Substance P-like immunoreactive nerve fibers in the pars distalis of the anterior pituitary in the dog

Summary The pars distalis of the anterior pituitary is known to be regulated by hypothalamic hormones. Recently, we have discovered the presence of substance P-like immunoreactive nerve fibers in the pars distalis of the monkeys. Substance P-like immunoreactivity in the pars distalis of the dog was investigated in this study. A substantial amount of substance P-like immunoreactive nerve fibers with a large amount of varicosities were found. They were widely distributed in the gland, more abundant along its periphery. Most of them were closely related to the glandular tissue, some were located on vascular walls. Substance P-like immunoreactive nerve fibers were also found in the meningeal sheath of the anterior pituitary. They could be followed into the parenchyma of the gland.     

Depicting the Spatial Distribution of Proteins in Human Tumor Tissue Combining SELDI and MALDI Imaging and Immunohistochemistry

Carcinoma tissue consists of not only tumor cells but also fibroblasts, endothelial cells or vascular structures, and inflammatory cells forming the supportive tumor stroma. Therefore, the spatial distribution of proteins that promote growth and proliferation in these complex functional units is of high interest. Matrix-assisted laser desorption/ionization imaging mass spectrometry is a newly developed technique that generates spatially resolved profiles of protein signals directly from thin tissue sections. Surface-enhanced laser desorption/ionization mass spectrometry (MS)combined with tissue microdissection allows analysis of defined parts of the tissue with a higher sensitivity and a broader mass range. Nevertheless, both MS-based techniques have a limited spatial resolution. IHC is a technique that allows a resolution down to the subcellular level. However, the detection and measurement of a specific protein expression level is possible only by semiquantitative methods. Moreover, prior knowledge about the identity of the proteins of interest is necessary. In this study, we combined all three techniques to gain highest spatial resolution, sensitivity, and quantitative information. We used frozen tissue from head and neck tumors and chose two exemplary proteins (HNP1–3 and S100A8) to highlight the advantages and disadvantages of each technique. It could be shown that the combination of these three techniques results in congruent but also synergetic data. (J Histochem Cytochem 58:929–937, 2010)     

Multiple mechanisms contribute to myenteric plexus ablation induced by benzalkonium chloride in the guinea-pig ileum

Ablation of rat myenteric plexus with benzalkonium chloride has provided a model of intestinal aganglionosis, but the degenerative responses are not well understood. We examined the effects of this detergent on neurons and glia, including expression of c-Myc, c-Jun, JunB, and c-Fos, and on immunocytes in the guinea-pig ileum. Benzalkonium chloride (0.1%) or saline was applied to the serosal surface of distal ileum. Tissues were analyzed 2, 3, or 7 days later and compared with cyclosporine-treated and untreated animals. More than 90% of myenteric neurons were destroyed in ileal segments 3–7 days after benzalkonium-chloride treatment. Glia withdrew processes from around neurons after 2 days and were mostly gone after 3 days. Neuronal c-Myc began to disappear while c-Fos, c-Jun, and JunB were evident in some neuronal nuclei after 2 or 3 days. After 3 days, widespread apoptosis was evident in the myenteric plexus. Populations of T cells, B cells, and macrophage-like cells in untreated and saline-treated myenteric plexuses were substantially increased 3 and 7 days after benzalkonium-chloride treatment. Cyclosporine delayed significant neuronal loss. We conclude that a variety of degenerative mechanisms may be active in this model, including an immune response which may actively contribute to tissue destruction. Received: 13 September 1996 / Accepted: 20 January 1997     

Ontogeny of gonadotropin releasing hormone and gonadotropin immunoreactivity in brain and pituitary of normal and estrogen-treated guppies,Poecilia reticulata Peters

Summary Gonadotropin releasing hormone (GnRH) and gonadotropic hormone (GTH) were identified by immunohistochemistry in the brains and pituitaries of neonate, juvenile and adult guppies. GTH was present in some cells of the pars intermedia (pi) and proximal pars distalis (ppd) of all animals. GnRH was found in the perikarya of the nucleus olfactoretinalis. In the pituitaries of juvenile 30-day-old guppies, GnRH-immunoreactive cells existed in a juvenile pattern, whereas in adult animals GnRH was recognized in only a few cells. GnRH-immunoreactive fibers were seen in the pituitaries of animals that were 30 days or older. In adult guppies, the ventral and lateral ppd (the gonadotropic region) contained a dense network of GnRH-immunoreactive fibers. Pituitary cells staining for either GnRH or GTH were located in different places. After immunohistochemical double staining of adult pituitaries, none of the GnRH-immunoreactive cells were LH-immunoreactive, although both cell types were often found in close proximity. After 20 days or more of ethinylestradiol treatment, less immunoreactive GnRH was detected in the pituitary cells of juvenile guppies, and fewer animals exhibited the juvenile pattern of GnRH-immunoreactive pituitary cells, when compared with untreated controls. The results indicate that GnRH-immunoreactive pituitary cells in the guppy are distinct from gonadotropes and that these cells are involved in regulatory processes along the juvenile brain-pituitary-gonad axis.     

