The development of identified neurosecretory cells in the tobacco hornworm, Manduca sexta

The prothoracicotropic hormone (PTTH) is a principal neuropeptide regulator of insect postembryonic molting and metamorphosis. In the tobacco hornworm, Manduca sexta, PTTH is produced by two neurosecretory cells (NSC) located in each protocerebral lobe of the brain. The development of these neurons, the L-NSC III, has been investigated immunocytologically to establish the time course of their morphological differentiation. PTTH may be one of the earliest neuropeptides expressed in insect embryos. PTTH-immunoreactivity was initially detected in the somata at 24 to 30% of embryonic development. Neurites sprouted shortly thereafter and began to grow medially through the brain anlage. By 42% embryonic development, the neurites had decussated to the contralateral brain lobe. As development progressed, the L-NSC III neurites grew along specific tracts through the contralateral brain lobe reaching the ventrolateral regions of the brain by approximately 60% development. The axons exited the brain through a retrocerebral nerve, the nervi corporis cardiaci I + II. At approximately 63% development, the axons innervated the corpus allatum and began branching to form neurohemal terminals for PTTH release. At 60% development, short collaterals began extending in the protocerebral neuropil. During the remainder of embryogenesis, both the dendritic collaterals and the terminal neurohemal varicosities continued to elongate and arborize. By 85% embryonic development, the basic architecture of the L-NSC III was established.     

The development of identified neurosecretory cells in the tobacco hornworm, Manduca sexta

The prothoracicotropic hormone (PTTH) is a principal neuropeptide regulator of insect postembryonic molting and metamorphosis. In the tobacco hornworm, Manduca sexta, PTTH is produced by two neurosecretory cells (NSC) located in each protocerebral lobe of the brain. The development of these neurons, the L-NSC III, has been investigated immunocytologically to establish the time course of their morphological differentiation. PTTH may be one of the earliest neuropeptides expressed in insect embryos. PTTH-immunoreactivity was initially detected in the somata at 24 to 30% of embryonic development. Neurites sprouted shortly thereafter and began to grow medially through the brain anlage. By 42% embryonic development, the neurites had decussated to the contralateral brain lobe. As development progressed, the L-NSC III neurites grew along specific tracts through the contralateral brain lobe reaching the ventrolateral regions of the brain by approximately 60% development. The axons exited the brain through a retrocerebral nerve, the nervi corporis cardiaci I + II. At approximately 63% development, the axons innervated the corpus allatum and began branching to form neurohemal terminals for PTTH release. At 60% development, short collaterals began extending in the protocerebral neuropil. During the remainder of embryogenesis, both the dendritic collaterals and the terminal neurohemal varicosities continued to elongate and arborize. By 85% embryonic development, the basic architecture of the L-NSC III was established.     

New features of the brain-retrocerebral neuroendocrine complex of the locust Schistocerca vaga (scudder)

Summary The techniques of axonal iontophoresis and cobalt sulfide precipitation were used to elucidate the relationships of the brain's neurosecretory cell groups and the retrocerebral complex of the locust Schistocerca vaga. The axons of the nervi corporis cardiaci I (NCC I) arise (1) from the medial neurosecretory cells of the protocerebrum, showing only limited branching, looping or spiraling; and (2) from a cell group previously undescribed for this species, located in the tritocerebrum. The axons project into the neurohemal and the glandular portions of the corpora cardiaca and into the hypocerebral ganglion, but not into the corpora allata. Axons of the NCC II arise from the lateral neurosecretory cells of the protocerebrum and project into the center of the corpora allata via the nervi corporis allati I (NCA I), as well as into the neurohemal and glandular portions of the corpora cardiaca. Axons of the NCC III arise from another newly described cell group in the tritocerebrum and end in both the corpora cardiaca and corpora allata. Axons of the NCA II arise from cells in the subesophageal ganglion and also end in the corpora allata.Supported by NIH Predoctoral Fellowship No. 5 F 01 GM 43816-03, NSF Grant GB-23033 and NIH Grant CA-05045 to H. A. Bern and USPHS Grant 1 R 01 NS09404 to C.H.F. Rowell.I wish to express my gratitude to Professors H.A. Bern and C.H.F. Rowell for unending encouragement and advice. I am indebted to Dr. Mick O'Shea for instruction in the cobalt/axonal iontophoresis method, and to Ms. Bea Bacher for excellent technical assistance.     

