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Biomedical subjects

S Shiosaka

Publications and source records attributed to S Shiosaka.

At least 19 recordsLinked to original sources
PubMed2001

Expression of the kallikrein gene family in normal and Alzheimer's disease brain.

The human kallikrein gene family consists of 15 serine proteases. We examined the expression of the kallikrein genes in human cerebral cortex and hippocampus by RT-PCR and compared their expression between Alzheimer's disease (AD) and control tissue. KLK1, 4, 5, 6, 7, 8, 10, 11, 13 and 14 are expressed in both cerebral cortex and hippocampus. KLK9 is expressed in cortex but not hippocampus, whereas KLK2, 3, 12 and 15 are not expressed in either tissue. We demonstrate an 11.5-fold increase in KLK8 mRNA levels in AD hippocampus compared to controls. The KLK8 gene product, neuropsin, processes extracellular matrix and is important for neuronal plasticity. Therefore, the increase in KLK8 could have detrimental effects on hippocampal function in AD.

Aged↗
PubMed2001

Dendritic aberrations in the hippocampal granular layer and the amygdalohippocampal area following kindled-seizures.

Amygdaloid kindling is a model of human temporal lobe epilepsy, in which excitability in limbic structures is permanently enhanced by repeated stimulations. We report here dendritic aberrations occurring in mice following kindled-seizures. Adult mice received a biphasic square wave pulse [495+/-25.5 (S.E.M.) microA 60 Hz, 200 micros duration, for 2 s] unilaterally in the basolateral amygdaloid complex once a day and mice with electrophysiologically and behaviorally verified seizures were used in the experiments. The hippocampus and amygdaloid complex contralateral to the lesions were observed by immunofluorescence histochemistry with a somatodendritic marker, microtubule-associated protein 2 (MAP2), showing that kindled-seizures caused hypertrophy of proximal dendrites in the granule cells of the dentate gyrus and in neurons of the amygdalohippocampal area. To further characterize the morphological changes of the dendrites, electron micrographic analysis was performed on the contralateral side. (1) In the granular layer of the dentate gyrus and the amygdalohippocampal area, kindled-seizures generated an increase in the number of dendrites containing polymerized microtubules and width of dendritic profiles showing the increase was in the range 0.2-3.0 and 0.2-1.4 microm, respectively. (2) In the granular layer, bundles between dendrites separated by the puncta adhaerentia increased. (3) In the granular layer, the seizure-induced dendritic aberration was more severe in the rostral than the caudal region. These results suggested that growth of dendrites with enriched-stable microtubules is part of the structural plasticity in response to seizure activity in specific areas of the adult brain.

Amygdala↗
PubMed2001

Serine proteinase inhibitor 3 and murinoglobulin I are potent inhibitors of neuropsin in adult mouse brain.

Extracellular serine protease neuropsin (NP) is expressed in the forebrain limbic area of adult brain and is implicated in synaptic plasticity. We screened for endogenous NP inhibitors with recombinant NP (r-NP) from extracts of the hippocampus and the cerebral cortex in adult mouse brain. Two SDS-stable complexes were detected, and after their purification, peptide sequences were determined by amino acid sequencing and mass spectrometry, revealing that target molecules were serine proteinase inhibitor-3 (SPI3) and murinoglobulin I (MUG I). The addition of the recombinant SPI3 to r-NP resulted in an SDS-stable complex, and the complex formation followed bimolecular kinetics with an association rate constant of 3.4 +/- 0.22 x 10(6) M(-1) s(-1), showing that SPI3 was a slow, tight binding inhibitor of NP. In situ hybridization histochemistry showed that SPI3 mRNA was expressed in pyramidal neurons in the hippocampal CA1-CA3 subfields, as was NP mRNA. Alternatively, the addition of purified plasma MUG I to r-NP resulted in an SDS-stable complex, and MUG I inhibited degradation of fibronectin by r-NP to 24% at a r-NP/MUG I molar ratio of 1:2. Immunofluorescence histochemistry showed that MUG I localized in the hippocampal neurons. These findings indicate that SPI3 and MUG I serve to inactivate NP and control the level of NP in adult brain, respectively.

