Intrapulmonary neuro-epithelial bodies in newborn rabbits

Summary Neonatal rabbit neuro-epithelial bodies (NEB) were investigated under various experimental conditions with light microscopy, microspectrography, morphometry and electron microscopy. (1) Hypoxia causes a decreased amine fluorescence intensity and an increased secretory exocytosis of dense core vesicles (DCV). Otherwise the NEB appear structurally normal. (2) Hypercapnia also produces a decreased fluorescence and an increased exocytosis; ultrastructurally, however, the dense core of DCV fragmentizes. (3) Hyperoxia does not appear to affect significantly either fluorescence or exocytosis. (4) The uptake of biogenic amines such as 5-HTP and L-DOPA was demonstrated by fluorometry and electron microscopy. (5) Reserpine, on the other hand, provokes an amine depletion with a decrease of the NEB fluorescence and an ultrastructural palor of the DCV. (6) Intratracheally administered nicotine is accompanied by a decreased fluorescence and a distinct exocytosis of fragmented DCV.The reaction of NEB to hypoxia and hypercapnia suggests that these corpuscles could be intrapulmonary chemoreceptors (in addition to the classically known central and peripheral chemoreceptors), inducing a reflex reaction through the liberation of DCV at the corpuscular sensible nerve endings and via the CNS. In addition, they may subserve a local intrapulmonary effect by modulating directly the hypoxic and hypercapnic pulmonary vasoconstriction and thus the V/Q ratio. Acknowledgment. This study was supported by a grant from The Council for Tobacco Research, U.S.A., and the Nationaal Fonds voor Wetenschappelijk Onderzoek, Belgium. We thank M.R. Van Hamme, R. Renwart and K. Armee for technical, G. Pison and St. Ons for photographical and N. Tyberghien and G. Verbeeck for secretarial assistanceMartin Deleersnyder (deceased May 11, 1976) was Candidate of the Nationaal Fonds voor Wetenschappelijk Onderzoek, Belgium     

Immunohistochemical localization of serotonin in intrapulmonary neuro-epithelial bodies

Summary A recently developed immunohistochemical technique for serotonin (Steinbusch et al. 1978) was used in the present investigation to study the occurrence of this indoleamine in the granulated epithelial cells of neuroepithelial bodies (NEB). Lungs from neonatal rabbits and pigs exhibit immunoreactive cell groups identical in morphology and their preferential location (i.e., at bronchiolar bifurcations) to the recently described intrapulmonary NEB. Moreover, in the trachea and lung of rabbits isolated immunoreactive cells, presumably of Kultschitsky type, were found in the lining respiratory mucosa. Such single cells were also frequently observed in the bronchial epithelium of pig lungs. It is concluded that the corpuscular cells, being modulated by the central nervous system, probably represent intrapulmonary neuro(chemo-)receptors with local secretory activities, one of the substances released being serotonin and reacting to the oxygen composition of the inhaled air. It is proposed that the cells of Kultschitsky type exert a more local effect upon the airways. Send offprint requests to: Katholieke Universiteit te Leuven, School of Medicine, Laboratory of Histopathology, B-3000 Leuven, Belgium     

The distribution of monoamines in the tel-, di-and mesencephalon of Xenopus laevis tadpoles,with special reference to the hypothalamo-hypophysial system

