Biomedical subjects

Y Hanahisa

Publications and source records attributed to Y Hanahisa.

12 recordsLinked to original sources

PubMed2001

Decrease in Ca2+-ATPase activity in the brain plasma membrane of rats with increasing age: involvement of brain calcium accumulation.

The alteration in Ca(2+)-ATPase activity in the brain plasma membrane of rats with increasing age was investigated. Calcium content in the brain tissues was significantly raised in aged rats (50 weeks old) as compared with that of young rats (5 weeks old). Increasing age caused a significant decrease in Ca(2+)-ATPase activity in the brain plasma membranes. The presence of N-ethylmaleimide (2.5 or 5 mM), a modifying reagent of thiol (SH)-groups, in the reaction mixture caused a significant decrease in the brain plasma membrane Ca(2+)-ATPase activity of young and aged rats, while dithiothreitol (2.5 or 5 mM), a protecting reagent of SH-groups, produced a significant increase in the enzyme activity, indicating that the SH-group is an active site of Ca(2+)-ATPase. The active site of Ca(2+)-ATPase may not be impaired by ageing. The brain plasma membrane Ca(2+)-ATPase activity of young rats was significantly reduced in the presence of dibutyryl cyclic AMP (10(-7)-10(-5) M) or inositol 1, 4, 5-trisphosphate (10(-7)-10(-5) M) in the reaction mixture. Such an decrease was not seen in aged rats. The responsibility for signaling factors seemed to be weakened by ageing. Calmodulin (2.5 and 5 microg/ml) or regucalcin (10(-8) and 10(-7) M), a Ca(2+)-regulating protein, did not have an effect on Ca(2+)-ATPase activity. This study demonstrates that ageing induces a decrease in Ca(2+)-ATPase activity in the brain plasma membranes. This finding suggests a cellular mechanism by which ageing causes calcium accumulation in brain.

Aging↗

PubMed1999

Inhibitory effect of regucalcin on Ca(2+)-dependent protein kinase activity in rat brain cytosol: involvement of endogenous regucalcin.

The effect of regucalcin, a Ca(2+)-binding protein, on Ca(2+)-dependent protein kinase activity in the brain cytosol of rats with different ages (5 and 50 weeks old) was investigated. The addition of calmodulin (10 microg/ml) or dioctanoylglycerol (5 microg/ml) in the enzyme reaction mixture caused a significant increase in protein kinase activity in the presence of CaCl2 (1 mM), indicating that Ca2+ calmodulin or protein kinase C is present in the cytosol. Such an increase was completely prevented by the addition of regucalcin (10(-7) M). Moreover, regucalcin (10(-7) M) significantly inhibited cytosolic protein kinase activity without Ca2+/calmodulin or dioctanoylglycerol addition. Meanwhile, the presence of anti-regucalcin monoclonal antibody (10-50 ng/ml) in the enzyme reaction mixture caused a significant elevation of protein kinase activity, suggesting an inhibitory effect of endogenous regucalcin. Brain cytosolic protein kinase activity was significantly elevated by increasing age (50-week-old rats). Also, regucalcin (10(-7) M) significantly decreased protein kinase activity without Ca(2+) addition in the brain cytosol of aged rats. However, the effect of anti-regucalcin monoclonal antibody (50 ng/ml) in elevating protein kinase activity was not seen in the brain cytosol of aged rats. These results suggest that regucalcin has an inhibitory effect on Ca(2+)-dependent protein kinase activity in rat brain cytosol, and that the effect of endogenous regucalcin may be weakened in the brain cytosol of aged rats.

Animals↗

PubMed1999

Expression of calcium-binding protein regucalcin and microsomal Ca2+-ATPase regulation in rat brain: attenuation with increasing age.

The expression of calcium-binding protein regucalcin and its effect on the microsomal Ca2+-ATPase activity in rat brain tissues was investigated. The expression of regucalcin mRNA was demonstrated by reverse transcription-polymerase chain reaction (RT-PCR) analysis in brain tissues using rat regucalcin-specific primers. Regucalcin concentration in the brain tissues was about 5 x 10(-9)) M as measured using enzyme-linked immunoadsorbent assay (ELISA), and this level was lowered with increasing age (50 weeks old). The presence of regucalcin (10(-9) to 10(-7) M) in the enzyme reaction mixture caused a significant decrease in Ca2+-ATPase activity in the brain microsomes of young rats (5 weeks old). Meanwhile, the enzyme activity was not significantly altered by the addition of calmodulin (1 or 50 microg/ml), calbindin (1 or 10 microg/ml), and S-100 A protein (5 or 25 microg/ml), which are other Ca2+-binding proteins in rat brain. The effect of regucalcin to inhibit microsomal Ca2+-ATPase activity was weakened in the brain of rats with increasing age (50 weeks old). The present study demonstrates that regucalcin is expressed in the brain, and that it can uniquely inhibit Ca2+-ATPase activity in the brain microsomes of rats. The findings suggest that regucalcin plays a role in the regulation of microsomal Ca2+-ATPase activity in rat brain tissues.

