Adrenocorticotropin-like immunoreactivity in the granules of neural complex cells of the ascidian Halocynthia roretzi

Immunohistochemical studies on the neural complex (neural gland, dorsal strand, and cerebral ganglion) of an ascidian, Halocynthia roretzi, were performed by using an antiserum against porcine ACTH. The antiserum recognized a considerable number of the cells scattered along the tubular structure of the dorsal strand and a few cells in the cerebral ganglion. Immunoelectron microscopic studies revealed that the ACTH-like substance resided within secretory granules with diameter of 300-500 nm. Furthermore, those ACTH-immunoreactive cells were demonstrated to be different from PRL-immunoreactive cells, the presence of which had previously been reported.     

An ultrastructural investigation of the hypocerebral organ of the adult Euperipatoides kanangrensis (Onychophora, Peripatopsidae)

The hypocerebral organs of Euperipatoides kanangrensis are a pair of spherical vesicles located ventral to the cerebral ganglia. They develop in the embryo from the most anterior pair of ventral organs, in the antennal segment. The wall of each hypocerebral organ is a dense epithelium of elongate cells with peripheral nuclei. The cytoplasm of the cells includes numerous mitochondria, Golgi bodies and microtubules. The small lumen, located eccentrically within the organ, contains concentrically layered electron-dense material resembling cuticle.Each hypocerebral organ is enclosed by a layer of extracellular matrix continuous with that surrounding the adjacent cerebral ganglion. There are no nerve connections between ganglion and organ, but cellular connections traverse the intervening matrix and could serve as a communication pathway. The ultrastructure of the hypocerebral organs indicates that they are glands.     

A new species of Lepidodasys (Gastrotricha, Macrodasyida) from Panama with a description of its peptidergic nervous system using CLSM, anti-FMRFamide and anti-SCPB

A new species of Lepidodasys (Gastrotricha: Macrodasyida: Lepidodasyidae) is described from sublittoral sediments in the Bocas del Toro archipelago in Panama and represents the first species of Lepidodasyidae described from the Caribbean. The new species possesses keeled scales that form a crossed-helical pattern across its dorsal and lateral surfaces and ventral scales that form a herringbone pattern between and lateral to the ciliary columns. A bilateral pair of three ventral adhesive tubes at the posterior end further differentiates this new species from its seven congeners. A confocal laser scanning microscope examination of the nervous system using antibodies to small cardioactive peptide B (SCP_B) and FMRFamides reveals a dumbbell-shape cerebral ganglion, paired pharyngeal neurites and paired posterior nerve cords. Expression patterns of immunoreactivity to both classes of neuropeptides show a high degree of similarity. Only within the lateral somata of the cerebral ganglion and a single median pharyngeal neurite is there a difference in immunoreactivity to FMRFamide (positive) compared to SCP_B (negative). Results from this investigation reveal that neuropeptides, among other neuronal markers, might provide phylogenetically informative characters in macrodasyidan gastrotrichs, especially regarding the topology of the cerebral ganglion.     

两种软体动物神经系统一氧化氮合酶的组织化学定位

运用一氧化氮合酶(NOS)组织化学方法研究了软体动物门双壳纲种类中国蛤蜊和腹足纲种类嫁Qi神经系统中NOS阳性细胞以及阳性纤维的分布。结果表明：在蛤蜊脑神经节腹内侧,每侧约有10-15个细胞呈强NOS阳性反应,其突起也呈强阳性反应,并经脑足神经节进入足神经节的中央纤维网中;足神经节内只有2个细胞呈弱阳性反应,其突起较短,进入足神经节中央纤维网中,但足神经节中,来自脑神经节阳性细胞和外周神经系统的纤维大多呈NOS阳性反应;脏神经节的前内侧部和后外侧部各有一个阳性细胞团,其突起分别进入后闭壳肌水管后外套膜神经和脑脏神经索。脏神经节背侧小细胞层以及联系两侧小细胞层的纤维也呈NOS阳性反应。嫁Qi中枢神经系统各神经节中没有发现NOS阳性胞体存在;脑神经节、足神经节、侧神经节以及脑—侧、脑—足、侧—脏连索中均有反应程度不同的NOS阳性纤维,这些纤维均源于外周神经。与已研究的软体动物比较,嫁Qi和前鳃亚纲其它种类一样,神经系统中NO作为信息分子可能主要存在于感觉神经。而中国蛤蜊的神经系统中一氧化氮作为信息分子则可能参与更广泛的神经调节过程。     

