Representation of primary plant odorants in the antennal lobe of the moth Heliothis virescens using calcium imaging

The primary olfactory centre, the antennal lobe of Heliothis virescens moths, contains 62 glomeruli which process plant odour information and four male-specific glomeruli which form the macroglomerular complex, involved in processing information about pheromone and interspecific signals. Using calcium imaging, we recorded the spatio-temporal activity pattern of the glomeruli in the anterior antennal lobe during stimulation with odorants produced by plants or insects. Each odorant elicited specific excitatory responses in one or a few glomeruli: the major pheromone component did so exclusively in the large glomerulus of the macroglomerular complex and the plant odours exclusively in the ordinary glomeruli. Eight glomeruli, with corresponding plant odour responses and positions, were identified within each sex. Glomeruli responded specifically to linalool, beta-ocimene/beta-myrcene or germacrene D/alpha-farnesene. Responses to two essential plant oils covered the response areas of their major constituents, as well as activating additional glomeruli. Stronger activation in the AL due to increased odour concentration was expressed as increased response strength within the odorant-specific glomeruli as well as recruitment of less sensitive glomeruli.     

Olfactory activation patterns in the antennal lobe of the sphinx moth,<Emphasis Type="Italic"> Manduca sexta</Emphasis>

The sphinx moth Manduca sexta is a well-studied insect with regard to central olfactory functions. Until now, the innervation patterns of olfactory receptor neurons into the array of olfactory glomeruli in the antennal lobe have, however, been unclear. Using optical imaging to visualize calcium dynamics within the antennal lobe we demonstrate specific patterns elicited by sex pheromone components and plant-derived odours. These patterns mainly reflect receptor neuron activity. Within the male-specific macroglomerular complex the two major pheromone components evoke stereotyped activity in either of two macroglomerular complex glomeruli. Based on previous knowledge of output neuron specificity, our results suggest a matching of information between input and output in the macroglomerular complex. Plant odours evoked activity in the sexually isomorphic glomeruli. Two major results were obtained: (1). terpenes and aromatic compounds activate different clusters of glomeruli with only minor overlapping, and (2). the position of certain key glomeruli is fixed in both males and females, which suggests that host-plant related odorants are processed in a similar way in both sexes.     

Differential interactions of sex pheromone and plant odour in the olfactory pathway of a male moth

Most animals rely on olfaction to find sexual partners, food or a habitat. The olfactory system faces the challenge of extracting meaningful information from a noisy odorous environment. In most moth species, males respond to sex pheromone emitted by females in an environment with abundant plant volatiles. Plant odours could either facilitate the localization of females (females calling on host plants), mask the female pheromone or they could be neutral without any effect on the pheromone. Here we studied how mixtures of a behaviourally-attractive floral odour, heptanal, and the sex pheromone are encoded at different levels of the olfactory pathway in males of the noctuid moth Agrotis ipsilon. In addition, we asked how interactions between the two odorants change as a function of the males' mating status. We investigated mixture detection in both the pheromone-specific and in the general odorant pathway. We used a) recordings from individual sensilla to study responses of olfactory receptor neurons, b) in vivo calcium imaging with a bath-applied dye to characterize the global input response in the primary olfactory centre, the antennal lobe and c) intracellular recordings of antennal lobe output neurons, projection neurons, in virgin and newly-mated males. Our results show that heptanal reduces pheromone sensitivity at the peripheral and central olfactory level independently of the mating status. Contrarily, heptanal-responding olfactory receptor neurons are not influenced by pheromone in a mixture, although some post-mating modulation occurs at the input of the sexually isomorphic ordinary glomeruli, where general odours are processed within the antennal lobe. The results are discussed in the context of mate localization.     

Spatial representation of odours in the antennal lobe of the moth Spodoptera littoralis (Lepidoptera: Noctuidae)

Glomeruli within the antennal lobe (AL) of moths are convergence sites for a large number of olfactory receptor neurons (ORNs). The ORNs target single glomeruli. In the male-specific cluster of glomeruli, the macroglomerular complex (MGC), the input is chemotypic in that each glomerulus of the MGC receives information about a specific component of the conspecific female sex pheromone. Little is known about how neurons that detect other odorants arborize in and amongst glomeruli. The present study focuses on how sex pheromones and biologically relevant semiochemicals are represented in the ALs of both sexes of the moth Spodoptera littoralis. To assess this, we optically measured odour-evoked changes of calcium concentration in the ALs. Foci of calcium increase corresponded in size and shape with anatomical glomeruli. More than one glomerulus was normally activated by a specific non-pheromonal odorant and the same glomerulus was activated by several odorants. All odorants and pheromone components tested evoked unique patterns of glomerular activity that were highly reproducible at repeated stimulations within an individual. Odour-evoked patterns were similar between individuals for a given odorant, implicating a spatial olfactory code. In addition, we demonstrated that activity patterns evoked by host-plant related volatiles are similar between males and females.     

