Fig. 1.

Confocal images demonstrate that PAC1 receptor endocytosis is decreased by pretreatment with Src inhibitor PP2, but not its inactive analog PP3. PAC1 receptors tagged with EGFP are expressed predominantly on the cell surface of untreated control (CTL) cells (A). After 25 nM PACAP exposure for 20 min, internalized GFP-PAC1 receptors are present in numerous endocytic vesicles (B). When cultures were pretreated for 15 min with 10 µM PP2, 25 nM PACAP exposure for 20 min did not initiate EGFP-PAC1 receptor endocytosis (C). When cultures were pretreated for 15 min with 10 µM PP3 and then exposed to PACAP, EGFP-PAC1 receptor endocytosis was evident (D). Each panel is a projection of three confocal slices through the midpoint of the cell. Calibration bar, 20 μm.