Positive Immunocytochemical Reaction Research Articles

Morphological features of thyroid benign focal neoplasms in Graves’ disease

Background. The morphology of the thyroid in Graves’ disease (GD) can be quite diverse: in most cases there is a diffuse enlargement of the thyroid gland (diffuse goiter), in some cases it is not increased, and in some patients there are focal neoplasms (single and multiple), including thyroid cancer. According to some studies, in patients with Graves’ disease, focal formations are observed in 10–31% of cases, and in half of the patients, they appear against the background of treatment with thionamides. The purpose of this paper was a retrospective study of the morphological features of benign thyroid neoplasms due to GD based on the results of cytological and pathomorphological studies. Materials and methods. The study was performed at the clinic of the State Institution “V.P. Komisarenko Institute of Endocrinology and Metabolism of the National Academy of Medical Sciences of Ukraine”. All patients with GD who were operated in the Department of Endocrine Surgery from 2008 to 2019 (1854 patients) were first selected, and then those who had benign focal formations according to the pathohistological examination were selected among them. Results. The frequency of benign thyroid neoplasms in GD is 22.06% in women aged 26 to 55 years. Cytological studies of thyroid neoplasm punctures in GD made it possible to clearly establish their benign nature (BSRTC-2) in 94.3% of cases. In 5.7% of cases, cytological categories BSRTC-3 and BSRTC-4 were established. Positive immunocytochemical reactions to thyroid peroxidase and thyroglobulin, and the absence of CK17 allow avoiding cytological overdiagnosis, characteristic of benign neoplasms of the thyroid gland in GD. Conclusions. Among pathohistologically confirmed benign neoplasms of the thyroid gland in GD, in 63.82% of cases a histological diagnosis was adenomatous or colloid goiter, in 34.47%— signs of nodular hyperplasia, in 1.71%— the presence of thyroiditis.

Three-dimensional organization of the cytoplasmic neuroglobin-immunopositive structures in the rat medulla oblongata neurons

Neuroglobin is an iron-containing protein, most abundant in the vertebrate nervous system. Since neuroglobin is able to bind oxygen reversibly owing to the heme prosthetic group, it was believed that its function is an intercellular transport of oxygen in the nervous system and accumulation of oxygen for energy supply of cells in hypoxic conditions. In this work, a three-dimensional reconstruction of the neuroglobin distribution in large neurons of the rat medulla oblongata was carried out by means of immunocytochemistry and confocal laser microscopy. Positive neuroglobin immunocytochemical reaction was observed mainly in the perinuclear areas of large nerve cells exhibiting a discrete staining of the cytoplasm. Examination under the microscope at a high magnification revealed some neuroglobin-immunopositive granules, ring-like objects 1–2 μm in diameter, as well as linear and branched structures in neuronal cytoplasm and, occasionally, in the proximal segments of neuronal processes. Three-dimensional reconstruction of the neuroglobin-immunopositive structures showed that they mainly have the form of continuous lines and curves interlaced in some sites, about 1.0–1.5 μm thick, forming a complex network in the cytoplasm. The neuroglobin-immunopositive complexes found for the first time in neuronal cytoplasm are not identical to any known cytoplasmic compartments of nerve cells, but the diameter of their elements, as well as the shape and location suggest a possible link of neuroglobin with a mitochondrial network.

Membrane Transformations in Degenerating Rodlet Cells in Fishes of Two Teleostean Families (Salmonidae, Cyprinidae)