An ultrastructural classification of the neuronal cell bodies of the rat dorsal root ganglion using zinc iodide-osmium impregnation

Summary Zinc iodide-osmium (ZIO) impregnation of rat dorsal root ganglia differentially stained various elements in the neuronal cells, particularly their Golgi bodies. On the basis of this differential ZIO staining dorsal root ganglion neurones have been classified into seven types. The ultrastructure of these is described and the numbers of each type in the L4 dorsal root ganglion have been determined. Prolonged nerve stimulation did not change the relative numbers of the different cell types suggesting that none of the differences between cell types represents differences in their state of activity. The possibility is discussed that differences in morphology may reflect differences in neurotransmitter function.I.R.D. is supported by the Medical Research Council; P.K. thanks the Mental Health Trust for a project grant     

Cellular and ultrastructural features of the adult and the embryonic eye in the marine gastropod,Ilyanassa obsoleta

Light and electron microscopic techniques show that the eye of the marine prosobranch gastropod, Ilyanassa obsoleta, is composed of an optic cavity, lens, cornea, retina, and neuropile, and is surrounded by a connective tissue capsule. The adult retina is a columnar epithelium containing three morphologically distinct cell types: photoreceptor, pigmented, and ciliated cells. The retina is continuous anteriorly with a cuboidal corneal epithelium. The neuropile, located immediately behind the retina, is composed of photoreceptor cell axons, accessory neurons, and their neurites. The embryonic eye is formed from surface ectoderm, which sinks inward as a pigmented cellular mass. At this time, the eye primordium already contains presumptive photoreceptor cells, pigmented retinal cells, and corneal cells. Several days later, just before hatching, the embryonic eye remains in intimate contact with the cerebral ganglion. It has no ciliated retinal cells, neuropile, optic nerve, or connective tissue capsule and its photoreceptor cells lack the electron-lucent vesicles and multivesicular bodies of adult photoreceptor cells. As the eye and the cerebral ganglion grow apart, the optic nerve, neuropile, and connective tissue capsule develop.     

Ganglion ultrastructure in phylactolaemate Bryozoa: evidence for a neuroepithelium

In contrast to other Bryozoa, members of the subtaxon Phylactolaemata bear a subepithelial cerebral ganglion that resembles a hollow vesicle rather than being compact. In older studies this ganglion was said to originate by an invagination of the pharyngeal epithelium. Unfortunately, documentation for this is fragmentary. In chordates the central nervous system also arises by an invagination-like process, but this mode is uncommon among invertebrate phyla. As a first attempt to gather more data about this phenomenon, cerebral ganglia in two phylactolaemate species, Fredericella sultana and Plumatella emarginata, were examined at the ultrastructural level. In both species the ganglion bears a small central lumen. The ganglionic cells are organized in the form of a neuroepithelium. They are polarized and interconnected by adherens junctions on their apical sides and reside on a basal lamina. The nerve cell somata are directed towards the central lumen, whereas the majority of nervous processes are distributed basally. Orientation of the neuroepithelial cells can be best explained by the possibility that they develop by invagination. A comparison with potential outgroups reveals that a neuroepithelial ganglion is at least derived. Since, however, a reliable phylogenetic system of the Bryozoa is missing, a decision on whether such a ganglion is apomorphic for Bryozoa or evolved within this taxon can hardly be made.     