Synchronous firing of antennal-lobe projection neurons encodes the behaviorally effective ratio of sex-pheromone components in male Manduca sexta

Olfactory stimuli that are essential to an animal’s survival and reproduction are often complex mixtures of volatile organic compounds in characteristic proportions. Here, we investigated how these proportions are encoded in the primary olfactory processing center, the antennal lobe, of male Manduca sexta moths. Two key components of the female’s sex pheromone, present in an approximately 2:1 ratio, are processed in each of two neighboring glomeruli in the macroglomerular complex (MGC) of males of this species. In wind-tunnel flight experiments, males exhibited behavioral selectivity for ratios approximating the ratio released by conspecific females. The ratio between components was poorly represented, however, in the firing-rate output of uniglomerular MGC projection neurons (PNs). PN firing rate was mostly insensitive to the ratio between components, and individual PNs did not exhibit a preference for a particular ratio. Recording simultaneously from pairs of PNs in the same glomerulus, we found that the natural ratio between components elicited the most synchronous spikes, and altering the proportion of either component decreased the proportion of synchronous spikes. The degree of synchronous firing between PNs in the same glomerulus thus selectively encodes the natural ratio that most effectively evokes the natural behavioral response to pheromone.     

Physiological mismatching between neurons innervating olfactory glomeruli in a moth

The primary olfactory centres of most vertebrates and most neopteran insects are characterized by the presence of spherical neuropils, glomeruli, where synaptic interactions between olfactory receptor neurons and second-order neurons take place. In the neopteran insect taxa investigated so far, receptor neurons of a specific physiological identity target one glomerulus and thus bestow a functional identity on the glomerulus. In moths, input from pheromone-specific receptor neurons is received in a male-specific structure of the antennal lobe, called the macroglomerular complex (MGC), which consists of a number of specialized glomeruli. Each glomerulus of the complex receives a set of peripheral sensory afferents that encode one of several compounds involved in sexual communication. The complex is also innervated by dendritic branches of antennal lobe output neurons called projection neurons, which transfer information from the antennal lobe to higher centres of the brain. A hypothesis stemming from earlier work on moths claims that the receptor neuron innervation pattern of the MGC should be reflected in the pattern of dendrites of projection neurons invading the different MGC glomeruli. In this study we show that in the noctuid moth Trichoplusia ni, as in several other noctuid moth species, this hypothesis does not hold. The degree of matching between axon terminals of receptor neurons and the dendritic branches of identified projection neurons that express similar physiological specificity is very low.     

Morphology of the reproductive system and antennal lobes of gynandromorphic and normal black cutworm moths,agrotis ipsilon (Hufnagel) (Lepidoptera : Noctuidae)

Sexually dimorphic characteristics of a bilaterally asymmetric gynandromorphic black cutworm moth, Agrotis ipsilon (Hufnagel), were compared with those of normal males and females. On one side of the body, the gynandromorphs wings were larger and darker than on the other side, and the antenna was filiform. On the other side of the body, the wings were smaller and lighter in color, and the antenna was pectinate. Females were usually larger and more pigmented than males. Female antenna were filiform and those of males pectinate. At the tip of the abdomen, the gynandromorph had 2 valves, as normal males do, but the one on the female side was smaller. The antennal lobes of the gynandromorphs brain included only ordinary glomeruli on the female side, and ordinary glomeruli plus a partially developed macroglomerular complex (MGC) at the base of the antenna on the male side. Normal female antennal lobes contained only ordinary glomeruli. Normal male antennal lobes contained ordinary glomeruli and a fully developed MGC, consisting of one large and 3 smaller glomeruli. In the gynandromorph, female reproductive organs were partially developed or absent. A portion of the oviduct was missing together with several ovarioles, and no spermatheca or seminal duct were found. The male reproductive track was complete, except for the paired structures, which in the gynandromorph were single. Normal females had paired accessory glands and ovaries, and single oviduct, bursa copulatrix, and spermatheca. Normal males had fused testes, paired accessory glands, and a single ejaculatory duct and aedeagus. The gynandromorph assumed a calling posture and attracted one male, but it did not respond to the pheromone from females.     

