Biomedical subjects

S Sashihara

Publications and source records attributed to S Sashihara.

7 recordsLinked to original sources

PubMed1995

Na+ channel beta 1 subunit mRNA: differential expression in rat spinal sensory neurons.

The brain Na+ channel beta 1 subunit (Na beta 1) mRNA has recently been localized within rat central nervous system where it is expressed at differing levels in different types of neurons. In the present study, we have studied the expression pattern of Na beta 1 mRNA in rat dorsal root ganglion (DRG) neurons using non-radioactive in situ hybridization histochemistry. Na beta 1 mRNA is differentially expressed in adult DRG, with higher levels in intermediate-to-large (> approximately 25 microns in diameter) DRG neurons than in small (< 25 microns) DRG neurons. This cell body size-related Na beta 1 mRNA expression is consistently observed beginning at postnatal day 4 and continues throughout development to adulthood. The present results indicate that (i) Na beta 1 mRNA is expressed in neurons in the peripheral nervous system and (ii) Na beta 1 gene expression is differentially regulated in DRG neurons in relation to their cell body sizes.

Animals↗

PubMed1995

Tissue-specific distribution of a novel C-terminal truncation retinoic acid receptor mutant which acts as a negative repressor in a promoter- and cell-type-specific manner.

A cDNA clone which encodes a truncation form of the gamma subtype of the retinoic acid receptor (RAR gamma) has been isolated. The mutant RAR gamma (RAR gamma Bm382) has lost its 65 C-terminal amino acids, thus truncating a part of the dimerization and activation domains. By using a reverse transcription-coupled PCR technique, it was shown that RAR gamma Bm382 is expressed at different levels in various mouse tissues and that the level of its expression does not correlate with that of normal RAR gamma B. Cotransfection studies revealed that RAR gamma Bm382 acts as a repressor of normal RARs in a promoter- and cell-type-specific manner. Transcription of beta RARE and TREinv promoters was inhibited by RAR gamma Bm382 in both HeLa and F9 cells. Unlike these two promoters, however, RAR gamma Bm382 did not inhibit transcription of the TREpal promoter in HeLa cells but did so in F9 cells. Moreover, while transcription of the lamRARE promoter was inhibited by RAR gamma Bm382 in both HeLa and F9 cells, the inhibition was not observed when F9 cells were induced to differentiate with retinoic acid and dibutyryl cyclic AMP. DNA-binding analysis revealed that RAR gamma Bm382 is able to form a heterodimer with the retinoid X receptor and bind to the different types of retinoic acid response elements with almost the same efficiency as normal RAR. By comparison with effects of other truncation mutants created in vitro, it was suggested that the C-terminal end of the ligand binding domain of RAR is crucial for determining the specificity of transactivation by RAR. Given these observations, we discuss the possibility that protein factors which mediate retinoic acid response element- and cell-type-specific transactivation by RAR are present.

Amino Acid Sequence↗

PubMed1994

In situ hybridization localization of the Na+ channel beta 1 subunit mRNA in rat CNS neurons.

Localization of Na+ channel beta 1 subunit (Na beta 1) mRNA was examined in adult rat hippocampus, cerebellum and spinal cord by in situ hybridization histochemistry. In hippocampus, Na beta 1 mRNA was strongly expressed by CA3 followed by CA1 pyramidal cells and dentate granule cells. In cerebellum, strong Na beta 1 mRNA expression was observed in Purkinje cells and moderate expression in granule cells and scattered cells of the molecular layer. In spinal cord, neurons in gray matter exhibited moderate to strong expression of Na beta 1 mRNA. These results provide the first localization study of Na beta 1 mRNA in the CNS, demonstrating a differential expression in different neurons.

Animals↗

PubMed1994

Differential up-regulation of voltage-dependent Na+ channels induced by phenytoin in brains of genetically seizure-susceptible (E1) and control (ddY) mice.

We investigated the effect of in vivo administration of an antiepileptic drug, phenytoin, on the saxitoxin binding capacity of receptor site 1 of the Na+ channel alpha-subunit, and the expression activity of the channel messenger RNA in epileptic El mouse brains, as compared with parental ddY mice. Subchronic treatment with phenytoin (25 mg/kg per day) for 14 days increased the [3H]saxitoxin binding to brain-derived synaptic membranes of both El and control ddY mice in a time dependent manner. This increase plateaued at 21 +/- 4% in El mice and 28 +/- 3% in ddY control mice after administration of phenytoin for seven days. After cessation of treatment with phenytoin, [3H]saxitoxin binding capacity returned to the basal level within two weeks in both ddY and El brains. Scatchard plot analysis revealed that the phenytoin treatment caused a 20-30% increase in maximum binding capacity of [3H]saxitoxin binding without any change in equilibrium dissociation constant in the brain cortical synaptic membranes of both epileptic El and control ddY mice. A single injection of phenytoin (25 mg/kg) elevated the level of Na+ channel messenger RNA within 1 h in ddY mouse brains. The increase in Na+ channel messenger RNA reached a peak (about 80% increase) after 5 h of phenytoin administration in a concentration-dependent manner (6.25-50 mg/kg). On the other hand, in El mouse brains, Na+ channel messenger RNA was not elevated until more than 5 h after phenytoin injection, and was increased by only about 33%.(ABSTRACT TRUNCATED AT 250 WORDS)