Novel, posterior sensory organ in the trochophore larva of Phyllodoce maculata (Polychaeta)

A new posterior sensory organ (PSO), located at the dorsal midline of the hyposphere, is described by immunocytochemical detection of acetylated alpha tubulin and serotonin (5-HT) in a laser-scanning microscope, as well as three-dimensional reconstructions after optical serial sectioning in the trochophore larva of the polychaete Phyllodoce maculata (Phyllodocidae). The unpaired PSO consists of five bipolar sensory cells, two of them being 5-HT immunopositive, which send axons to the cerebral ganglion and prototroch nerve. The dendrites of these cells project to the surface and bear one cilium each. A single neuronal fibre from the apical sensory organ innervates the PSO.     

Trigeminal Projections to Contralateral Dorsal Horn Originate in Midline Hairy Skin

The present study tested the hypothesis that the trigeminal (V) primary afferent projection to the contralateral dorsal horn originates in midline hairy skin. A prior study (Jacquin et al., 1990) showed that this crossed projection is heaviest to ophthalmic regions of medullary and cervical dorsal horns, and that it does not arise from V ganglion cells that innervate cornea, nasal mucosa, or cerebral dura mater. Here, retrograde double-labeling methods were used to show that many ophthalmic ganglion cells that innervate midline hairy skin via the supraorbital nerve project to the contralateral medullary and upper cervical dorsal horns. Diamidino yellow injections into the right dorsal horn labeled an average of 104 cells in the left V ganglion. Of these contralaterally projecting ganglion cells, an average of 45% were also labeled by horseradish peroxidase (HRP) injections into the left supraorbital nerve, and 25% were also labeled by HRP injections into the midline opthalmic hairy skin. However, only 2% were labeled by HRP injections restricted to left supraorbital vibrissae follicle nerves. Almost all of the double-labeled cells were located in the dorsal one-half of the V ganglion, and they did not differ in size from single-labeled cells.

On the basis of these and prior data, we conclude that a high percentage of contralaterally projecting V ganglion cells originate in midline hairy skin. It is also likely that the contralaterally projecting V ganglion cells serve a low-threshold mechanoreceptive function, given the relatively large ganglion cells and axons giving rise to this pathway and their central terminations in dorsal horn laminae III-V.     

Early development of the hypoglossal nerve in the chick embryo as observed by the whole-mount nerve staining method

The developmental morphology of the hypoglossal nerve and associated structures were studied in the chick embryo (Hamburger and Hamilton stages 16-27) stained by the immunohistochemical technique. Ventral rootlets of the occipital nerves, including O1, were seen at stage 16. The distal ends of these nerves anastomosed to form the hypoglossal nerve at stage 20. At stage 23, four occipital and the first three cervical nerves were observed to be involved. The transient contribution of C3 at this stage seemed to be correlated with the formation of the longitudinal anastomosis of the distal end of the spinal nerves which begins around stage 23. The anterior hypoglossal roots appeared between O1 and the abducens nerve at stage 20. These rootlets were observed to arise as the rostral continuation of the occipital sequence and were found to be arranged in a straight line from O1 to the abducens nerve. The recurrent branch of the abducens was also observed. The posterior end of the ganglion crest produced dorsal root ganglion (DRG)-like structures transiently at the level of C2, and sometimes at the level of C1 also. The ganglion crest developed descending processes in the occipital region seemingly related to the spinal dorsal root formation. These phenomena seemed to represent the potential of the ganglion crest to produce the spinal nerve components which are depressed in the occipital region.     

