The effect of cyclic AMP and cyclic GMP phosphodiesterase inhibitors on the superoxide burst of guinea‐pig peritoneal macrophages

1 The cyclic nucleotide phosphodiesterase (PDE) activity of guinea‐pig peritoneal macrophages was partially characterized and the effects of selective and non‐selective inhibitors of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP PDE) and guanosine 3′:5′‐cyclic monophosphate (cyclic GMP PDE) phosphodiesterases on superoxide generation were investigated using peritoneal macrophages from horse‐serum pretreated guinea‐pigs. 2 The non‐selective PDE inhibitor, 3‐isobutyl‐1‐methylxanthine (IBMX) and the PDE I/V selective inhibitor, zaprinast, inhibited spontaneous superoxide generation with IC50s of 30.7 ± 11.3 μm and 145 ± 17 μm respectively (n = 6 and 5). The concentration‐response curves for the PDE IV selective inhibitors rolipram and Ro20–1724 were biphasic; mean maximum inhibitions were 56.9 ± 5.9% and 66.8 ± 10.5% respectively at 300 μm, but in 2 out of 6 (rolipram) and 2 out of 5 (Ro20–1724) experiments inhibition was < 50%. The PDE III inhibitor SK&F 94120 was without effect. Spontaneous superoxide generation was reduced 57 ± 10% by 1 μm prostaglandin E2 (PGE2) and 62.6 ± 3.76% by 1 μm salbutamol. 3 The increase in superoxide generation elicited by FMLP (10−9−10−5 m) was unaffected by any of the PDE inhibitors studied. Inhibition of FMLP‐stimulated superoxide generation by PGE2 was enhanced in the presence of 10 μm IBMX. 4 Macrophages were found to contain a predominantly membrane bound cyclic AMP PDE (90% of total activity) which was unaffected by cyclic GMP or calcium/calmodulin. The cyclic AMP PDE activity in the cytosolic fraction was enhanced in the presence of calcium/calmodulin. Selective inhibitors of PDE IV inhibited the particulate cyclic AMP PDE activity (IC50s rolipram 1.5 ± 0.3 μm, Ro 20–1724 4.1 ± 0.6 μm) as did the non‐selective inhibitor IBMX (IC50 22 ± 8 μm). The macrophage particulate PDE activity was resistant to inhibition by the PDE III inhibitor SK&F 94836 and the PDE I/V inhibitor, zaprinast. The cytosolic calcium/calmodulin stimulated cyclic AMP hydrolytic activity was inhibited by zaprinast (IC50 ‐ calcium/calmodulin 123 ± 39 μm; + calcium/calmodulin IC50 17.7 ± 6.3 μm). 5 The results indicate that guinea‐pig peritoneal macrophages contain a type IV cyclic AMP PDE which is predominantly membrane associated and a predominantly cytosolic calcium/calmodulin stimulated cyclic AMP PDE. Functional studies suggest that both of these PDE activities contribute to cyclic AMP hydrolysis and regulation of superoxide generation in these ce