Amphibian Proteins↗

PubMed1992

Overproduction of voltage-dependent Na+ channels in the developing brain of genetically seizure-susceptible E1 mice.

We used E1 mice, a ddY mouse-derived, autosomal mutant strain and a model of hereditary sensory-precipitated epilepsy, to test the hypothesis that epileptic susceptibility may be associated with the activity of voltage-dependent ion channels. We examined the saxitoxin binding capacity of the receptor site 1 of the Na+ channel alpha-subunit, the expression activity of the Na+ channel mRNA, the veratridine-induced 22Na+ influx in the brain synaptosomes, and the regional distribution of Na+ channels in the brain. Compared with control ddY mice, in E1 mice which have not experienced seizures, the number of Na+ channels in the brain synaptosomes increased by approximately 20% starting at the fourth postnatal week through the adult stage as determined by [3H]saxitoxin binding assay. Northern blot hybridization analysis showed excess expression of Na+ channel mRNA (by 30-40%) coincidentally with Na+ channel increases. Regional analysis using the saxitoxin binding assay demonstrated approximately 1.3-fold denser distribution of Na+ channels in the cortex and cerebellum but not the hippocampus and midbrain including thalamus of E1 mice compared to ddY mice. Scatchard plot analysis for saxitoxin binding in the cortex of E1 mouse brains revealed higher maximum binding capacity (Bmax) values (ddY, 4.43 +/- 0.28 pmol/mg protein; E1, 5.43 +/- 0.25 pmol/mg protein) without a change in Kd (ddY, 1.05 +/- 0.03 nM; E1, 1.03 +/- 0.01 nM). Lastly, veratridine-evoked 22Na+ influx, sensitive to tetrodotoxin, was increased approximately 45% in the cortical synaptosomes in six-week-old E1 mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗

PubMed1991

Unusual biochemical development of genetically seizure-susceptible El mice.

Looking for the 'epilepsy gene', we used ddY derived, genetically seizure-susceptible El mice. To find biochemical abnormalities, we examined the amino acid metabolism and gene activity, including poly(A)+ RNA and sodium channel mRNA expressions, in the developmental growth of El mice. At the early postnatal stage, abnormalities in amino acid metabolism were aberrant free amino acid fluctuations. Almost all free amino acids in the liver of newborn El mice showed considerably lower levels than did ddY mice. Among those amino acids, Asp, Glu and Tyr were extremely low, but rapidly recovered to the ddY level within a week. During the successive growth period, we observed no significant difference in hepatic amino acid levels between El and ddY mice. No such drastic changes were noted in the amino acid levels in the brains of ddY and El mice; only the Gly level was greater in El mice than in ddY mice on the day of birth. Rotatory stimulation which evokes convulsions in El mice but not in ddY mice was applied to adult mice and changes in the amino acid level were assessed. The level of Glu and Tyr in seizure-induced El mice was approximately twice that noted in the liver and brain of El mice, which did not experience seizures. It was also somewhat increased in ddY mice subjected to rotational stress which did not induce seizures in that strain. Gene activity that expresses poly(A)+ RNAs, including sodium channel mRNA, was determined by Northern blot analysis, which reveals unscheduled mRNA synthesis by the appearance of an extra band approximately 3 kb in size.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗

PubMed1991

Biochemical abnormalities in developing E1 mouse.

Complex biochemical abnormalities were found in the early developmental stage of the E1 mouse. First, the E1 mouse has abnormal levels of specific amino acid concentration within a week from birth. Second, an unusual expression of poly(A)+ RNA from the one-day newborn liver of the E1 mouse was detected by use of Cot 100 DNA as a probe. Third, sodium channels are increased in synaptosomes and at the mRNA expression level of the 3 or 4-week-old E1 mouse brains, compared with the ddY mouse. These results suggest that the biochemical abnormalities described in this study may affect greatly the epileptogenesis of E1 mouse.

Age Factors↗