Animals↗
PubMed2001

Abnormalities of synapses and neurons in the hippocampus of neuropsin-deficient mice.

In the present study, we produced null-mutant mice of neuropsin, an extracellular matrix serine protease, to examine the neural functions of this protein particularly in the hippocampus. Golgi-Cox impregnation and Nissl-staining revealed morphological change of cell soma in the mutant mice compared to wild-type mice. However, Golgi-Cox impregnation revealed no apparent change in the dendritic arborization and spine density. Quantitative electronmicroscopic analysis revealed that number of asymmetrical synapses were significantly decreased in the stratum radiatum, the major terminal field of Schaffer-collaterals, whereas free boutons still holding synaptic vesicles but with no synaptic specialization were increased in number in the same microscopic fields. An increased number of parvalbumin-immunoreactive cells (known as fast spiking cells) in mutant was also observed. These results strongly suggest that neuropsin is involved in connectivity of a group of CA1 synapses and consequently in the hippocampal networking.

Animals↗
PubMed2001

Effects of fibronectin cleaved by neuropsin on cell adhesion and migration.

Neuropsin is a serine protease cloned from the mouse hippocampus. Since neuropsin is a secreted protein which effectively cleaves fibronectin, it may affect cell adhesion or cell migration by modulating the content and/or chemical characteriscs of fibronectin in extracellular matrix (ECM). In adhesion assays, alpha5B2 cells expressing integrin alpha5beta1 bound less effectively to fibronectin teated with neuropsin than intact fibronectin. In Boyden chamber chemotaxis assays, the fibronectin-induced migration of alpha5B2 cells was not affected by neuropsin treatment. These findings suggest that neuropsin regulates the local microenvironment by modulating the interaction between cells and fibronectin in ECM.

Animals↗
PubMed2001

Expression of neuropsin in oligodendrocytes after injury to the CNS.

Proteases are involved in a variety of processes including demyelination after injury to the central nervous system. Neuropsin is a serine protease, which is constitutively expressed in the neurons of the limbic system. In the present study, intrahippocampal kainate injection and enucleation were performed on adult mice. Neuropsin mRNA and protein expression was detected by in situ hybridization and immunohistochemistry. Double in situ hybridization confirmed that the mRNA expression was induced in oligodendrocytes. One day after kainate injection to the hippocampus, neuropsin mRNA was expressed, peaking 4-8 days postoperatively and disappearing at 14 days. Immunohistochemistry and immunoelectron microscopy revealed that neuropsin was expressed in the cell body of oligodendrocytes and myelin. To see if neuropsin degrades myelin protein, purified myelin was incubated with recombinant neuropsin. A decrease in the intensity of the bands of myelin basic protein was observed. These results indicate that neuropsin is involved in demyelination.

Animals↗
PubMed2001

Differential expression of mRNAs for sialyltransferase isoenzymes induced in the hippocampus of mouse following kindled seizures.

Sialic acids play important roles in various biological functions. In the brain, evidence suggests that sialylation of glycoproteins and glycolipids affects neural plasticity. While the 18 sialyltransferase isoenzymes (STs) identified to date synthesize individual sialyl-oligosaccharide structures, they each exhibit activity toward more than one substrate and can overlap in their specificity. Therefore, the distribution of STs is a secondary factor in the study of specific sialylation. Here, seven STs; ST3Gal I-IV, ST8Sia IV, ST6Gal I and ST6GalNAc II, the expressions of which were identified in the adult hippocampus by RT-PCR, showed diverse localization patterns in the hippocampus on in situ hybridization, suggesting that the individual cells expressed relevant STS: Furthermore, to assay activity-related changes in ST expression, we used amygdaloid-kindling among models of neural plasticity. Differential expression of the STs participating in the kindling, notably, up-regulation of ST3Gal IV and ST6GalNAc II mRNAs, and down-regulation of ST3Gal I and ST8Sia IV mRNAs, were observed in the hippocampus following kindled seizures. These results indicate that ST expressions are regulated by physiological activity and may play a role in neural plasticity.

Animals↗
PubMed2000

Phosphorylation of neurofilament-L during LTD.

