Increased cGMP phosphodiesterase activity mediates renal resistance to ANPin rats with bile duct ligation

Background. Liver disease resulting from common bile duct ligation (CBDL) c auses abnormal sodium metabolism that is manifested by resistance to the na triuretic action of atrial natriuretic peptide (ANP). This resistance is co rrected both in vitro and in vivo by zaprinast, a selective inhibitor of a guanosine cyclic-3'-5'-monophosphate (cGMP)-specific phosphodiesterase (PDE 5). Several other PDEs with affinity for cGMP are expressed in kidney and c ould also be involved in this response. Methods. We measured cGMP hydrolysis in inner medullary collecting duct (IM CD) cell homogenates from kidneys of sham-operated and CBDL rats and quanti tated the amount of PDES protein by Western blotting and immunoprecipitatio n studies. We also characterized ANP responsiveness in vivo of kidneys of a nesthetized sham and CBDL rats by measuring sodium excretion before and aft er volume expansion (VE). Results. Kinetic analysis of PDE5 activity in homogenates of IMCD cells iso lated from kidneys of sham-operated rats indicated a V-max of 85.3 +/- 1.7 versus 157 +/- 2.9 pmol/mg/min from CBDL rats (P < 0.01), without a differe nce in K-m. Enzyme activity was inhibited competitively by 1,3-dimethyl-6-( 2-propoxy-5-methanesulfonylamidophenyi)pyrazol[3,4d] -pyrimidin-4-(5H)-one (DMPPO): a potent and specific inhibitor of PDE5, with an apparent K-i of 4 .5 <plus/minus> 0.7 and 4.9 +/- 0.7 nmol/L and an IC50, of 6.1 +/- 0.8 and 8.7 +/- 0.7 nmol/L in sham and CBDL rats, respectively (P = NS). DMPPO exhi bited very poor inhibitory activity against the calcium-calmodulin-dependen t PDE1 in IMCD homogenates from sham rats (K-i 1.3 +/- 0.1 mu mol/L and IC5 0 1.9 +/- 0.2 mu mol/L). Western analysis using an antiserum made against b ovine lung PDE5 revealed a twofold increase in PDES protein in cytosolic ex tracts from IMCD of CBDL rat kidneys compared with sham-operated controls, and immunoprecipitation studies indicated that the increase in PDES protein accounted for the observed increase in cGMP hydrolysis. DMPPO (10 nmol/L) normalized the blunted ANP-dependent cGMP accumulation by IMCD cells from C BDL rats in vitro. Intrarenal infusion of DMPPO (0.5 nmol/min) in CBDL rats corrected both the impaired natriuretic response to VE and the blunted VE- related increase in urinary cGMP excretion from the infused, but not the co ntralateral kidney. Conclusion. These results demonstrate that renal resistance to ANP in CBDL rats is accompanied by heightened activity of PDES, which is due largely to an increase in PDES protein. Other PDEs could contribute only a minor part to the enhanced cGMP hydrolysis observed in kidneys of CBDL rats. This PDE 5-dependent ANP resistance may represent an important contributor to the so dium retention of liver disease.