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Biomedical subjects

M Tohyama

Publications and source records attributed to M Tohyama.

At least 199 records · Page 11Linked to original sources
PubMed1994

Effect of dibutyryl cyclic AMP on morphologic features and marker production of human cervical argyrophil small-cell carcinoma cell line.

The effect of dibutyryl cyclic AMP (dB-cAMP) on the morphologic features and marker production of a human cervical argyrophil small-cell carcinoma (ASCC) cell line was examined. Following 1-5 days' exposure to 5 mM dB-cAMP, morphologic differentiation as defined by the expression of cytoplasmic processes (stellate cells) was observed. The number of stellate cells depended on the dose of dB-cAMP and incubation time. Shortly after removal of dB-cAMP from the culture medium, the treated cells returned to their original spherical shape. dB-cAMP caused a reduction in the growth rate of cells which recovered after removal of the agent. The morphological changes appeared not to be the result of growth inhibition by dB-cAMP, because the cells maintained in a serum-free medium did not show any change in shape. Electron microscopic study revealed the development of intracytoplasmic microtubules, microfilaments, and an increase in the number of neurosecretory granules in the treated cells. The levels of neuron-specific enolase, serotonin and gastrin in treated cells were significantly higher than those in untreated controls. These findings indicate that a reversible differentiation of cultured ASCC cells into neuroendocrine cells occurs in a growth medium containing dB-cAMP.

Biomarkers, Tumor↗
PubMed1994

Genes for members of the GATA-binding protein family (GATA-GT1 and GATA-GT2) together with H+/K(+)-ATPase are specifically transcribed in gastric parietal cells.

mRNAs for novel DNA-binding proteins (GATA-GT1 and GATA-GT2) recognizing the (G/C)PuPu(G/C)NGAT(A/T)PuPy sequence and H+/K(+)-ATPase (proton pump) alpha subunit were detected in parietal cells of the rat gastric body mucosa by in situ hybridization. These results suggest that GATA-GT1 and GATA-GT2 together with H+/K(+)-ATPase are transcribed specifically in gastric parietal cells and that the two DNA-binding proteins may have important roles in cell specific gene regulation. Furthermore, we could detect parietal cells in different states of gene expression.

Animals↗
PubMed1994

Two-dimensional demonstration of myenteric nerve plexus: application for pseudo-Hirschsprung's disease.

The use of two-dimensional demonstration of myenteric nerve plexus for the morphological estimation of pseudo-Hirschsprung's disease is introduced. By fluorescent immunohistochemistry for S-100 protein on flat-mounted frozen sections, a decrease in the amount of ganglionic plexuses in the small intestinal segments in cases of pseudo-Hirschsprung's disease was able to be distinguished from normoganglionic plexuses in control cases. Moreover, a morphological difference between the two types of cases was clearly demonstrated. This method may be an indispensable tool in the reliable diagnosis of pseudo-Hirschsprung's disease.

Female↗
PubMed1994

Distribution of corticotropin-releasing factor and calcitonin gene-related peptide in the developing mouse cerebellum.

Corticotropin-releasing factor (CRF)-like immunoreactive (IR) fibers were investigated ontogenically in the mouse cerebellum. CRF-IR was detected in the climbing fiber and mossy fibers as in other species. In addition, CRF-IR dense fiber plexuses were detected from postnatal day (PD) 2 to 9, in the developing Purkinje cell layer of the vermal lobules, paraflocculus, flocculus and crus 1 ansiform lobule, gradually forming a pericellular nest around the Purkinje cell somata. Immunoelectron-microscopical analysis showed that dense fibers made synaptic contacts with the Purkinje cell somata on PD 7. In the lobules mentioned above, CRF-IR dense fibers showed parasagittal banded patterns. Calcitonin gene-related peptide (CGRP)-IR showed similar fiber bands at these stages. Interestingly, these two patterns of peptidergic fiber bands were complementary in distribution. From around PD 9, CRF-IR fibers lost the immunoreactive dots in the Purkinje cell layer. Immunoreactivity at this stage was observed in the axons projecting to the molecular layer, and thin CRF-IR fibers began to appear in the neighboring area. Numerous typical climbing fiber-like CRF-IR fibers were found throughout the cerebellar cortex from PD 16 to adult. The inferior olivary complex (the origin of climbing fibers) appears to be the origin of these dense fiber plexuses as CRF-IR cells were already present from PD 2 in the dorsal cap nucleus, beta subnucleus and caudomedial part of the accessory olivary nucleus. No neurons containing both CRF and CGRP immunoreactivities were observed. These results suggest that CGRP- and CRF-IR developing climbing fibers innervate different compartments of Purkinje cells, especially in the vestibular cerebellar cortex in mice. Furthermore, CRF-IR fibers gradually changed to become typical climbing fibers, while CGRP-IR disappeared altogether.