Neuroendocrine organs of the lemon-butterfly, Papilio demoleus L. II. The corpora cardiaca-allata complex of the adult.

The retrocerebral endocrine organs of the adult lemon-butterfly; Papilio demoleus have been described. The organs are subaortic lying closely behind the brain. While the nervi corporis cardiaci I (NCCI) originate from the protocerebrum of the brain, the NCCII seem to take their origin in the tritocerebrum in common with another nerve named earlier as the tegumentary nerve. The corpora cardiaca (CC) and corpora allata (CA) are closely approximated to each other obliterating the nervi corporis allati (NCA) which are conspicuous in the larva of the same species. An intercardiacal bridge (ICB) connects the CC of the two sides and acts as a possible centre of distribution for the neurosecretory material (NSM) to the gut. Histological evidence suggests that the NSM inside the CC remains intraaxonal without being primarily unloaded in the organs for storage. The intrinsic secretory cell of the CC are intimately associated with the neurosecretory fibres from the brain and bear fairly thick axons. No NSM could be detected in the CA of this insect.     

Studies on the neurosecretory system and retrocerebral endocrine glands of Nezara viridula Linn. (Heteroptera: Pentatomidae)

The neurosecretory system and retrocerebral endocrine glands of Nezara viridula Linn. have been described on the basis of in situ preparations and histological sections employing the paraldehyde fuchsin (PF) and performic acid-victoria blue (PAVB) techniques. In the brain of N. viridula, there are two medial groups–each consisting of five neurosecretory cells which belong to A-type. The lateral neurosecretory cells are absent. The axons of the two groups of medial neurosecretory cells (MNC) compose the two bundles of neurosecretory pathways (NSP) that decussate in the anterodorsal part of the protocerebrum. The two pathways, after the cross-over, run deep into the protocerebrum and deutocerebrum and emerge as NCC-I from the tritocerebrum. The nervi corporis cardiaci-I (NCC-I) of each side which are heavily loaded with NSM terminate in the aorta wall. Thus, the neurosecretory material (NSM), elaborated in the medial neurosecretory cells of the brain, is stored in the aortic wall and nervi corporis cardiaci-I (NCC-I). The NCC-II are very short nerves that originate from the tritocerebrum and terminate in the corpora cardiaca (CC) of their side. Below the aorta, but dorsal to the oesophagus, lie two oval or spherical corpora cardiaca. A corpus allatum (CA) lies posterior to the corpora cardiaca (CC). The corpora cardiaca do not contain NSM; only the intrinsic secretion of their cells has been occasionally observed which stains orange or green with PF staining method. The corpus allatum sometimes exhibits PF positive granules of cerebral origin. A new connection between the corpus allatum and aorta has been recorded. The suboesophageal ganglion contains two neurosecretory cells of A-type which, in structure and staining behaviour, are similar to the medial neurosecretory cells of the brain. The course and termination of axons of suboesophageal ganglion neurosecretory cells, and the storage organ for the secretion of these cells have been reported. It is suggested that the aortic wall and NCC-I axons function as neurohaemal organ for cerebral and suboesophageal secretions.     

Histology of the neurosecretory system and the retrocerebral endocrine glands of the adult migratory grasshopper, Melanoplus sanguinipes (Fab.) (Orthoptera: Acrididae)

Neurosecretory cells of only one type (A, sub type A₂) are seen in adult Melanoplus. Two groups of about 400 cells each are located dorsally in the pars intercerebralis medialis; four cells are located deep within the protocerebrum. We found no neurosecretory cells in other parts of the central or sympathetic nervous systems. In about 10% of the specimens, there was marked asymmetry in the location of the dorsal cell groups, with both of these groups and their axons located in one lobe of the protocerebrum. The nervi corporis cardiaci 1 cross-over in the corpus cardiacum, with the result that material produced by neurosecretory cells on one side of the brain is transported along axons that undergo two chiasmata to the corpus cardiacum of the same side. Stainable secretory material could be traced clearly from the cerebral cells to the corpus cardiacum, and even into the oesophageal nerves from the hypocerebral ganglion. However, stainable neurosecretory material is never present in the corpus allatum or along any of the nerves to this gland.     