Rodlet cells (RCs) of teleosts are identified by their fibrillar capsule and peculiar inclusions, the rodlets, consisting of a club-like sac and a central dense core. Former ultrastructural studies showing signs of hypertrophy of endoplasmic reticulum (ER) were followed up in Salmonids (Oncorhynchus mykiss, Salmo trutta L.) and compared with Cyprinids (Cyprinus carpio L., Carassius auratus L., Alburnus alburnus). Focusing on membrane transformations, unusual undulations of the membranes of rodlet sacs and often apposed ER-membranes, which were observed in mature or discharging cells, increased continuously in degenerating stages and ejected cytoplasmic packages or rodlets. Tubular elements (ø 25-30 nm or 30-50 nm) or small vesicles appeared partly derived from them. Terminal stages of this development were represented by RCs retained in the epithelium, which were completely filled by stacks of tubules and cores. Convoluted membranes were also found persisting between mostly undissolved rodlets at the epithelial surfaces. In Cyprinid species, the membrane changes were less conspicuous but essentially similar, including stages with confluent ER reported only in trout up to now. The membrane transformations resemble structures known as "crystalloid ER" indicating a disturbance in the protein production. The positive immunocytochemical reaction for calreticulin in the rodlet sacs, a luminal ER chaperone mediating recycling of misfolded proteins and upregulated during stress, supports this interpretation. The ER stress-reaction is an evolutionary conservative cytoprotective mechanism during physiological, environmental, and genetic aberrations and fits the increase of RCs reported in quite different situations, although details of its triggering need further investigation.

Establishment of a Multipotent Neural Stem Cell Line from the Adult Mouse Cerebrum

Study Design: Establishment of a multipotent neural stem cell line from the adult mouse cerebrum. Objectives: To establish a daughter cell line, B2A1, from B2 cells through the limiting dilution method, and to determine if the cells have the characteristics of neural stem cell (NSCs) using immunocytochemistry and RT-PCR methods. Summary and Literature Review: In the development of NSCs, differentiated organ or tissue-derived multipotent stem cells have attracted considerable interest because of the lack of ethical issues. Previously, a glial precursor cell line (B2 cells) was generated from the primary cultures of oligodendrocytes/ astrocytes in an adult BALB/c mouse brain. These cells exhibited the cell-type specific markers for immature neuroectodermal cells, astrocytes, and oligodendrocytes in serum-contained media. Materials and Methods: The primary cultures of oligodendrocytes/astrocytes were established from the whole brains of 12 to 16-week-old BALB/c mice from either gender. After 6 months with 25 serial passages, the culture consisted of a morphologically homogeneous cell population, which was designated as B2 cells. A subclone, B2A1, was isolated from B2 cells through two consecutive limiting dilutions. Results: More than 90% of B2A1 cells showed immunopositivity for nestin, a specific marker for NSC. The cells also showed immunopositivity for the neuronal, astrocytic and oligodendroglial markers. These cells expressed the genotypic mRNA messages for both neural progenitor cells and differentiated neuronoglial cells. These positive immunocytochemical reactions and mRNA messages for neuronoglial cells varied according to the extrinsic growth factors used. However, the treatment of extrinsic growth factors did not produce any significant differences in the nestin-immunopositive cells. Conclusions: B2A1 cells have the immunocytochemical and cytogenetic properties of NSCs, and the capacity to differentiate into neuronoglial cells.

