Development of the rat sublingual gland: A light and electron microscopic immunocytochemical study

Cell differentiation in the rat sublingual gland occurs rapidly and is largely complete by birth. To study differentiation of the serous and mucous cells of the sublingual gland, we used antibodies to the secretory proteins CSP‐1, SMGB, PSP, and SMGD, and sublingual mucin as specific cell markers. Glands from rats at ages 18, 19, and 20 days in utero, and postnatal days 0, 1, 5, 9, 14, 18, 25, 40, and 60 were fixed and prepared for morphological analysis and immunocytochemical labeling. At age 18 days in utero, a few cells in the developing terminal bulbs contained mucous‐like apical granules that labeled with anti‐mucin. Other cells had mixed granules with a peripheral lucent region and a dense core of variable size that occasionally labeled with anti‐SMGD. Additionally, presumptive serous cells with small dense granules that contained CSP‐1 and SMGB were present. At age 19 days in utero, the dense granules of these cells also labeled with anti‐SMGD. By age 20 days in utero, mucous cells were filled with large, pale granules that labeled with anti‐mucin, and serous cells had numerous dense granules containing CSP‐1, SMGB, PSP, and SMGD. Fewer cells with mixed granules were seen, but dense regions present in some mucous granules (MGs) labeled with anti‐SMGD. After birth, fewer MGs had dense regions, and serous cells were organized into well‐formed demilunes. Except for PSP, which was undetectable after the fifth postnatal day, the pattern of immunoreactivity observed in glands of neonatal and adult animals was similar to that seen by age 20 days in utero. These results suggest that mucous and serous cells have separate developmental origins, mucous cells differentiate earlier than serous cells, and cells with mixed granules may become mucous cells. Anat Rec 266:30–42, 2002. © 2002 Wiley‐Liss, Inc.     

A GABA immunocytochemical study of rat motor thalamus: light and electron microscopic observations.

A light and electron microscopic study of GABA-immunoreactive neurons and profiles in the ventroanterior-ventrolateral and ventromedial nuclei of rat dorsal thalamus was conducted using antiserum raised against GABA. Less than 1% of the neurons in these motor-related nuclei exhibited GABA immunoreactivity, confirming previous reports that these nuclei are largely devoid of interneurons. Immunoreactive neurons in the ventral anterior-ventral lateral complex and ventromedial nucleus were bipolar or multipolar in shape, and tended to be smaller than non-immunoreactive neurons. GABA immunoreactivity in the neuropil consisted of labeled axon terminals and myelinated and unmyelinated axons, and was lower in the ventral anterior-ventral lateral complex and ventromedial nucleus than in neighboring thalamic nuclei. The density of neuropil immunolabeling was slightly higher in ventral anterior-ventral lateral complex than in ventromedial nucleus. GABA-immunoreactive axon terminals, collectively termed MP boutons for their medium size and pleomorphic vesicles (and corresponding to "F" profiles of some previous studies of thalamic ultrastructure), formed symmetric synapses and puncta adhaerentia contacts predominantly with large and medium-diameter (i.e. proximal) non-immunoreactive dendrites. Approximately 12 and 18% of boutons in the ventral anterior-ventral lateral complex and ventromedial nucleus, respectively, were GABA-immunopositive. Many of these immunoreactive profiles probably arose from GABAergic neurons in the thalamic reticular nucleus, substantia nigra pars reticulata and entopeduncular nucleus. Two types of non-immunoreactive axon terminals were distinguished based on differences in morphology and synaptic termination sites. Boutons with small ovoid profiles and round vesicles that formed prominent asymmetric synapses onto small-diameter dendrites were observed. Mitochondria were rarely observed within these boutons, which arose from thin unmyelinated axons. These boutons composed approximately 82 and 74% of boutons in the ventral anterior-ventral lateral complex and ventromedial nucleus, respectively, and were considered to arise predominantly from neurons in the cerebral cortex. In contrast, boutons with large terminals that contained round or plemorphic vesicles and formed multiple asymmetric synapses predominantly with large-diameter dendrites were also observed. Puncta adhaerentia contacts were also common. Mitochondria were numerous within large boutons with round vesicles, which arose from myelinated axons. Many of the large boutons were likely to have originated from neurons in the cerebellar nuclei. Approximately 6% of the boutons in the ventral anterior-ventral lateral complex and 8% in ventromedial nucleus were of the large type.(ABSTRACT TRUNCATED AT 400 WORDS)     

