POSTTRANSLATIONAL PROCESSING OF ATRIAL-NATRIURETIC-FACTOR BY ADULT-RAT ATRIAL CARDIOCYTES IN CULTURE

Post-translational processing of the cardiac polypeptide hormone atria l natriuretic factor (ANF) was studied using primary cultures of cardi ocytes derived from adult rat atria. Atrial cardiocytes attached to mi crocarrier beads were maintained for up to 15 days under continuous su perfusion in minichromatographic columns. The cultures were characteri zed for their ability to store, process, and release ANF and by immuno fluorescence microscopy for ANF, desmin, and myosin. Nuclear staining using the fluorescent DNA stain Hoechst 33258 was carried out to deter mine the total number of cells in culture. Column eluates were assayed for ANF by radioimmunoassay and analyzed by reverse phase high-perfor mance liquid chromatography. For comparison purposes, superfusion expe riments using freshly isolated cardiocytes supported in Bio-Gel P2 wer e carried out. Freshly isolated atrial cardiocytes stored high molecul ar weight ANF (5.2 +/- 1.9 pmol/mug DNA) and released mostly (83.3 +/- 6.7%) low molecular weight ANF, at an average rate of 97 +/- 18 fmol min-1 . mug-1 DNA. The cell content and the rate of release of ANF aft er 15 days in culture were 1.3 +/- 0.4 pmol/mug DNA and 1.7 +/- 0.4 fm ol . min-1 . mug-1 DNA, respectively, and 62.7 +/- 6.3% of the release d peptide was of a low molecular weight. There was no correlation betw een changes in cell population and the extent of processing. Cultures of noncardiocytes, superfused with exogenous proANF, did not significa ntly process proANF to a lower molecular weight peptide. The present i nvestigation shows that adult rat atrial cardiocytes, maintained super fused in microcarrier culture and in a serum-supplemented medium for u p to 15 days, retain phenotypic and biochemical characteristics normal ly associated with the dual contractile-endocrine nature of mammalian atrial cardiocytes in vivo. The results obtained in the present work s trongly support die view that ANF post-translational processing is an intrinsic property of die atrial cardiocytes and is independent of any other cell type.