Summary In an attempt to further our knowledge about the structure and function of the recently identified intrapulmonary Neuro-epithelial Bodies (NEB's) (Lauweryns et al., 1972a, 1972b), lungs of 84 neonatal rabbits and 6 neonatal mice were studied along four different lines of investigation. Several routine and silver staining methods, Falck's fluorescent amine technic and histochemical and electron microscopical techniques were performed.1. In order to test the probable chemoreceptor function of the NEB's, animals were exposed to hypoxia. Under such circumstances, the corpuscular cells of the NEB's secrete their dense-cored, serotonin-containing vesicles at their basal vascular pole. 2. After reserpine pretreatment, the NEB's of otherwise normal animals reveal a distinct amine depletion, the corpuscular cells exhibiting a decreased yellow fluorescence and ultrastructurally a clearing up of their dense-cored vesicles. 3. Studied on serial sections with the electron microscope, various types of morphologically afferent-like and efferent-like nerve endings, making contact as well with the corpuscular cells as amongst themselves have been detailed. 4. Cytochemically the corpuscular cells react positively with alpha-glycerophosphate dehydrogenase, acetylcholinesterase and Solcia's lead hematoxylin stain for endocrine cells producing polypeptides and amines.It is proposed that the NEB's provide an intrapulmonary, hypoxia-sensitive neuro(chemo-) receptor system in addition to the well established central and peripheral (e.g. carotid body) chemoreceptors. They contain and secrete serotonin and probably also related amines or peptides, which could influence the pulmonary vasoconstrictor response. According to classic morphologic criteria, they possess a dual innervation, both afferent and efferent.Various other possible functions of the NEB's in normal and diseased lungs are briefly proposed.This study has been supported by a grant from The Council for Tobacco Research — U.S.A. and the Nationaal Fonds voor Wetenschappelijk Onderzoek — Belgium. We thank P. Theunynck for performing some of the histochemical studies, B. Van Rijkel, B. Emanuel and R. Renwart for technical, G. Pison and St. Ons for photographical and N. Tyberghien for secretarial assistance.     

Neuro-epithelial bodies in organ cultures of fetal rabbit lungs

Lung explants from fetal rabbit at the late glandular stage of development (20 days' gestation) and near term (31 days' gestation) were maintained in organ culture for up to 22 days. They were studied by light and electron microscopy to determine whether neuro-epithelial bodies (NEB) of the lung retain structural integrity in vitro. Cultured NEB retained argyrophilia and specific amine fluorescence after formaldehyde condensation. Their ultrastructural morphology showed some differences from that of uncultured NEB: the terminal axons had degenerated and the secretory granules (dense-core vesicles, DCV) were slightly larger, more pleomorphic, more electron-dense, and redistributed throughout the cytoplasm rather than being confined chiefly to the basal regions. These changes, together with hypertrophy of Golgi zones, suggest increased synthesis and storage of secretory products in the DCV during culture. In NEB from near-term explants cultured for 7 days and incubated with reserpine, the core of DCV decreased in size and electron-density and became finely granular, a sign of amine release. Ca++ ionophore No. A-23187, also, induced changes in the ultrastructure of DCV, suggesting that the secretory process in lung neuro-endocrine cells, as in other secretory cells, is Ca++-dependent.     

Morphometric analysis of hypoxia-induced synaptic activity in intrapulmonary neuroepithelial bodies

Summary A morphometric analysis has demonstrated ultrastructural changes induced by hypoxia in the epithelial cells and the intracorpuscular nerve endings of the presumed chemoreceptive intrapulmonary neuroepithelial bodies (NEB) of neonatal rabbits.Acute hypoxia stimulates an exocytosis of epithelial dense-core vesicles (DCV) at the level of the morphologically afferent or sensory (type 1 a) intracorpuscular nerve endings of the NEB. Assuming the epithelial cells to be chemoreceptive, this phenomenon could represent a transduction of sensory stimuli.In the morphologically efferent or motor (type 2 and type 1 b) intracorpuscular nerve endings of the NEB, acute hypoxia causes a depletion of synaptic vesicles and an increase in the amount of membrane-bounded cisternae and multivesicular bodies, suggestive of an enhanced synaptic activity of these nerve endings. It is proposed that the chemoreceptor cells could thus in turn be modulated centrifugally by their efferent-like intracorpuscular nerve endings.It has been proposed in our earlier studies that the NEB probably are intrapulmonary chemoreceptors with local secretory activities, reacting to the composition of the inhaled air. By the release of serotonin and peptide substances they may produce a local vasoconstriction in hypoxically aerated lung areas, enabling an intrapulmonary regulation of the V/Q ratio. The present study provides evidence that, in addition to this local effect, NEB could generate centripetal nerve impulses via exocytosis of epithelial DCV at the afferent-like intracorpuscular nerve endings. At the same time they could be modulated by the CNS via their efferent-like intracorpuscular nerve endings.With respect to their innervation, numerous similarities appear to exist morphologically and functionally between the carotid body and the intrapulmonary NEB.     