We recently reported that CaMKII-dependent phosphorylation of the neurofilament-L (NF-L) head domain was induced in the apical dendrites during long-term potentiation. Long-term depression (LTD) is another cellular model for neuronal plasticity. In the present study, we examined the phosphorylation of NF-L during hippocampal LTD using a series of phospho-specific antibodies against the NF-L head domain. During LTD, these antibodies visualized NF-L phosphorylation at Ser57 in the apical dendrites of the hippocampal pyramidal neurons. The assembly and disassembly of NF-L filaments are regulated by phosphorylation of its head domain. Thus, our results indicated that NF-L phosphorylation might be associated with alterations of the neuronal structure during LTD.

Animals↗
PubMed2000

Synaptic microenvironments--structural plasticity, adhesion molecules, proteases and their inhibitors.

Proteolytic regulation might be essential in neural plasticity in mature brain as well as the developing brain. An increasing number of studies support the idea that structural changes in the synapses are closely associated with synaptic plasticity. Proteases and their inhibitors in a synaptic microenvironment are important in the regulation of dynamic changes in the extracellular matrix components associated with synaptic plasticity. In the present article, the possible roles of neuronal proteases, protease inhibitors and extracellular macromolecules are reviewed.

Animals↗
PubMed2000

Neuropsin regulates an early phase of schaffer-collateral long-term potentiation in the murine hippocampus.

We found that neuropsin, an extracellular matrix serine protease, has a regulatory effect on Schaffer-collateral long-term potentiation (LTP) in the mouse hippocampus. Bath application of 1-170 nM recombinant neuropsin modulated early phase LTP in the Schaffer-collateral pathway with a 'bell-shape' dose-response curve. The maximum enhancing activity (134% of control LTP) was found at approximately 2.5 nM. Bath application of a neutralizing antibody against neuropsin in the hippocampal slice resulted in a marked inhibition of the tetanus-induced early phase of LTP. The in vivo continuous intraventricular infusion of an antisense oligonucleotide against neuropsin significantly reduced the amplitude of the tetanus-induced early phase of LTP in vitro. Neuropsin did not directly change the N-methyl D-aspartate (NMDA) current. Thus, neuropsin appears to act as a regulatory molecule in the early phase of LTP via its proteolytic function on extracellular matrix rather than affecting NMDA receptor-mediated calcium increase.

Animals↗
PubMed2000

Site-specific phosphorylation of neurofilament-L is mediated by calcium/calmodulin-dependent protein kinase II in the apical dendrites during long-term potentiation.

Neurofilament-L (NF-L), one subunit of the neuronal intermediate filaments, is a major element of neuronal cytoskeletons. The dynamics of NF-L are regulated by phosphorylation of its head domain. The phosphorylation sites of the NF-L head domain by protein kinase A, protein kinase C, and Rho-associated kinase have been previously identified, and those by calcium/calmodulin-dependent protein kinase II (CaMKII) were identified in this study. A series of site- and phosphorylation state-specific antibodies against NF-L was prepared to investigate NF-L phosphorylation in neuronal systems. Long-term potentiation (LTP) is a cellular model of neuronal plasticity that is thought to involve the phosphorylation of various proteins. NF-L is considered a possible substrate for phosphorylation. During LTP stimulation of mouse hippocampal slices, the series of antibodies demonstrated the increase in the phosphorylation level of Ser(57) in NF-L and the visualization of the localized distribution of Ser(57) phosphorylation in a subpopulation of apical dendrites of the pyramidal neurons. Furthermore, Ser(57) phosphorylation during LTP is suggested to be mediated by CaMKII. Here we show that NF-L is phosphorylated by CaMKII in a subpopulation of apical dendrites during LTP, indicating that Ser(57) is a novel phosphorylation site of NF-L in vivo related to the neuronal signal transduction.

Amino Acid Sequence↗
PubMed1999

Protease M/neurosin mRNA is expressed in mature oligodendrocytes.

