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.     

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.     

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     

Organization of the antennal motor system in the sphinx moth Manduca sexta

The antennae of the sphinx moth Manduca sexta are multimodal sense organs, each comprising three segments: scape, pedicel, and flagellum. Each antenna is moved by two systems of muscles, one controlling the movement of the scape and consisting of five muscles situated in the head capsule (extrinsic muscles), and the other system located within the scape (intrinsic muscles) and consisting of four muscles that move the pedicel. At least seven motoneurons innervate the extrinsic muscles, and at least five motoneurons innervate the intrinsic muscles. The dendritic fields of the antennal motoneurons overlap one another extensively and are located in the neuropil of the antennal mechanosensory and motor center. The density of motoneuronal arborizations is greatest in the lateral part of this neuropil region and decreases more medially. None of the motoneurons exhibits a contralateral projection. The cell bodies of motoneurons innervating the extrinsic muscles are distributed throughout an arching band of neuronal somata dorsal and dorsolateral to the neuropil of the antennal mechanosensory and motor center, whereas the cell bodies of motoneurons innervating the intrinsic muscles reside mainly among the neuronal somata situated dorsolateral to that neuropil. Received: 30 March 1996 / Accepted: 23 June 1996     

Local interneurons and information processing in the olfactory glomeruli of the moth Manduca sexta

Intracellular recordings were made from the major neurites of local interneurons in the moth antennal lobe. Antennal nerve stimulation evoked 3 patterns of postsynaptic activity: (i) a short-latency compound excitatory postsynaptic potential that, based on electrical stimulation of the antennal nerve and stimulation of the antenna with odors, represents a monosynaptic input from olfactory afferent axons (71 out of 86 neurons), (ii) a delayed activation of firing in response to both electrical- and odor-driven input (11 neurons), and (iii) a delayed membrane hyperpolarization in response to antennal nerve input (4 neurons).Simultaneous intracellular recordings from a local interneuron with short-latency responses and a projection (output) neuron revealed unidirectional synaptic interactions between these two cell types. In 20% of the 30 pairs studied, spontaneous and current-induced spiking activity in a local interneuron correlated with hyperpolarization and suppression of firing in a projection neuron. No evidence for recurrent or feedback inhibition of projection neurons was found. Furthermore, suppression of firing in an inhibitory local interneuron led to an increase in firing in the normally quiescent projection neuron, suggesting that a disinhibitory pathway may mediate excitation in projection neurons. This is the first direct evidence of an inhibitory role for local interneurons in olfactory information processing in insects. Through different types of multisynaptic interactions with projection neurons, local interneurons help to generate and shape the output from olfactory glomeruli in the antennal lobe.Abbreviations AL antennal lobe - EPSP excitatory postsynaptic potential - GABA -aminobutyric acid - IPSP inhibitory postsynaptic potential - LN local interneuron - MGC macroglomerular complex - OB olfactory bulb - PN projection neuron - TES N-tris[hydroxymethyl]methyl-2-aminoethane-sulfonic acid     

An accessory olfactory pathway in Lepidoptera: the labial pit organ and its central projections in Manduca sexta and certain other sphinx moths and silk moths

Summary In the hawkmoth, Manduca sexta, the third segment of each labial palp contains a pit, which houses a densely packed array of sensilla. We have named this structure the labial pit organ (LPO). The sensilla within the pit are typical of olfactory receptors, characterized by a grooved surface, wall pores, and pore tubules. Axons arising from receptor cells that innervate these sensilla project bilaterally to a single glomerulus in each antennal lobe. We have compared this central projection with that in three other species of Manduca (M. quinquemaculata, M. dilucida, and M. lanuginosa) and in the silkmoths Antheraea polyphemus and Bombyx mori. A bilateral projection to a single glomerulus in each antennal lobe is present in all cases. We suggest that the LPO serves as an accessory olfactory organ in adult Lepidoptera.     