Trigeminal projections to contralateral dorsal horn originate in midline hairy skin

The present study tested the hypothesis that the trigeminal (V) primary afferent projection to the contralateral dorsal horn originates in midline hairy skin. A prior study (Jacquin et al., 1990) showed that this crossed projection is heaviest to ophthalmic regions of medullary and cervical dorsal horns, and that it does not arise from V ganglion cells that innervate cornea, nasal mucosa, or cerebral dura mater. Here, retrograde double-labeling methods were used to show that many ophthalmic ganglion cells that innervate midline hairy skin via the supraorbital nerve project to the contralateral medullary and upper cervical dorsal horns. Diamidino yellow injections into the right dorsal horn labeled an average of 104 cells in the left V ganglion. Of these contralaterally projecting ganglion cells, an average of 45% were also labeled by horseradish peroxidase (HRP) injections into the left supraorbital nerve, and 25% were also labeled by HRP injections into the midline opthalmic hairy skin. However, only 2% were labeled by HRP injections restricted to left supraorbital vibrissae follicle nerves. Almost all of the double-labeled cells were located in the dorsal one-half of the V ganglion, and they did not differ in size from single-labeled cells. On the basis of these and prior data, we conclude that a high percentage of contralaterally projecting V ganglion cells originate in midline hairy skin. It is also likely that the contralaterally projecting V ganglion cells serve a low-threshold mechanoreceptive function, given the relatively large ganglion cells and axons giving rise to this pathway and their central terminations in dorsal horn laminae III-V.     

K252a Modulates the Expression of Nerve Growth Factor-Dependent Capsaicin Sensitivity and Substance P Levels in Cultured Adult Rat Dorsal Root Ganglion Neurones

Abstract: K252a, an inhibitor of trk phosphorylation and nerve growth factor signal transduction in PC12 cells, blocked nerve growth factor-induced responses in cultured adult rat dorsal root ganglion sensory neurones. The nerve growth factor-dependent appearance of capsaicin sensitivity and accumulation of the neuropeptide substance P were inhibited when dorsal root ganglion neurones were grown in the presence of low concentrations (100 n M ) of K252a. At higher concentrations (3 µ M ), however, K252a stimulated the development of capsaicin sensitivity and the accumulation of substance P even in the absence of nerve growth factor. By using a wide dose range, therefore, we showed that K252a could either inhibit or mimic nerve growth factor's actions on sensory neurones. These results may explain the apparent paradox in the literature that some groups show a blocking effect of K252a on nerve growth factor-dependent survival of dorsal root ganglion sensory neurones, whereas others report that K252a can substitute for nerve growth factor or other trophic factors and promote neuronal survival.     

Scanning electron microscope observations on the muscle innervation of Oikopleura dioica fol (appendicularia,tunicata) with notes on the arrangement of connective tissue fibres

Critical point dried and fractured appendicularia of the species Oikopleura dioica have been examined in the scanning electron microscope. The dorsal nerve cord with ganglion cells and peripheral nerve fibres could easily be observed. Thick peripheral nerve fibres leave the nerve cord as bilateral pairs at constant intervals along the tail. Most of these fibres branch from the naked nerve cord, but some evidently originate in ganglion perikarya bulging out from the nerve cord itself. These paired peripheral nerves always have elaborate end-arborizations on the medial surface of the lateral muscle cells. They are accordingly interpreted as motor axons. Some thinner peripheral nerve fibres originate at irregular intervals from both the nerve cord and the ganglion cells. Due to the numerous extracellular fibrils that connect the bilateral layers of the epidermal fins and the muscle cells to each other, these thin nerve fibres can seldom be traced to their termination. A few ones can, however, be traced ventrally between the notochord and the muscle cells and seem to end in singular bulb-like expansions. Clusters of synaptic vesicles are present in transmission electron micrographs of such nerves, and they are accordingly believed to carry efferent impulses. The extracellular fibrils are arranged in a highly ordered pattern with thick bundles crossing the gap between the structures to be interconnected and with numerous radiating insertions on the surface of the tissues.     