Serotonergic innervation of the dorsal horn of rat spinal cord: light and electron microscopic immunocytochemical study

Summary The ultrastructure of serotonergic projections to the dorsal horn of the rat spinal cord has been investigated, using a highly specific polyclonal antiserum. The highest concentrations of immunoreactive profiles were found in lamina I and the outer part of lamina II (IIo). Intermediate concentrations were found in laminae III and IV, while the inner part of lamina II (IIi) was almost devoid of immunoreactivity. Whereas 60% of the profiles show at least one varicosity studded with synaptic vesicles, only one-fifth of the latter contributes to classical synapses, the remaining profiles being devoid of a facing postsynaptic density. The results are compared with those in the literature and our own results relative to other regions of the cord. It is concluded that the pauci-synaptic projections to the dorsal horn could correspond to a diffuse influence of serotonin, the targets for which are determined by the corresponding serotonergic receptors.     

Presence of glial cells in the rat pineal gland: a light and electron microscopic immunohistochemical study

Immunoperoxidase methods for the demonstration of three glial antigens, vimentin, glial fibrillary acidic protein, and S-100 protein, were applied to routine-fixed paraffin sections of rat pineal gland. A pre-embedding electron microscope immunoperoxidase method was also used to study the ultrastructural localization of S-100 protein in pineal cells. Light and electron microscopic results showed the presence of these antigenic glial markers in the second pineal cell type. The term glial cell is proposed for the second of parenchymatous cell in rat pineal gland.     

Regeneration of rat submandibular gland following partial extirpation. A light and electron microscopic study

A wedge of parenchymal tissue was excised from the left submandibular gland of six week old male Sprague-Dawley rats. The animals were randomly grouped by body weight and killed at intervals of one day to five weeks following the operation. Tissue from and adjacent to the site of injury was removed and prepared for routine light and electron microscopy. Light microscopic findings consisted of degeneration and necrosis of the parenchymal tissue during the first 24 hours, followed by hyperemia and endothelial as well as epithelial proliferation from one to three days. Extensive epithelial proliferation occurred during the next two weeks, followed by regeneration of new lobules, beginning at the periphery of the injured lobes. Ultrastructurally, the new parenchymal tissue appeared to have regenerated from residual duct cells. Dedifferentiated epithelial cells gave rise to two different cell lines: one line which transformed into terminal tubules and acinar cells, and another which became striated ducts. These differentiating cells were organizing into lobules as early as three weeks after the operation. Because of their proximity to cells of regenerating striated ducts as well as intercalated ducts and acini, the myoepithelial cells appeared to be of epithelial origin.     

Age-related alterations in the lacrimal gland of adult albino rat: A light and electron microscopic study

BackgroundAge related changes in the lacrimal gland are associated with alterations in the structural organization and functional response in the gland of diverse mammalian species. Dry eye syndrome is one of the most common ocular problems in the world especially in old age. It results when the lacrimal gland fails to secrete proteins and fluid in sufficient quantity or appropriate composition.Aim of the workThe present study is designed to demonstrate the influence of aging on the structure of the lacrimal gland of albino rat and to provide a morphological basis to explain the pathogenesis of the dry eye syndrome with ageing. It also aims to carry out a comparative analysis of age-dependent changes in male and female rats and to address how the lacrimal gland ages in each sex.Material and MethodsEighty albino rats were used in this study. The animals were divided into two age groups, young adult and senile. Tear secretion was measured using a modified Schirmer test. Corneal impression cytology of the anesthetized rats was done. The glands were subjected to gross morphologic examination, microscopic examination using H&E, PAS, Masson's trichrome and Giemsa stains. Electron microscopic examination was done in addition to quantitative histomorphometric estimations included acinar density, ductal count and mast cell count.ResultsLight microscopic examination of the lacimal glands of the senile rats revealed different pathological changes. These included acinar, ductal as well as stromal changes. Electron microscope examination of the lacrimal gland of the senile group showed a decrease in the electron dense secretory vesicles, mitochondrial swelling and lipofuscin-like inclusions were frequently seen in the cytoplasm of acinar cells in senile rats.ConclusionThe structural changes in the lacrimal glands of senile rats were associated with reduction in tear secretion as well as alterations in corneal epithelium. Gender difference in lacrimal gland structure was recorded.     

