Unscheduled DNA synthesis in rat adult myenteric neurons: an immunohistochemical study.

Unscheduled DNA synthesis refers to DNA synthesis not followed by cell division. Previous studies have suggested that this phenomenon may occur in neurons from peripheral myenteric ganglia in conditions of functional hyperstimulation. In order to verify these observations, we have carried on an immunohistochemical study on myenteric neurons from the hypertrophic intestinal loops upstream from a partial obstruction (an experimental condition that induces a relevant increase of the neuronal workload) after labelling with two different markers: the proliferating cell nuclear antigen (PCNA), that is specifically expressed in cell nuclei during the S-phase, and the protein gene product 9.5 (PGP 9.5), a specific marker of nerve cells. While no myenteric neuron immunopositive for the anti-PCNA antibody was found in the control intestine, in the hypertrophic myenteric ganglia some neurons were positive for PCNA. These results provide an unequivocal evidence on the existence of unscheduled DNA synthesis in myenteric neurons from the hypertrophic intestine.

Effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on ultrastructure of nigral neuromelanin in Macaca fascicularis.

In neurons of the substantia nigra (SN) of Macaca fascicularis the administration of parkinsongenic doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) caused morphological changes of the neuromelanic granules. Under light microscopy, the granules appeared more dispersed and larger. Electron microscopy revealed coalescence of granules in large masses with loss of the electrodense component. Phagocytosis of neuromelanin by glial cells was also observed. In several neurons the neuromelanic changes were evident in the presence of morphologically intact mitochondria. These data suggest an interaction between MPTP and neuromelanin that may have relevance to the nigrotropic toxicity of MPTP and are in agreement with observations on neuromelanin in parkinsonian patients.

Relations between subcellular events in the degenerating notochord and histopathological features of the spinal chordomas.

The authors focused their attention on the cytology of the degenerating notochord at the level of the vertebral anlagen and compared it with data of the literature on the histopathology of the spinal chordomas. The purpose of the research was to investigate the notochordal histological features justifying the fact that the remnants of a structure destined to atrophy during prenatal life, maintain a proliferative potential in postnatal life. Therefore, from the earliest stages (24th incubation hour) to the terms of development, the notochord was studied by electron microscope in chick embryo. The data obtained show that, in most notochordal cells, signs of metabolic damage and progressive degeneration coexist with signs of secretory and mitotic activity, from which the proliferative potential of these cells seems to derive. Vacuolar degeneration is also likely due to the progressive metabolic segregation of the notochord, owing to the absence of blood-vessels and the appearance of perichordal sheath.

Morphometrical analysis of types and sub-types of neurons in dorsal root ganglia of the lizard Podarcis sicula.

In the present study, we conducted a morphometrical analysis of the different types and sub-types of lizard DRG neurons at various spinal levels. This analysis demonstrated significant differences in size distribution among the various neuron types and sub-types, as well as a significant shift to greater values in neurons from the dorsal root ganglia at the cervical and the lumbar spinal levels. The results are critically evaluated in relation to methodological issues, and the implications of these findings are discussed.

Notochord degeneration in chick embryo: a particular case of developmental cell death?

To define the type of cell death occurring in notochordal tissue, the cytological features of degenerating notochord were studied by transmission electron-microscope in thirty chick embryos from the 20th hour to the 15th incubation day. During the first two days the notochordal cells show nuclei with large nucleoli and cytoplasm with yolk granules, lipid droplets, phagolysosomes and deposits of glycogen. From the 3rd to the 5th incubation day, besides the peculiar vacuolization, disaggregation of the endoplasmic reticulum, transformation of the mitochondrial morphology, and disintegration of the cell membrane are detectable. Nuclei are normal up to advanced stages of cytoplasmic degeneration. On the 6th day a large number of cells are dying and, later on, the tissue disintegrates at the level of the vertebral bodies. Cell death in the notochord does not seem to be classifiable as one of the types of developmental cell death described in literature: the comparison with similar cytological features referred by pathologists as a consequence of metabolic damage, suggests that the degeneration of the notochord may be related to its morphological isolation and thus to trophic deprivation.

Types and sub-types of neurons in dorsal root ganglia of the lizard Podarcis sicula: a light and electron microscope study.

