Tamiko Tachibana

Ultrastructural evidence for a possible secretory function of Merkel cells in the barbels of a teleost fish, Cyprinus carpio

SummaryExamination of barbels of the carp (Cyprinus carpio) revealed cells showing the characteristics of Merkel cells. Some ultrastructural features of these cells suggest a secretory function.

Ultrastructural localization of calcium in mechanoreceptors of the oral mucosa

SummaryCytochemical localization of Ca2+ in Meissner corpuscles and Merkel cell-neurite complexes in the palatine mucosa of the Mongolian gerbil was studied by a combined oxalate antimonate-microwave irradiation procedure. The reaction products obtained were identified as calcium antimonate by EGTA solubility and X-ray microanalysis. Meissner corpuscles in the normal palatine rugae could be roughly classified into three types by amount and localization of Ca2+. Type I corpuscles were characterized by a high Ca2+ content in both the terminal axoplasm and caveolae of the lamellar plates, type II, by a low Ca2+ content in the terminal axoplasm and a high Ca2+ content in the lamellar cytoplasm. Type III corpuscles showed intermediate characteristics. Palatine rugae stimulated mechanically during fixation contained an increased number of type I corpuscles. On the other hand, two patterns were distinguished in the distribution of Ca2+ in Merkel cells in palatine rugae fixed under normal conditions. One showed abundant Ca2+ dispersed throughout the cell, while in the other, Ca2+ was specifically localized in the Golgi apparatus and mitochondria. Similar distribution patterns also were observed in palatine rugae that had received mechanical stimulus during fixation. Axon terminals of most Merkel cell-neurite complexes in normal palatine rugae were poor in axoplasmic Ca2+, whereas those in most Merkel cell-neurite complexes in mechanically stimulated palatine rugae contained abundant Ca2+ in their axoplasm.

Migration of Merkel cells in the labial mucous epithelium of adult rabbits following mental nerve resection

SummaryMerkel cells in the lower labial mucosa of adult rabbits were studied electron microscopically, 9, 21, 28, and 50 days after resection of the mental nerves. By day 9, nerve fibers were completely retracted from the epithelial layer of the mucosa. On and after day 21, Merkel cells were located not only in the basal layer but also in the prickle or more superficial cell layers. The ultrastructure of the migrating Merkel cells was unchanged, both as to the amount and location of the specific cored granules in the cytoplasm, until the cells reached the granular cell layer. The position of the migrating Merkel cells differed from cell to cell, and migration continued for at least 50 days. A remarkably large number of immature Merkel cells was observed in the basal and suprabasal cell layers of the denervated epithelium even by day 50. Therefore, the possibility of the reproduction of Merkel cells exists. The migrating Merkel cells, as well as the keratinocytes in the same cell layer, had degenerated drastically in the parakeratinized cell layer. This seems to indicate that the Merkel cells belong to the line of keratinocytes.

Postnatal differentiation of Merkel cells in the rat palatine mucosa, with special reference to the timing of peripheral nerve development and the potency of cell mitosis.

The origin and mechanism of the differentiation and proliferation of Merkel cells are enigmatic. A preliminary study in our laboratory showed that Merkel cells in the rat palatine mucosa emerge after birth. This is in contrast to the case of similar cells in the skin that differentiate during the embryonic period prior to the establishment of peripheral nerve innervation. We studied immunohistochemically the developmental timings of Merkel cells and peripheral nerves in the rat palatine mucosa using antibodies to cytokeratins 18 and 20, PGP 9.5, and CGRP using developing palates of prenatal and postnatal rats. We also studied the potency of mitosis in Merkel cells by immunohistochemistry using antibodies for a cell proliferation marker Ki67 and cyclin D-kinase inhibitors p16, p21 and p27. It was shown that Merkel cells in the rat palatine mucosa differentiate postnatally, after the development of peripheral nerve fiber terminals was almost established. The emergence and increase in number of Merkel cells progressed in an anterior-to-posterior wave. Newly appearing Merkel cells were usually negative for anti-cytokeratin 20 antibody but gained affinity for the antibody with progress of maturation. All Merkel cells in the palatine mucosa were negative for anti-Ki67 antibody but positive for anti-p27 antibody. These results indicate that Merkel cells in the rat palatine mucosa are not responsible for the development of peripheral nerve fiber terminals and that these cells differentiate in situ from intraepithelial stem cells.

Hemopoietic sites and development of eosinophil granulocytes in the loach, Misgurnus anguillicaudatus

SummaryUnique eosinophils, each of which contained only one eosinophilic granule, have been found in the peripheral blood of the loach (itMisgurnus anguillicaudatus). Several loach organs have been studied by light and electron microscopy to determine the hemopoietic site of this cell type. Eosinophils are produced mainly in the spleen and to a small extent in the kidney, but not in other organs.Presumed myeloblasts are identified as large lymphoid cells containing a number of small-dense granules (diameter, 0.12–0.16 μm) in the cytoplasm. These granules have been observed throughout eosinophilopoiesis but they are most abundant in the promyelocyte stage. The largest cells have been identified as myelocytes which contain a number of large granules (diameter, 0.7–1.4 μm) with electron-dense crystalline cores. These large granules are present from the myelocyte to metamyelocyte stage. Metamyelocytes differ from myelocytes in having more large granules. Mature eosinophils are morphologically similar to metamyelocytes but are characterized by the presence of only one very large electron-dense granule (diameter, 2.5–2.8 μm) with a crystalline core.The nature of these granules has been studied by enzyme digestion using pepsin and trypsin. The results indicate that the crystalline cores are almost pure protein.