Aging↗
PubMed1994

Marked increase in glutamate-aspartate transporter (GLAST/GluT-1) mRNA following transient retinal ischemia.

We have demonstrated the cellular localization of glutamate-aspartate transporter (GLAST/GluT-1) mRNA in the rat retina and its induction after ischemia by in situ hybridization. GLAST mRNA was expressed in the inner two-thirds of the inner nuclear layer (INL) and in sparse small cells in the inner portion of the ganglion cell layer (GCL) of the adult rat retina. GLAST mRNA was also found in about 90% of cells in the optic nerve head where more than 90% of cells express glial fibrillary acidic protein (GFAP) mRNA. Moreover, experimental occlusion of the central retinal artery followed by reperfusion for 48 h resulted in degeneration of neurons and a marked increase in GLAST mRNA expression in the INL. These findings suggest that GLAST may be expressed in Müller cells and astrocytes in the retina, and may play an important role in regulation of extracellular glutamate concentration especially under ischemic conditions.

Amino Acid Transport System X-AG↗
PubMed1994

Differential expression of sgk mRNA, a member of the Ser/Thr protein kinase gene family, in rat brain after CNS injury.

We cloned genes the expression of which were induced 3 days after cortical injury of rat brain by a differential display technique, and four novel and known sequences were isolated. Among these sequences, the sgk gene which was recently identified as a novel member of the serine/threonine protein kinase gene family, was selected for analysis of its expression patterns in rat brain by northern blotting and in situ hybridization, because hybridization signals were strong at the lesion sites. Expression of sgk mRNA was induced within 3 days after injury, and was maintained at a high level for at least 14 days. The cells which strongly expressed the sgk gene were in the deep layers of the cortex and in the corpus callosum. In situ hybridization analysis for sgk and myelin proteolipid protein mRNA using serial sections showed that the distribution of both signals was very similar at the damaged regions. Therefore, it is likely that the sgk transcript is expressed by oligodendrocytes after brain injury. Investigation of the developmental expression of the sgk gene showed that neurons in layers I and II of the cortex, lateroposterior and laterodorsal thalamic nucleus, and ventral posterolateral and posteromedial thalamic nucleus strongly expressed sgk mRNA at postnatal day 1 and day 7, but these neurons showed no expression in fetal or adult brain. These results suggest that the induction of sgk gene may be associated with a series of axonal regenerations after brain injury, and in addition, the sgk gene may also play important roles in the development of particular groups of neurons in the postnatal brain.

Aging↗
PubMed1994

Cloning and expression of a neural differentiation-associated gene, p205, in the embryonal carcinoma cell line P19 and in the developing mouse.

Mouse P19 embryonal carcinoma cells can be reproducibly differentiated into neurons and glial cells upon treatment with high concentration of retinoic acid (RA). In order to understand the molecular mechanisms that control early neural differentiation, we screened a cDNA library made from 24-h RA-treated P19 cells with subtracted cDNA probes. One clone was positive in the secondary screening and was designated as p205. This clone (1.1 kb) has an open reading frame of 317 amino acids with homology to G-protein beta subunit. This protein sequence was identical to chicken and human genes previously identified as a major histocompatibility complex-associated gene. The complete conservation of its amino acid sequence between mouse, human and chicken provides strong evidence that the p205 protein fulfills a fundamental function. Developmental Northern blot analysis revealed that a p205 mRNA is expressed at high levels in the embryonic mouse brain, decreasing as development proceeds. In situ hybridization revealed that p205 mRNA is strongly and ubiquitously expressed in the embryonic and early postnatal mouse brain. This expression decreased during postnatal development and was localized in the dentate gyrus, habenula, piriform cortex, paraventricular nucleus of the hypothalamus and supraoptic nucleus of the adult brain. These results suggest that this protein plays an important role in the developing brain and neuronal differentiation.

