We have recently reported that store-operated Ca2+ entry (SOCE) in non-excitable cells is likely to be mediated by a reversible interaction between Ca²⁺-channels in the plasma membrane and the endoplasmic reticulum, a mechanism known as secretion-like coupling. As for secretion, in this model the actin cytoskeleton plays a key regulatory role. Here we have explored the involvement of the secretory proteins SNAP-25 and VAMPs in SOCE in pancreatic acinar cells. Cleavage of SNAP-25 and VAMPs by treatment with botulinum toxin (BoNT) A and tetanus toxin (TeTx), respectively, effectively inhibited amylase secretion stimulated by the physiological agonist CCK-8. BoNT A significantly reduced Ca²⁺ entry induced by store depletion using thapsigargin (TG) or CCK-8. In addition, treatment with BoNT A once SOCE had been activated reduced Ca²⁺ influx, indicating that SNAP-25 is needed for both the activation and maintenance of SOCE in pancreatic acinar cells. VAMP-2 and VAMP-3 are expressed in mouse pancreatic acinar cells. Both proteins associate with the cytoskeleton upon Ca²⁺ store depletion although only VAMP-2 seems to be sensitive to TeTx.. Treatment of pancreatic acinar cells with TeTx reduced the activation of SOCE without affecting its maintenance. These findings support a role for SNAP-25 and VAMP-2 in the activation of SOCE in pancreatic acinar cells and show parallels between this process and secretion in a specialized secretory cell type.