1 The effects of zaprinast (M&B 22948), a selective guanosine 3':5'-cyclic monophosphate (cyclic GMP) phosphodiesterase inhibitor, and sodium nitroprusside on cyclic GMP content, phosphoinositide hydrolysis and airway smooth muscle tone were examined in flurbiprofen pretreated bovine tracheal smooth muscle (BTSM). 2 Anion-exchange chromatography of the soluble fraction of BTSM homogenates resolved three peaks of Ca2+/calmodulin-independent phosphodiesterase (PDE) activity that corresponded to type Ia (cyclic GMP-specific, zaprinast-inhibitable), type II (cyclic GMP-stimulated) and type IV (Ro 20 1724-inhibitable) PDE isoenzymes. Zaprinast caused a selective inhibition of the type Ia PDE isoenzyme (IC50 0.94-mu-M) with respect to the type II and IV (IC50 s 93-mu-M and 197-mu-M respectively) isoenzymes. 3 Pretreatment of BTSM strips with zaprinast (10-mu-M) for 20 min affected neither the initial rate of force development, nor the resultant magnitude of contraction induced by methacholine (10-mu-M). In addition, zaprinast (10-mu-M; 20 min) did not affect the cumulative concentration-response relationship induced by methacholine. In contrast, sodium nitroprusside (300-mu-M) either alone, or in combination with zaprinast (10-mu-M), significantly attenuated tone induced by low, but not high concentrations of methacholine. This resulted in a non-parallel, rightwards shift of the methacholine concentration-response curves (nitroprusside: 4.0 fold; nitroprusside/zaprinast: 4.8 fold at the EC50 values), without a reduction in the maximum tone generated. 4 In BTSM slices, zaprinast (10 or 100-mu-M) did not influence basal or methacholine (10-mu-M)-stimulated cyclic GMP accumulation or inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) mass accumulation over a 60 s incubation period, although it did significantly increase cyclic GMP content over longer (30 min) stimulation periods. 5 In [H-3]-inositol prelabelled BTSM slices, stimulated in the presence of 5 mM LiCl, methacholine (10-mu-M) caused a marked increase in total [H-3]-inositol phosphate accumulation. This effect was not inhibited by zaprinast (10-mu-M), sodium nitroprusside (300-mu-M), or a combination of these drugs despite these agents markedly increasing tissue cyclic GMP content. 6 These findings demonstrate that despite zaprinast being a potent and selective inhibitor of the type Ia PDE isoenzyme in a cell-free system, this drug only increases cyclic GMP content in BTSM following prolonged agonist-stimulation. This may explain its lack of inhibitory effect on methacholine-induced tone. The inability of drugs which increase tissue cyclic GMP content and exhibit anti-spasmogenic activity to inhibit methacholine-stimulated Ins(1,4,5)P3 formation suggests that, unlike vascular smooth muscle, cyclic GMP-dependent mechanisms do not regulate receptor-mediated phosphoinositide hydrolysis in BTSM.