Aging↗
PubMed1994

Coexpression of GABAA receptor gamma 1 and gamma 2 subunits in the rat trigeminal ganglion.

We examined the expression of gamma-aminobutyric acid (GABA)A receptor gamma 1 and gamma 2 subunit mRNAs in the rat trigeminal ganglion using in situ hybridization histochemistry. Most ganglion cells expressed both gamma 1 and gamma 2 mRNAs simultaneously. These findings are a marked contrast to the findings in the central nervous system where areas expressing both subunits are rare. In addition, we demonstrated using immunohistochemistry that the gamma 1 subunit is also expressed at the protein level in trigeminal ganglion neurons and fibers in the trigeminal spinal nucleus.

Animals↗
PubMed1994

Messenger RNA and protein expression of basic fibroblast growth factor receptor after cortical ablation.

In a previous report we demonstrated that basic fibroblast growth factor (bFGF), as a multipotent neurotrophic factor, could prevent retrograde degeneration of the thalamic neurons after ablation of the somatosensory cortex. To elucidate the mechanism of this bFGF action, we examined changes in FGF receptor (FGFR) mRNA (flg) expression with in situ hybridization. The FGF receptor protein was detected with the immunoblotting method. The FGFR mRNA expression was found to be diffusely increased in the affected cortex. Microscopic observation indicated that FGFR mRNA was expressed in several types of cortical cells including neurons and non-neuronal cells. This increase could be observed as early as 6 hours after surgery and lasted for 48 hours. In the thalamus, however no change in FGFR mRNA signals was observed. Western blotting detected a protein immunoreactive to anti-FGFR antibody. Samples from the periablated cortex showed an increase in FGFR protein. Samples from the thalamus, however, showed no difference in FGFR protein level between the lesion side and the contralateral side. Application of exogenous bFGF in Gelfoam to the cortical ablation cavity did not show any effect on the gene expression or protein level of FGFR. These results suggest that FGFR is diffusely induced throughout the injured cortex in the early phase after injury and that bFGF may play an important role after injury. Topically applied bFGF might thus modulate cellular responses in the cortex and have a neurotrophic effect on the affected thalamic neurons.

Animals↗
PubMed1994

GAP-43 (B50/F1) gene regulation by axonal injury of the hypoglossal nerve in the adult rat.

The expression of mRNA encoding the growth associated protein, GAP-43, was investigated in rat hypoglossal motor neurons when the hypoglossal nerve was either resected or crushed unilaterally. For the detection of GAP-43 mRNA, a histochemical in situ hybridization method. using an alkaline phosphatase labeled probe, was used. The temporal profiles of GAP-43 mRNA expression were not identical following the two types of injuries. Increased expression in the hypoglossal nucleus contralateral to the injured nerve was observed from 1 day to 4-6 weeks after nerve crush, but lasted up to 7-8 weeks after resection. The magnitude and duration of increased GAP-43 mRNA expression were significantly greater following resection than crush injury. Local treatment with vinblastine, which is known to disturb the fast axonal flow by depolymerizing tubulin, also induced GAP-43 mRNA expression. The patterns of gene regulation following these nerve injuries may be due to the extent of nerve damage, to tubulin disturbance, or to some other factors derived from outside the nerve.

Animals↗
PubMed1994

Marginal topography of neurons expressing the substance P receptor in the rat suprachiasmatic nucleus.

Neurons expressing the substance P (SP) receptor (NK1 receptor) in the suprachiasmatic nucleus of the hypothalamus (SCN) have been topographically identified using radioactive in situ hybridization histochemistry. In the anterior hypothalamic area, clustered labeled neurons of small size and exhibiting low levels of gene expression are observed exclusively at the dorsolateral margin of the SCN, straddling cytoarchitectural boundaries of the nucleus. The marginal topography of neurons putative target of a SP-containing retinal input to the ventral SCN indicates that their dendrites bearing the receptor extend towards the retinorecipient part of the nucleus, where they can be modulated by overlapping inputs from the intergeniculate leaflet and the raphe. Eventual interactions between glutamatergic and putative tachykininergic retinal pathways for a coherent photic control of circadian rhythms may therefore occur mainly via intrinsic neuronal connections between their distinct target populations. In addition, since glutamate and SP induce electrophysiological responses in ventrolateral neurons with no interactive effect, neurons integrating both chemical signals, subsequently to their modulation by several influences, may be not located within the ventrolateral SCN. Alternatively but not exclusively, marginal neurons could be a target of SP-containing neurons within the SCN or nearby the nucleus, or from ascending projections from the raphé where serotonin and SP colocalize. The marginal topography of neurons expressing the SP receptor supports the view of the involvement of neurons located in the vicinity of the nucleus in the regulation of circadian rhythms.