An electron microscopic study of the endocrine dorsal bodies in reproductively active and inactive Siphonaria pectinata (Pulmonata: Mollusca)

The fine structure of the dorsal bodies of the pulmonate limpet Siphonaria pectinata is described in the context of female reproduction involving egg production. In reproductively-active (egg-laying) animals, the ciliated dorsal body cells are filled with lipid droplets and mitochondria. Gap junctions are commonly seen between the cells. The Golgi complexes and the smooth endoplasmic reticulum constitute the other prominent cell organelles. In reproductively-inactive (non-egg-laying) animals, there is a significant reduction in the number of lipid droplets and evidence of reduced synthetic activity in the dorsal bodies. About 12 dorsal body cells are present immediately underneath the perineurium of each cerebral ganglion of the central nervous system. These internal cells are structurally similar to those outside the central nervous system. Cell processes of some of these cells exit the central nervous system at a minimum of three locations on each side and they come in close proximity to the dorsal body cells outside the cerebral ganglia. Like the external cells, the internal cells also communicate via gap junctions and exhibit structural differences according to whether or not the animals are reproductively active. The dorsal body cells, inside and outside the central nervous system, appear to be innervated by neurosecretory axons suggesting neuronal control of dorsal body activity.     

河北环毛蚓神经系统 一氧化氮合酶的组织化学定位

用依赖还原型辅酶Ⅱ的黄酶组织化学方法,研究了环节动物门寡毛纲种类河北环毛蚓（Pheretima tschiliensis)神经系统k 一氧化氮合酶(NOS)阳性细胞及阳性纤维的分布,结果表明,河北环毛蚓神经系统中脑神经节背侧有大量细胞呈现NO强阳性反应,胞体和突起染色明显。咽下神经中偶尔能见少数染色较浅的神经元。在脑神经节腹内侧、围咽神经、 咽下神经节外侧部及腹神经链中都有一氧化氮合酶阳性纤维存在脸染色很深,实验结果表明,在环节动物中作为信息分子的一氧化氮已广泛存在于神经系统中。     

Metabolism of histamine in the CNS of Aplysia californica: cellular distribution of gamma-glutamylhistamine synthetase

1. The cellular distribution of the histamine-metabolizing enzyme, gamma-glutamylhistamine synthetase, was studied in the CNS of Aplysia californica. 2. Enzyme activity was assayed in single, re-identifiable neuronal cell bodies, clusters of nerve cells and neuropil and capsule tissue surrounding the ganglia. 3. The "histaminergic" C-2 cells and all other single nerve cell bodies contained measurable gamma-glutamylhistamine synthetase activity. 4. The cerebral E cluster, which houses the C-2 cells and several of its post-synaptic neurons, had an apparently higher specific enzyme activity than other neuronal clusters. 5. The finding of measurable enzyme activity in the histamine-rich C-2 cell bodies and in clusters of cells responsive to this imidazoleamine supports the hypothesis that one function for gamma-glutamylhistamine synthetase is in the disposal of neuronally released histamine. 6. The average specific activity of gamma-glutamylhistamine synthetase in single cells was 3.64 +/- 0.32 mumol g protein hr. This represented only 5% of the enzyme activity measured in the whole ganglion. 7. The bulk of synthetase activity was found to reside in the capsule tissue.     

Engraftment and differentiation of embryonic stem cell-derived neural progenitor cells in the cochlear nerve trunk: growth of processes into the organ of Corti

Hearing loss in mammals is irreversible because cochlear neurons and hair cells do not regenerate. To determine whether we could replace neurons lost to primary neuronal degeneration, we injected EYFP-expressing embryonic stem cell-derived mouse neural progenitor cells into the cochlear nerve trunk in immunosuppressed animals 1 week after destroying the cochlear nerve (spiral ganglion) cells while leaving hair cells intact by ouabain application to the round window at the base of the cochlea in gerbils. At 3 days post transplantation, small grafts were seen that expressed endogenous EYFP and could be immunolabeled for neuron-specific markers. Twelve days after transplantation, the grafts had neurons that extended processes from the nerve core toward the denervated organ of Corti. By 64-98 days, the grafts had sent out abundant processes that occupied a significant portion of the space formerly occupied by the cochlear nerve. The neurites grew in fasciculating bundles projecting through Rosenthal's canal, the former site of spiral ganglion cells, into the osseous spiral lamina and ultimately into the organ of Corti, where they contacted hair cells. Neuronal counts showed a significant increase in neuronal processes near the sensory epithelium, compared to animals that were denervated without subsequent stem cell transplantation. The regeneration of these neurons shows that neurons differentiated from stem cells have the capacity to grow to a specific target in an animal model of neuronal degeneration.     