A morphological analysis of types and sub-types of neurons from dorsal root ganglia at different spinal levels was carried out by combined light and electron microscopy in Podarcis sicula. Two neuron types were recognized: small dark cells (type D) and large light cells (type L). Type L cells were further sub-divided into three sub-types (L1, L2, L3) on the basis of entity and distribution of neurofilaments. Percentage distribution of neuron types did not vary in relation to the spinal level. On the contrary, differences were found in the percentage of the three type L neuron sub-types; a higher percentage of cells very rich in neurofilaments (L2, L3) was found in dorsal root ganglia from the cervical and the lumbar spinal levels. Results are discussed in relation to the spinal-level-related extent of innervation territory of dorsal root ganglia.

First appearance and distribution of calretinin-immunoreactive neurons in the early development of the chick central nervous system.

Silver impregnation studies in chick embryos have shown that, by the 2nd day of incubation, the earliest neurofibrillar differentiation occurred in neuroblasts located at the diencephalic-mesencephalic junction and in the rhombencephalon; some of these neuroblasts were believed to become reticular neurons. Since calretinin, a cytosolic calcium-binding protein of the "E-F hand" family, occurs in reticular neurons, the present study investigated immunohistochemically whether the early differentiating reticular neurons are also the first neurons to express this marker during chick embryo development. The first calretinin-immunoreactive neuroblasts appeared at stage 11 (40-45 h of incubation according to the series of Hamburger and Hamilton), and were located in the basal plate of the diencephalic-mesencephalic junction and of rhombomeres adjacent to the otic placode and in the alar plate and intermediate zone of the cervical spinal cord. In bromodeoxyuridine-injected embryos, these earliest calretinin-immunoreactive neurons were shown to express the calcium-binding protein 11-16 h after their last mitosis. By stage 11 up to the 14th day of incubation (stage 40), the calretinin-immunostained neurons increased in number and ultimately formed a chemically defined subset of neurons belonging to the tegmental reticular formation and raphe region of the brainstem. In the meantime, early calretinin-immunostained nerve processes were shown to form two conspicuous longitudinal bundles which run in the ventral and lateral margins of the brainstem and spinal cord.

[Peptidergic innervation of the human dental pulp: neuropeptide Y]. / Innervazione peptidergica della polpa dentaria umana: il neuropeptide Y.

The localization and distribution of the Neuropeptide Y (NPY) has been studied with immunofluorescent methods in the human dental pulp. Immunofluorescence for the NPY has been observed in nervous fibers running in medium and big nerves associated in vascular structures, and in single fibers scattered in the pulpar connective or organized in the subdontoblastic plexus.

Cytodifferentiation of the notochord in early chick embryo related to the development of the axial and paraxial structures.

The cytology of the notochord was investigated by means of L.M. and T.E.M. in fifty chick embryos from the 7th HH stage to hatching, at different levels of the longitudinal axis and at different levels in each somite. The cytodifferentiation of the notochord was compared with the one of the surrounding axial and paraxial structures (neuro-epithelium and somites). Close segmental relations among notochord, neural groove and sclerotome, i.e. reciprocal contacts of cell processes and filopodia, where observed in 7-9 HH stages embryos. During the following stages in the notochordal cells signs of secretory activity are detectable, but at the same time degenerative changes are evident and they will increase up to the total regression of the notochord. These morphological observations seem to suggest the presence of reciprocal morphogenetic influences among notochord, neuro-epithelium and sclerotome, only before the 9th stage, rather than a determinant role of the notochord in the surrounding structures at later developmental stages.

Notochordal histogenesis: preliminary report of ultrastructural observations in chick embryo.

Transverse, frontal and sagittal notochord sections of chick embryos at different developmental stages were ultrastructurally examined. The notochord tissue organization appears to be more likely to build a stiff but flexible body axis than to have an inducting role on the surrounding embryonic structures.

Chondrogenic gradients in the developing vertebral body.

Since in literature the question of the spatio-temporal sequence of the cartilage maturation in the developing vertebra is still controversial, the authors studied by light and electron microscope, the chondrification of the vertebral body in chick embryo from the 6th to the 13th incubation days, in order to define the correlations between morphology and distribution of the cartilage cells in this phase of vertebral development. The results show that the chondrogenesis follows spatio-temporal gradients, starting at about the 8th incubation day from a zone located between notochord and neural tube, slightly cranially to the midvertebral level. From this starting point the chondrification proceeds with dorso-lateral and radial progression, and at the same time extends towards the cranial and caudal plates of the developing vertebra. These data are compared to the findings obtained by other authors on the ultrastructural and biochemical aspects of the vertebral development.

