Innervation regulates the metamorphic fates of larval abdominal muscles in the moth, Manduca sexta

 With the onset of metamorphosis, the abdominal muscles of the moth, Manduca sexta, follow one of three developmental fates: maintenance, respecification, or death. The maintained muscles retain their larval size and morphology throughout adult development. The respecified and dying muscles dedifferentiate, which involves regression, nuclear degeneration, and myofibril breakdown. Nuclei in both dying and respecified muscles also proliferate. The amount of nuclear degeneration is greater in the dying muscle fibers, and the amount of nuclear proliferation is greater in the respecified muscles. Four to ten days after pupation, the sizes of the respecified muscles stabilize while the dying muscles are lost. During regression, a subset of the respecified muscle fibers die. The surviving respecified muscle fibers grow and differentiate during the last half of adult development. In respecified muscles, denervation triggers an increased amount of nuclear degeneration and a decreased amount of nuclear proliferation. As a result, denervated respecified fibers experience increased muscle regression including an increased loss of muscle fibers and sometimes muscle death. Surviving respecified fibers still grow and differentiate yet are only 5 to 12% of the control size. Denervation triggers dedifferentiation in maintained muscles, resulting in fiber loss and occasionally muscle death. The percentage of fibers which dedifferentiate varies between different muscles. Denervation also triggers nuclear proliferation, with the amount of nuclear proliferation correlated with the extent of dedifferentiation of the individual muscle fibers. The dedifferentiated maintained fibers subsequently undergo differentiation in the absence of muscle growth. Received: 10 July 1997 / Accepted: 21 April 1998     

Projection pattern of sensory neurons in the central nervous system of a homeotic mutation of the moth Manduca sexta

Octopod (Octo) is a mutation of the moth Manduca sexta, which transforms the first abdominal segment (A1) in the anterior direction. Mutant animals are characterized by the appearance of homeotic thoracic-like legs on A1. We exploited this mutation to determine what rules might be used in specifying the fates of sensory neurons located on the body surface of larval Manduca. Mechanical stimulation of homeotic leg sensilla did not cause reflexive movements of the homeotic legs, but elicited responses similar to those observed following stimulation of ventral A1 body wall hairs. Intracellular recordings demonstrated that several of the motoneurons in the A1 ganglion received inputs from the homeotic sensory hairs. The responses of these motoneurons to stimulation of homeotic sensilla resembled their responses to stimulation of ventral body wall sensilla. Cobalt fills revealed that the mutation transformed the segmental projection pattern of only the sensory neurons located on the ventral surface of A1, resulting in a greater number with intersegmental projection patterns typical of sensory neurons found on the thoracic body wall. Many of the sensory neurons on the homeotic legs had intersegmental projection patterns typical of abdominal sensory neurons: an anteriorly directed projection terminating in the third thoracic ganglion (T3). Once this projection reached T3, however, it mimicked the projections of the thoracic leg sensory neurons. These results demonstrate that the same rules are not used in the establishment of the intersegmental and leg-specific projection patterns. Segmental identity influences the intersegmental projection pattern of the sensory neurons of Manduca, whereas the leg-specific projections are consistent with a role for positional information in determining their pattern. © 1995 John Wiley & Sons, Inc.     

Hormonal control of ventral diaphragm myogenesis during metamorphosis of the moth, Manduca sexta

 Both the proliferation and differentiation of ventral diaphragm myoblasts are controlled by ecdysteroid during metamorphosis of the moth, Manduca sexta, but the responses have different hormonal requirements. Tonic exposure to moderate levels of ecdysteroid are required to stimulate myoblast proliferation. This is due to the presence of an ecdysteroid-dependent control point in the G₂ phase of the cell cycle. As a result, proliferation can be repeatedly turned on or off simply by adjusting the concentration of ecdysteroid to be above or below a critical threshold concentration. In contrast, high levels of ecdysteroid trigger irreversible proliferative arrest and differentiation of myofibers. Myoblast proliferation and differentiation also differ in their response to the juvenile hormone mimic, methoprene. Ecdysteroid-dependent proliferative arrest and differentiation are blocked by coculture with methoprene but methoprene has no effect on ecdysteroid-dependent proliferation. In the animal, premature exposure to high levels of ecdysteroid in the absence of juvenile hormone triggers precocious differentiation of the myoblasts, resulting in the formation of several thin bands of muscle rather than a complete diaphragm. Thus, ecdysteroid and juvenile hormone collaborate to determine the size and shape of the adult musculature. Received: 12 November 1998 / Accepted: 23 December 1998     