Crustacean cardioactive peptide-immunoreactive neurons innervating brain neuropils,retrocerebral complex and stomatogastric nervous system of the locust,Locusta migratoria

The distribution and morphology of crustacean cardioactive peptide-immunoreactive neurons in the brain of the locust Locusta migratoria has been determined. Of more than 500 immunoreactive neurons in total, about 380 are interneurons in the optic lobes. These neurons invade several layers of the medulla and distal parts of the lobula. In addition, a small group of neurons projects into the accessory medulla, the lamina, and to several areas in the median protocerebrum. In the midbrain, 12 groups or individual neurons have been reconstructed. Four groups innervate areas of the superior lateral and ventral lateral protocerebrum and the lateral horn. Two cell groups have bilateral arborizations anterior and posterior to the central body or in the superior median protocerebrum. Ramifications in subunits of the central body and in the lateral and the median accessory lobes arise from four additional cell groups. Two local interneurons innervate the antennal lobe. A tritocerebral cell projects contralaterally into the frontal ganglion and appears to give rise to fibers in the recurrent nerve, and in the hypocerebral and ingluvial ganglia. Varicose fibers in the nervi corporis cardiaci III and the corpora cardiaca, and terminals on pharyngeal dilator muscles arise from two subesophageal neurons. Some of the locust neurons closely resemble immunopositive neurons in a beetle and a moth. Our results suggest that the peptide may be (1) a modulatory substance produced by many brain interneurons, and (2) a neurohormone released from subesophageal neurosecretory cells.     

Studies on the neuroendocrine system of the mangohopper, Idiocerus atkinsoni Leth. (Homoptera : Jassidae)

The neuroendocrine system of the homopteran, Idiocerus atkinsoni has been described, employing a neurosecretory stain. Two groups of medial neurosecretory cells (NSC) of one tinctorial type are present in the pars intercerebralis of the brain. Processes believed to be dendrites of the neurosecretory neurons lie superficially underneath the neurilemma and enclose neurosecretory material (NSM). Both the nervi corporis cardiaci, NCCI and NCCII, are branched. The branches of the former join to form an oesophageal nerve that runs on the oesophageal surface and terminates on the midgut, and those of the latter, innervate the oesophageal dilator muscles. Besides being present in the dendrite-like processes and NSC, the NSM is also seen in the NCCI, anterior part of the aorta and oesophageal nerve but not in the NCCII, corpora cardiaca (CC) and the corpus allatum (CA). It is suggested that the release of NSM into the circulation in this insect occurs through two main routes: the dendrites and the aorta. The evolution of the aorta as an exclusive neurohaemal organ in Hemiptera is discussed.     

Brain-derived neurotrophic factor-like neuropeptide is secreted as a neurohormone from specific brain neurons into the corpus allatum in the silkworm Bombyx mori

The aim of this study was to investigate the secretion of brain‐derived neurotrophic factor (BDNF)‐like neuropeptide in the silkworm, Bombyx mori , by using immunocytochemical techniques on the brain and retrocerebral complex of fifth instar larvae. In the brain, four pairs of median neurosecretory cell (MNC) bodies and six pairs of lateral neurosecretory cell (LNC) bodies had distinct immunoreactivities to this peptide, suggesting that this peptide is produced from two types of brain neuron. These reactivities were much stronger in the MNC than in the LNC. Labeled MNC projected their axons into the contralateral corpora allata, to which axons of labeled MNC were eventually innervated, through decussation in the median region, contralateral nerve corporis cardiaci I and nerve corpora allata I. Labeled LNC extended their axons into the ipsilateral corpora allata to be innervated through the ipsilateral nerve corporis cardiaci II and nerve corpora allata I. These results suggest that BDNF is secreted as a neurohormone from MNC and LNC of the brain into the corpora allata.     