177 ISOLATION AND CULTURE OF MOUSE ENDOMETRIAL CELLS IN A THREE-DIMENSIONAL CULTURE SYSTEM

Implantation of mammalian embryos occurs only during a restricted narrow window. The endometrium becomes highly receptive for the embryos during this period of time. The objective of this study was to establish an in vitro culture system for pre- and post-implantation mouse embryos. In Experiment 1, mouse uterine horns were excised at Day 3.5 post-coitus. After being washed with Dulbecco's phosphate-buffered solution (DPBS), the uterine horns were cut open and incubated with 0.05% trypsin in DPBS at 4°C for 2 h. After trypsinization, the tissues were incubated at 37°C for an additional 30 min. The solution containing epithelial cell suspension was recovered after 30 s of vortexing. The trypsinized uterine horns were then cut into 1-mm3 pieces and digested with collagenase (type I, 1 mg mL-1 in DPBS) at 37°C for 3 h with vigorous shaking. At the end of digestion, the solution was filtered through 40-µm nylon mesh and the flowthrough containing stromal cells was collected. The isolated epithelial and stromal cells were characterized by their morphology and immunocytochemistry. Both types of cells showed positive immunocytochemical reaction with desmin antibody. The cultured epithelial cells formed polyhedral shapes, and more than 95% expressed epithelium-specific protein, cytokeratin-18. On the other hand, most of the spindle-like stromal cells had no signal for cytokeratin-18 expression, although a few scattered cells were positively labeled. In Experiment 2, for construction of the 3-dimensional culture system, epithelial cells obtained by the method described in Experiment 1 were seeded on an artificial basal membrane (ECMatrixTM; Millipore/Upstate/Chemicon, Temecula, CA, USA) with underlying stromal cells embedded in the type I collagen matrix. The whole system was settled in a Millicell® (Millipore) hanging in a 24-well culture plate, and immersed in DMEM medium supplemented with 10% fetal bovine serum, 20 ng mL-1 epidermal growth factor, 63.5 nmol progesterone, and 7.14 nmol 17β-estradiol. The morphology of epithelial cells on the matrix became cuboidal after 48 h of culture. Additionally, the columnar cells with a basal nucleus were observed on the paraffin wax sections. In Experiment 3, mouse E3.5 embryos were recovered and cultured in this established culture system. Normal hatching and/or attachment of the blastocysts were observed after 2 days of culture. In conclusion, our results showed that epithelial cells formed morphologically columnar monolayers and apparently interacted with blastocyst embryos. Successful construction of this model system would facilitate the study of early embryo development through the implantation stage.

Evidence of Prolactin Immunoreactivity in the Bone Marrow of Untreated Multiple Myeloma Patients

The role of prolactin (PRL) in the physiological regulation of the immune system and in hematopoiesis is well known. There is also evidence of the significance of PRL in several pathological conditions such as autoimmune diseases and some malignancies, e.g. colon and breast carcinomas and also B cell malignancies. Multiple myeloma is known as a B cell malignancy. It is the result of malignant transformation of a single clone of neoplastic plasma cells that synthesize abnormal amounts of monoclonal immunoglobulins or immunoglobulin fragments. In our present studies, the possible expression of PRL in bone marrow cells obtained from diagnosed multiple myeloma (17 cases) or nonmyeloma (5 cases) patients was examined by the method of immunocytochemistry. Samples obtained from those multiple myeloma patients (13 cases) who had not received chemotherapy for 6 months prior to these studies showed a positive immunocytochemical reaction for PRL. Bone marrow smears of patients diagnosed with multiple myeloma who had received chemotherapy within 6 months of the study and also the smears of patients without diagnosed multiple myeloma failed to show a positive immune reaction for PRL. In the case of a patient who was examined prior to and also after a period of 3 months of chemotherapy, the PRL-immunopositive bone marrow cells had disappeared due to the treatment. According to the light microscopic analysis of the cell morphology, PRL-immunopositive cells in the bone marrow were mainly, but not exclusively, plasma cells. There was no correlation between the positive PRL staining of cells and the type of monoclonal immunoglobulin or the ratio of plasma cells detected in the bone marrow. Taken together, our results indicate a possible role of PRL in multiple myeloma. Further experiments are necessary to identify the prognostic value of PRL in multiple myeloma.

Deltins: immunochemical evidence for a novel population of peptides of the D cells of the gastro-entero-pancreatic endocrine system.

Differences between the immunocytochemical behaviour of antisera to partially purified porcine gastrins and antisera to either synthetic human gastrin-17-I or highly purified porcine gastrin-17-I raised the hypothesis that hog antral gastrin extracts contain peptides different from somatostatin and gastrin that are responsible for the immunocytochemical reaction of the former antisera in the D (delta) cells of the gastro-entero-pancreatic (GEP) endocrine system. This study was performed to prove this hypothesis. A discard fraction obtained after gel filtration of hog antral gastrin extracts on Sephadex G-50 Superfine was employed to immunize five rabbits. The discard fraction is highly heterogeneous on two-dimensional electrophoresis and contains merely traces of somatostatin and gastrin in RIA. However, rabbit antisera to the discard fraction give strongly positive immunocytochemical reactions exclusively in the D cells of the human antroduodenal mucosa and of the pancreatic islets. Absorption of the antisera with the lyophilized discard fraction abolishes the staining of the D cells, whereas absorption of the antisera with several somatostatins does not affect the staining. Vice versa, staining of the D cells with antisera to cyclic somatostatin-14 is abolished by absorption of the antisera with somatostatin-14 but not by absorption with excess of the discard fraction. In RIA, antisera to the discard fraction do not bind radiolabelled (Tyr(1))-somatostatin-14, Tyr-somatostatin-28 or synthetic human gastrin-17-I. Two-dimensional electrophoresis of acid extracts of isolated canine pancreatic islets followed by Western blotting shows different patterns of distribution of immunoreactive spots obtained with antisera to the discard fraction, to somatostatin-14, and to human proinsulin respectively. These results indicate the existence of a novel population of peptides of the D cells of the GEP endocrine system, for which we propose the term deltins.