Animals↗
PubMed1994

Differential regulation of phospholipase C isozymes in the rat facial nucleus following axotomy.

The expression of phospholipase C isozymes and phosphatidylinositol 4-kinase in the rat facial nucleus was studied using in situ hybridization at various times after unilateral crushing and resectioning the facial nerve. The level of phospholipase C alpha messenger RNA increased from three days to one week after the operation. On the other hand, an apparent reduction in the level of phospholipase C beta 1 occurred from three days to one week after resection. After either crushing or resection, phospholipase C gamma 1 messenger RNA levels were not noticeably changed. As phosphatidylinositol 4-kinase is the rate-limiting enzyme for the production of phosphatidylinositol 4,5-bisphosphate, which is the preferred substrate for phospholipase C, we investigated the expression of phosphatidylinositol 4-kinase messenger RNA. The level of phosphatidylinositol 4-kinase messenger RNA was decreased one day after axonal injury. Among phospholipase C isozymes, phospholipase C alpha is up-regulated. As the structure of phospholipase C alpha is different from other isozymes, phospholipase C alpha is supposed to have a different function. The present unique up-regulation of phospholipase C alpha may suggest a novel function in nerve regeneration. Phospholipase C beta 1 is down-regulated, as is phosphatidylinositol 4-kinase. This suggests that the signal transmission system using a G-linked receptor is broken down after nerve injury. On the other hand, phospholipase C gamma 1, which is related to the receptor tyrosine kinase, does not demonstrate any transcriptional regulation after nerve injury.

1-Phosphatidylinositol 4-Kinase↗
PubMed1994

Projections of tachykinin- and glutaminase-containing rat retinal ganglion cells.

Glutamate (Glu) and the tachykinin substance P (SP) have been proposed as neurotransmitters or neuromodulators of the retinal projection to the brain. In the present study, we demonstrate that tachykinin-like (TK) immunoreactivity (IR) accumulates in rat retinal axons following electrical lesions to the optic tract, indicating that SP is conveyed in the optic nerve to its central targets. In addition, we show that eye enucleation causes a dramatic decrease in TK-IR fibers in the pretectal olivary nucleus (PON), but not in other retinorecipient nuclei of the thalamus and the midbrain, and that Fluorogold injected into the pretectum is retrogradely transported to the somata of TK-IR retinal ganglion cells (RGCs), indicating an important projection of TK-IR RGCs to the PON. We also show that most rat RGCs are labeled with antibodies against phosphate-activated glutaminase, an enzyme considered to generate the transmitter pool of glutamate. Unlike TK-IR fibers, phosphate-activated glutaminase-IR structures disappear in most retinorecipient nuclei following eye enucleation. The present results give neuroanatomical support to the idea that glutamate is a neurotransmitter in the retinal projection and suggest an important role for TK-IR RGCs in the relay of visual information to the PON.

Animals↗
PubMed1994

Distribution of cocaine recognition sites in rat brain: in vitro and ex vivo autoradiography with [125I]RTI-55.