Ultrastructural observations suggesting merocrine secretion in the initial segment of the mammalian epididymis

Summary Principal cells in the initial segment of the epididymis in horses, cattle, pigs, sheep, dogs, cats, and rabbits have an abundant, partly rough, endoplasmic reticulum and a large Golgi complex. Small vacuoles with opaque content seem to be formed by the Golgi complex and move to the cell apex, where they empty their contents into the lumen by a merocrine mechanism.Financial support for this study was received from The Swedish Council for Forestry and Agricultural Research     

Neuro-epithelial bodies in the respiratory mucosa of various mammals

Summary Lungs of fetal, neonatal and adult rabbits and of various other adult mammals were investigated. Several routine and silver staining methods, Falck's fluorescent amine technic and various electron microscopic techniques were performed.As in the human infant lung (Lauwerynset al., 1972) each of these techniques revealed the occurrence throughout the intrapulmonary airways of intramucosal corpuscles or so-called Neuroepithelial Bodies, composed of more or less parallely oriented, non-ciliated cylindrical cells which reach from the basement membrane to the airway lumen and display a prominent cytoplasmic argyrophilia, a less pronounced argentaffinity and an intense yellow fluorescence. Ultrastructurally, they are granulated, containing mainly two types of dense-cored vesicles of which the first type exhibits a positive reaction for serotonin (technic of Jaim-Etcheverryet al., 1968). They contain intracorpuscular nerve endings which form synaptic end formations upon the granulated cells.Though the functions of these serotonin producing Neuroepithelial Bodies remain furthermore unsettled, they seem to be related also to the recently reported AFG (Argyrophil, Fluorescent and Granulated) cells (Lauwerynset al., 1969, 1970a) and might be involved in various mucosal bronchial and bronchiolar neurosecretory processes; most probably they are chemo-, stretch- and/or tactile neuroreceptor organs modulated by the central nervous system.This study has been supported by a grant from The Council for Tobacco Research—U.S.A. and the Nationaal Fonds voor Wetenschappelijk Onderzoek—Belgium. We thank the Koninklijke Maatschappij voor Dierkunde of the Zoological Garden in Antwerp and Prof. Dr. J. Mortelmans for providing us with some rare animal species as well as Dr. J. Peuskens for interest in our studies. We thank B. Van Rijkel, J. Van Reempts, B. Emmanuel and R. Janssens for technical, G. Pison and St. Ons for photographic and N. Tyberghien for secretarial assistance.     

Adrenergic nerves and 5-hydroxytryptamine-containing cells in the pulmonary vasculature of the aquatic file snake Acrochordus granulatus

Summary The adrenergic innervation of the pulmonary vasculature of the file snake Acrochordus granulatus was examined by use of glyoxylic acid-induced fluorescence. Perivascular plexuses of blue-green fluorescent nerves are observed around the common pulmonary artery, the anterior and posterior pulmonary arteries, the arterioles leading to the gas exchange capillaries of the lung, the venules draining the lung, and the anterior and posterior pulmonary veins. Adrenergic nerves are also associated with the visceral smooth muscle of the lung septa and other tissues. Thus, adrenergic control of pulmonary blood flow may occur either at the common pulmonary artery or more regionally within the lung. Regional control of blood flow in the elongate lung of this snake may be important in matching pulmonary perfusion with the distribution of respiratory gas. Glyoxylic acid-histochemistry and immunohistochemistry revealed that populations of cells located in the common pulmonary artery contain the indoleamine 5-hydroxy-tryptamine. Many of the cells are intimately associated with varicose blue-green fluorescent nerves. It is proposed that the 5-hydroxytryptamine-containing cells may be involved in intravascular chemoreception.     

Hypoxia increases susceptibility of non-small cell lung cancer cells to complement attack

The complement system can be specifically targeted to tumor cells due to molecular changes on their surfaces that are recognized by complement directly or via naturally occurring antibodies. However, tumor cells often overexpress membrane-bound complement inhibitors protecting them from complement attack. We have previously shown that non-small cell lung cancer (NSCLC) cells, additionally to membrane-bound inhibitors, produce substantial amounts of soluble regulators such as factor I (FI) and factor H (FH). Since low oxygen concentration is associated with rapidly growing solid tumors, we studied how NSCLC cells protect themselves from complement attack under hypoxic conditions. Unexpectedly, mRNA levels and secretion of both FI and FH were significantly decreased already after 24 h exposure to hypoxia while cell viability measured by XTT assay and annexin V/7-AAD staining was affected only marginally. Furthermore, we observed decrease of mRNA level and loss of membrane-bound complement inhibitor CD46 and increased deposition of early (C3b) and terminal (C9) complement components on hypoxic NSCLC cells. All three complement pathways (classical, lectin and alternative) were employed to deposit C3b on cell surface. Taken together, our results imply that under hypoxic conditions NSCLC give up some of their available defense mechanisms and become more prone to complement attack.     