Expression of ciliary neurotrophic factor is maintained in spinal motor neurons of amyotrophic lateral sclerosis

Ciliary neurotrophic factor (CNTF) was originally identified as a potent survival factor for a variety of neuronal cell types in vitro and in vivo and in particular in spinal motor neurons of embryonic chick and rat. Using a monoclonal antibody against CNTF (clone 4–68) we analysed the expression of CNTF in paraffin sections of seven human brains and spinal cords immunocytochemically using the ABC method and compared these results with sections of the spinal cords of patients suffering from amyotrophic lateral sclerosis (ALS). In normal human tissue of the central nervous system CNTF immunoreactivity was found in most of the motor neurons of the motor cortex and ventral horn, neurons of the nucleus oculomotorius, intermediolateralis, thoracicus, ependymal cells as well as in smooth muscle cells and endothelial cells of small arteries. A reduced number of astrocytes showed a positive immunocytochemical reaction. In peripheral nerves and nerve roots of the spinal cord we also found a positive staining of Schwann cells and some axons. These immunoreactions could be confirmed by Western blot analyses. Next we analysed postmortem paraffin sections of the spinal cord of seven patients suffering from ALS (age range 30–76 years, median age 46 years, female/male = 4:3). We found CNTF immunoreactivity in most of the motor neurons of the ventral horn in 5 cases. In two cases the number of positively stained motor neurons was less. From these results we conclude that CNTF is expressed in a high number of upper and lower motor neurons in the human CNS and that its expression is maintained in ALS patients.

Gastrin/cholecystokinin-like peptides in a bivalve mollusc Pecten maximus. Immunochemical characterisation and post-prandial variations

1. Gastrin/cholecystokinin-like peptides have been detected in the juveniles of Pecten maximus. 2. For the first time, positive immunocytochemical reactions have been observed in nervous tissue, neurosecretory cells of the cerebral ganglia, and digestive tissues: gastric shield and digestive gland. 3. Radioimmunoassays have demonstrated the presence of G/CCK material in the haemolymph; this suggests a neuroendocrine secretion of this peptide. 4. Heterogeneity of the G/CCK immunoreactive material was shown after molecular sieving of crude extracts of the three tissues. 5. A high molecular weight peptide was found to be present in ganglia and stomach, while only small peptides were common to the ganglia, stomach and haemolymph. 6. After a test meal, an increase of the G/CCK material has been noted in total extracts of Pecten maximus juveniles. 7. Such variations suggest a role of these substances in the regulation of molluscan digestive processes.

Detection of ras oncoprotein in liver cells of flatfish (Dab) from a contaminated site in the North Sea

A positive immunocytochemical reaction for the N-terminal peptide of N-ras oncoprotein was detected in small groups of hepatocytes and circular foci of altered cells in all the livers of dab ( Limanda limanda) sampled from a contaminated site in the German Bight. Fish sampled from less contaminated sites showed a low or zero incidence of N-ras positive epitope. Livers of fish from the contaminated site were known to have a high incidence of putative preneoplastic lesions and the possible use of ras oncoprotein as a tumour marker in environmental toxicologic pathology is discussed.

ETN-1: a new human endometrial carcinoma cell line producing ascites and distant metastases in nude mice.