Localization of leucokinin VIII in the cockroach,Leucophaea maderae,using an antiserum directed against an achetakinin-I analog

An antiserum against an achetakinin analog selectively localized leucokinin VIII (LKVIII) in the CNS ofLeucophaea maderae. Preabsorption studies of the achetakinin antiserum with either preimmune serum or LKVIII prevented a positive reaction in both ELISA and immunocytochemical procedures. LKVIII immunoreactive neurons were found in the brain, frontal, and subesophageal ganglion, all 3 thoracic ganglia and the terminal ganglion. Nerves originating from the thoracic and terminal abdominal ganglia contain LKVIII material. Lateral and medial neurosecretory cells synthesizing LKVIII-like products contribute axons to the nervi corporis cardiaci that terminate in neurohemal sites in the corpora cardiaca and nervi corporis allati. Thus, leucokinin VIII, like leucokinin I (LKI) and leucomyosuppressin (LMS), appears to have both a neurohemal and neurotransmitter mode of regulating target cells inL. maderae.     

A comparative ultrastructural and physiological study on melanophores of wild-type and periodic albino mutants of Xenopus laevis

Summary The central and visceral nervous systems of the cockroach Periplaneta americana were studied by means of the peroxidase-antiperoxidase immunocytochemical method, with the use of antibody to bovine pancreatic polypeptide (PP). PP-like immunoreactive neuron somata are most numerous in the brain; at least 6 pairs of cell groups occur in clearly defined regions. Three pairs of cells each are also present in the suboesophageal ganglion and the thoracic ganglia, one pair of a single cell each in the first abdominal and the frontal ganglia, and 4 to 6 pairs of single cells in the terminal ganglion. No reactive cells were found in the retrocerebral complex and the second to the fifth abdominal ganglia. The axons containing PP-like immunoreactivity issue many branches that are distributed in the entire brain-retrocerebral complex, ventral cord, and visceral nervous system. PP-like immunoreactive material produced in the brain seems to be transported by three routes: protocerebrum to corpora cardiaca (-allata) through the nervi corporis cardiaci, tritocerebrum to visceral nervous system through frontal commissures, and to ventral cord through circumoesophageal connectives.A possible homology between the mammalian brain-GEP (gastro-enteropancreatic) system and the brain-midgut system of this insect is discussed.     

Life cycle expression of a bombyxin-like neuropeptide in the tobacco hornworm,Manduca sexta

Immunocytochemistry was used to investigate the developmental expression of the insulin-like neuropeptide bombyxin in the tobacco hornworm, Manduca sexta. A mouse monoclonal antibody raised against a synthetic peptide corresponding to bombyxin's A-chain N-terminus was used to localize a bombyxin-like peptide to a group of cerebral medial neurosecretory cells, the M-NSC IIa(2). Immunostaining was first detected on day 0 of the second larval instar, localized in the M-NSC IIa(2) somata and in the neurohemal organ, the corpora allata (CA). By day 0 of the fourth larval instar, the peptide was present throughout the M-NSC IIa(2) somata, axons, dendritic fields and CA. Between days 7 and 9 of the fifth instar, a dramatic reduction in the dendritic fields and CA staining occurred, suggesting the peptide is released. After day 2 of the pupal period, only M-NSC IIa(2) somata immunostained, a pattern that persisted through day 2 of the adult stage. The specificity of immunostaining was demonstrated by using a synthetic bombyxin peptide to block staining. These developmental data reveal times of potential Manduca bombyxin-like peptide release which should provide insight into the peptide's function.     

THE NEUROENDOCRINE SYSTEM AND STOMATOGASTRIC NERVOUS SYSTEM OF THE ADULT TSETSE FLY GLOSSINA MORSITANS

The brain of Glossina morsitans Westwood contains four groups of neurosecretory cells which are stainable with chrome haematozylin and phloxin. The axons of these cells form a pair of nervi corporis cardiaci which pass posteriorly from the brain and innervate the corpora cardiaca and corpus allatum before uniting with a small ganglion posterior to the corpora cardiaca. This ganglion is considered to represent the fusion of the fusion of the hypocerebral and ventricular ganglia which remain separate in other insects.
There is no frontal ganglion in the adult Glossina and the recurrent nerve fuses with one of the nervi corporis cardiaci immediately behind the brain. The oesophageal nerves arising from the fused hypocerebral and ventricular ganglia innervate the oesophagus in the anterior part of the thorax, the proventriculus and the posterior extension of the oesophagus close to the crop. These nerves possess both sensory and motor nerve endings. The differences which exist between Glossina and other cyclorrhaphous Diptera with respect to their neuroendocrine/stomatogastric system are noted and considered in terms of the control of neuroendocrine function.     

