Fig. 1. Ca²⁺-sensitive adenylyl cyclase and PDE4 activities dominate membrane cAMP synthesis and hydrolysis in PMVECs. (A) Using membranes obtained from a 30-40% sucrose gradient, increasing buffer Ca²⁺ concentration from 100 nM Ca²⁺ to 10 µM Ca²⁺ inhibited 97% of adenylyl cyclase activity in PMVECs compared with 22% in PAECs. (B) In buffer containing 100 nM Ca²⁺, rolipram (10 µM, EC₁₀₀) inhibited sufficient PDE4 cAMP hydrolysis to increase cAMP accumulation by 330% in PMVEC membranes, but had no effect on cAMP accumulation in PAEC membranes. Whole cell (C) and membrane-specific (D) cAMP hydrolysis is similar in PMVECs and PAECs. However, rolipram (10 µM, EC₁₀₀) inhibited 75% of cAMP PDE activity in PMVECs compared with 20% inhibition in PAECs (D). Data are means ± s.e.m.^*P<0.05, n=3.