Aging↗

PubMed1999

Effect of calcium-binding protein on adenosine 5'-triphosphatase activity in the brain cytosol of rats of different ages: the inhibitory role of regucalcin.

Adenosine 5'-triphosphatase (ATPase) plays a role in the process of energy conversion in the controlled hydrolysis of ATP. The effect of Ca2+ -binding protein on ATPase activity in the brain cytosol of rats of different ages was investigated. ATPase activity in the brain cytosol of 50-week-old rats was significantly decreased as compared with that of 5-week-old rats. The presence of calcium chloride (10(-5) and 10(-4) M) in the enzyme reaction mixture caused a significant increase in ATPase activity in the brain cytosol of rats of different ages. This increase was not altered by trifluoperazine (2x10(-5) M), an antagonist of calmodulin. Calmodulin (5 microg/ml), calbindin (5 microg/ml) or S-100A protein (10 microg/ml), a Ca2+ -binding protein, had no effect on ATPase activity. Meanwhile, regucalcin (10(-9) M), which is present in brain, significantly decreased ATPase activity in young and older rats. However, the effect of regucalcin was weakened by the presence of Ca2+ (10(-5) M). The addition of anti-regucalcin monoclonal antibody in the reaction mixture caused a significant elevation of ATPase activity; this increase was completely abolished by addition of regucalcin (10(-9) M). The present study suggests that regucalcin plays an inhibitory role in the regulation of ATPase activity in the brain cytosol of rats of different ages.

Adenosine Triphosphatases↗

PubMed1999

Brain microsomal calcium accumulation in rats with increasing age: involvement of thapsigargin-sensitive Ca2+-ATPase.

The alteration in calcium content and Ca2+-ATPase activity in the brain microsomes of rats with increasing age was investigated. Brain microsomal calcium content was significantly increased in aged rats (50 weeks old) as compared with that of young rats (5 weeks old). Such an increase was not seen in the brain mitochondria of aged rats. Increasing age caused a significant elevation of Ca2+-ATPase activity in the brain microsomes. The microsomal Ca2+-ATPase activity of young rats was significantly decreased by the addition of thapsigargin (10-7-10-5 M), a specific inhibitor of microsomal Ca2+ pump enzyme (Ca2+-ATPase), in the enzyme reaction mixture. The inhibitory effect of thapsigargin (10-5 M) was also seen in the brain microsomes of aged rats. Moreover, the effect of calcium (5 and 10 microM) addition in elevating Ca2+-ATPase activity was not revealed in the microsomes of aged rats, whereas the metal could increase the enzyme activity in young rats, suggesting an involvement of activatory factor on the enzyme with increasing age. The present study demonstrates that ageing induces an increase in thapsigargin-sensitive Ca2+-ATPase activity and a corresponding elevation of calcium content in the brain microsomes. This finding suggests a cellular mechanism by which ageing causes calcium accumulation in brain.

Aging↗

PubMed1998

Increase of Ca2+-ATPase activity in the brain microsomes of rats with increasing ages: involvement of protein kinase C.

A possible mechanism of aging-induced increase in brain microsomal Ca2+-adenosine triphosphatase (ATPase) activity of rats was investigated. Calcium content in the brain tissues and Ca2+-ATPase activity in the brain microsomes of aging rats (50 weeks of age) increased significantly as compared with those of young rats (5 weeks of age). Brain microsomal Ca2+-ATPase activity in aging rats was decreased significantly by treatment of ethyleneglycol-bis-(aminoethylether) N,N,N',N'-tetraacetic acid (EGTA) (2.7 mM) or digitonin (10(-3)%), while such decrease was not seen in the enzyme activity of young rats. Microsomal Ca2+-ATPase activity in aging rats was markedly decreased by the presence of staurosporine (10(-8) and 10(-7) M), an inhibitor of protein kinase C, in the enzyme reaction mixture, although the enzyme activity of young rats was not inhibited. Meanwhile, dibucaine (10(-6) and 10(-5) M), an inhibitor of Ca2+/calmodulin-dependent protein kinase, did not have an effect on Ca2+-ATPase activity in the brain microsomes of young and aging rats. The addition of protein kinase C (100 and 200 mU/ml) in the reaction mixture caused a significant increase in brain microsomal Ca2+-ATPase activity of young rats. These results suggest that protein kinase C is partly involved in the elevation of brain microsomal Ca2+-ATPase activity in rats with increasing ages.

