Localization of alkaline phosphatase (AlPase) in the spinal cord ependyma of newborn and adult rats was studied using cytochemical techniques. When the reaction products were precipitated on the plasma membrane of the ependymal cells, the most intense AlPase activity was apparent on the membrane of the luminal surface of tissues from the newborn rats and this activity was also detected in the neuropil around the central canal. In the axon terminals, reaction products were precipitated on the plasma membrane of the presynapse. These observations warrant further studies on AlPase activity in the ependyma to determine if this enzyme plays a role in the membrane transport of the substances from the cerebrospinal fluid to the extracellular spaces of the spinal cord.
Noradrenaline terminals in the periventricular hypothalamic region of the rat were visualized electron microscopically using a KMnO4 fixation technique. Alterations of these terminals were examined following chronic treatment of chlorpromazine (CPZ; 20 mg/kg) for one month. In the control rats, noradrenaline terminals with a typical synaptic contact accounted for only 7% of the total number of noradrenaline terminals. After 12 days of daily administration of CPZ, noradrenaline terminals with typical synapses increased markedly to approximately 3.5 times over those seenL in the controls. In addition, these terminals with a synapse-like formation also increased up to 2.5 times the number in the controls 30 days after chronic injections of CPZ. On some occasions, daily administration of CPZ for over 20 days resulted in degenerative changes of the noradrenaline terminals which contained membranous structures.
Neoplastic endocrine cells in the carcinomas of the stomach and small intestine of Wistar male rats, induced by MNNG or ENNG, were examined by histochemical, immunohistological and ultrastructural methods. Detection rates of argyrophil and argentaffin cells in the gastric cancers were 60% and 34.3%, respectively. MNNG induced small intestinal carcinomas contained argyrophil cells in 78.3% and argentaffin cells in 65.2%. Of 205 carcinomas of the small intestine induced by ENNG, 115 lesions (56.1%) contained argyrophil cells and 69 lesions (33.7%) contained argentaffin cells. In some of the cases, gastrin, glucagon and somatostatin were demonstrated in the tumor cells by the immunoperoxidase technique. Ultrastructurally, intracytoplasmic granules were observed, and a special form of endocrine cell which contained two different types of intracytoplasmic granules was also found (endocrine-exocrine cell). This latter finding indicates a multidifferentiating potentiality of tumor cell. Moreover, it would appear that these endocrine cells can arise from undifferentiated stem cells, and that the endocrine cells of the gastrointestinal tract are of the endodermal origin.
In order to clarify the distribution of cholesterol in the plasma-and cytomembranes of the oviduct epithelium of mice, freeze-fracture images of the filipin-treated tissues were observed. The filipin-sterol complexes were distributed in high density on the fractured plasma membranes of both secretory and ciliated cells except for special areas. The limiting membranes of secretory granules of the secretory cell were very rich in complexes. In both types of cells, membranes of nucleus and rough endoplasmic reticulum were virtually free of the complexes, though very small aggregates of a few complexes were infrequently scattered. Most Golgi stacks were labeled slightly with filipin, but small vesicles and a stack in the trans-side showed a fairly large number of the complexes to form small aggregates. No complexes were recognized on the outer and inner mitochondrial membranes of both type cells. Although most of the ciliary membrane contained numerous complexes, only the ciliary necklace portion lacked the complexes. The area of zonula occludens was generally free of the complexes. The membrane enclosed by the strands of zonula occludens was devoid of the complexes, though filipin extensively labeled the membrane surrounding the zonula occludens. Round patches studded beneath the zonula occludens, which might correspond to desmosomes, did not bear the complexes.
Following intrastriatal microinjection of haloperidol, the binding sites were demonstrated using ultraimmunohistochemistry. Haloperidol immunoreactive neurons began to appear 1 hr after the injection and dispersed far from the injection site as time passed. Electron microscopically, haloperidol binding sites were visualized as a soluble form. The immunoreaction products were seen throughout the cytoplasm but avoided the nuclear membrane, cisternal structures and most of the mitochondria. The haloperidol immunoreaction products showed shading differences in each cytoplasm, which was understood to be due to various states of affinity to the ligand. Based on these findings, a new concept of receptors is proposed: There are spare receptors before functioning and active receptors having a higher affinity for theligand. Four out of six neuronal types in the rat neostriatum, according to the classification by Kaiya et al. (13, 15), showed haloperidol immunoreaction products.