Concurrent modulation of neuronal and behavioural olfactory responses to sex and host plant cues in a male moth

Mating has profound effects on animal physiology and behaviour, not only in females but also in males, which we show here for olfactory responses. In cotton leafworm moths, Spodoptera littoralis, odour-mediated attraction to sex pheromone and plant volatiles are modulated after mating, producing a behavioural response that matches the physiological condition of the male insect. Unmated males are attracted by upwind flight to sex pheromone released by calling females, as well as to volatiles of lilac flowers and green leaves of the host plant cotton, signalling adult food and mating sites, respectively. Mating temporarily abolishes male attraction to females and host plant odour, but does not diminish attraction to flowers. This behavioural modulation is correlated with a response modulation in the olfactory system, as shown by electro-physiological recordings from antennae and by functional imaging of the antennal lobe, using natural odours and synthetic compounds. An effect of mating on the olfactory responses to pheromone and cotton plant volatiles but not to lilac flowers indicates the presence of functionally independent neural circuits within the olfactory system. Our results indicate that these circuits interconnect and weigh perception of social and habitat odour signals to generate appropriate behavioural responses according to mating state.     

Representation of a mixture of pheromone and host plant odor by antennal lobe projection neurons of the silkmoth Bombyx mori

Pheromone-source orientation behavior can be modified by coexisting plant volatiles. Some host plant volatiles enhance the pheromonal responses of olfactory receptor neurons and increase the sensitivity of orientation behavior in the Lepidoptera species. Although many electrophysiological studies have focused on the pheromonal response of olfactory interneurons, the response to the mixture of pheromone and plant odor is not yet known. Using the silkmoth, Bombyx mori, we investigated the physiology of interneurons in the antennal lobe (AL), the primary olfactory center in the insect brain, in response to a mixture of the primary pheromone component bombykol and cis-3-hexen-1-ol, a mulberry leaf volatile. Application of the mixture enhanced the pheromonal responses of projection neurons innervating the macroglomerular complex in the AL. In contrast, the mixture of pheromone and cis-3-hexen-1-ol had little influence on the responses of projection neurons innervating the ordinary glomeruli whereas other plant odors dynamically modified the response. Together this suggests moths can process plant odor information under conditions of simultaneous exposure to sex pheromone.     

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     

Glomerular targets of Heliothis subflexa male olfactory receptor neurons housed within long trichoid sensilla

We used single-sensillum recordings to characterize male Heliothis subflexa antennal olfactory receptor neuron physiology in response to compounds related to their sex pheromone. The recordings were then followed by cobalt staining in order to trace the neurons' axons to their glomerular destinations in the antennal lobe. Receptor neurons responding to the major pheromone component, (Z)-11-hexadecenal, in the first type of sensillum, type-A, projected axons to the cumulus of the macroglomerular complex (MGC). In approximately 40% of the type-A sensilla, a colocalized receptor neuron was stained that projected consistently to the posterior complex 1 (PCx1), a specific glomerulus in an 8-glomerulus complex that we call the Posterior Complex (PCx). We found that receptor neurons residing in type-B sensilla and responding to a secondary pheromone component, (Z)-9-hexadecenal, send their axons to the dorsal medial glomerulus of the MGC. As in the type-A sensilla, we found a cocompartmentalized neuron within type-B sensilla that sends its axon to a different glomerulus of the PCx4. One neuron in type-C sensilla tuned to a third pheromone component, (Z)-11-hexadecenol, and a colocalized neuron responding to (Z)-11-hexadecenyl acetate projected their axons to the anteromedial and ventromedial glomeruli of the MGC, respectively.     

Axons of olfactory receptor cells of transsexually grafted antennae induce development of sexually dimorphic glomeruli in Manduca sexta

The influence of olfactory receptor cell (ORC) axons from transsexually grafted antennae on the development of glomeruli in the antennal lobes (ALs), the primary olfactory centers, was studied in the moth Manduca sexta. Normally during metamorphic adult development, the pheromone-specific macroglomerular complex (MGC) forms only in the ALs of males, whereas two lateral female-specific glomeruli (LFGs) develop exclusively in females. A female AL innervated by ORC axons from a grafted male antenna developed an MGC with three glomeruli, like the MGC of a normal male AL. Conversely, a male AL innervated by ORC axons from a grafted female antenna lacked the MGC but exhibited LFGs. ORC axons from grafted male antenna terminated in the MGC-specific target area, even in cases when the antennal nerve (AN) entered the AL via an abnormal route. Within ectopic neuromas formed by ANs that had become misrouted and failed to enter the brain, male-specific axons were not organized and formed terminal branches in many areas. The results suggest the presence of guidance cues within the AL for male-specific ORC axons. Depending on the sex of the antennal innervation, glial borders formed in a pattern characteristic of the MGC or LFGs. The sex-specific number of projection neurons (PNs) in the medial group of AL neurons remained unaffected by the antennal graft, but significant changes occurred in the organization of PN arborizations. In gynandromorphic females, LFG-specific PNs extended processes into the induced MGC, whereas in gynandromorphic males, PNs became restricted to the LFGs. The results indicate that male-and female-specific ORC axons play important roles in determining the position, anatomical features, and innervation of sexually dimorphic glomeruli.     