Juvenile hormone modulates 20-hydroxyecdysone-inducible ecdysone receptor and ultraspiracle gene expression in the tobacco hornworm, Manduca sexta

 Insect molting and metamorphosis are orchestrated by ecdysteroids with juvenile hormone (JH) preventing the actions of ecdysteroids necessary for metamorphosis. During the molt and metamorphosis of the dorsal abdominal epidermis of the tobacco hornworm, Manduca sexta, the isoforms involved in the ecdysone receptor (EcR)/Ultraspiracle (USP) complex change with the most dramatic switch being the loss of USP-1 and the appearance of USP-2 during the larval and pupal molts. We show here that this switch in USP isoforms is mediated by high 20-hydroxyecdysone (20E) and that the presence of JH is necessary for the down-regulation of USP-1 mRNA. The decrease of USP-1 mRNA in day 2 fourth instar larval epidermis in vitro required exposure to a high concentration (10^–5 M) of 20E equivalent to the peak ecdysteroid concentration in vivo, whereas the increase of USP-2 mRNA occurred at lower concentrations (effective concentrations, EC₅₀=6.3×10^–7 M). During the pupal molt of allatectomized larvae which lack JH, USP-2 mRNA increased normally with the increasing ecdysteroid titer, whereas USP-1 mRNA remained high until pupation. When day 2 fifth instar larval epidermis was exposed to 500 ng/ml 20E in the absence of JH to cause pupal commitment of the cells by 24 h, USP-1 RNA remained at its high preculture level for 12 h, then increased two- to threefold by 24 h. The increase was prevented by the presence of 1 μg/ml JH I which also prevents the pupal commitment of the cells. By contrast, USP-2 mRNA increased steadily with the same EC₅₀ as in fourth stage epidermis, irrespective of the presence or absence of JH. Under the same conditions, mRNAs for both EcR-B1 and EcR-A isoforms were up-regulated by 20E, each in its own time-dependent manner, similar to that seen in vivo. These initial mRNA increases were unaffected by the presence of JH I, but those seen after 12 h exposure to 20E were prevented by JH, indicating a difference in response between larvally and pupally committed cells. The presence of JH which maintained larval commitment of the cells also prolonged the half-life of the EcR proteins in these cells. These results indicate that both EcR and USP RNAs are regulated by 20E and can be modulated by JH in a complex manner with only that of USP-2 apparently unaffected. Received: 16 July 1998 / Accepted: 5 August 1998     

Neural correlates of habituation of the proleg withdrawal reflex in larvae of the hawk moth, Manduca sexta

The larval proleg withdrawal reflex of the hawk moth, Manduca sexta, exhibits robust habituation. This reflex is evoked by deflecting one or more mechanosensory planta hairs on a proleg tip. We examined neural correlates of habituation in an isolated proleg preparation consisting of one proleg and its segmental ganglion. Repeated deflection of a single planta hair caused a significant decrease in the number of action potentials evoked in the proleg motor nerve (which carries the axons of proleg retractor motor neurons). Significant response decrement was seen for interstimulus intervals of 10 s, 60 s and 5 min. Response decrement failed to occur in the absence of repetitive stimulation, the decremented response recovered spontaneously following a rest, and electrical stimulation of a body wall nerve facilitated the decremented response (a neural correlate of dishabituation). Adaptation of sensory neuron responses occurred during repeated hair deflections. However, when adaptation was eliminated by direct electrical stimulation of sensory neurons, the response in the proleg motor nerve still decreased significantly. Muscle recordings indicated that the response of an identified proleg retractor motor neuron decreased significantly during habituation training. Thus, habituation of the proleg withdrawal reflex includes a central component that is apparent at the level of a single motor neuron. Accepted: 20 December 1996     