Glial cells in adult and developing prothoracic ganglion of the hawk moth Manduca sexta

The glial cells of the prothoracic ganglion of the hawk moth Manduca sexta were studied in histological sections of several postembryonic stages and classified according to cell morphology, size, staining properties, and topographical relationships. In general, each glial cell type was found to be confined to one of the major ganglionic domains and each of these domains (i.e., perineurium, cell body rind, glial cover of the neuropil, and neuropil) was found to comprise specific cell types. Some types of glia were recognized in both larval and later stages, but other types were found exclusively from late pupal stages. It is proposed that the higher morphological diversity expressed by the glia of the pharate adult is attained by differentiation of new cell types during metamorphosis. Before the differentiation of new cell types, extensive cell death and cell proliferation seem to occur within some glial subpopulations.     

Development of A-type allatostatin immunoreactivity in antennal lobe neurons of the sphinx moth <Emphasis Type="Italic">Manduca sexta</Emphasis>

The antennal lobe (AL) of the sphinx moth Manduca sexta is a well-established model system for studying mechanisms of neuronal development. To understand whether neuropeptides are suited to playing a role during AL development, we have studied the cellular localization and temporal expression pattern of neuropeptides of the A-type allatostatin family. Based on morphology and developmental appearance, we distinguished four types of AST-A-immunoreactive cell types. The majority of the cells were local interneurons of the AL (type Ia) which acquired AST-A immunostaining in a complex pattern consisting of three rising (RI–RIII) and two declining phases (DI, DII). Type Ib neurons consisted of two local neurons with large cell bodies not appearing before 7/8 days after pupal ecdysis (P7/P8). Types II and III neurons accounted for single centrifugal neurons, with type II neurons present in the larva and disappearing in the early pupa. The type III neuron did not appear before P7/P8. RI and RII coincided with the rises of the ecdysteroid hemolymph titer. Artificially shifting the pupal 20-hydroxyecdysone (20E) peak to an earlier developmental time point resulted in the precocious appearance of AST-A immunostaining in types Ia, Ib, and III neurons. This result supports the hypothesis that the pupal rise in 20E plays a role in AST-A expression during AL development. Because of their early appearance in newly forming glomeruli, AST-A-immunoreactive fibers could be involved in glomerulus formation. Diffuse AST-A labeling during early AL development is discussed as a possible signal providing information for ingrowing olfactory receptor neurons.This work was supported by a DFG grant (Scha 678/3-3) to J.S.     

Developmental distribution of CaM kinase II in the antennal lobe of the sphinx moth <Emphasis Type="Italic">Manduca sexta</Emphasis>

The antennal lobe (primary olfactory center of insects) is completely reorganized during metamorphosis. This reorganization is accompanied by changing patterns of calcium signaling in neurons and glial cells. In the present study, we investigated the developmental distribution of a major calcium-dependent protein, viz., calcium/calmodulin-dependent protein kinase II (CaM kinase II), in the antennal lobe of the sphinx moth Manduca sexta by using a monoclonal antibody. During synaptogenesis (developmental stages 6–10), we found a redistribution of CaM kinase II immunoreactivity, from a homogeneous distribution in the immature neuropil to an accumulation in the neuropil of the glomeruli. CaM kinase II immunoreactivity was less intense in olfactory receptor axons of the antennal nerve and antennal lobe glial cells. Western blot analysis revealed a growing content of CaM kinase II in antennal lobe tissue throughout metamorphosis. Injection of the CaM kinase inhibitor KN-93 into pupae resulted in a reduced number of antennal lobe glial cells migrating into the neuropil to form borders around glomeruli. The results suggest that CaM kinase II is involved in glial cell migration.This work was supported by the DFG LO779/2.     

Distribution of FMRFamide-like immunoreactivity in the brain and suboesophageal ganglion of the sphinx mothManduca sexta and colocalization with SCPB-, BPP-, and GABA-like immunoreactivity