No proteases have been identified to be oligodendrocyte-specific though there are phenomena in which proteases may be committed. We have cloned a cDNA for mouse homologue of protease M/neurosin, serine protease, and localized its mRNA in the mouse brain. The cDNA and the deduced amino acid sequences were 66% identical to those of human protease M/neurosin. In situ hybridization histochemistry revealed that the mRNA was localized in the mature oligodendrocytes which expressed proteolipid protein mRNA. Developmentally, protease M/neurosin mRNA was expressed by oligodendrocytes 2 to 7 days after the maturation. Thus, protease M/neurosin may be important to the processes occurring after the maturation of oligodendrocytes such as myelination or turnover of the proteins in the myelin.

Amino Acid Sequence↗
PubMed1999

cDNA cloning and expression of a novel serine protease in the mouse brain.

A cDNA for a novel serine protease, termed brain type granzyme K (B-GRK) was cloned from the mouse brain. The cDNA codes a protein similar to granzyme K (GRK) but completely different at the N-terminus. Genomic Southern and PCR analysis of the gene suggests B-GRK is the alternative transcription form of GRK. B-GRK and GRK have a different organ-specific expression pattern: B-GRK is expressed in the brain, while GRK is expressed in the spleen. The recombinant fusion protein was detected in the neuro2a cells transfected with a plasmid containing B-GRK sequence. The mRNA for B-GRK/GRK was detected in cerebral cortex, hippocampus and diencephalon of the mouse brain. In situ hybridization for B-GRK/GRK revealed that several regions in the forebrain and hypothalamus express the mRNA. Developmental analysis showed that in the prenatal stage, the mRNA was expressed also in pituitary and pineal body in addition to the brain.

Amino Acid Sequence↗
PubMed1999

Injury induces neuropsin mRNA in the central nervous system.

We have shown that neuropsin is expressed in the neurons of the limbic system in the adult mouse. After the central nervous system was injured by incision or intraperitoneal kainate injection, neuropsin mRNA was induced in the peri-lesioned region. The cells in which neuropsin mRNA was induced were localized mainly in axon fiber pathways and closely associated to proteolipid protein (PLP) mRNA expressing oligodendrocytes.

Animals↗
PubMed1999

Crystal structure of neuropsin, a hippocampal protease involved in kindling epileptogenesis.

Neuropsin is a novel serine protease, the expression of which is highly localized in the limbic areas of the mouse brain and which is suggested to be involved in kindling epileptogenesis and hippocampal plasticity. The 2.1-A resolution crystal structure of neuropsin provides the first three-dimensional view of one of the serine proteases highly expressed in the nervous system, and reveals a serine protease fold that exhibits chimeric features between trypsin and nerve growth factor-gamma (NGFgamma), a member of the kallikrein family. Neuropsin possesses an N-glycosylated "kallikrein loop" but forms six disulfide bonds corresponding to those of trypsin. The ordered kallikrein loop projects proline toward the active site to restrict smaller residues or proline at the P2 position of substrates. Loop F, which participates in forming the S3/S4 sites, is similar to trypsin rather than NGFgamma. The unique conformations of loops G and H form an S1 pocket specific for both arginine and lysine. These characteristic loop structures forming the substrate-binding site suggest the novel substrate specificity of neuropsin and give a clue to the design of its specific inhibitors.

Amino Acid Sequence↗
PubMed1999

Effect of 12-O-tetradecanoyl-phorbol ester and incisional wounding on neuropsin mRNA and its protein expression in murine skin.

The expression of neuropsin mRNA in vivo in mouse skin was examined by in situ hybridization and Northern blotting under stimulated conditions. Two kinds of epidermal stimuli, a topical application of a chemical tumor promoter and incisional wounding, were used. A single topical application of 12-O-tetradecanoyl-phorbol 13-acetate induced epidermal hyperplasia and simultaneously induced an extensive increase in neuropsin mRNA in the suprabasal cells. A full-thickness skin incision also induced a profound increase in neuropsin mRNA in the suprabasal cells surrounding the wound but not in actively proliferating basal cells. The increases in neuropsin mRNA occurred rather late and were limited to the site of drug application or around the incision. Interestingly, neuropsin mRNA was not expressed in the epithelial tongue migrating toward the wound during re-epithelialization. Thus, neuropsin might participate in accelerated epidermal differentiation rather than in the proliferation or migration of keratinocytes in the wound.

Animals↗