The exchange proteins directly activated by cyclic AMP (Epac or cAMP-GEF) are a family of cAMP-regulated guanine nucleotide exchange factors (GEF) [1]. Binding of cAMP to Epac activates the GEF activity thus stimulating the exchange of GTP for GDP on the monomeric G proteins Rap1/2 [1]. The discovery of the Epac proteins (Epac1/2) has raised the possibility of novel signalling pathways for cAMP that are independent of its traditional target, protein kinase A (PKA). It has previously been reported using yeast two-hybrid screening that Epac interacts with sulphonylurea receptors (SUR) [2], the regulatory subunits of ATP-sensitive potassium (K_ATP) channels. This suggests that Epac may play a role in the regulation of K_ATP channel activity. Indeed, we found that antibodies directed against Epac1 co-immunoprecipitate SUR2B, the dominant SUR subtype found in vascular smooth muscle, from rat aortic homogenates. Also, using the Epac-specific cAMP analogue 8-pCPT-2`-O-Me-cAMP at concentrations that activate Epac but not PKA, we show cAMP-mediated but PKA-independent modulation of vascular K<sub>ATP</sub> channels. Application of 8-pCPT-2`-O-Me-cAMP (5 µM) caused a 41.6 ± 4.7 % inhibition (mean ± SEM; n = 7) of pinacidil-evoked whole-cell K_ATP currents recorded in isolated rat aortic smooth muscle cells. Consistent with the idea that Epac mediates some of its effects by inducing a rise in intracellular Ca²⁺ [3], we found that the activation of Epac via application of 8-pCPT-2`-O-Me-cAMP (5 µM) caused a transient 171.0 ± 18.0 nM (n = 5) increase in intracellular Ca<sup>2+</sup> in Fura-2-loaded rat aortic smooth muscle myocytes. Inclusion of the Ca<sup>2+</sup> chelator BAPTA (20 µM) in the pipette-filling solution significantly reduced the ability of 8-pCPT-2`-O-Me-cAMP (5 µM) to inhibit whole-cell K_ATP currents (inhibition 8.7 ± 4.4 %; n = 4; p < 0.001; student’s t-test). Preincubation with cyclosporin A (10 µM) and ascomycin (5 µM), inhibitors of the Ca²⁺-sensitive protein phosphatase 2B (PP-2B, calcineurin), also significantly reduced the ability of 8-pCPT-2`-O-Me-cAMP to inhibit whole-cell K_ATP currents (inhibition 10.8 ± 2.8 %; n = 9; p < 0.001; student’s t-test and 7.3 ± 1.6 %; n = 8; p < 0.001; student’s t-test, respectively). These findings suggest cAMP-mediated activation of Epac inhibits aortic K_ATP channels via a Ca²⁺-dependent mechanism involving the activation of calcineurin, and highlight a potentially important role for Epac in regulating vascular tone.