cAMP-activated anion conductance is associated with expression of CFTR in neonatal mouse cardiac myocytes

ALAN S. LADER, YIHAN WANG, G. ROBERT JACKSON, JR., STEVEN C. BORKAN AND HORACIO F. CANTIELLO

AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY

Año: 2000 p. 348 - 353

0363-6143

cAMPactivated anion conductance is associated with expression of CFTR in neonatal mouse cardiac myocytes. Am. J. Physiol. Cell Physiol. 278: C436–C450, 2000.—In this study, patchclamp techniques were applied to cultured neonatal mouse cardiac myocytes (NMCM) to assess the contribution of cAMP stimulation to the anion permeability in this cell model. Addition of either isoproterenol or a cocktail to raise intracellular cAMP increased the whole cell currents of NMCM. The cAMP-dependent conductance was largely anionic, as determined under asymmetrical (low intracellular) Cl2 conditions and symmetrical Cl2 in the presence of various counterions, including Na1, Mg21, Cs1, and N-methyl-D-glucamine. Furthermore, the cAMP-stimulated conductance was also permeable to ATP. The cAMP-activated currents were inhibited by diphenylamine-2-carboxylate, glibenclamide, and an anticystic fibrosis transmembrane conductance regulator (CFTR) monoclonal antibody. The anti-CFTR monoclonal antibody failed, however, to inhibit an osmotically activated anion conductance, indicating that CFTR is not linked to osmotically stimulated currents in this cell model. Immunodetection studies of both neonatal mouse heart tissue and cultured NMCM evealed that CFTR is expressed in these preparations. The implication of CFTR in the cAMP-stimulated Cl2- and ATP-permeable conductance was further verified with NMCM of CFTR knockout mice [cftr(2/2)] in which cAMP stimulation was without effect on the whole cell currents. In addition, stimulation with protein kinase A and ATP induced Cl2-permeable single-channel activity in excised, inside-out patches from control, but not cftr(2/2) NMCM. The data in this report indicate that cAMP stimulation of NMCM activates an anion-permeable conductance with functional properties similar to those expected for CFTR, thus suggesting that CFTR may be responsible for the cAMP-activated conductance. CFTR may thus contribute to the permeation and/or regulation of Cl2- and ATP-permeable pathways in the developing heart. Am. J. Physiol. Cell Physiol. 278: C436–C450, 2000.—In this study, patchclamp techniques were applied to cultured neonatal mouse cardiac myocytes (NMCM) to assess the contribution of cAMP stimulation to the anion permeability in this cell model. Addition of either isoproterenol or a cocktail to raise intracellular cAMP increased the whole cell currents of NMCM. The cAMP-dependent conductance was largely anionic, as determined under asymmetrical (low intracellular) Cl2 conditions and symmetrical Cl2 in the presence of various counterions, including Na1, Mg21, Cs1, and N-methyl-D-glucamine. Furthermore, the cAMP-stimulated conductance was also permeable to ATP. The cAMP-activated currents were inhibited by diphenylamine-2-carboxylate, glibenclamide, and an anticystic fibrosis transmembrane conductance regulator (CFTR) monoclonal antibody. The anti-CFTR monoclonal antibody failed, however, to inhibit an osmotically activated anion conductance, indicating that CFTR is not linked to osmotically stimulated currents in this cell model. Immunodetection studies of both neonatal mouse heart tissue and cultured NMCM evealed that CFTR is expressed in these preparations. The implication of CFTR in the cAMP-stimulated Cl2- and ATP-permeable conductance was further verified with NMCM of CFTR knockout mice [cftr(2/2)] in which cAMP stimulation was without effect on the whole cell currents. In addition, stimulation with protein kinase A and ATP induced Cl2-permeable single-channel activity in excised, inside-out patches from control, but not cftr(2/2) NMCM. The data in this report indicate that cAMP stimulation of NMCM activates an anion-permeable conductance with functional properties similar to those expected for CFTR, thus suggesting that CFTR may be responsible for the cAMP-activated conductance. CFTR may thus contribute to the permeation and/or regulation of Cl2- and ATP-permeable pathways in the developing heart.