Fetal neostriatal transplants in the rat: a light and electron microscopic Golgi study.

Fetal striatal neurons were transplanted into the ibotenic acid lesioned rat striatum. Three months after transplantation the grafted tissue was Golgi-impregnated and examined at the light microscopic level to determine the morphological characteristics of the transplanted neurons. Golgi-impregnated neurons were then gold-toned and examined at the electron microscopic level. The transplanted neurons were classified by both somatic size and somatic and dendritic morphology, which demonstrated that at least seven distinct cell types are present in striatal grafts. Type I large neurons had aspinous somata, sparsely spined dendrites, and indented nuclei, whereas type II large neurons displayed somatic spines, sparsely spined dendrites, and indented nuclei. Type I medium neurons exhibited aspinous somata and proximal dendrites, heavily spined distal dendrites, and unindented nuclei. Type II medium neurons had somatic spines, sparsely spined dendrites, and indented nuclei. Type III medium neurons had aspinous somata, poorly branched and sparsely spined dendrites, and indented nuclei, while type IV medium neurons had aspinous somata, highly branched and sparsely spined dendrites, and indented nuclei. Type V medium neurons displayed aspinous somata, varicose dendrites, and indented nuclei. These results demonstrate that transplanted fetal striatal neurons differentiate into morphologically and ultrastructurally distinct striatal cell types.     

Light and electron microscopic immunocytochemical localization of glutamine synthetase in the superficial pineal gland of the rat.

Glutamine synthetase (L-glutamate:ammonia ligase; EC 6.3.1.2), an enzyme catalysing the ATP-dependent formation of glutamine from glutamate and ammonia, was detected immunocytochemically only in glial (interstitial) cells of the superficial pineal gland of the rat. The results show the important role of pineal glial cells in the metabolism of the presumptive neurotransmitters, glutamate and gamma-aminobutyric acid (GABA) as well as in detoxification of ammonia.     

The adrenergic innervation of the rat central amygdaloid nucleus: a light and electron microscopic immunocytochemical study using phenylethanolamine N-methyltransferase as a marker

Using immunocytochemistry of phenylethanolamine N-methyltransferase for light and electron microscopy, investigations were carried out to document the morphology of adrenergic afferents innervating the rat central amygdaloid nucleus and to analyse the manner in which contacts with neurons of the nucleus are formed. With the light microscope, dense terminal plexus of phenylethanolamine N-methyltransferase-immunoreactive axons with typical large boutons (diameter>1 m) were found in the medial central nucleus, extending into its ventral subdivision and the adjacent intra-amygdaloid portion of the bed nucleus of the stria terminalis. Electron microscopy of the medial central nucleus showed phenylethanolamine N-methyltransferase-immunoreaction product in the cytoplasm of intervaricose axons and boutons. Large adrenergic boutons contained numerous small clear vesicles and, occasionally, large dense-cored vesicles. In serial sections, most boutons formed synaptic contacts. Synapses of immunoreactive terminals were mainly of the asymmetric type and localized preferentially on medium sized to small dendrites and dendritic spines. Structures postsynaptic to adrenergic boutons were often additionally contacted by non-labelled terminals. The study gives evidence that adrenergic afferents exert a direct synaptic influence on medial central nucleus neurons. The peripheral localization of the majority of adrenergic synapses, their asymmetric configuration, and the presence of non-adrenergic synapsing terminals in their immediate vicinity indicate that the major component of the adrenergic input is of an excitatory nature, and is integrated with innervation from other sources.Abbreviations BNST _i Intra-amygdaloid portion of the bed nucleus of the stria terminalis - CL lateral central amygdaloid nucleus - CLc lateral capsular central amygdaloid nucleus - CM medial central amygdaloid nucleus - CN central amygdaloid nucleus - CV ventral central amygdaloid nucleus - DAB diaminobenzidine - EM electron microscopy - ir immunoreactive - LM light microscopy - NPY neuropeptide tyrosine - PBS phosphate buffered saline - PFA paraformaldehyde - PNMT phenylethanolamine N-methyltransferase     

Peripheral neuroepithelioma: a light microscopic, immunocytochemical, and ultrastructural study.