Stamenless, a tomato mutant with homeotic conversions in petals and stamens

A tomato (Lycopersicon esculentum Mill.) monogenic semidominant mutation, stamenless (sl), which results in homeotic conversions in two adjacent floral whorls, was studied. When grown at standard temperature, flowers of sl/sl plants showed sepaloid petals in the second whorl and strong transformation of stamens to carpels in whorl three. These transformed carpels were fused with each other and with the genuine carpels in the fourth whorl to form a unique gynoecium. The mutation is semidominant since heterozygous plants showed a phenotype intermediate between that of the wild type (WT) and that of homozygous mutant plants, with nearly WT petals but with feminized stamens bearing naked ovules on the base of their adaxial face. The initiation and position of organ primordia in sl/sl flowers were not altered when compared with WT primordia although development of organ primordia in the second and third whorls deviated from WT at an early stage as observed by scanning electron microscopy. The mutant phenotype is temperature sensitive and when sl/sl plants were cultured at low temperature, the morphology of some flowers resembled that of the WT. This reversion of the mutant phenotype is also induced by treatment of young sl/sl plants with gibberellic acid, providing evidence that gibberellin synthesis or sensitivity could mediate the effect of low temperature on the mutant phenotype. Southern blot analyses using a Deficiens-homologous gene from Solanum tuberosum as a probe showed a restriction-fragment-length polymorphism (RFLP) linked to the sl mutation. This result indicates that the mutation affects a Deficiens-like gene that controls the identity of petals and stamens. Received: 10 December 1998 / Accepted: 29 March 1999     

The innervation of the salivary gland of the moth,Manduca sexta

Summary The salivary glands of the moth, Manduca sexta (Insecta: Sphingidae), are unlike most other salivary glands in that they are innervated from one source only. Vital staining of nerves with methylene-blue reveals numerous fine nerves extending to the glands from the oesophageal nerve, a part of the stomatogastric or visceral nervous system. Light and electron microscopy confirm that only the fluid-secreting cells, confined to a discrete region in these glands, are innervated. Axons with or without glial wrappings are found in intercellular spaces between fluid-secreting cells. Axons lacking a glial sheath contain, after glutaraldehyde-osmium tetroxide fixation, large granular and small agranular vesicles. In nerve endings in glands fixed with permanganate these smaller vesicles are granular, having the electron-dense cores characteristic of monoamine-containing neurons. These nerve endings with synaptoid areas are in close (direct) contact with the fluid-secreting cells.I am grateful to Professor T. Weis-Fogh for accommodation in the department of Zoology and to Dr. Nancy Lane for use of A.R.C. facilities and advice. Thanks are also due to Drs. M. J. Berridge, S.H.P. Maddrell, and W. T. Prince and Mr. R. A. Leslie for helpful discussion. Financial assistance from Clare College, Cambridge is gratefully acknowledged.     

Indolic constituents and indole-3-acetic acid biosynthesis in the wild-type and a tryptophan auxotroph mutant of Arabidopsis thaliana

 The tryptophan auxotroph mutant trp3-1 of Arabidopsis thaliana (L.) Heynh., despite having reduced levels of l-tryptophan, accumulates the tryptophan-derived glucosinolate, glucobrassicin and, thus, does not appear to be tryptophan-limited. However, due to the block in tryptophan synthase, the mutant hyperaccumulates the precursor indole-3-glycerophosphate (up to 10 mg per g FW). Instability of indole-3-glycerophosphate leads to release of indole-3-acetic acid (IAA) from this metabolite during standard workup of samples for determination of conjugated IAA. The apparent increase in “conjugated IAA” in trp3-1 mutant plants can be traced back entirely to indole-3-glycerophosphate degradation. Thus, the levels of neither free IAA nor conjugated IAA increase detectably in the trp3-1 mutant compared to wild-type plants. Precursor-feeding experiments to shoots of sterile-grown wild-type plants using [²H]₅-l-tryptophan have shown incorporation of label from this precursor into indole-3-acetonitrile and indole-3-acetic acid with very little isotope dilution. It is concluded that Arabidopsis thaliana shoots synthesize IAA from l-tryptophan and that the non-tryptophan pathway is probably an artifact. Received: 1 March 2000 / Accepted: 10 April 2000     