Aging↗

PubMed1998

Stimulatory effect of calcium-binding protein regucalcin on phosphatase activity in the brain cytosol of rats with different ages.

The effect of calcium-binding protein regucalcin on phosphatase activity in the brain cytosol of rats with different ages was investigated. The presence of regucalcin (10(-8) and 10(-7) M) in the enzyme reaction mixture caused a significant increase of neutral p-nitrophenylphosphatase activity in the brain cytosol obtained from 5- and 50-week-old rats. This increase was seen in the absence or presence of calmodulin (2 microg/mL) and calcium chloride (100 microM). Brain cytosolic phosphatase activity was not significantly altered by S-100A (10(-6) M) or calbindin (10(-7) M), which is a calcium-binding protein. Regucalcin-increased phosphatase activity was clearly decreased by N-ethylmaleimide, a modifying reagent of thiol(SH)-group, suggesting that regucalcin acts on the SH-group of the enzyme. Moreover, the presence of anti-regucalcin monoclonal antibody (50-200 ng/mL) in the reaction mixture caused a significant decrease of brain cytosolic phosphatase activity, suggesting that the endogenous regucalcin has a stimulatory effect on the enzyme activity. These results suggest that regucalcin plays a role in the regulation of protein phosphatase in rat brain cytosol.

Aging↗

PubMed1998

Streptozotocin-induced diabetes increases (Ca2+-Mg2+)-ATPase activity in hepatic plasma membranes of rats: involvement of protein kinase C.

The alteration in calcium transport in the liver of rats with streptozocin(STZ)-diabetic state was investigated. STZ (6 mg/100 g body weight) was subcutaneously administered in rats, and 1 or 2 weeks later they were sacrificed by bleeding. STZ administration caused a remarkable elevation of serum glucose concentration. Liver calcium content was significantly increased by STZ administration. Hepatic plasma membrane (Ca2+-Mg2+)-ATPase activity was markedly elevated by STZ administration. This increase was completely abolished by the presence of staurosporine (10(-7)-10(-5) M), an inhibitor of protein kinase C, in the enzyme reaction mixture, suggesting an involvement of protein kinase C signalling. Moreover, the STZ-induced increase in liver plasma membrane (Ca2+-Mg2+)-ATPase activity was significantly raised by the presence of okadaic acid (10(-5) and 10(-4) M). Meanwhile, the STZ-increased (Ca2+-Mg2+)-ATPase activity was not appreciably altered by the presence of anti-regucalcin IgG in the reaction mixture, indicating that the activatory protein regucalcin does not participate in the elevation of the enzyme activity. The present study demonstrates that STZ-induced diabetes causes the increase in hepatic plasma membrane (Ca2+-Mg2+)-ATPase activity of rats.

Animals↗

PubMed1998

Inhibitory effect of zinc and copper on phosphatase activity in the brain cytosol of rats: involvement of SH groups.

The effect of various metal ions on neutral phosphatase activity in the brain cytosol of rats was investigated. p-Nitrophenylphosphate was used as the substrate for the enzyme assay. Brain cytosolic phosphatase activity was significantly decreased by the presence of zinc or copper in the range of 10(-5) to 10(-3) M in the enzyme reaction mixture, while it was significantly increased by manganese, nickel or cobalt (10(-5) to 10(-3) M). Calcium (10(-5) to 10(-3) M) did not have an appreciable effect on brain cytosolic phosphatase activity. The inhibitory effect of zinc (10(-5) M) or copper (10(-5) M) on phosphatase activity was seen in the presence of manganese, nickel or cobalt with 10(-4) M concentration, indicating a potent inhibitory effect of these metals. Zinc or copper-decreased phosphatase activity was clearly prevented by dithiothreitol (5 mM), although the effect of each metal was further enhanced by N-ethylmaleimide (5 mM), suggesting that the metals act on the SH-groups of enzyme. Moreover, the inhibitory effect of zinc (10(-5) M) was significantly enhanced by S-100A (10(-6) M) or calbindin (10(-7) M), although neither protein had any effect in the presence of calcium (10(-5) M) or copper (10(-5) M). These results suggests that zinc has an unique role in the regulation of neutral phosphatase activity in rat brain cytosol.

Animals↗

PubMed1997

Increase in calcium content and Ca(2+)-ATPase activity in the brain of fasted rats: comparison with different ages.

