Figure 4.

Effect of fractalkine (FK) and gp120_IIIB on Akt phosphorylation and neuronal survival. (A) Protein extracts (40 μg per lane) were obtained from neurons treated 15 min with 100 nM fractalkine or 200 pM gp120_IIIB (or with fractalkine + gp120 for the indicated time) 4 h after glial deprivation. Immunoblots were conducted with antibodies to total Akt or P-Akt. (B) Densitometric analysis of the immunoblots treated with fractalkine and/or gp120_IIIB for 5–45 min showing the increase in P-Akt induced by fractalkine (*, P < 0.05 vs. control, n = 3). (C) Activation of Akt in HEK293 cells expressing CX₃CR1 and treated with fractalkine (100 nM) for the indicated time (30 μg per lane). Densitometric analysis of P-Akt bands from two separate experiments showed a twofold increase in the band density after 5- and 15-min treatments with fractalkine, and of more than 400% after 45 min. (D) Calcium responses to fractalkine (100 nM) in CX₃CR1-expressing HEK293 cells (n = 87). No response was observed after pertussis toxin (PTX) treatment (200 nM for 16–18 h, n = 56). Representative traces from single cells are shown.