A human carcinoma cell line (ETN-1) has been established from a skin metastasis of a moderately differentiated adenocarcinoma of endometrial origin. The cell line has been so far maintained for 27 months through 55 passages, growing as a monolayer as well as in 3-dimensional clusters with a population doubling time of 72 hr. The number of chromosomes per cell varied from 39 to 107 (average number 61.0 +/- 19.8), with a modal number of 46-48. Seven clonal marker chromosomes were detected. Flow cytometric analysis revealed a population of pseudo-tetraploid cells (DNA index 2.1) next to a pseudo-diploid population (DNA index 1.1). The epithelial character of the cells was confirmed by a positive immunocytochemical reaction using monoclonal antibodies (MAbs) to different keratins, the epithelial cell markers BW 495/36 and HMFG-2, as well as by the presence of many junctional complexes. The tumour cells retained a positive reaction with the anti-ovarian carcinoma OV-TL 3, OV-TL 10 and OC 125 MAbs, although the reaction was markedly diminished in comparison with the original tumour. Tumour cells inoculated subcutaneously in nude mice produced well differentiated adenomatous tumour nodules with formation of glandular lumina and basal lamina. Tumour cells injected intraperitoneally produced malignant ascites and regional as well as distant metastases of adenomatous character.

Expression of simple epithelial type cytokeratins in stratified epithelia as detected by immunolocalization and hybridization in situ.

Multi-layered ("stratified") epithelia differ from one-layered ("simple") polar epithelia by various architectural and functional properties as well as by their cytoskeletal complements, notably a set of cytokeratins characteristic of stratified tissue. The simple epithelial cytokeratins 8 and 18 have so far not been detected in any stratified epithelium. Using specific monoclonal antibodies we have noted, in several but not all samples of stratified epithelia, including esophagus, tongue, exocervix, and vagina, positive immunocytochemical reactions for cytokeratins 8, 18, and 19 which in some regions were selective for the basal cell layer(s) but extended into suprabasal layers in others. In situ hybridization with different probes (riboprobes, synthetic oligonucleotides) for mRNAs of cytokeratin 8 on esophageal epithelium has shown, in extended regions, relatively strong reactivity for cytokeratin 8 mRNA in the basal cell layer. In contrast, probes to cytokeratin 18 have shown much weaker hybridization which, however, was rather evenly spread over basal and suprabasal strata. These results, which emphasize the importance of in situ hybridization in studies of gene expression in complex tissues, show that the genes encoding simple epithelial cytokeratins can be expressed in stratified epithelia. This suggests that continual expression of genes coding for simple epithelial cytokeratins is compatible with the formation of squamous stratified tissues and can occur, at least in basal cell layers, simultaneously with the synthesis of certain stratification-related cytokeratins. We also emphasize differences of expression and immunoreactivity of these cytokeratins between different samples and in different regions of the same stratified epithelium and discuss the results in relation to changes of cytokeratin expression during fetal development of stratified epithelia, in response to environmental factors and during the formation of squamous cell carcinomas.

Detection of peroxisomes in human liver and kidney fixed with formalin and embedded in paraffin: the use of catalase and lipid beta-oxidation enzymes as immunocytochemical markers.

We describe the immunocytochemical localization of four peroxisomal enzymes by light microscopy in human liver and kidney processed routinely by formalin fixation and paraffin embedding. Monospecific antisera against catalase and three enzymes of peroxisomal lipid beta-oxidation (acyl-CoA oxidase, bifunctional protein (enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase) and 3-ketoacyl-CoA thiolase) were used in conjunction with either the indirect immunoperoxidase method or the protein A-gold technique followed by silver intensification. The sections of formalin-fixed paraffin-embedded tissue had to be deparaffinized and subjected to controlled proteolysis in order to obtain satisfactory immunostaining. Under the conditions employed, peroxisomes were distinctly visualized in liver parenchymal cells with no reaction in bile duct epithelial or sinusoidal lining cells. In the kidney, peroxisomes were confined to the proximal tubular epithelial cells with negative staining of glomeruli, distal tubules and collecting ducts. A positive immunocytochemical reaction was obtained even in paraffin blocks stored for several years. The method offers a simple approach for detection of peroxisomes and evaluation of their various enzyme proteins in material processed routinely in histopathology laboratories and should prove useful in the investigation of the role of peroxisomes in human pathology for both prospective and retrospective studies.