Immunohistochemical observations of the endocrine pancreas during metamorphosis of the sea lamprey,Petromyzon marinus L.

Summary Light-microscopic immunohistochemistry was used to localize insulin- and somatostatin-immunoreactive cells within developing endocrine pancreatic tissue of metamorphosing lampreys, Petromyzon marinus. The extrahepatic common bile duct and a portion of the intrahepatic bile duct develop into the caudal portion of the endocrine pancreas. The cranial pancreas is composed of follicles originating in the intestinal and diverticular epithelia, thus following the method of formation of pancreatic follicles from gut epithelium in larvae. In both the cranial and caudal portions, and in an intermediate cord of isolated follicles which connect these two major masses, insulin-immunoreactive cells appear first and are followed by cells showing somatostatin-immunoreactivity. In all stages of metamorphosis individual endocrine cells demonstrate immunoreactivity to a single hormone. Biliary atresia in lamprey may have some adaptive significance in providing cells that produce a caudal endocrine pancreas.Supported by NSERC of Canada grant No. A5945 and MRC of Canada grant No. MA8629 to JHY     

Mucosal epithelial cells and Langerhans cells are targets for infection by the immunosuppressive type D retrovirus simian AIDS retrovirus serotype 1

Simian acquired immune deficiency syndrome (SAIDS) caused by the type D retrovirus SRV-1 results in opportunistic infections and a spectrum of oral lesions similar to those seen in humans with AIDS. To better understand the pathogenesis of these oral lesions we have retrospectively examined the oral mucosa from ten rhesus monkeys that died with SAIDS and prospectively examined the oral mucosa of ten additional animals inoculated with SRV-1 to determine at what time, and in what cells SRV-1 infection of the oral mucosa occurs. Using single and double label immunohistologic techniques, and electron microscopy we detected SRV-1 in clusters of oral epithelial cells and rare Langerhans cells as early as 1 month postinoculation.     

Localization and characterization of neuropeptide Y-like peptides in the brain and islet organ of the anglerfish (Lophius americanus)

Summary Results from a previous report demonstrate that more than one molecular form of neuropeptide Y-like peptide may be present in the islet organ of the anglerfish (Lophius americanus). Most of the neuropeptide Y-like immunoreactive material was anglerfish peptide YG, which is expressed in a subset of islet cells, whereas an additional neuropeptide Y-like peptide(s) was localized in islet nerves. To learn more about the neuropeptide Y-like peptides in islet nerves, we have employed immunohistochemical and biochemical methods to compare peptides found in anglerfish islets and brain. Using antisera that selectively react with either mammalian forms of neuropeptide Y or with anglerfish peptide YG, subsets of neurons were found in the brain that labelled with only one or the other of the antisera. In separate sections, other neurons that were labelled with either antiserum exhibited similar morphologies. Peptides from brains and islets were subjected to gel filtration and reverse-phase high performance liquid chromatography. Radioimmunoassays employing either the neuropeptide Y or peptide YG antisera were used to examine chromatographic eluates. Immunoreactive peptides having retention times of human neuropeptide Y and porcine neuropeptide Y were identified in extracts of both brain and islets. This indicates that peptides structurally similar to both of these peptides from the neuropeptide Y-pancreatic polypeptide family are expressed in neurons of anglerfish brain and nerve fibers of anglerfish islets. The predominant form of neuropeptide Y-like peptide in islets was anglerfish peptide YG. Neuropeptide Y-immunoreactive peptides from islet extracts that had chromatographic retention times identical to human neuropeptide Y and porcine neuropeptide Y were present in much smaller quantities. These results are consistent with the hypothesis that peptides having significant sequence homology with human neuropeptide Y and porcine neuropeptide Y are present in the nerve fibers that permeate the islet.     