Proctolin-like immunoreactivity in the central and peripheral nervous systems of the locust, Locusta migratoria

Proctolin-like immunoreactivity (PLI) was widely distributed in the locust, Locusta migratoria, within the central, peripheral and stomatogastric nervous systems, as well as the digestive system and retrocerebral complex. Proctolin-like immunoreactivity was observed in cells and processes of the brain and all ganglia of the ventral nerve cord. Of interest, PLI was found in the lateral neurosecretory cells, which send axons within the paired nervi corporis cardiaci II (NCC II) to the corpus cardiacum (CC). The CC contained extensive processes displaying PLI, which continued on within the paired nervi corporis allata (NCA) to the paired corpora allata (CA) where the axons entered and branched therein. The frontal and hypocerebral ganglia of the stomatogastric nervous system contained PLI within processes, resulting in a brightly staining neuropile. Each region of the gut contained PLI in axons and processes of varying patterns and densities. The paired ingluvial ganglia contained PLI, including an extensively stained neuropile and immunoreactive axons projecting through the nerves to the foregut. The hindgut contained PLI within longitudinal tracts, with lateral projections originating from the 8th abdominal ganglion via the proctodeal nerve. The midgut contained PLI in a regular latticework pattern with many varicosities and blebs. No difference in PLI in cells and processes of the central nervous system (CNS) was found between males and females.     

Development of blood-cerebrospinal fluid barrier to horseradish peroxidase in the avian choroidal epithelium

Summary Four neurons in the brain of the migratory locust were immunohistologically identified with an anti-met-enkephalin antiserum. The perikarya of two of these cells are located in the center of each of the two groups of lateral protocerebral neurosecretory cells. The fibres coming from these perikarya terminate in numerous immunoreactive ramifications visible at the periphery of both tractus I to the corpora cardiaca, through which pass the neurosecretory products of the pars intercerebralis. The other two cell bodies are located at the bases of the two optic lobes; their fibres enter the posterior part of the protocerebrum and ramify around the root of the nervus corporis cardiaci II, another area through which neurosecretory products pass. The topographic distribution of these met-enkephalin arborizations suggests that these four neurons may act as neuromodulators of the acitivity of the major neurosecretory cells in the brain of this insect.     

Neurons important for the photoperiodic control of diapause in the bean bug, <Emphasis Type="Italic">Riptortus pedestris</Emphasis>

The morphology and functions of the brain neurons projecting to the retrocerebral complex were examined in terms of photoperiodic control of adult diapause in the bean bug, Riptortus pedestris. Backfills through the nervi corporis cardiaci stained 15-20 pairs of somata in the pars intercerebralis (PI) with contralateral axons, and 14-24 pairs in the pars lateralis (PL) with ipsilateral axons to the nervi corporis cardiaci. In the PL, two clusters of somata, PL-d and PL-v, were found. Forwardfills showed neurons in the PI terminated in the aorta, and those in the PL at the corpus cardiacum, corpus allatum, and aorta. Removal of the PI did not cause effects on diapause incidence both under short-day (12 h:12 h, light:dark) and long-day conditions (16 h:8 h, light:dark) at 25 degrees C. Under short-day conditions, diapause incidence was significantly lower than the controls after removal of the PL. Either removal of PL-d or PL-v did not reduce diapause incidence. It decreased only when both the PL-d and PL-v were ablated. The PI is not indispensable for diapause in R. pedestris, and both PL-d and PL-v neurons are suggested to be involved in photoperiodic inhibition of ovarian development.     