Synapses on the mauthner cell of the goldfish: Thin section, freeze‐fracture, and immunocytochemical studies

AbstractLarge myelinated club ending and small‐vesicle bouton synapses on the distal part of the lateral dendrite of the goldfish Mauthner cell were investigated with thin section, freeze‐fracture, and immunocytochemical electron microscopic methods. Large myelinated club endings form mixed synapses, having both gap junctions and chemical synaptic junctions. The correlation of the number of gap junction particles (connexons) and the data from electrophysiological studies of single large myelinated club ending synapses suggest that only a small fraction of gap junction channels are open at any given time during electrical synaptic transmission. The chemical synaptic junctions at the large myelinated club ending synapse have large, round synaptic vesicles, indicating that they are excitatory. This result is in agreement with electrophysiological data demonstrating the excitatory nature of this chemical synapse. Freeze‐fracture of these excitatory chemical synaptic junctions reveals the presence of the intramembrane particle aggregates in the postsynaptic E face.Small‐vesicle boutons form chemical synaptic junctions with small, flat or oval synaptic vesicles. These structural data, in combination with previous electrophysiological studies, suggest that the small‐vesicle bouton synapses are inhibitory. In support of this theory, the cytoplasmic side of the postsynaptic membrane of some of these synapses show positive immunocytochemical reaction to monoclonal antibodies against the rat glycine receptor. Freeze‐fracture data reveal intramembrane particle aggregates in the postsynaptic P face of some small‐vesicle bouton synapses which could possibly represent glycine receptor aggregates.

Studies on immunocytochemical localization of inhibin-like material in human prostatic tissue: Comparison of its distribution in normal, benign and malignant prostates

A specific antiserum has been generated against inhibin-like material (ILM) of prostatic origin. Using the immunoperoxidase technique, localization of ILM has been examined in a total of 114 prostates including normal (4 specimens), malignant (46) and hyperplastic (55) tissues. ILM positive immunocytochemical reactions were confined to the cytoplasm and not the nucleus of the prostatic acinar cells in the three categories of prostate, whereas the stroma showed negative reactions. The intensity of positive reactions decreased in the following order: Hyperplasia, incidental and moderately differentiated carcinomas, poorly differentiated carcinomas, whereas metaplasia and granulomatous prostatitis gave negative reactions for ILM. Using this experimental protocol, 200 non-prostatic tissue were found to be completely negative, demonstrating the specificity of the test for prostatic epithelium. These findings indicate a potential use of ILM as a marker of prostatic tissue.

Primary oat cell carcinoma of the kidney

A malignant renal neoplasm with all the morphologic attributes of oat cell (small cell, neuroendocrine) carcinoma is presented. It metastasized widely to regional lymph nodes and resulted in the death of the patient. Ultrastructurally, the tumor contained dense-core endocrine-type secretory granules. It had a cell component which was argyrophilic and which gave a positive immunocytochemical reaction for calcitonin. To the best of our knowledge, this is the first documentation of this tumor type in the kidney.

Immunocytochemical localization of a substance in the eyestalk of the prawn, Palaemon serratus, reactive with an anti-FMRF-amide rabbit serum.

By use of a specific antiserum against the molluscan cardio-excitatory tetrapeptide FMRF-amide in combination with the PAP-method it was possible to obtain positive immunocytochemical reactions in several neurosecretory regions of the eyestalk of the prawn Palaemon serratus. FMRF-amide-like material was found in perikarya and nerve fibers of the medulla terminalis and in neurons in the lamina ganglionaris. The immunoreactivity observed in the glandular tissue located at the basal insertion of the eyestalk muscles must be ascribed to a non-specific reaction. The identification of immunopositive nerve fibers, ending on a nerve bundle in the medulla terminalis, and the fact that immunoreactive material was absent in the neurohemal sinus gland seem to indicate a neurotransmitter/neuromodulator function.