The effect of fasting on calcium content and Ca(2+)-ATPase activity in the brain tissues of 5 weeks and 50 weeks old rats was investigated. Brain calcium content and Ca(2+)-ATPase activity in the microsomal and mitochondrial functions of the brain homogenate from young and elderly rats were significantly increased by overnight-fasting. These increases were appreciably restored by a single oral administration of glucose solution (400 mg/100 g body weight) to fasted rats. In comparison with young and elderly rats, brain calcium content and microsomal Ca(2+)-ATPase activity were significantly elevated by increasing ages. The effect of ageing was not seen in the brain mitochondrial Ca(2+)-ATPase activity. When calcium (50 mg/100 g) was orally administered to young and elderly rats, brain calcium content was significantly elevated. The calcium administration-induced increase in brain calcium content was greater in elderly rats than in young rats. Also, calcium administration caused a significant increase in Ca(2+)-ATPase activity in the microsomal and mitochondrial fractions of brain homogenates from young rats. In aged rats, the microsomal Ca(2+)-ATPase activity was not further enhanced by calcium administration, although the mitochondrial enzyme activity was significantly raised. The present study demonstrates that the fasting-induced increase in brain calcium content is involved in Ca(2+)-ATPase activity raised in the brain microsomes and mitochondria of rats with different ages, supporting a energy-dependent mechanism in brain calcium accumulation.

Age Factors↗

PubMed1996

Characterization of calcium accumulation in the brain of rats administered orally calcium: the significance of energy-dependent mechanism.

The characterization of calcium accumulation in the brain of rats administered orally calcium chloride solution was investigated. Rats received a single oral administration of calcium (15-50 mg/100 g body weight), and they were sacrificed by bleeding between 15 and 120 min after the administration. The administration of calcium (50 mg/100 g) produced a significant increase in serum calcium concentration and a corresponding elevation of brain calcium content, indicating that the transport of calcium into the brain is associated with the elevation of serum calcium levels. The increase in brain calcium content by calcium administration was not appreciably altered by the pretreatment with Ca2+ channel blockers (verapamil or diltiazem with the doses of 1.5 and 3.0 mg/100 g). In thyroparathyroidectomized rats, the administration of calcium (50 mg/100 g) caused a significant increase in brain calcium content, indicating that calcium-regulating hormones do not participate in the brain calcium transport. Now, brain calcium content was clearly elevated by fasting (overnight), although serum calcium level was not significantly altered. Calcium administration to fasted rats induced a further elevation of brain calcium content as compared with that of control (fasted) rats. The fasting-induced increase in brain calcium content was appreciably restored by refeeding. This restoration was also seen by the oral administration of glucose (0.4 g/100 g) to fasted rats. The present study demonstrates that serum calcium is transported to brain, and that the increased brain calcium is released promptly. The release of calcium from brain may be involved in energy metabolism, and this release may be weakened by the reduction of glucose supply into brain. The finding suggests a physiological significance of energy-dependent mechanism in the regulation of brain calcium.

Animals↗

PubMed1996

Effect of vitamin K2 (menaquinone-7) on bone metabolism in the femoral-metaphyseal tissues of normal and skeletal-unloaded rats: enhancement with zinc.

The effect of vitamin K2 (menaquinone-7) on bone metabolism in the femoral-metaphyseal tissues of normal and skeletal-unloaded rats was investigated. Skeletal unloading was designed using a model of hindlimb suspension; the rats were fed for the 4 days of unloading. The metaphyseal tissues obtained from normal and skeletal-unloaded rats were cultured for 48 h in medium containing either vehicle or vitamin K2 (10(-6) and 10(-5) M). The presence of vitamin K2 (10(-5) M) caused a significant increase in alkaline phosphatase activity and calcium content in the metaphyseal tissues from normal rats. Such an effect was not seen in the bone tissues from skeletal-unloaded rats. Additionally, the presence of zinc sulfate (10(-5) M) in effective concentration produced a significant increase in alkaline phosphatase activity and calcium content in the metaphyseal tissues from normal and skeletal-unloaded rats. In the presence of vitamin K2 (10(-5) M), the stimulatory effect of zinc sulfate on bone calcium content was appreciably enhanced; although this effect was completely abolished by cycloheximide (10(-6) M), an inhibitor of protein synthesis. This study demonstrates that the effect of vitamin K2 (menaquinone-7) on trabecular bone calcification in rats with skeletal unloading-induced osteopenia is enhanced by zinc in vitro. The enhancement with zinc may be based on a newly synthesized protein in the bone tissues.

Alkaline Phosphatase↗