Forty-two cases of peripheral neuroepithelioma (PN) retrieved from the files of the National Cancer Institute (Bethesda, MD) and the Pathology Department of Padua University, Italy, were reviewed. No sex predilection was observed (25M/17F) and ages ranged from 7 to 54 yr (median 22 yr). Roughly a third of the tumors were thoracopulmonary ("Askin tumor"), a third were axial, and a third were in extremities. A lobular pattern with rosettes or pseudo-rosettes characterized PN. Seventeen cases showed a strong diastase-sensitive PAS positivity. Transitional areas with an Ewing's-like appearance and, in one case, transition to malignant nerve sheath tumor have been documented. The presence of neuron specific enolase (NSE), S-100 protein, HNK-1, neurofilaments, vimentin, keratin (AE1-AE3), beta 2-microglobulin, chromogranin A, and synaptophysin was investigated using the avidin-biotin technique. Immunocytochemically, NSE (95% of cases), beta 2-microglobulin (77.5%), synaptophysin (73.3%), and S-100 protein (67.5%) were the most consistently positive markers. Ultrastructurally, PN is characterized by a primitive appearance, although it was routinely possible to recognize neural features such as primitive neuritic extensions and dense core granules, either in the cytoplasm or in the cellular processes. In our experience, a light microscopic picture of a primitive round cell tumor with a lobular pattern, and particularly with rosettes when present, with NSE and beta 2-microglobulin positivity by immunocytochemistry, ideally with positive synaptophysin, along with supportive electron microscopy, is required for the diagnosis of PN. Conversely, no one feature alone is generally sufficient for diagnosis, but does allow distinction from extraosseous Ewing's, which (like osseous Ewing's) lacks features of neural differentiation.     

Esthesioneuroblastoma: a light and electron microscopic study

The electron microscopic features of an olfactory esthesioneuroblastoma are described. The tumor cells had well developed neuritic processes containing neurotubules and neurofibrils. In addition, neurosecretory granules, similar to those seen in adrenal neuroblastomas, were present in the main cell body as well as in the neuritic processes. The neurites were also well demonstrated by light microscopic examination when stained by the Palmgren silver method. The importance of demonstration of these processes in establishing a firm diagnosis is stressed. Electron microscopic demonstration of neurites and secretory granules in a nasal tumor is diagnostic of an esthesioneuroblastoma.     

Micronodular adrenal disease: a light and electron microscopic study.

A case of Cushing's syndrome due to micronodular adrenal disease in a 17-year-old girl is presented. The adrenals showed both black and yellow nodules. Histologically the cells contained lipofuschin and either had a clear cytoplasm or an eosinophilic cytoplasm with a prominent nucleus. Lymphocytes were a prominent feature. No cells of the zona reticularis were identified. The cell of origin of these nodules appeared to be from the inner layer of the zona fasciculata. We postulate that the disease is caused by an abnormality in the migration and ultimate destruction of cells from the zona fasciculata to the zona reticularis with a build up of cells at the interface zone.     

Retroperitoneal leiomyosarcoma: a light and electron microscopic study

Retroperitoneal leiomyosarcoma arising in a spermatic blood vessel is described. In addition to typical leiomyosarcoma, areas of atypical histiocyte-like cells and storiform pattern bearing a close resemblance to malignant fibrous histiocytoma, were seen. Ultrastructurally, cells with features of smooth muscle, partially differentiated cells with marginal densities and basal lamina and histiocyte-like giant cells were present. Phagocytosis of inflammatory cells by tumour cells with marginal densities and an investment of basal lamina was observed. It is suggested that the precursor cell in this lesion was a primitive mesenchymal cell showing varying degrees of smooth muscle differentiation and giant cell formation.     

Scar adenocarcinoma of the lung: a light and electron microscopic study.

Five well differentiated peripheral adenocarcinomas of the lung were investigated, using light and electron microscopy. Each tumour contained a central nidus of fibrous tissue and fulfilled the criteria for "scar cancer." One tumour also had a focus of lamellated collagenous tissue, suggestive of an old tuberculous granuloma. Electron microscopy showed the features of Clara cells, with characteristic dense bodies in the apical cytoplasm and scattered microvilli on the luminal surface. It was concluded that this variant of scar cancer was a carcinoma of Clara cells, which was sufficiently distinctive in appearance to be recognised on light microscopy alone. It remains uncertain, however, whether the central fibrous area is a desmoplastic response to tumour growth or a pre-existing scar.     

