Morphological and functional peculiarities of mesenchymal cells in the pars tuberalis of the pituitary gland

Summary Following injection of high doses of horseradish peroxidase (HRP), mesenchymal cells distributed in the perisinusoidal space of the pars tuberalis of the hypophysis in cats, rabbits and Japanese quails, sequester the exogenously administrated peroxidase intensively. These cells are designated by the authors as horseradish peroxidase-uptake cells (HRP-uptake cells or HUC). HRP-uptake cells constitute a system of macrophages in the pars tuberalis of mammals and birds, and are located around the hypophysial portal veins. HRP-uptake cells differ in morphological and functional characteristics from similar cells in other parts of hypophysis. They are thought to play a role in the hypothalamic control of adenohypophysial secretion.Supported by grants (No. 144022, 237002) from the Ministry of Education, Science and Culture, Japan     

Morphological studies on neuroglia

Summary The silver-impregnation procedure of Tsujiyama is suitable for demonstration of all three classical types of neuroglial cells; in the present study it was used for electron microscopic identification of neuroglial cells in the brain of the cat. The aim of the present study was 1) to determine impregnated structural correlates of neuroglial cells at the light- and electron-microscopic levels, and 2) to determine whether the method of Tsujiyama is applicable for the electron microscopic identification of the single types of neuroglial cells. Silver deposits were observed over the cytoplasm and processes of astrocytes where numerous glial filaments were present. Oligodendrocytes and microglial cells may be precisely differentiated by use of Tsujiyama's silver impregnation method at the electron microscopic level due to the pattern of silver-deposition in these two basic types of cells. This silver-impregnation method combined with electron microscopy is thus suitable for a precise identification of neuroglial cells; the technique may prove to be very helpful in identification of such categories of neuroglial cells that encompass also the images of cells which cannot be classified by use of the standard methods.Supported by a grant (No. 437002) from the Ministry of Education, Science and Culture, Japan     

Thiaminepyrophosphatase activity in the plasma membrane of microglia

An intense thiaminepyrophosphatase (TPPase) activity was demonstrated in glial cells and blood vessels in the central nervous system (CNS), when incubation was carried out with thiaminepyrophosphte (TPP, cocarboxylase), using the method of Novikoff and Goldfischer (1961). Glial cells with TPPSase activity were identified as microglia because they were morphologically similar to microglial cells in the sections stained with silver impregnation. TPPase activity was localized in the microglial perikaryon and in the processes, as viewed under a light microscope. Electron microscopically, enzyme activity was localized in the plasma membrane of microglia. We consider this activity to be a true TPPase activity hydrolyzing TPP, and we then went on to examine the substrate specificity, optimum pH, effect of chemical inhibitors and activators, and the effect of glutaraldehyde fixation. Our data are reported herein.     

Cells capable of uptake of horseradish peroxidase in some circumventricular organs of the cat and rat

Summary The structure of mesenchymal cells distributed in some of the hypendymal organs of the circumventricular system in the cat and rat was demonstrated after intravenous injection of high doses of horseradish peroxidase. These cellular elements were observed in the vicinity of blood vessels of the organon vasculosum laminae terminalis, subfornical organ and area postrema. Electron-microscopically, these cells located between the basal laminae of the brain parenchyma and the blood capillaries show long cellular processes encircling fenestrated capillaries. Light and electron-microscopic examination revealed that this cell type is identical with the horseradish peroxidase-uptake cells, previously reported in the vicinity of the hypophysial portal system. Such phagocytic cells may be considered as a cellular component intervening between the brain parenchyma and the blood stream, playing a role in selective barrier functions in the above-mentioned circumventricular organs where a blood-brain barrier in the classical sense of the definition is lacking.This work was supported by grant No. 437002 from the Ministry of Education, Science and Culture, Japan     

Morphological studies on neuroglia

Summary Electron-microscopic survey of selectively stained microglial cells in the cerebral cortex of the rat reveals that the processes of this cell type often encircle axo-dendritic synapses. Enzyme-histochemical methods for thiamine pyrophosphatase (TPPase) or nucleoside diphosphatase (NDPase) were used for the selective marking of the microglial cells; TPPase and NDPase activities were observed in the plasma membrane of microglial cells. The synapses encircled by microglial processes displayed presynaptic structures containing round clear vesicles (50 nm in diameter) and a prominent thickening of the postsynaptic membrane. In vitro, the above-mentioned enzymatic activities were completely suppressed by neuroactive agents such as catecholamines and phenothiazine derivatives. Examination using enzyme-histochemical techniques suggests that a single enzyme may be responsible for both above-mentioned enzymatic reactions. The functional significance of microglial cells in the normal central nervous tissue is discussed.This work was supported by grant No. 437002 from the Ministry of Education, Science and Culture, Japan     

Adhesive papillae on the brachiolar arms of brachiolaria larvae in two starfishes, Asterina pectinifera and Asterias amurensis, are sensors for metamorphic inducing factor(s)

It has been hypothesized by Barker that starfish brachiolaria larvae initiate metamorphosis by sensing of metamorphic inducing factor(s) with neural cells within the adhesive papillae on their brachiolar arms. We present evidence supporting Barker's hypothesis using brachiolaria larvae of the two species, Asterina pectinifera and Asterias amurensis. Brachiolaria larvae of these two species underwent metamorphosis in response to pebbles from aquaria in which adults were kept. Time-lapse analysis of A. pectinifera indicated that the pebbles were explored with adhesive papillae prior to establishment of a stable attachment for metamorphosis. Microsurgical dissections, which removed adhesive papillae, resulted in failure of the brachiolaria larvae to respond to the pebbles, but other organs such as the lateral ganglia, the oral ganglion, the adhesive disk or the adult rudiment were not required. Immunohistochemical analysis with a neuron-specific monoclonal antibody and transmission electron microscopy revealed that the adhesive papillae contained neural cells that project their processes towards the external surface of the adhesive papillae and they therefore qualify as sensory neural cells.     

