1. UMDNJ-Robert Wood Johnson Medical School
Strain Differences in Stem/Progenitor Cell Proliferation and Survival During Adult Neurogenesis in the Dentate Gyrus
Richard S. Nowakowski
Nancy L. Hayes
UMDNJ-Robert Wood Johnson Medical School
Piscataway, New Jersey

2. Why do we need stem cells?

3. Adult Neuronogenesis in the Mouse
Two proliferative populations in adult brain
Dentate gyrus (intrahilar)
Subventricular zone (olfactory bulb)
Let’s examine the properties of the proliferative cells in the dentate gyrus in vivo.
Some new cells become neurons and seem to become integrated into the circuitry of the dentate gyrus, i.e., exactly what we need stem cells to do.

4. 0.5 hours

5. 12 hours

6. 24 hours

7. 36 hours

8. 12 days

9. Productive Capacity of a Proliferating Population
Proportional to the number of proliferating cells
Inversely proportional to the length of the cell cycle
Expansion vs production controlled by cell fate decisions, i.e. re-entry into the cell cycle (P) vs exit to differentiate (Q)

10. Saturate and Survive Method: Principle

11. Saturate and Survive Method: Application
12 hour saturate with BUdR + sacrifice
Note increase and then decrease in # of labeled cells

12. Estimated Cell Cycle Parameters in C57Bl/6J

13. Strain Difference in S-phase Cell Number

14. A Genetic Difference

15. BALB/c has fewer proliferating cells (800 vs 1800), but same kinetics as C57

17. Number of S-phase Cells Differs in Inbred Strains
C57Bl/6J is high; others are low

18. Size of the Hippocampus also varies
129X1/SvJ and DBA/2J are significantly smaller than the others

19. No correlation between Number of S-phase cells and Size of Hippocampus

22. Maximum Likelihood Analysis shows 3 Clusters with Greatest Distance between C57BL/6J and 129X1/SvJ
3 Clusters --> at least 2 genes
Not different?

23. Strain Differences in Survival

24. Strain Differences in S-phase and Survival

25. Survival/S-phase

26. NeuN is Neuronal Marker

27. BUdR + NeuN

28. Double and Single Labeled Cells

29. Measure of Neuron Production

30. Summary of Strain Differences

31. Neuronogenesis Development vs Adult
Transient proliferative population
VZ/PVE produces mostly neurons
Most neurons persist
Cell Cycle length changes
Changing P/Q
Permanent proliferative population
Intrahilar zone produces mixture of cell types
Most cells are transient
Cell cycle length constant (*to be determined*)
Constant P/Q (*to be determined*)
P=Q=0.5 or, more accurately, P=Q+D=0.5
Output and/or the behavior of the proliferating cells are affected by drugs, behavior, etc. (Question: How are cell cycle vs P/Q changed?)

32. Translation: It is now possible to get a provisional map location of the genes producing a phenotype by characterizing that phenotype in several inbred strains of mice.

33. QTL Candidates:
4, 7, 8, 13
7,12,15, X
2, 8, 13, X

34. Strain Differences in Stem/Progenitor Cell Proliferation in the Mouse Dentate Gyrus
Number of S-phase cells (size of proliferating population)
Survival of “new” cells for 5-6 weeks
Septo-temporal distribution of the S-phase cells
Size of the dentate gyrus
Proportion of cells that become neurons?
Behavior?
Tools for Genetic Mapping: Inbred Strains, Recombinant Inbred Strains, F2 hybrids, SNP's, Sequenced Genomes

37. Effect of Low (50 ug/g) vs High (300 ug/g) Dosage

38. Conclusion: Low dose is more controlled and avoids issue of toxicity.

39. NeuN Montage

40. NeuN in Neocortex

41. Through-focus series