Fig. 3. SDF-1
induces chemotactic responses in granule cells. (A) Increasing concentrations of SDF-1 (
) or MIP-1 (
) were added to the bottom wells of a modified Boyden chemotaxis chamber. The chemotactic response of purified granule cells was determined by counting migrating cells per high-powered field (Cells/HPF ±s.e.m) in five fields from replicate wells. Data shown is combined from 5 separate experiments using cells derived from P8-P10 mice where each data point was performed in triplicate. Chemotactic responses of granule cells to SDF-1 are significantly above background (0 concentration) and significantly greater than chemotactic responses to MIP-1 at every concentration (P<0.05). Chemotactic responses to MIP-1 are not significantly greater than background (negative control). (B) Granule cells derived from P8 mice were pretreated with 200 ng/ml pertussis toxin for 1 hour, 10 µg/ml polyclonal antibodies to CXCR4 for 15 minutes, or medium alone before being placed in a modified Boyden chemotaxis chamber. For some wells, 1000 ng/ml SDF-1 was added to both the top wells with cells and the bottom wells to test for SDF-1 mediated random migration (chemokinesis). The chemotactic response of purified granule cells was determined by counting migrating cells per high-powered field (Cells/HPF) in five fields from replicate wells and results are expressed ± s.e.m. All cells were examined for chemotactic responses to 1000 ng/ml SDF-1, which was deemed to produce maximal chemotactic responses in dose response experiments (see A). Data shown is combined from 2 separate experiments using cells derived from P8 mice where each data point was performed in triplicate. (*P<0.002; #P<0.005). (C) The chemotactic response of purified granule cells derived from P3-4 vs. P7-8 rats to 0 and 1000 ng/ml SDF-1 was determined by counting migrating cells per high-powered field (Cells/HPF ±s.e.m.) in five fields from replicate wells. Data shown is combined from 2 separate experiments where each data point was performed in triplicate. (**P<0.05).