Developmental expression of Ultraspiracle proteins in the epidermis of the tobacco hornworm, Manduca sexta, during larval life and the onset of metamorphosis

 During the final two larval instars, a changing pattern of three Ultraspiracle (Usp) proteins (50.5, 52.5, and 57 kDa) was detected in immunoblots of the dorsal abdominal epidermis of the tobacco hornworm, Manduca sexta, by a monoclonal antibody against Drosophila Usp that was shown to detect MsUsp. The 57- and 52.5-kDa bands were present during the intermolt periods and the 50.5- and 52.5-kDa bands during the molting phases. The antibody detected a nuclear antigen present in epidermis, muscle, fat body, and the central nervous system from the time of hatching. In the epidermis Usp was present in all cell nuclei but was especially prominent in the tormogen and trichogen cells immediately after ecdysis in both the penultimate and final instars. This latter immunoreactivity disappeared within 12 h whereas the remainder of the epidermis retained high levels throughout the feeding period. During the molt immunostaining reappeared in the hair cell nuclei. During the wandering stage at the onset of metamorphosis and just before pupal ecdysis, immunoblots showed high levels of Usp, but nuclei showed little or no staining. This discrepancy is likely due to the loss of one Usp isoform from the nucleus and its dispersal in the cytoplasm in preparation for the appearance of the second isoform. Received: 10 June 1997 / Accepted: 22 August 1997     

Early events in adult eye development of the moth, Manduca sexta

The eye imaginal disc of Manduca sexta is created early in the final larval instar from the adult eye primordium, which is composed of fully differentiated cells of the larval head capsule epidermis. Concomitant with the down-regulation of the larval epidermal program, expression of broad, a marker of pupal commitment, is activated in the primordium. The cells then detach from the cuticle, fold inward, and begin to proliferate at high levels to produce the inverted, eye imaginal disc. These and other events that begin on the first day of the final larval instar appear to mark the initiation of metamorphosis. Little is known about the endocrine control of the initiation of metamorphosis in any insect. The hemolymph titer of juvenile hormone (JH) declines to low levels during this period and the presence of JH is sufficient to repress development in cultured eye primordia. However, maintenance of JH at high levels in vivo by treatment with long-lasting JH mimics has no apparent effect on early steps in eye imaginal disc development. We discuss our findings in the context of the endocrine control of metamorphosis. The initiation of metamorphosis in Manduca, and perhaps a wide range of insect species, appears to involve the overcoming of JH repression by an unidentified, nutrient-dependent, hormonal factor.     

Acetylcholinesterase activity in antennal receptor neurons of the sphinx moth Manduca sexta

Summary We have used a cytochemical technique to investigate the distribution of acetylcholinesterase (AChE) activity in the antenna of the sphinx moth Manduca sexta. High levels of echothiophate-insensitive (presumably intracellular) AChE activity were found in six different types of antennal receptors localized in specific regions of the three antennal segments of the adult moth. Mechanosensory organs in the scape and pedicel, the Böhm bristles and Johnston's organ, are innervated by AChE-positive neurons. In each annulus of the antennal flagellum, AChE-positive neurons are associated with six sensilla chaetica and a peg organ, probably a sensillum styloconicum. At least 112 receptor neurons (8–10 per annulus) innervating the intersegmental membranes between the 14 distalmost annuli also exhibit high levels of echothiophate-resistant AChE. In addition, each annulus has more than 30 AChE-positive somata in the epidermis of the scale-covered (back) side of the flagellum, and 4 AChE-positive somata reside within the first annulus of the flagellum. Since none of the olfactory receptor neurons show a high level of echothiophateresistant AChE activity, and all known mechanoreceptors are AChE-positive, apparently intracellular AChE activity in the antenna correlates well with mechanosensory functions and is consistent with the idea that these cells employ acetylcholine as a neurotransmitter.     