促红细胞生成素对神经干细胞缺氧性损伤保护作用的实验研究

目的观察外源性EPO对神经干细胞缺氧性损伤的保护作用,为缺氧缺血性脑损伤的治疗提供新思路。方法从孕11.5d（E11.5d）大鼠获得神经干细胞,经无血清培养基悬浮培养并传代,对所获细胞的自我增殖、自我更新及其多分化潜能进行检测。取传3代神经干细胞中添加不同剂量的EPO,在5%O2培养箱中培养120h。通过计数干细胞克隆形成率和MTT法检测神经干细胞的增殖情况。于含血清分化培养基中加入不同剂量的EPO,用NSE和GFAP免疫细胞化学染色观察神经干细胞的分化情况。采用AnnexinⅤ-FITC/PI染色,激光扫描共聚焦显微镜观察、检测细胞凋亡率。结果加入EPO后神经干细胞的克隆形成率和MTT检测的OD值显著增高,细胞凋亡率显著下降,NSE阳性细胞的比例明显升高,其作用随剂量增加而增大,50U/ml时作用最大。结论EPO对神经干细胞缺氧性损伤具有明显的保护作用,并可促进神经干细胞向神经元方向分化。EPO的这种作用随剂量增加而增大,50U/ml时达高峰。     

Ascorbyl palmitate augments hypoxic respiratory response in the cat

The redox signaling is germane for the hypoxia-sensing mechanisms at the carotid body. This raises the strong possibility that agents possess reducing and antioxidant attributes, such as ascorbate, could influence the hypoxic respiratory response. However, water solubility of ascorbate makes its effectiveness at membrane-associated target sites dubious. In this study, we sought to determine the effect of ascorbyl-6-palmitate (AP), a lipidsoluble derivative of ascorbate which penetrates biomembranes, on hypoxic respiration in the anesthetized, paralyzed and ventilated cat. AP was given by gavage: 600 mg/kg daily for 6 days before the beginning of the acute experiment. Respiration was then assessed from the phrenic electroneurogram, from which peak phrenic amplitude, a surrogate of tidal component, respiratory frequency, and their product, the minute phrenic output, were quantified. The response to normocapnic hypoxia, 7% O₂ in N₂, in the AP-treated cats was compared with that in controls. We found that AP augmented hypoxic respiration, delayed the appearance of hypoxic depression and decreased it, although the stimulatory/depressant character was preserved. The results suggest that the ascorbate moiety of AP interacts with the hypoxiasensing mechanisms. Ascorbate may affect hypoxic respiration at multiple stages of chemotransduction pathways, which are subject to continuing uncertainties. The study highlights the augmentative effect of AP, a redox modulator, on hypoxic respiration, which may have a therapeutic potential.     

Comparative study of gill neuroepithelial cells and their innervation in teleosts and Xenopus tadpoles

Peripheral O₂ chemoreceptors initiate adaptive cardiorespiratory responses to hypoxia in vertebrates. Morphological and physiological evidence suggests that, in fish, neuroepithelial cells (NECs) of the gill perform this role. We conducted a comparative examination in three species of teleosts (zebrafish, goldfish and trout) and larvae of the amphibian Xenopus laevis, using whole-mount gill preparations and confocal immunofluorescence, to elucidate the distribution, morphology and innervation of gill NECs. Nerve fibres were immunolabelled with the neuronal marker zn-12 and were associated with serotonin-immunoreactive NECs in the gills of all species tested. With the exception of trout, innervated NECs were present on all gill arches in the filaments and respiratory lamellae in fish and on homologous structures in Xenopus (i.e. gill “tufts”, including respiratory terminal branches). Thus, the distribution and innervation of NECs of the internal gills of amphibians and teleosts are relatively well conserved, suggesting an important role for gill NECs as O₂ chemoreceptors in aquatic vertebrates. Furthermore, the size and density of gill NECs is variable among teleosts and developmental stages of Xenopus larvae and may be dependent on general gill dimensions or environmental conditions. This report constitutes the first comparative study of gill NECs in fish and amphibians and highlights the significance of gill NECs as an evolutionary model for studying O₂ sensing in vertebrates. We acknowledge the Natural Sciences and Engineering Research Council (NSERC) of Canada for funding through an operating grant to C.A.N., and the NSERC and the Ontario Graduate Scholarship (OGS) program for postgraduate scholarships to M.G.J.     