The distribution of binding sites of [125I]RTI-55 (3 beta-(4-iodophenyl)tropan-2 beta-carboxylic acid methyl ester), a phenyl tropane analog of cocaine, and the selective labeling of the dopamine transporter (DAT) were studied by in vitro and ex vivo autoradiography in the rat whole brain. Recent evidence has shown that RTI-55 binds to not only DAT but also serotonin transporter (5HTT). In the present study, in vitro autoradiography revealed that [125I]RTI-55 bound to the olfactory tubercle, the caudate putamen, the accumbens nucleus, the midline and lateral geniculate nuclei of the thalamus, the hypothalamic nuclei, the substantia nigra compact part, the subthalamic nucleus, the ventral tegmental area, the superior colliculus, the dorsal raphe nucleus, and the facial nucleus. Further, in the presence of clomipramine, a selective ligand for 5HTT, [125I]RTI-55 binding was remarkably inhibited in the midline and lateral geniculate nuclei of the thalamus, the hypothalamic nuclei, the superior colliculus, the dorsal raphe nucleus, and the facial nucleus, while [125I]RTI-55 binding remained in the olfactory tubercle, the caudate putamen, the accumbens nucleus, the substantia nigra compact part, the subthalamic nucleus, and the ventral tegmental area. These findings suggest that [125I]RTI-55 binds to 5HTT in the former areas and to DAT in the latter areas. It is therefore concluded that RTI-55 is a suitable ligand for studying the action of cocaine in whole brain regions, including the thalamus, the hypothalamus and the dorsal raphe nucleus, regions in which cocaine is thought to act evoking several neurological effects, e.g., analgesia and elevation of adrenocorticotropic hormone. DAT was also labelled selectively both in vitro and in vivo using [125I]RTI-55 combined with clomipramine. Therefore, radiolabelled RTI-55, combined with unlabelled clomipramine, which displaces its binding to 5HTT, also appears to be suitable for the selective imaging of DAT in vivo.

Animals↗
PubMed1994

Growth factors and extracellular signal-regulated kinases (mitogen-activated protein kinase) in the rat pineal gland.

The present study revealed that the pineal gland expressed basic fibroblast growth factor (bFGF) and FGF-receptor 1 (FGFR1/flg), suggesting that bFGF in the pineal gland acts in an autocrine or paracrine manner, which is mediated by FGFR1/flg. The present study also examined gene expression of the extracellular signal-regulated kinase (ERK) family (ERK1-3) which may be intracellular signal mediators of growth factors. ERK1 [mitogen-activated protein kinase (MAP-kinase)] was strongly expressed throughout the pineal gland, while expression of ERK2 and ERK3 was not found. These findings suggest the presence of a signal pathway from bFGF to ERK1 via FGFR1/flg in the pineal gland.

Animals↗
PubMed1994

Localization of perforin in viral vesicles and erythema multiforme.

BACKGROUND: Perforin (Pf), a pore-forming protein, is a cytolytic protein of killer cells. Its deposition in lesioned skin has not been studied. OBJECTIVE: The purpose of this study is to show Pf deposition in the lesioned skin and Pf expression in the dermal infiltrates of various inflammatory skin diseases. METHODS: Frozen specimens obtained from 29 patients with 5 different diseases were immunohistochemically stained. RESULTS: Granular deposition of Pf was found in the lesioned skin in 2 out of 5 cases of viral vesicles and in 3 out of 7 cases of erythema multiforme. In the cases with Pf deposition, the percentages of Pf+ cells in the dermis were higher than in those without deposition. CONCLUSION: Pf released from natural killer cells or cytotoxic T lymphocytes may play a role in tissue damage.

Adolescent↗
PubMed1994

Electron microscopic observation of bFGF immunoreactivity in the hippocampus.

Light and electron microscopic observation showed two types of neuronal immunostaining for basic fibroblast growth factor in the hippocampal CA2 subfield, where the densest immunoreactive neurons were localized in the brain. One neuronal type showed intense nuclear (eu- and heterochromatin) immunostaining but weak cytoplasmic immunostaining (N-type), and the other showed intense cytoplasmic but no or only faint nuclear immunoreactivity (C-type). The N-type also showed weak immunoreactivity in the perinuclear rough endoplasmic reticulum and contained bFGF mRNA as observed by in situ hybridization histochemistry, showed that this type can produces the bFGF protein. The N-type localized exclusively in the CA2 subfield. The C-type showed strong immunoreactivity on the rER, free ribosomes, and Golgi apparatus, although no clear evidence for bFGF production was observed. The multivesicular bodies, a pathway of endocytosis in hippocampal neurons showed apparent immunoreactivity under EM observation of both of types neurons (Parton et al. J. Cell Biol. 119: 123-137, 1992) suggesting a receptor-mediated type of incorporation of the bFGF.

Amino Acid Sequence↗