|Budget Amount *help
¥3,100,000 (Direct Cost : ¥3,100,000)
Fiscal Year 1998 : ¥1,100,000 (Direct Cost : ¥1,100,000)
Fiscal Year 1997 : ¥2,000,000 (Direct Cost : ¥2,000,000)
Analysis of cell cycle regulation in neocortical neuronogenesis has been undertaken in vitro in the embryonic day 13 mouse. Explants of the dorsomedial region of the cerebral wall, implanted in collagen gel, were incubated for 1 hr in minimal medium and then in minimal medium containing BUdR.After 30 mm with BUdR (1.5 hr in vitro), the labeling index (LI, labeled/total cells) was 0.41 as compared to the previously determined LI of 0.38 in vivo. The in vitro LI remained at 0.4 over the next 7 hr of continued exposure to BUdR and then ascended linearly to asymptote at 0.73. These observations establish (1) that there is a flux of cells from G1 to S for the first 1.5 hr in vitro, (2) that this G1 to S flux is then interrupted for the following 7 hr but (3) that G1 to S flux is subsequently reestablished. In this same system, mitotic figures were first labeled only after a 5 hr exposure to BUdR but then over 2 more hours all mitoses became labeled. In vivo, by contrast, all mitoses become
labeled during 2 hr of BUdR exposure. By implication there is a transient arrest in the progression from S to M phase which is evidently overcome within 5 hr of explantation. The duration of the reestablished cell cycle and those of G1, Sand combined G2-M phases were estimated to be 19.2, 6.3 -8.3, 8.8, and 2.0 -4.0 hr, respectively. The leaving (Q) fraction of the cycle (0,64) was twice the in vivo value. Two thirds of the Q fraction cells remained in the ventricular epithelium, resulting in a substantially low growth fraction of 0.73 as compared with 1.0 in vivo.
Also examined in a cerebral wall explant in vitro was the consequences of basic fibroblast growth factor (b-FGF) and the gap junction uncoupling agent, 1-octanol. LI was determined in the sector of the medial and lateral cortical zones, corresponding to areas 1 and 40, respectively. In area 40, the overall LI in b-FGF-exposed explants was higher and that in 1-octanol-exposed explants was lower than the LI in control explants. Those observations suggest that b-FGF reduces Q fraction while 1-octanol increases the fraction.
すべての系においてGrowth Fraction(GF)がin vivoの1.0という値に比して明らかに低いのは、おそらくQ Fractionの細胞が脳室層内に留まっているためと考えられる(平成9年度実績)。したがってArea40におけるFGF、OctanolによるGFの更なる低下は、Q Fractionの増加(分化の誘導)を反映していると考えられる。一方、Gl期の長さに関しては、Area1、Area40ともにFGF、Octともに効果は認められなかった。今後は、分化が進行した部位(Area40)におけるFGFのQ Fractionに及ぼす影響について、検討を進める予定である。 Less