Summary Using an antiserum against the tetrapeptide FMRFamide, we have studied the distribution of FMRFamide-like substances in the brain and suboesophageal ganglion of the sphinx mothManduca sexta. More than 2000 neurons per hemisphere exhibit FMRFamide-like immunoreactivity. Most of these cells reside within the optic lobe. Particular types of FMRFamide-immunoreactive neurons can be identified. Among these are neurosecretory cells, putatively centrifugal neurons of the optic lobe, local interneurons of the antennal lobe, mushroom-body Kenyon cells, and small-field neurons of the central complex. In the suboesophageal ganglion, groups of ventral midline neurons exhibit FMRFamide-like immunoreactivity. Some of these cells have axons in the maxillary nerves and apparently give rise to FMRFamide-immunoreactive terminals in the sheath of the suboesophageal ganglion and the maxillary nerves. In local interneurons of the antennal lobe and a particular group of protocerebral neurons, FMRFamide-like immunoreactivity is colocalized with GABA-like immunoreactivity. This suggests that FMRFamide-like peptides may be cotransmitters of these putatively GABAergic interneurons. All FMRFamide-immunoreactive neurons are, furthermore, immunoreactive with an antiserum against bovine pancreatic polypeptide, and the vast majority is also immunoreactive with an antibody against the molluscan small cardioactive peptide SCP_B. Therefore, it is possible that more than one peptide is localized within many FMRFamide-immunoreactive neurons. The results suggest that FMRFamide-related peptides are widespread within the nervous system ofM. sexta and might function as neurohormones and neurotransmitters in a variety of neuronal cell types.Abbreviations AL antennal lobe - BPPLI bovine pancreatic polypeptide-like immunoreactivity - FLI FMRFamide-like immunoreactivity - GLI GABA-like immunoreactivity - NSC neurosecretory cell - SCP _B LI small cardioactive peptide_B-like immunoreactivity - SLI serotonin-like immunoreactivity - SOG suboesophageal ganglion     

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.     

Serotonin-immunoreactive neurons in the suboesophageal ganglion of the caterpillar of the hawk moth Manduca sexta

Summary Serotonin-immunoreactivity is mapped in wholemounts and slices of the suboesophageal ganglion (SOG) of larval Manduca sexta by means of immunocytochemistry. An extensive meshwork of serotonin-immunoreactive nerve fibres on some peripheral nerves of the SOG has been demonstrated. This meshwork appears to belong to a serotonergic neurohemal system, probably supplied by two pairs of bilateral serotonin-immunoreactive neurons with big cell bodies on the dorsal side near the midline in the mandibular neuromere. Intracellular recording and staining revealed their physiology and morphology. These neurons produce long lasting (50 msec) action potentials, which suggest that they are neurosecretory cells. Two pairs of bilateral serotonin-immunoreactive interneurons similar to those of other insects are stained in the labial and maxillar neuromeres, but not in the mandibular neuromere. Their ventrolaterally located cell bodies project through a ventral commissure into the contralateral hemiganglion and then cross back again through a dorsal commissure. The axons project into the contralateral circumoesophageal connective.     

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     

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.     

Ecdysteroid binding sites localized by autoradiography in the central nervous system of precommitment fifth-stadium Manduca sexta larvae

Summary Brains and subesophageal ganglia from day 3.5 fifth stadium larvae of Manduca sexta were incubated in vitro with 4 nM tritiated ponasterone A, a 20-hydroxyecdysone analog, to determine whether uptake and specific binding of ecdysteroids occur at a cellular level. These tissues, which were taken just prior to the commitment peak in the hemolymph-ecdysteroid titer, showed saturable uptake of ³H-ponasterone A after 40–60 min of incubation. Uptake was blocked by the addition of 400 nM unlabelled ponasterone A, or of 500 nM or 1000 nM 20-hydroxyecdysone. RH 5849, a synthetic 20-hydroxyecdysone agonist with a long half-life, for which ecdysteroid receptors have low affinity, also reduced ponasterone A uptake at a concentration of 10 M. Autoradiographs of 4 m sections of brains revealed distinct nuclear concentrations of silver grains over cell populations in the pars intercerebralis, pars lateralis, and ventral tritocerebrum. Nuclear labelling was also found in many small cells around the mushroom bodies and the neuropil, and between the inner and outer larval optic lobes. Nuclear labelling of cells in the subesophageal ganglion was observed in the fronto-medial and lateral regions, in small cells around the neuropil, and caudally in a few large neurons. In addition to cells with nuclear labelling, both brains and ganglia at this development stage contained cells with exclusively cytoplasmic or both nuclear and cytoplasmic labelling. None of these apparent binding sites were observed in the competition experiments, suggesting that the binding is specific.     