The distribution of the neurofilament triplet proteins within individual neurones

The three proteins of the mammalian neurofilament ‘triplet’ were purified from rat sciatic nerve as individual polypeptides. Antibodies were raised in rabbits and in guinea pigs. When tested by the very sensitive immune-blotting technique some of the antibodies proved to be completely specific for the peptide to which they had been raised. Others, however, exhibited weak cross-reactivity with other proteins of the triplet. Cross-reacting IgGs could be removed by appropriate antigen affinity chromatography. Thus a series of rabbit and guinea pig antibodies specific for each of the triplet proteins was obtained. The antibodies were used in immunofluorescence microscopy on cultured rat dorsal root ganglion cells. Only cells with a neuronal morphology were stained by these antibodies, some very strongly and some extremely weakly. When double immunofluorescence was performed it was found that cells stained in an equivalent manner with any combination of antibodies. Neurones which stained strongly with any one antibody could be stained strongly with any other and the converse was true for weakly staining cells. When fine profiles in the growth cones of positive cells were examined it was found that these profiles, representing single or small numbers of neurofilaments, were stained in an identical manner in double immunofluorescence. The results show that the distribution of the three proteins is identical at the level of resolution of the light microscope in rat dorsal root ganglion neurones in tissue culture, and lend support to the supposition that all three triplet polypeptides are contained within each individual neurofilament.     

Expression of mRNAs for PPT, CGRP, NF-200, and MAP-2 in cocultures of dissociated DRG neurons and skeletal muscle cells in administration of NGF or NT-3

Both neurotrophins (NTs) and target skeletal muscle (SKM) cells are essential for the maintenance of the function of neurons and nerve-muscle communication. However, much less is known about the association of target SKM cells with distinct NTs on the expression of mRNAs for preprotachykinin (PPT), calcitonin-gene related peptide (CGRP), neurofilament 200 (NF-200), and microtubule associated protein 2 (MAP-2) in dorsal root ganglion (DRG) sensory neurons. In the present study, a neuromuscular coculture model of dissociated dorsal root ganglion (DRG) neurons and SKM cells was established. The morphology of DRG neurons and SKM cells in coculture was observed with an inverted phase contrast microscope. The effects of nerve growth factor (NGF) or neurotrophin-3 (NT-3) on the expression of mRNAs for PPT, CGRP, NF-200, and MAP-2 was analyzed by real time-PCR assay. The morphology of DRG neuronal cell bodies and SKM cells in neuromuscular coculture at different conditions was similar. The neurons presented evidence of dense neurite outgrowth in the presence of distinct NTs in neuromuscular cocultures. NGF and NT-3 increased mRNA levels of PPT, CGRP, and NF-200, but not MAP-2, in neuromuscular cocultures. These results offer new clues towards a better understanding of the association of target SKM cells with distinct NTs on the expression of mRNAs for PPT, CGRP, NF-200 and MAP-2, and implicate the association of target SKM cells and NTs with DRG sensory neuronal phenotypes.     

Gene Delivery into Neuronal Cells by Calcium Phosphate-Mediated Transfection

This paper describes a method for introducing DNA constructs into primary adult dorsal root ganglion (DRG) neuron cultures. The method uses a modified calcium phosphate technique and enables relatively small numbers of cells to be used. We have used this method to study the promoter sequences responsible for mediating gene activity and nerve growth factor responsiveness in DRG neurons. It can also be used, however, for other purposes such as testing the effect on neuronal function of overexpressing a specific gene product.