Immunohistochemical localization of tissue-type plasminogen activator in ovaries before and after induced and spontaneous ovulation in the rat

Summary The observation that tissue-type plasminogen activator (tPA) activity increased dramatically in preovulatory follicles has led to the hypothesis that plasminogen activation is causally related to follicle rupture. With immunohistochemistry, we have studied the appearance of tPA in ovaries of immature rats induced to ovulate and in adult cycling rats. Treatment of immature female rats with a single dose of pregnant mare serum gonadotropin (PMSG) induced follicular maturation. A subsequent human chorionic gonadotropin (hCG) injection resulted in follicle rupture 12–14 h later. PMSG treatment alone did not induce appearance of tPA-immunoreactive cells in any ovarian compartment. After hCG stimulation, however, theca cells, granulosa cells, and oocytes of pre- and postovulatory follicles displayed distinct tPA immunoreactivity. Fibroblastlike cells in the theca layers and tunica albuginea of the follicle apex also demonstrated localized cytoplasmic tPA reactivity. In addition to tPA synthesis in preovulatory follicles, hCG also induced tPA staining in the theca (but not granulosa) layers of non-ovulatory follicles. At 24 h after hCG treatment, there was a marked tPA staining in developing corpora lutea, ovulated ova, and oviductal epithelium. Ovaries from regularly cycling adult rats displayed a similar ovulation-related pattern of tPA immunostaining. The appearance of tPA in different cell types of the preovulatory follicle and in the fibroblast-like cells at the follicle apex, strengthens the hypothesis of a direct involvement of tPA in follicle rupture. Presence of tPA in postovulatory oocytes, cumulus cells, and surrounding oviductal epithelium may also indicate a role for tPA in the transfer of eggs in the oviduct.This work was supported by NIH Research Grants HD-14084; 12303     

Immunohistochemical study on fetal raphe samples transplanted into the leptomeningeal tissue of 5,6-dihydroxytryptamine-treated adult rats

Summary Pieces of fetal midbrain raphe containing serotonergic and dopaminergic neurons were transplanted into the leptomeningeal tissue (see Fig. 3) of adult host rats that had previously been denervated by treatment with 5,6-dihydroxytryptamine. One, 2 and 5 months after transplantation, the rate of neuronal survival in the grafted tissue and the extent of axonal outgrowth into the host brain were studied by use of serotonin and tyrosine hydroxylase (TH) immunohistochemistry. The survival rate of the grafts in the 1-month group was approximately 70%. Neurons containing either serotonin or catecholamine were demonstrated by means of immunocytochemical procedures in the grafts. Two and 5 months after transplantation, serotonin-immunoreactive nerve fibers were densely distributed throughout the graft tissue, while TH-immunoreactive fiber elements were restricted to an area near the somata of TH-positive neurons. Numerous serotonin-immunoreactive fibers derived from the transplant were found in the leptomeningeal tissue surrounding the graft, on the wall of neighboring blood vessels, and also in the adjacent parenchyma of the host brain. Outgrowing TH-immunoreactive nerve fibers were not observed in the host brain, although such elements occurred in the leptomeningeal tissue and the wall of the larger blood vessels. These results suggest that the serotonergic and catecholaminergic (dopaminergic) neurons located in transplants of the raphe nuclei show different patterns when reinnervating the host tissue.     

Immunohistochemical localization of glial fibrillary acidic protein (GFAP) and vimentin in the subcommissural organ of the Mongolian gerbil (Meriones unguiculatus)

Summary The chemical composition of intermediate filaments (IF's) in the ependyma of the subcommissural organ (SCO) of the Mongolian gerbil (Meriones unguiculatus) was investigated immunohistochemically in paraffin-embedded tissue. Antibodies against glial fibrillary acidic protein (GFAP), vimentin, neurofilament proteins and cytokeratins were used. Only GFAP and vimentin were detected in the non-specialized diencephalic ependyma and in the ependymocytes of the SCO. Staining could be observed in apical and basal processes of the SCO-cells. The latter processes extended into the posterior commissure up to the subpial surface, thus establishing a well-developed leptomeningeal route of ependymal projections. In contrast to the homogeneous vimentin-labeling, the SCO was particularly immunoreactive for GFAP in its lateral aspects and in the supraand precommissural parts. The coexpression of GFAP and vimentin in a subclass of SCO-ependymocytes was demonstrated on differentially immunostained semithin sections. The present study confirms the glial nature of the SCO-ependyma, which has been a matter of debate recently. It appears from this investigation that the high degree of secretory activity in the SCO does not necessarily lead to the disappearance of glial IF proteins. Moreover, the SCO-cells belong to the expanding group of mature astroglia, which is characterized by coexpression of GFAP and vimentin. The morphological similarity between SCO-ependymocytes and tanycytes is underscored by their common immunoreactivity against these two IF proteins. In view of the absence of GFAP from the rat SCO, interspecific differences must be considered in the evaluation of the IF protein composition.