| Co-Investigator(Kenkyū-buntansha) |
HIGASHI Hideho Kurume University, Pharmacology, Professor, 医学部, 教授 (10098907)
NISHI Akirori Kurume University, Pharmacology, Associate Professor, 医学部, 助教授 (50228144)
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| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2005: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2004: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
Chondroitin sulfate (CS) proteoglycans play important roles in neuronal development and regeneration. CS is divided into four subclasses, known as CS-A, CS-C, CS-D and CS-E, on the basis of the sulfation patterns of the disaccharide unit. In dopaminergic neurons, CS is known to modulate neurite outgrowth during development and cell adhesion after injury. Although the biological importance of CS is well established, the precise structure and the molecular mechanisms underlying its influence on dopaminergic neurons are poorly understood due to the complexity and heterogeneity of CS. In this study, we investigated the biological activity of synthetic CS as well as CS polysaccharides to promote the neurite outgrowth of mesencephalic dopaminergic neurons and the intracellular mechanisms underlying the action of CS. CS-E polysaccharide, but not CS-A, -C or -D polysaccharide, facilitated the neurite outgrowth of dopaminergic neurons. The stimulatory effect of CS-E on neurite outgrowth was com
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pletely abolished by digestion of CS-E polysaccharide into disaccharide units with chondroitinase ABC. Similarly to CS-E polysaccharide, synthetic CS-E tetrasaccharide stimulated the neurite outgrowth of dopaminergic neurons, but synthetic CS-E disaccharide or unsulfated tetrasaccharide did not affect the neurite outgrowth. Analysis of intracellular mechanisms revealed the involvement of phospholipase C (PLC) signaling in the action of CS-E tetrasaccharide. The effect of CS-E tetrasccharide was abolished by the inhibitor of PLC, protein kinase C (PKC), inositol 1, 4, 5-triphosphate (IP_3) receptors, Ca^<2+>/calmodulin-dependent kinase II (CaMKII) or calcineurin. These results indicate that the specific sulfation motif of CS-E and the CS-E tetrasaccharide unit represent the structural determinants for the neuritogenic activity of CS in dopaminergic neurons, and that the action of CS-E is mediated through PLC/PKC and PLC/IP_3/IP_3 receptor signaling, leading to [Ca^<2+>]_i-dependent activation of CaMKII and calcineurin. Less
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