Anatomical and physiological characteristics of individual neurones in the central antennal pathway of insects

Signals of tens up to hundreds of thousands of (mostly olfactory) receptor cells on an insect antenna are switched to a comparatively low number of neurones in the antennal lobe of the deutocerebrum in circumscribed units of neuropile, the glomeruli. Each glomerulus is connected via its output neurone to two separate neuropiles (calyces of mushroom body, and lateral lobe) of the protocerebrum. Local interneurones interconnect between the glomeruli. Certain modes of convergence between receptors and central neurones provide for a very high sensitivity of the latter to certain odours and their sensitivity for complex odour stimuli, and in many cases for a marked multimodality. Anatomical and physiological data are given especially for pheromone sensitive neurones and their projections.     

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.     

Modelling efficiency in insect olfactory information processing

The olfactory system of insects is essential for the search of food and mates, and weak signals can be detected, amplified and discriminated in a fluctuating environment. The olfactory system also allows for learning and recall of odour memories. Based on anatomical, physiological, and behavioural data from the olfactory system of insects, we have developed a cross-scale dynamical neural network model to simulate the presentation, amplification and discrimination of host plant odours and sex pheromones. In particular, we model how the spatial and temporal patterns of the odour information emerging in the glomeruli of the antennal lobe (AL) rely on the glomerular morphology, the connectivity and the complex dynamics of the AL circuits. We study how weak signals can be amplified, how different odours can be discriminated, based on stochastic (resonance) dynamics and the connectivity of the network. We further investigate the spatial and temporal coding of sex pheromone components and plant volatile compounds, in relation to the glomerular structure, arborizing patterns of the projection neurons (PNs) and timing patterns of the neuronal spiking activity.     

Sex pheromone and plant-associated odour processing in antennal lobe interneurons of male Spodoptera littoralis (Lepidoptera: Noctuidae)

Antennal lobe interneurons of male Spodoptera littoralis (Boisd.) were investigated by using intracellular recording and staining techniques. Physiological and morphological characteristics of local interneurons and projection neurons responding to sex pheromone and plant-associated volatiles are described. The interneurons identified were divided into three groups, depending on their physiological response characteristics. Both types of interneurons, local interneurons and projection neurons, were described in all three groups. 1. Interneurons responding exclusively to sex pheromone stimuli, displayed different degrees of specificity. These neurons responded to either one, two, three or all four of the single sex pheromone or sex pheromone-like compounds tested. Most of these neurons also responded to a mixture of the two pheromone components present in the female S. littoralis blend. Two local interneurons and one projection neuron were identified as blend specialists, not responding to the single female produced sex pheromone components, but only to their mixture. Five pheromone specific projection neurons arborized in one or more subcompartments of the macroglomercular complex (MGC) and some of them had axonal branches in the calyces of the mushroom body and in different parts of the lateral protocerebrum. 2. Interneurons responding only to plant-associated volatiles varied highly in specificity. Neurons responding to only one of the stimuli, neurons responding to a variety of different odours and one neuron responding to all stimuli tested, were found. Three specialized local interneurons had arborizations only in ordinary glomeruli. One specialized and three less specialized local interneurons had arborizations within the MGC and the ordinary glomeruli. The projection neurons responding only to plant-associated volatiles had mostly uni- or multiglomerular arborizations within the ordinary glomeruli. 3. Interneurons responding to both sex pheromones and plant-associated stimuli varied in specificity. Individual interneurons that responded to few plant-associated odours mostly responded to several pheromone stimuli as well. Projection neurons responding to most of the plant-associated volatiles also responded to all pheromone stimuli. Two local interneurons responding to both stimulus groups, arborized within the MGC and the ordinary glomeruli. Projection neurons mostly arborized in only one ordinary glomerulus or in one compartment of the MGC. The variation in specificity and sensitivity of antennal lobe interneurons and structure-function correlations are discussed.     

Odour perception in honeybees: coding information in glomerular patterns

Major advances have been made during the past two years in understanding how honeybees process olfactory input at the level of their first brain structure dealing with odours, the antennal lobe (the insect analogue of the mammalian olfactory bulb). It is now possible to map physiological responses to morphologically identified olfactory glomeruli, allowing for the creation of a functional atlas of the antennal lobe. Furthermore, the measurement of odour-evoked activity patterns has now been combined with studies of appetitive odour learning. The results show that both genetically determined components and learning-related plasticity shape olfactory processing in the antennal lobe.     