Spindle cell carcinoma (pseudosarcoma) of the anus: a light, electron microscopic and immunocytochemical study of a case

We report a case of polypoid spindle cell squamous carcinoma (pseudosarcoma) occurring in the anal canal. Electron microscopic findings and the demonstration of keratin by an immunoperoxidase method, gave clear cut evidence of the epithelial nature of the sarcomatoid cells forming this tumour. To our knowledge, this is the first reported case in the literature.     

Kappa opioid receptors are expressed by interneurons in the CA1 area of the rat hippocampus: a correlated light and electron microscopic immunocytochemical study

A local GABA-system is known to have a mediatory function between several afferents and the principal cells of the hippocampus. This study examines the distribution and fine structure of kappa opioid receptor-immunoreactive elements in the CA1 subfield and reveals some new aspects concerning the structural basis of opioid-GABA interaction in the rat hippocampal formation. Kappa receptors were visualized immunocytochemically with a previously produced and characterized monoclonal antibody, the mAb KA8 (Maderspach, K., Németh, K., Simon, J., Benyhe, S., Szûcs, M., Wollemann, M., 1991. A monoclonal antibody recognizing kappa-, but not mu- and delta-opioid receptors. J. Neurochem. 56, 1897–1904). The antibody selectively recognizes the kappa opioid receptor with preference to the kappa₂ subtype. Neuronal cell bodies, proximal dendrites and occasionally glial processes surrounding neuronal perikarya were labelled in the CA1 area. The immunopositive cells were present mainly in the stratum oriens, followed by the stratum pyramidale in a rostrocaudally increasing number. Their shape was fusiform, or multipolar. Occasionally kappa receptor-immunoreactive boutons surrounding weakly immunopositive somata were also observed. Electron microscopy of immunopositive neurons showed that the DAB labelling was intensive in the perinuclear cytoplasm. The widths and electron densities of the postsynaptic densities of some axosomatic synapses were remarkably increased. Similar increase of postsynaptic densities were observable at some axodendritic and axospinous synapses. On the basis of their location and fine structural properties the labelled cells are suggested to be GABAergic inhibitory interneurons, probably belonging to the somatostatinergic sub-population. The axons of these inhibitory interneurons are known to arborize in the stratum lacunosum-moleculare where the entorhinal afferents terminate. A modulatory effect of opioids on the entorhinal input, mediated by somatostatinergic interneurons is suggested     

The localisation of urocortin in the adult rat cerebellum: a light and electron microscopic study

Light and electron microscopic immunocytochemistry was used to identify the cellular and subcellular localisation of urocortin in the adult rat cerebellum. Urocortin immunoreactivity (UCN-ir) was visualised throughout the cerebellum, yet predominated in the posterior vermal lobules, especially lobules IX and X, the flocculus, paraflocculus and deep cerebellar nuclei. Cortical immunoreactivity was most evident in the Purkinje cell layer and molecular layer. Reaction product, though sparse, was found in the somata of Purkinje cells, primarily in the region of the Golgi apparatus. Purkinje cell dendritic UCN-ir was compartmentalised, with it being prevalent in proximal regions especially where climbing fibres synapsed, yet absent in distal regions where parallel fibres synapsed. In the Purkinje cell layer, the labelling was also contained in axonal terminals, synapsing directly on Purkinje cell somata. These were identified as axon terminals of basket cells based on their morphology. Terminals of stellate cells in the upper molecular layer also expressed the peptide. Whilst somata of inferior olivary neurones showed intense immunoreactivity, axonal labelling was indistinct, with only the terminals of climbing fibres containing reaction product. UCN-ir in the mossy fibre-parallel fibre system was restricted to mossy fibre rosettes of mainly posterior lobules and the varicose terminals of parallel fibres. Furthermore, labelling also was prevalent in glial perikarya and their sheaths.The current study shows, firstly, that urocortin enjoys a close ligand-receptor symmetry in the cerebellum, probably to a greater degree than corticotropin-releasing factor since corticotropin-releasing factor itself is found exclusively in the two major cerebellar afferent systems. Its congregation in excitatory and inhibitory axonal terminals suggests a significant degree of participation in the synaptic milieu, perhaps in the capacity as a neurotransmitter or effecting the release of co-localised neurotransmitters. Finally, its unique distribution in the Purkinje cell dendrite might serve as an anatomical marker of discrete populations of dendritic spines.