The activity of pleurodorsal muscles during flight and at rest in the moth Manduca sexta (L.)

In the moth Manduca sexta, the paired mesothoracic flight steering muscle II PD2m takes part in the generation of the flight rhythm and is spontaneously active in the non-flying animal. This spontaneous activity is modulated by optomotor stimuli and directionally selective. The directional response characteristics are analyzed. Another spontaneously active steering muscle pair, the III PD2c, is situated in the metathorax. The activities of this pair and of a third muscle pair, the III PD3 are also influenced by visual stimulation.The responses of all 6 muscles to optomotor stimuli which simulate the flight situations yaw, roll, thrust and lift are analyzed. Each situation elicits a unique pattern of activation/deactivation within this set of muscles. The activity pattern in non-flying animals allows the prediction of flight steering mechanisms such as changes of wing area in flight turns and provides a useful basis for the analysis of visuo-motor pathways.     

Anatomy of antenno-cerebral pathways in the brain of the sphinx moth Manduca sexta

Summary In the moth Manduca sexta, the number and morphology of neuronal connections between the antennal lobes and the protocerebrum were examined. Cobalt injections revealed eight morphological types of neurons with somata adjacent to the AL neuropil that project in the inner, middle, and outer antenno-cerebral tracts to the protocerebrum. Neurons innervating the macroglomerular complex and many neurons with fibers in the inner antennocerebral tract have uniglomerular antennal-lobe arborizations. Most neurons in the middle and outer antenno-cerebral tracts, on the other hand, seem to innervate more than one glomerulus. Protocerebral areas receiving direct input from the antennal lobe include the calyces of the mushroom bodies, and circumscribed areas termed olfactory foci in the lateral horn of the protocerebrum and several other regions, especially areas in close proximity to the mushroom bodies. Fibers in the inner antenno-cerebral tract that innervate the male-specific macroglomerular complex have arborizations in the protocerebrum that are distinct from the projections of sexually non-specific neurons. Protocerebral neurons projecting into the antennal lobe are much less numerous than antennal-lobe output cells. Most of these protocerebral fibers enter the antennal lobe in small fiber tracts that are different from those described above. In the protocerebrum, these centrifugal cells arborize in olfactory foci and also in the inferior median protocerebrum and the lateral accessory lobes. The morphological diversity of connections between the antennal lobes and the protocerebrum, described here for the first time on a single-cell level, suggests a much greater physiological complexity of the olfactory system than has been assumed so far.     

Induced responses in Nicotiana attenuata affect behavior and growth of the specialist herbivore Manduca sexta

Many plants employ induced responses against generalist herbivores. Specialist herbivores, however, may employ several mechanisms to overcome the negative effects of induced plant defenses. Here we test how the behavior and development of specialist Manduca sexta larvae are affected by induced responses in their natural host plant Nicotiana attenuata. On a spatial scale relevant to both the plant and the herbivore, we first determined how methyl jasmonate (MeJA)-induced responses, such as increased nicotine production, affect the tendency of larvae to leave induced plants. When larvae were allowed to move between two plants planted in one pot, they left an MeJA-treated plant faster than a control plant. When both plants in the pot were MeJA-treated, the larvae developed more slowly than when both plants were uninduced, or when the larvae had the opportunity to move to an uninduced neighbor. The sooner larvae moved from an MeJA-treated plant to an untreated neighbor, the larger the body mass they attained. This demonstrates that M. sexta larvae can compensate behaviorally for the deleterious effects of induced plant responses. These effects were observed in plants grown under both low and high N supply rates, though the effects were more pronounced under high N. To examine the consequences of the timing and the direction of the host plant switching behavior for larval development, neonate larvae were fed leaves excised from induced and uninduced plants. Larvae confined to MeJA-treated leaves had higher mortality rates and grew slower than larvae fed only control leaves. This demonstrates that MeJA-induced responses decrease growth and development of specialist herbivores that do not have the behavioral option of moving to an uninduced plant. The sooner the larvae were switched to MeJA-treated leaves, the slower their development compared to larvae fed only uninduced leaves. In contrast, the sooner larvae fed MeJA-treated leaves were switched to control leaves, the faster they developed. Again the effects of MeJA treatment were stronger in plants grown under high N supply. We propose that induced plants growing in close competition with an uninduced conspecific may offset the fitness costs of these induced responses and perhaps obtain a fitness benefit by motivating herbivores to move to their neighboring competitors. Received: 25 March 1999 / Accepted: 8 October 1999     

