The role of IP₃ in the heart is currently unclear, though reports have suggested a role for IP₃ in atrial muscle through a variety of mechanisms (Lipp et al. 2000 Wang et al. 2005). An important possibility yet to be considered is the involvement of Ca²⁺ stimulated adenylyl cyclases (AC 1 and 8), which have an important role in the sino-atrial node (SAN), particularly AC 1 (Mattick et al. 2007). The aim of this study was to investigate whether IP₃ and cAMP may mediate the effects of phenylephrine on guinea-pig atrial myocyte Ca²⁺ transients. Male guinea-pigs (300-400 g) were killed by cervical dislocation following stunning. Atrial myocytes were isolated using collagenase, loaded with indo-5F-AM (as the Ca²⁺ probe) and field stimulated (3ms pulse width, 1Hz; 36°C) to fire action potentials. Data are presented as mean±S.E.M. and P<0.05 (paired Student’s t-tests) was taken to indicate statistical significance. Immunocytochemistry revealed that the type 2 IP₃ receptor (IP₃R) is located sub-sarcolemmally while AC1 and AC8 are associated with the sarcolemma (n>6). Phenylephrine (10 μM) increased atrial myocyte Ca²⁺ transients by 35±9% (% change in fluorescence ratio for emission at 410 and 490 nm; P<0.01, n=8). This effect was prevented by pre-treatment with prazosin (1μM, n=6), an α₁-adrenoceptor antagonist. The effects of phenylephrine were also prevented by 2-APB (2.5μM, n=5), a cell-permeant inhibitor of IP₃Rs, suggesting that IP₃ mediates the response to phenylephrine. The possibility of constitutive production of cAMP in atrial myocytes, as has been shown in SAN myocytes (Mattick et al. 2007), was also investigated. MDL (10μM), an AC inhibitor, reduced Ca²⁺ transient amplitude by 48±8% (P<0.001, n=7). H89 (1μM), a protein kinase A inhibitor, reduced Ca²⁺ transient amplitude by 37±5% (P<0.001, n=6). IBMX (100μM), a phosphodiesterase inhibitor, increased Ca²⁺ transient amplitude by 85±11% (P<0.001, n=6). These data support the suggestion of constitutive cAMP production. In the presence of MDL, phenylephrine produced a 12±4% (P<0.05, n=5) increase in Ca²⁺ transient amplitude. This increase was smaller than phenylephrine alone (unpaired Student’s t-test, P<0.05). These data are consistent with phenylephrine stimulating the production of IP₃ which then acts to release Ca²⁺ from the SR. This sub-sarcolemmal Ca²⁺ stimulates the activity of sarcolemmal AC1 and perhaps AC8 to enhance cAMP synthesis, thereby activating protein kinase A to phosphorylate proteins involved in the increase in Ca²⁺ transient amplitude.