Ontogeny of calbindin-D28K and calretinin in developing chick kidney.

The ontogeny of two calcium-binding proteins (calbindin-D28k and calretinin) was studied by immunohistochemical techniques in developing chick kidney. This study showed the presence of calbindin on the 5th incubation day and calretinin on the 7th incubation day in mesonephric distal and connecting tubules, and in the medial wall of the Wolffian duct. At later stages, immunostaining for these two proteins, in particular for calretinin, was also demonstrated in some metanephric proximal tubules. Glomeruli and Bowman's capsules were negative both in the mesonephros and metanephros. The presence of calretinin in the developing kidney has thus been demonstrated for the first time. The early expression of calbindin and calretinin in mesonephric distal tubules suggests their role in regulating the final excretion of calcium. The different patterns of immunoreactivity of the walls of the Wolffian duct can be correlated with their different histogenetic and histological features.

Distribution of substance P immunoreactive cell bodies and fibers in cranial sensory and autonomic ganglia of the chick.

Substance P-immunoreactive neurons were demonstrated in chick embryonic and adult trigeminal ganglion and jugular-superior ganglionic complex using FITC-immunohistochemical methods. Both small-size and large ganglion cells exhibited SP immunoreactivity, without apparent changes during embryonic and post-hatching development. SP-positive fibers could be detected in a good number in the sympathetic cranial cervical ganglion, either during embryonic development or in adult chick. No immunoreactive perikarya were observed in this ganglion. In the ciliary ganglion, both choroidal and ciliary neurons were SP-negative, whereas SP immunoreactive fibers surrounded the perikarya of both cell populations.

Distribution and development of substance P immunoreactive axons in the chick cornea and uvea.

A study was made of the distribution of Substance P-immunoreactive fibers in chick cornea and uvea in whole mount preparation, using the indirect immunofluorescence technique. The development of these fibers during embryogenesis was also investigated. SP-fibers were present in all chick eye structures, in the various prenatal and postnatal stages examined. Their distribution was comparable with that observed by other workers in mammals. Transformation of the iris musculature from smooth to striated, during development, is not accompanied by significant changes in SP-ergic innervation.

Differentiation of cardiac conducting cells from the neural crest.

Some of the cells that migrate to the dorsal myocardium of the chick embryo in stage 21 H-H begin to synthesize desmin. They retain this property even when they reach the subendothelial layers of the heart bud, i.e. the characteristic site of the cardiac conducting cells.

Chick heart peptidergic innervation: localization and development.

Localization and development of chick heart peptidergic innervation (Substance P, VIP and Somatostatin) were investigated by means of immunofluorescence technique. The peptidergic component of the heart innervation was observed, for the first time, in older than 11 day chick embryos, i.e., subsequently to the appearance of the cholinergic component. The peptidergic structures achieve nearly full development in about 16-17 day embryos. Substance P is the most represented of the three peptides. It is localized both in nerve bundle fibers and in isolated fibers within the myocardium, the pericardium, the vessel walls; it is also present in fibers of some heart base ganglia. VIP is mostly contained in some thick single fibers travelling along the vessel walls of the heart base, the myocardium and the pericardium. Some VIP immunoreactive cells were also observed in the base ganglia. Somatostatin is mostly contained in some ganglia cells, whilst thin Somatostatin-immunoreactive fibers form a rich plexus among the atrial and ventricular myofibers, without contacting the vessel walls.

Origin of the iris sphincter muscle in chick embryo.

The origin of the iridial sphincter muscle in chick embryo was investigated by means of immunohistochemistry. Desmin immunoreactive cells are shown in the mesenchymal stroma overlying the anterior epithelial layer of the iris in 4 1/2-day chick embryos. In 9-11-day chick embryos also some cells of the posterior epithelium near the pupillary margin, and of the iridial lamella show a slighter desmin-immunoreactivity. This finding agrees with a double origin of the iridial sphincter muscle: an early mesenchymal one and a later epithelial other.