Neuroepithelial bodies in the lung of the tree frog,Hyla arborea L.

Summary The endocrine-like cells (ELC), which together with nerve endings form the neuroepithelial bodies, are located on primary and secondary septa in the non-ciliated epithelium of the lung of Hyla arborea. ELC protrude markedly toward the lumen of the lung and are surrounded by pneumocytes, which separate ELC from the lumen by thin cytoplasmic processes. ELC possess a light cytoplasm containing two types of granules: (i) numerous small granules, 50–110 nm in diameter, and (ii) single large granules, 290–860 nm in diameter. Numerous nerve fibers, often forming synaptic junctions, can be observed in contact with ELC.This study was supported by a grant No. 476/II from the Polish Academy of Sciences     

Gill chemoreceptors and cardio-respiratory reflexes in the neotropical teleost pacu, <Emphasis Type="Italic">Piaractus mesopotamicus</Emphasis>

This study examined the location and distribution of O₂ chemoreceptors involved in cardio-respiratory responses to hypoxia in the neotropical teleost, the pacu (Piaractus mesopotamicus). Intact fish and fish experiencing progressive gill denervation by selective transection of cranial nerves IX and X were exposed to gradual hypoxia and submitted to intrabuccal and intravenous injections of NaCN while their heart rate, ventilation rate and ventilation amplitude were measured. The chemoreceptors producing reflex bradycardia were confined to, but distributed along all gill arches, and were sensitive to O₂ levels in the water and the blood. Ventilatory responses to all stimuli, though modified, continued following gill denervation, however, indicating the presence of internally and externally oriented receptors along all gill arches and either in the pseudobranch or at extra-branchial sites. Chemoreceptors located on the first pair of gill arches and innervated by the glossopharyngeal nerve appeared to attenuate the cardiac and respiratory responses to hypoxia. The data indicate that the location and distribution of cardio-respiratory O₂ receptors are not identical to those in tambaqui (Colossoma macropomum) despite their similar habitats and close phylogenetic lineage, although the differences between the two species could reduce to nothing more than the presence or absence of the pseudobranch.     

大鼠肺内表皮生长因子(EGF)之分布及缺氧对其含量的影响

本文用免疫体金银法对大白鼠肺内表皮生长因子(Epidermal growth factor,EGF)进行了定位研究,并观察缺氧时肺内 EGF 含量的变化。结果显示正常大鼠肺泡细胞、支气管粘膜及肺内血管壁上均含有 EGF 免疫阳性反应物质;缺氧时,肺内 EGF 含量增加。这提示 EGF 不仅对肺具有一定的生理学影响,而且可能参与缺氧时肺的某些病理学过程。     

Adaptation, specificity and host-parasite coevolution in mites (ACARI)

Parasitism by mites is widespread and involves all the classes of vertebrates, from fishes to mammals. Owing to their small size and their great plasticity, mites are able to adapt to a wide range of habitats. Most of the species are ectoparasites but endoparasitism, especially in the respiratory tract, is common in birds and mammals. The morphological modifications appearing during the process adaptation to parasitic life, especially in Myobiidae, are analysed. Two kinds of characters are particularly important: the constructive specialized characters, consisting of the production of new structures, especially attachment organs allowing the mite to attach to the skin and the hair of the host, and regressive characters. Regression of the external structures is the most important phenomenon appearing in the process of evolution of parasitic mites. The importance of the regression in the parasite is correlated with the degree of evolution of the host. Host and parasite have a parallel evolution, but they go in opposite directions. The author surmises that the regressive evolution is related to the immunological reactions of the host that tend to reject the parasite. To escape from this rejection the parasite tends to select the less antigenic and therefore the most regressed phenotype. Specificity is generally strict in permanent parasites. Coevolution of host and parastie is studied-in the family Myobiidae which parasitizes marsupials, insectivores, bats and rodents. The concordance between the radiations of the mites and that of their hosts is very high.