Normal glomerular organization of the antennal lobes is not necessary for odor-modulated flight in female moths

A prominent hypothesis for the function of the glomerular structures in the primary olfactory neuropil of many groups of vertebrate and invertebrate animals is that they enable the processing and coding of information about the chemical compounds that compose complex odors. Previous studies have indicated that various degrees of glomerulus formation in the antennal lobes of the brain of the moth Manduca sexta can be effected by reducing the number of olfactory sensory axons that grow from the antenna into the antennal lobe during metamorphosis. To test the hypothesis that the presence of glomerular structure is necessary to process and identify odors, we substantially reduced, by surgery, the number of antennal segments in developing moths and upon metamorphosis we observed and quantified behavioral responses known to be elicited by odors. Intact and lesioned adult female moths were challenged to fly upwind to the source of an attractive host-plant odor in a wind tunnel. Some of the moths that had developed with reduced olfactory input flew upwind to the odor source. The flight behavior of these individuals was similar to the odor-mediated flight typically observed in moths that had developed normally. Histological analysis of the moths antennal lobes revealed that the lobes of more than half of the respondents that had been lesioned during development lacked normal glomerular organization. The neuropil of these abnormally developed antennal lobes was mostly aglomerular, but with a few isolated, clearly abnormal glomerulus-like structures. This suggests either that even a few abnormal glomeruli are sufficient to mediate this specific behavior or that canonical glomerular organization per se is not necessary for this odor-mediated behavior.     

Standard three-dimensional glomeruli of the <Emphasis Type="Italic">Manduca sexta</Emphasis> antennal lobe: a tool to study both developmental and adult neuronal plasticity

The metamorphosing antennal lobe (AL) of the sphinx moth Manduca sexta serves as an established model system for studying neuronal development. To improve our understanding of mechanisms involved in neuronal plasticity, we have analyzed the size, shape, and localization of ten identified glomeruli at three different time points during development and in the adult, viz., (1) 13 days after pupal eclosion (P13), which reflects a time when the basic glomerular map has formed, (2) immediately after adult eclosion (A0), which represents a time when the newly formed glomeruli are uninfluenced by external odors, and (3) 4 days after adult eclosion (A4), which reflects a time when the animals have been exposed to surrounding odors. Our data from normally developing ALs of male M. sexta from P13 to A0 revealed an increase in size of all examined glomeruli of between 40% and 130%, with the strongest increases occurring in two of the three sex-specific glomeruli (cumulus, toroid). From A0 to A4, the cumulus and toroid increased significantly when correlated to AL volume, whereas the other glomeruli reached the sizes gained after A0. This study was based on antibody staining against the ubiquitous synaptic vesicle protein synaptotagmin, confocal laser scan microscopy, and the three-dimensional (3D) analysis tool AMIRA. Tissue permeability and therefore reliability of the staining quality was enhanced by using formalin/methanol fixation. The standard 3D glomeruli introduced in this study can now be used as basic tools for further examination of neuronal plasticity at the level of the identified neuropil structures, viz., the glomeruli of the AL of M. sexta.     

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.     

Sensory organs of the thoracic legs of the moth Manduca sexta

Summary The thoracic legs of the moth Manduca sexta acquire a new form and develop a new complement of sensory organs and muscles during metamorphosis from larva to adult. Because of our interest in the reorganization of neural circuitry and the acquisition of new behaviors during metamorphosis, we are characterizing sensory elements of larval and adult legs so that we may determine the contribution of new sensory inputs to the changes in behaviors. Here we describe the sensory structures of adult legs using scanning electron microscopy to view the external sensilla and cobalt staining to examine innervation by underlying sensory neurons. We find that, in contrast to larval legs, the adult legs are covered with a diverse array of sensilla. All three pairs of thoracic legs contain scattered, singly innervated scalelike sensilla. Campaniform sensilla occur singly or in clusters near joints. Hair plates, consisting of numerous singly innervated hairs, are also present near joints. Other more specialized sensilla occur on distal leg segments. These include singly innervated spines, two additional classes of singly innervated hairs, and three classes of multiply innervated sensilla. Internal sensory organs include chordotonal organs, subgenual organs, and multipolar joint receptors.