The fine structural localization of the cholinesterase (ChE) activity was studied in the presumed baroreceptors of the cat atrial endocardium. The unit of baroreceptors belonging to one myelinated fiber was about 60×10μm, in which the axon terminals were delicately arborized and intermingled with collagen fibrils. Such axon terminals were identified by forming various-sized varicosities ranging from 1 to 3μm in diameter, containing numerous mitochondria and a few dense bodies. A large portion of the axolemma of the varicosities was covered only by basement membranes, being in direct contact with the connective tissue. Such ultrastructural characteristics were closely similar to those of the baroreceptors in the carotid sinus, aortic arch and endocardium, as reported by other investigators. The enzyme activity was found mainly in the spaces between the axon terminals and the surrounding Schwann cells. The nuclear envelope in some Schwann cells also displayed reaction products. No definite enzyme activity was demonstrated in any of the organelles in the axon terminals. It is assumed that ChE in the baroreceptors might be synthesized and released by the Schwann cells which surround the axon terminals. The specific localization of ChE activity in the baroreceptors suggests that, similar to other mechanoreceptors, ChE has some important functions in the baroreceptors.
Immunohisto-and immunocytochemical localizations of tyrosine hydroxylase (TH) were studied in the area postrema of hamster brain. By immunohistochemistry, TH-positive neurons with thick, smooth nerve processes were seen to be situated in close apposition to the blood vessels, and they were of oval-shaped, almost unipolar types. By immunocytochemistry, TH-positive reaction products were observed within the cytoplasm and granular vesicles but not in either the nucleus nor Golgi area of the TH-positive neurons. From the size of the granular vesicles, TH-positive cells in the area postrema were divided into two types of neurons, cells containing small granular vesicles (about 100 nm in diameter), and cells containing both small and large granular vesicles (larger than 150 nm in diameter). Small numbers of neurons with TH-negative reaction, were also found. These TH-negative neurons were agranular.
The present investigation is aimed at showing the localization of human chorionic gonadotropin (hCG) and alpha-fetoprotein (AFP) in the same section of human testicular carcinoma using successive double staining by the indirect immunoperoxidase method. Specimens from 3 cases of human testicular tumors with elevated serum hCG and AFP values were fixed in 95% ethanol or 10% buffered formalin. Both anti-hCG-β and anti-AFP rabbit sera were absorbed with several materials in order to avoid cross reaction in the double staining. Indirect immunostaining was performed using anti-hCG-β or anti-AFP serum. HRP activity was demonstrated by reacting with either 3, 3′-diaminobenzidine, 4-Cl-1 naphthol and 4-Cl-1 naphthol-pyronin. HCG- and AFP-localizing cells could be completley differentiated from each other. HCG was localized in the cytoplasm of the syncytiotrophoblastic cells, syncytiotrophoblastic giant cells and embryonal carcinoma cells. AFP was observed at endodermal sinus tumor elements, embryonal carcinoma cells and teratoma elements.
Hepatic gamma-glutamyl transpeptidase (GGTP) induced by the administration of 0.2% phenobarbital (PB) for 20 weeks in rats was investigated histochemically. By the PB treatment, centrolobular and midzonal hepato-cytes became hypertrophic and periportal hepatocytes proliferated when examined on sections stained with hematoxylin and eosin. Histochemically, GGTP activity was demonstrated only in ductular structure and in bile canali-culi of occasional periportal hepatocytes of non-treated rats, whereas with PB treatment, the majority of proliferated periportal hepatocytes showed markedly increased activity of GGTP in bile canaliculi. Centrolobular and midzonal hepatocytes did not show any activity of GGTP, even with the PB treatment. Thus, the induction of GGTP by PB was only evident in the periportal hepato-cytes.
A functional aspect of gamma-glutamyl transpeptidase (GGT) activity in human thyroid tissues was immunohistochemically analysed mainly in malignant thyroid neoplasms. Fresh thyroid tissues were removed and fixed in cold acetone and paraffin sections were used for GGT staining (11), and for immune staining for thyroglobulin (Tg), triiodothyronine (T3) and thyroxine (T4) with the immunoperoxidase technique. In most sections, double stain (GGT and each immunostain) was performed. A high degree of correlation in positivity between GGT activity and immunoreactivities for Tg and T3 was demonstrated in well-differentiated adenocarcinomas, whereas both reactions were almost negative in anaplastic and medullary carcinomas. Squamous cell carcinoma showed positive staining for GGT in spite of negligible immuno-reactivities. Thus, in malignant thyroid neoplasms of follicular cell origin except squamous cell carcinoma, it was concluded that GGT activity was closely related to the immunohistochemically detected functions of follicular epithelium of the thyroid as well as cell proliferation rate (10).