Multisegmental motor activity in the segmentally restricted gin trap behavior in Manduca sexta pupae

Stimulation of sensory neurons innervating hairs in the gin traps on the abdomen of Manduca sexta pupae evokes a rapid bending of the abdomen that is restricted to one or more of the three articulating posterior segments. However, electrical stimulation of the gin trap sensory nerve in an isolated abdominal nerve cord evokes characteristic motor neuron activity in every abdominal segment. To determine if the segmentally distributed motor activity also occurred in intact animals and how it contributed to the segmentally restricted reflex movement, mechanical stimulation of the sensory hairs in intact animals was used to evoke reflex responses that were recorded as electromyograms synchronized with video recordings of the behavior. Motor activity was monitored during movements to determine if there was activity in many segments when the movement was restricted to one segment. Coordinated muscle activity was evoked throughout the abdomen in response to stimulation of any of the three gin traps, even when movement was restricted to one segment. Differences in the timing of ipsilateral and contralateral motor activity among segments allowed the closing of gin traps to be segmentally restricted. These findings suggest that the neural circuit underlying the gin trap reflex is distributed throughout the abdominal nerve cord. This network generates a complex, yet coordinated, motor pattern with muscular activity in many abdominal segments that produces a localized bending reflex. Accepted: 10 January 1997     

Altered expression of flowering class B and class C genes in the Appendix tobacco mutant

 In the tobacco (Nicotiana tabacum) Appendix mutant, anthers are tipped by a miniature style and stigma. The outgrowth appears on the anther when it is already differentiating and follows the developmental timing of the central carpel. The Appendix mutation thus represents a late homeotic transformation suggesting that the APPENDIX (APX) gene either could be a misregulated organ identity gene or could be involved in regulating the expression of such genes. RFLP analysis with two class B (TM6 and NTGLO) and a class C (NAG) probes revealed that the Appendix phenotype is not caused by a mutation in one of these genes. However, in situ hybridization showed important changes in the expression of NTGLO and NAG in the mutant when compared with wild-type tobacco. Surprisingly, although no phenotypic alteration other than the style and stigma outgrowth is observed in the Appendix mutant, changes in class B and class C gene expession were not restricted to the anther tip cells from which the outgrowth originates. As expected, NAG was expressed in the Appendix outgrowth but it was also overexpressed in the normal third and fourth whorl organs at the time the outgrowth, as well as the central styles and stigmas, differentiated. Overexpression of a class C gene is probably responsible for the Appendix phenotype. In normal and mutant flowers, NTGLO was expressed in the second, third and fourth whorls up to the time of carpel fusion. Expression of this class B gene then ceased in the fourth whorl organs but was reactivated at later stages only in the styles and stigmas as well as in the outgrowths of the mutant. It thus seems that the function of the APX gene is either to regulate the late expression of organ identity genes or to control cell proliferation in such a way that, in the mutant, some cells are in a state where they respond in an unusual way to developmental signals. Received: 17 October 1997 / Revision accepted: 24 March 1998     

Antennal receptive fields of pheromone-responsive projection neurons in the antennal lobes of the male sphinx moth Manduca sexta