Gadd45a, the gene induced by the mood stabilizer valproic acid, regulates neurite outgrowth through JNK and the substrate paxillin in N1E-115 neuroblastoma cells

Valproic acid (VPA), a mood stabilizer and anticonvulsant, has a variety of neurotrophic functions; however, less is known about how VPA regulates neurite outgrowth. Here, using N1E-115 neuroblastoma cells as the model, we show that VPA upregulates Gadd45a to trigger activation of the downstream JNK cascade controlling neurite outgrowth. VPA induces the phosphorylation of c-Jun N-terminal kinase (JNK) and the substrate paxillin, while VPA induction of neurite outgrowth is inhibited by JNK inhibitors (SP600125 and the small JNK-binding peptide) or a paxillin construct harboring a Ser 178-to-Ala mutation at the JNK phosphorylation. Transfection of Gadd45a, acting through the effector MEKK4, leads to the phosphorylation of the JNK cascade. Conversely, knockdown of Gadd45a with siRNA reduces the effect of VPA. Taken together, these results suggest that upregulation of Gadd45a explains one of the mechanisms whereby VPA induces the neurotrophic effect, providing a new role of Gadd45a in neurite outgrowth.     

Estimation of the embryotoxic effect of CBZ using an ES cell differentiation system

Carbamazepine (CBZ) is one of the most commonly prescribed antiepileptic drugs (AEDs). However, a higher rate of congenital anomalies has been found in infants of mothers treated with CBZ during early pregnancy. Here, we characterize the effects of CBZ using a mouse ES cell differentiation system. The analysis of tissue-specific gene markers showed that CBZ induced early endodermal and mesodermal differentiation but inhibited differentiation of later stages. CBZ also induced ectodermal development, and there was evidence of neural differentiation as ES cells with an immature neuronal phenotype were observed. In contrast, valproic acid (VPA), another anticonvulsant drug, was previously shown to be able to induce ES cells to differentiate into neurons with a mature appearance. CBZ was less cytotoxic to ES cells than VPA. The in vitro ES cell assay system has the potential to provide a rapid and accurate approach for estimating the in vivo embryotoxicity of therapeutic drugs.     

Nervous system development of the sea cucumber Stichopus japonicus

The nervous system development of the sea cucumber Stichopus japonicus was investigated to explore the development of the bilateral larval nervous system into the pentaradial adult form typical of echinoderms. The first nerve cells were detected in the apical region of epidermis in the late gastrula. In the auricularia larvae, nerve tracts were seen along the ciliary band. There was a pair of bilateral apical ganglia consisted of serotonergic nerve cells lined along the ciliary bands. During the transition to the doliolaria larvae, the nerve tracts rearranged together with the ciliary bands, but they were not segmented and remained continuous. The doliolaria larvae possessed nerves along the ciliary rings but strongly retained the features of auricularia larvae nerve pattern. The adult nervous system began to develop inside the doliolaria larvae before the larval nervous system disappears. None of the larval nervous system was observed to be incorporated into the adult nervous system with immunohistochemistry. Since S. japonicus are known to possess an ancestral mode of development for echinoderms, these results suggest that the larval nervous system and the adult nervous system were probably formed independently in the last common ancestor of echinoderms.     

Morphological studies on neuroglia

The postnatal development of microglial cells was investigated in the neonatal rat brain by use of light- and electron microscopy, including enzyme-histochemical techniques. Microglial cells were selectively stained by demonstration of their nucleoside diphosphatase (NDPase) activity and classified into three types: 1) In the early postnatal period "primitive microglial cells" showing scantily ramified processes were found in the cerebral cortex, the hippocampal formation, and the hypothalamus. During the course of the first postnatal week the processes of this cell type developed gradually and the cells were transformed into typical ramified microglial cells, called "resting microglial cells". 2) "Amoeboid microglial cells "showing typical features of macrophages were characteristic of the cerebral white matter. 3) "Round microglial cells" possessing a round soma and few pseudopodia but no characteristic processes occurred in large numbers in the subventricular zone of the lateral ventricle and as single elements in the vicinity of blood vessels. Histochemically, thiamine pyrophosphatase (TPPase) was demonstrated only in the fully developed, ramified microglial cells ("resting microglial cells"), which could be readily observed in the central nervous tissue from the age of 14 day. "Round and amoeboid microglial cells" did not show TPPase activity and disappeared after 14 days of postnatal life. By use of electron microscopy, in neonatal rats NDPase activity was apparent in the plasma membrane of the three types of microglial cells ("primitive, round, and amoeboid" types). They showed basically similar submicroscopic characteristics, i.e., well-developed Golgi apparatus, long strands of rough-surfaced endoplasmic reticulum, single dense bodies and vacuoles, and numerous ribosomes. "Amoeboid microglial cells" were characterized by their well-developed cytoplasmic vacuoles and phagocytic inclusion bodies. The present study strongly suggests a mesodermal origin for these microglial elements.