male-specific projection neurons may signal general pheromone-triggered arousal, whereas a smaller number can actively integrate inputs from the two know receptor classes (Bal- and C-15-selective) and may operate as 'mixture detectors' at this level of the olfactory subsystem that processes information about sex pheromones.