A comparative electron microscopic analysis of mechanoreceptors in the hard palate of the mouse (Mus musculus; Rodentia) and the musk shrew (Suncus murinus; Insectivora).

These mechanoreceptors were studied with particular regard to their type and the distribution. In both species they were concentrated in the crests of transverse palatine rugae. In the mouse, Meissner corpuscles, glomerular corpuscles, and Merkel cell nerve endings were seen in every palatine ruga, though the first antemolar ruga also contained simple and atypical lamellated corpuscles. In contrast, the palatine rugae of the musk shrew contained only simple corpuscles and Merkel cell nerve endings. Meissner corpuscles in the mouse palate were frequently aggregated to form a palisade-like, huge corpuscle. Anterior intermolar rugae especially contained a larger number of such huge corpuscles. Simple corpuscles in the palate of the musk shrew were more densely distributed in the anterior than in the posterior rugae. In both species Merkel cell nerve endings were localized within epithelial downgrowths of the rugal crest. These downgrowths tended to increase in number anteroposteriorly.

Intraperineural localization of lamellated sensory corpuscles in the skin and oral mucosae of the adult cat and miniature pig

SummaryLamellated sensory corpuscles in the perioral tissues of the adult cat and the adult miniature pig were studied by light and electron microscopy. In the cat lip, over half of the lamellated corpuscles existed in an aggregate form associated with other nerve elements and blood capillaries in peripheral nerve swellings, while the remainder existed in an isolated form. Conversely, almost all the lamellated corpuscles in the miniature pig existed in an aggregate form in nerve swellings. Most of the intraperineural lamellated corpuscles showed ultrastructural characteristics of simple corpuscles. However, some of the intraperineural lamellated corpuscles exhibited interlaced arrangements of tortuous axon terminals and cytoplasmic lamellae resembling the arrangement in Meissner corpuscles. Immunohistochemical staining using anti-neuron specific enolase also revealed the presence of intraperineural, Meissner-like corpuscles in the cat lip. This study indicates that intraperineural localization of lamellated corpuscles is common in carnivora and artiodactyla and that the intraperineural lamellated corpuscles are heterogeneous in their ultrastructural pattern.

Localization of Signal Transduction Proteins in the Merkel Cell Axon Complex

The function of the Merkel cell (MC) is still enigmatic. Immunohistochemically, we investigated the localization of signal transduction proteins in MC-axon complexes. Axon terminals connected to MCs in the skin and oral mucosa of the rat and monkey showed positive immunoreactivities for Gαo, Gαi-1 and Gαs, whereas Gαi-, Gαo-, Gαq-, Gαs- and Gαz-like immunoreactions were localized on MCs of both animals. Since Gαq is known to couple with group 1 metabotropic glutamate receptors and activate beta-type phospholipase C, we immunohistochemically investigated the localization of these substances in rat MCs. It was found that MCs express mGluR5-like immunoreactions. However, intensely positive immunoreactions for PLCs4 were found to be localized in type 1 sensory nerve terminals connected to MCs, but not in MCs themselves.

Effects of trifluoperazine and amiloride on the extrusion of axoplasmic Ca++ by rapidly adapting and slowly adapting type I cutaneous mechanoreceptors: an electron-microscopic study using the oxalate-antimonate method.

Influences of Ca++ transport inhibitors on the extrusion of cytosolic Ca++ from axon terminals of oral mucosal mechanoreceptors were cytochemically studied through the use of trifluoperazine (TFP), a Ca++ pump inhibitor, and amiloride, a Na+/Ca++ exchanger inhibitor. Palates of Mongolian gerbils were isolated after perfusion with normal, TFP-containing, or amiloride-containing HEPES-buffered saline, and then mechanically stimulated in the same buffer. Cytosolic Ca++ in axon terminals at 2 min after the stimulation was visualized by means of an oxalate-antimonate method using microwave fixation, and then evaluated electron-microscopically. TFP was found to inhibit the Ca++ extrusion from axon terminals of Meissner corpuscles, but not from those of Merkel cell-axon complexes. Conversely, amiloride severely reduced the Ca++ extrusion from axon terminals of Merkel cell-axon complexes, but not from those of Meissner corpuscles. These results suggest that cytosolic Ca++ extrusion from axon terminals of rapidly adapting and slowly adapting type I mechanoreceptors is regulated by a Ca++ pump and an Na+/Ca++ exchanger, respectively.

Immunohistochemical and cytochemical evidences for a possible localization of leucine aminopeptidase in the Merkel cell granule

Localization of leucine aminopeptidase (LAP) in the Merkel cell-axon complex was studied immunohistochemically and cytochemically in the labial tissues of the mouse, rat, dog, and monkey. Anti-LAP was obtained in rabbits by the injection of commercially supplied swine LAP which was confirmed as electrophoretically pure. The Merkel cells of the mouse, rat, and monkey were positively stained by treatment with anti-LAP but the Merkel cells of the dog were negative. When ultrathin sections of the hair follicle from the rat whisker pad, which contain an abundance of Merkel cells, were processed by immuno-peroxidase or by the immuno-gold method, the reaction products were predominantly deposited on the Merkel cells granules. Furthermore, an immuno-blot assay revealed that an extract of the hair follicles from murine whisker pads contained a molecule of relative molecular mass Mr = 60,000 which is similar in size to a subunit of swine LAP. Thus, it appears that Merkel cell granules of rodents and the monkey contain a protein which resembles lucine aminopeptidase.