Stimulation of the antenna of the male moth, Manduca sexta, with a key component of the female's sex pheromone and a mimic of the second key component evokes responses in projection neurons in the sexually dimorphic macroglomerular complex of the antennal lobe. Using intracellular recording and staining techniques, we studied the antennal receptive fields of 149 such projection neurons. An antennal flagellum was stimulated in six regions along its proximo-distal axis with one or both of the pheromone-related compounds while activity was recorded in projection neurons. These neurons fell mainly into two groups, based on their responses to the two-component blend: neurons with broad receptive fields that were excited when any region of the flagellum was stimulated, and neurons selectively excited by stimulation of the proximal region of the flagellum. Projection neurons that were depolarized by stimulation of one antennal region were not inhibited by stimulation of other regions, suggesting absence of antennotopic center-surround organization. In most projection neurons, the receptive field was determined by afferent input evoked by only one of the two components. Different receptive-field properties of projection neurons may be related to the roles of these neurons in sensory control of the various phases of pheromone-modulated behavior of male moths. Accepted: 30 January 1998     

Serotonin-immunoreactive neurons in the median protocerebrum and suboesophageal ganglion of the sphinx moth Manduca sexta

Summary Serotonin-immunoreactive neurons in the median protocerebrum and suboesophageal ganglion of the sphinx moth Manduca sexta were individually reconstructed. Serotonin immunoreactivity was detected in 19–20 bilaterally symmetrical pairs of interneurons in the midbrain and 10 pairs in the suboesophageal ganglion. These neurons were also immunoreactive with antisera against DOPA decarboxylase. All major neuropil regions except the protocerebral bridge are innervated by these neurons. In addition, efferent cells are serotonin-immunoreactive in the frontal ganglion (5 neurons) and the suboesophageal ganglion (2 pairs of neurons). The latter cells probably give rise to an extensive network of immunoreactive terminals on the surface of the suboesophageal ganglion and suboesophageal nerves. Most of the serotonin-immunoreactive neurons show a gradient in the intensity of immunoreactive staining, suggesting low levels of serotonin in cell bodies and dendritic arbors and highest concentrations in axonal terminals. Serotonin-immunoreactive cells often occur in pairs with similar morphological features. With one exception, all serotonin-immunoreactive neurons have bilateral projections with at least some arborizations in identical neuropil areas in both hemispheres. The morphology of several neurons suggests that they are part of neuronal feedback circuits. The similarity in the arborization patterns of serotonin-immunoreactive neurons raises the possibility that their outgrowing neurites experienced similar forces during embryonic development. The morphological similarities further suggest that serotonin-immunoreactive interneurons in the midbrain and suboesophageal ganglion share physiological characteristics.Abbreviations CNS central nervous system - DDC DOPA decarboxylase - LAL lateral accessory lobe - SLI serotonin-like immunoreactivity - SOG suboesophageal ganglion - VLP ventro-lateral protocerebrum     

Anatomical identification of glomeruli in the antennal lobes of the male sphinx moth Manduca sexta

Summary Computer-assisted neuroanatomical methods have been used to demonstrate unique identities of the glomeruli of the antennal lobes (ALs) in males of the sphinx moth Manduca sexta. The glomerular neuropil consists of the male-specific macroglomerular complex, which comprises two closely apposed bulky subunits, and 64±1 ordinary glomeruli arrayed in a shell around a central region of coarse neuropil. Computergenerated maps show the exact locations of all glomeruli and adjacent groups of neuronal somata in a constant Cartesian coordinate system, such that these can be accurately identified in any individual. The glomeruli belong to three classes according to the number and type of identification criteria they satisfy. The larger class comprises glomeruli (n=44) identified only in the computer-generated maps on the basis of their relative positions. The other two classes include glomeruli that were also identified in sections, either directly from their proximity to readily identifiable structures and their shape and size (n=10, including the labial-palp-pit-organ (LPO) glomerulus), or indirectly from their positions relative to the former (n=9). Two very small glomeruli were present in only one AL, demonstrating the existence of anomalous glomeruli, whereas another glomerulus had no homologue in both ALs of one individual. The true number of ordinary glomeruli (per male AL) was thus estimated to be 64. The uncertainty in delineating some glomeruli might affect this number without implying modification of the homologies proposed. The locations of tracts and cell groups, both within and near the AL, are also invariant with respect to glomeruli, as shown in the computer maps. The methods employed are general and might be useful to researchers in related fields. The results obtained call for more attention to the precise geometry of neural structures.