Physiology and morphology of projection neurons in the antennal lobe of the male mothManduca sexta

1. We have used intracellular recording and staining, followed by reconstruction from serial sections, to characterize the responses and structure of projection neurons (PNs) that link the antennal lobe (AL) to other regions of the brain of the male sphinx moth Manduca sexta. 2. Dendritic arborizations of the AL PNs were usually restricted either to ordinary glomeruli or to the male-specific macroglomerular complex (MGC) within the AL neuropil. Dendritic fields in the MGC appeared to belong to distinct partitions within the MGC. PNs innervating the ordinary glomeruli had arborizations in a single glomerulus (uniglomerular) or in more than one ordinary glomerulus of one AL (multiglomerular) or in one case, in single glomeruli in both ALs (bilateral-uniglomerular). One PN innervated the MGC and many or all ordinary glomeruli of the AL. 3. PNs with dendritic arborizations in the ordinary glomeruli and PNs associated with the MGC typically projected both to the calyces of the ipsilateral mushroom body and to the lateral protocerebrum, but some differences in the patterns of termination in those regions have been noted for the two classes of PNs. One PN conspicuously lacked branches in the calyces but did project to the lateral protocerebrum. The PN innervating the MGC and many ordinary glomeruli projected to the calyces of the ipsilateral mushroom body and the superior protocerebrum. 4. Crude sex-pheromone extracts excited all neurons with arborizations in the MGC, although some were inhibited by other odors. One P(MGC) was excited by crude sex-pheromone extract and by a mimic of one component of the pheromone blend but was inhibited by another component of the blend. 5. PNs with dendritic arborizations in ordinary glomeruli were excited or inhibited by certain non-pheromonal odors. Some of these PNs also responded to mechanosensory stimulation of the antennae. 6. The PN with dendritic arborizations in the MGC and many ordinary glomeruli was excited by crude sex-pheromone extracts and non-pheromonal odors and also responded to mechanosensory stimulation of the antenna.     

Evolution and ultrastructure of the bovine spermatogonia precursor cell line

We have used a cytochemical technique to investigate the distribution of acetylcholinesterase (AChE) activity in the deutocerebrum of the brain of the sphinx moth Manduca sexta. To distinguish between extra-and intracellular pools of the enzyme, some brains were treated prior to histochemical staining with echothiophate, an irreversible AChE inhibitor which penetrates cell membranes very slowly and, therefore, inhibits only extracellular AChE. In the antennal nerve, fascicles of presumably mechanosensory fibers show echothiophateinsensitive AChE activity. They bypass the antennal lobe and project to the antennal mechanosensory and motor center of the deutocerebrum. In the antennal lobe, fibers in the coarse neuropil, cell bodies in the lateral cell group, and all glomeruli exhibit AChE activity. In most ordinary glomeruli, echothiophate-sensitive AChE activity is concentrated in the outer cap regions, corresponding to the terminal arborizations of olfactory afferents. A previously unrecognized glomerulus in the ventro-median antennal lobe shows uniform and more intense AChE-specific staining that the other glomeruli. No AChE activity appeared to be associated with malespecific pheromone-sensitive afferents in the macro-glomerular complex. About 67 interneurons with somata in the lateral cell group of the antennal lobe show echo-thiophate-insensitive AChE activity. These neurous seem to be members of two types of antennal-lobe projection neurons with fibers passing through the outer-antenno-cerebral tract to the protocerebrum. AChE-stained arborizations of these neurons appear to invade all glomeruli, including three distinguishable subunits of the male-specific macroglomerular complex. In echothiophate-treated animals, the projections of one of these types of fiber form large terminals in the lateral horn of protocerebrum, which partly protrude into the adjacent glial cell layer. The results suggest that extracellularly accessible AChE is associated with ordinary olfactory receptor terminals but apparently not with pheromone-sensitive afferents. Intracellular AChE appears to be present in antennal mechanosensory fibers and in two types of olfactory projection neurons of the antennal lobe. The study provides further evidence for cholinergic neurotransmission of most antennal afferents. The AChE-containing interneurons might be cholinergic as well or use the enzyme for functions unrelated to hydrolysis of acetylcholine.Abbreviations ACh acetylcholine - AChE acetylcholinesterase - AL antennal lobe - AMMC antennal mechanosensory and motor center - ChAT choline acetyltransferase - IACT inner antenno-cerebral tract - MGC macroglomerular complex     

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.