The effect of age, pairing, copulation, and 2-Br-alpha-ergocryptine in the male Mongolian gerbil on the prolactin cells in the anterior pituitary identified by immunocytochemical and Herlant's tetrachrome methods.

Near-adjacent, midsagittal sections of pituitaries of infantile and adult male Mongolian gerbils in several experimental groups (bachelor, paired, tartaric acid-injected, postcopulatory gerbil, and postcopulatory animal injected with 2-bromo-alpha-ergocryptine [CB-154]), were stained by a modification of Herlant's tetrachrome and by the peroxidase-labeled antibody method for prolactin. Cell counts of erythrosinophils and cells immunoreactive for prolactin were made. Sequential staining procedures and thin adjacent sections were used to correlate the staining results. Erythrosinophils were very rare in the infant pituitaries; they increased (P less than .01) in numbers in bachelor pituitaries, remained at the same level (P greater than .05) in paired animals, increased (P less than .01) in the postcopulatory gerbils that were injected with tartaric acid, and increased much less (P less than .05) in postcopulatory males that were injected with CB-154. Prolactin cells were present in modest numbers in infant pituitaries; they increased (P less than .01); in bachelors and reached their highest number in paired animals (P less than .001); they remained unchanged (P greater than .05) in tartaric acid-injected postcopulatory animals but declined (P less than .001) in CB-154-injected, postcopulatory animals. The number of prolactin cells was always significantly greater (P less than .001) than the number of erythrosinophils. Correlative studies revealed the erythrosinophils, some of the light blue cells, and some of the chromophobes gave positive immunocytochemical reactions for prolactin. Apparently, CB-154 inhibited the erythrosinophils and the immunoreactive prolactin cells in the postcopulatory male gerbil, as indicated by a reduction in the number, size, and staining intensity of the cells.

Preparation and discharge of secretion in the subcommissural organ of the rat. An electron-microscopic immunocytochemical study.

The secretion of the subcommissural organ (SCO) of the rat was studied by means of immunocytochemistry at the electron-microscopic level with the use of (1) the polar embedding medium Lowicryl K4M at -30 degrees C, (2) the protein A-gold technique, and (3) a rabbit antiserum against bovine Reissner's fiber (see Sterba et al. 1981). Two different substructures of the ependymal and the hypendymal SCO-cells display a positive immunocytochemical reaction: (1) sacs containing flocculent secretion, which originate from the granular endoplasmic reticulum, and (2) vacuoles filled with fine granular secretion, which are pinched off from the Golgi apparatus. The secretory material of the sacs and the vacuoles is discharged both (i) apically into the cerebrospinal fluid and (ii) basally into intercellular spaces of the SCO-hypendyma. The apically released secretion is condensed to a lamina-like formation, which more caudally assumes the form of Reissner's fiber. The route of the basally released secretion remains, however, vague. The "periodically striated bodies", which were thought to be morphological mediators of the discharge of the secretion into the capillaries, are never labeled by gold particles.

Immunocytochemical Investigations of the Infundibular Organ in Amphioxus (Branchiostoma lanceolatum; Cephalochordata)

AbstractReissner's fibre (RF) is an extraordinary ancient feature of the chordate central nervous system. In vertebrates the RF material is released by the cells of the subcommissural organ (SCO) situated in the roof of the brain. In cephalochordates the RF forming apparatus is the infundibular organ (IFO), a structure in the floor of the brain vesicle. Both the SCO and the IFO are areas of specialized secretory ependymal cells. Recently the SCO‐secretion of many vertebrate species has been studied immunocytochemically by use of different antisera against RF‐material from cows. The present investigation with the PAP‐technique showed that these antisera give a positive immunocytochemical reaction with the secretion of the IFO of amphioxus. The results indicate molecular similarities of compounds present in both the SCO‐secretion and the IFO‐secretion. The conception that the SCO and the IFO are descendants from a single large glandular area in the brain vesicle of an ancient chordate ancestor is emphasized by these findings