Diapause-dependent changes in prothoracicotropic hormone-producing neurons of the tobacco hornworm,Manduca sexta

The prothoracicotropic hormone (PTTH), which stimulates ecdysteroid synthesis in the prothoracic glands, is produced, in the dorso-lateral protocerebrum of Manduca sexta, by paired peptidergic neurons, the lateral neurosecretory cell group III (L-NSC III). Our study revealed ultrastructural features of L-NSC III, identified by immunogold labeling, and compared developing and diapause states. In developing and early-diapause pupae, L-NSC III soma ultrastructure is similar and is characterized by numerous clusters of neurosecretory granules (NSG) and an extensive trophospongium formed by satellite-glial cells. However, as diapause progresses, the ultrastructure changes, with the NSG becoming concentrated into large clusters separated by highly organized rough endoplasmic reticulum. Most conspicuous is a substantial reduction in the number of Golgi complexes and the glial trophospongium, and the presence of stacked plasma membrane separating the glia and neuron somata. The deep-diapause soma also has abundant glycogen deposits and autophagic vacuoles. With diapause termination, this morphology reverts to the nondiapause ultrastructure within three days, i.e. just before PTTH release that evokes development to the adult. During PTTH release the abundance of NSG in the soma does not change, suggesting that NSG depletion in the perikarya is not a marker for neurosecretion by the L-NSC III.     

The neurosecretory cells of the brain and suboesophageal ganglion of Chironomus riparius

The neurosecretory cells of the supra- and suboesophageal ganglia of young, unmated, adult male midges, Chironomus riparius, have been examined by both light and electron microscopy. The 5 cell types recognized have been placed in three major categories on the basis of their ultrastructural characteristics:—α₁ cells, of which there are 8 in each medial neurosecretory cell (MNC) group and 3 in each group of ventral neurosecretory cells (VNC), contain electron-dense granules, 150 to 200 nm in diameter; α₂ cells containing irregular, electron-dense granules, 70 to 120 nm in diameter comprise the remaining 3 cells in each VNC group and the 2 or 3 cells in each outer neurosecretory cell (ONC) group; α₃ cells, of which there are 1 or 2 on each side of the midline in the ventral cortex of the sub-oesophageal ganglion (SNC₂), contain electron-lucent, spherical granules, 70 to 120 nm in diameter. The β cells contain spherical or ellipsoidal, electron-lucent granules, 80 to 100 nm in diameter, and make up the lateral neurosecretory cell (LNC) groups, each of three or four cells. The γ cells contain both spherical and flattened, electron-dense granules, 130 to 160 nm in diameter and 150 to 250 by 70 to 150 nm in size respectively, only 1 cell of this category being found in each half of the suboesophageal ganglion in the dorsal cortex (SNC₁). Axons from the MNC and VNC form the nervi corporis cardiaci I (NCCI) and those of the LNC and ONC, the nervi corporis cardiaci II (NCCII). Those of the SNC₁ appear to enter the wall of the stomodaeum but axons of the SNC₂ could not be traced.     

The neurosecretory system of the adult Melanogryllus desertus pall. (Orthoptera,Gryllidae)

Summary The cerebral neurohemal area of Melanogryllus desertus is located posteriorly among the neurons of nervus corporis cardiaci I (NCCI) on the ventral median surface of the protocerebrum where axons penetrate the neural lamella and terminate on its outer surface. Numerous neurosecretory fibers containing three different types of granule occur within and on the outer surface of the neural lamella.The release of neurosecretory granules is accomplished by exocytosis and the formation of synaptoids. It can also take place as a mass release of granules into the stroma.     

Dynamics, following axotomy, of the A1 and A2 median protocerebral neurosecretory cells of the African locust. I. Histophysiological study

In the African locust, the secretory dynamics of the A1 and A2 median protocerebral neurosecretory cells (M-NSC) is evaluated by the product content and the gonadotropic action of these cells. The axotomy of the A1 and A2 M-NSC disturbes their secretory dynamics. The axonal regeneration, if it leads to a reconnexion with the corpora cardiaca (CC), restores a normal secretory dynamics of the M-NSC. Without reconnexion to the CC, the nervi corpori cardiaci interni (NCCI) do note regenerate or regenerate to form new nerve endings. In these cases, the secretory dynamics of the M-NSC remains more or less inhibited specially that of the A2 M-NSC which can be completely suppressed. A functional state of the A2 M-NSC could be necessary for the beginning of the vitellogenesis. In the CC, separated from the regenerating M-NSC, the A1 and A2 neurosecretory products are transformed into large globules which became intensively basophil before disappearance.