Abstract
We examined the effects of intracellular perfusion of cyclic adenosine monophosphate (cAMP) on the sodium current (I Na) of guinea-pig ventricular myocytes, using the whole-cell clamp technique. I Na was elicited by depolarizing voltage steps (−20 mV) from a variety of holding potentials (−120 to −50 mV), under conditions of 60 mM extracellular Na+ concentration ([Na+]0) and at the temperature of 24–26°C.
Intracellular perfusion of cAMP decreased the I Na elicited from the holding potentials less negative than −90 mV. In the presence of 1 mM cAMP, for example, the peak I Na elicited from −80 mV decreased from 6.0±2.0 nA to 4.0±2.2 nA (mean±SD, P<0.02, n=7) within 3–6 min. In the presence of extracellular 3-isobutyl-1-methylxanthine (IBMX, 20 μM), much lower concentrations of cAMP (0.2 mM) yielded a comparable effect. On the other hand, intracellular perfusion of cAMP increased the I Na elicited from very negative holding potentials (<−100 mV). For instance, the application of cAMP (1 mM) increased the I Na elicited by step depolarizations from −120 mV (to −20 mV), from 9.9±2.1 nA to 11.0±3.1 nA (P<0.05, n=5).
The former effect was attributed to a marked shift of the steady-state inactivation curve of I Na to the negative direction; the voltage of half-inactivation shifted from −77.9±1.0 to −83.5±1.4 mV, or by −5.6 mV. The latter effect may be explained by increases in maximum available conductance of I Na. Extracellular application of isoproterenol (1 μM) also decreased the I Na evoked from a holding potential of −80 mV, whereas it increased the I Na elicited from more negative potentials of −120 mV. These effects of isoproterenol were reversible and markedly attenuated in the presence of a specific inhibitor of cAMP-dependent protein kinase, H-89 {N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulphonamide}, an isoquinolinesulphonamide derivative, in the extracellular medium (2–10 μM) and a protein kinase inhibitor (Walsh inhibitor) in the pipette solution (40 μM). H-89 (10 and 30 μM) affected neither the adenylate cyclase activity prepared from rabbit ventricular muscles, nor the isoproterenol-mediated increases in the cAMP content in guinea-pig ventricular muscles. Our observations suggest that the increase in intracellular cAMP modulates the function of cardiac Na channels, preferentially by stimulating cAMP-dependent protein kinase, with subsequent phosphorylation of the channel protein.
Similar content being viewed by others
References
Arita M, Kiyosue T, Aomine M, Imanishi S (1983) Nature of “residual fast channel” dependent action potentials and slow conduction in guinea pig ventricular muscle and its modification by isoproterenol. Am J Cardiol 51:1433–1440
Asano T, Hidaka H (1977) Purification of guanylate cyclase from human platelets and effect of arachidonic acid peroxide. Biochem Biophys Res Commun 78:910–918
Ashby CD, Walsh DA (1973) Characterization of the interaction of a protein inhibitor with adenosine 3′,5′-monophosphate-dependent protein kinases. J Biol Chem 248:1255–1261
Bahinski A, Nairn AC, Greengard P, Gadsby DC (1989) Chloride conductance regulated by cAMP-dependent protein kinase in cardiac myocytes. Nature 340:718–721
Catterall WA (1992) Cellular and molecular biology of voltage-gated sodium channels. Physiol Rev 72:s 15-s 48
Cheng YN, Aomine M, Arita M (1991) Acetylcholine reverses isoproterenol-induced depression of Vmax in residual Na channel dependent action potentials of guinea-pig ventricular muscles. J Mol Cell Cardiol 23:537–549
Chijiwa T, Mashima A, Hagiwara M, Sano M, Hayashi K, Inoue T, Naito K, Toshioka T, Hidaka H (1990) Inhibition of forskolin-induced phosphorylation by a newly synthesized selective inhibitor of cyclic AMP-dependent protein kinase, N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89), of PC12D pheochromacytoma cells. J Biol Chem 265:5267–5272
Fernandez J, Fox AP, Krasne S (1984) Membrane patches and whole-cell membranes: a comparison of electrical properties in rat clonal pituitary (GH3) cells. J Physiol (Lond) 356:565–585
Fleming JW, Wisler DL, Watanabe AM (1992) Signal transduction by G protein in cardiac tissues. Circulation 85:420–433
Gonoi T, Sherman SJ, Catterall WA (1985) Voltage clamp analysis of sodium channel in normal and scorpion toxin-resistant neuroblastoma cells. J Neuroscience 4:2836–2842
Hanck DA, Sheets MF (1992) Extracellular divalent and trivalent cation effects on sodium current kinetics in single canine cardiac Purkinje cells. J Physiol (Lond) 454:267–298
Harvey RD, Hume JR (1989) Autonomic regulation of delayed rectifier K+ current in mammalian heart involves G proteins. Am J Physiol 257:H 818-H 823
Harvey RD, Hume JR (1989) Autonomic regulation of a chloride current in heart. Science 244:983–985
Hisatome I, Kiyosue T, Imanishi S, Arita M (1985) Isoproterenol inhibits residual fast channel via stimulation of β-adrenoceptors in guinea pig ventricular muscle. J Mol Cell Cardiol 17:657–665
Irisawa H, Kokubun S (1983) Modulation by intracellular ATP and cyclic AMP of the slow inward current in isolated single ventricular cells of the guinea-pig. J Physiol (Lond) 338:321–337
Irisawa H, Sato R (1986) Intra- and extracellular actions of proton on the calcium current of isolated guinea pig ventricular cells. Circ Res 59:348–355
Kameyama M, Hoffmann F, Trautwein W (1985) On the mechanism of β-adrenergic regulation of the Ca channel in the guinea pig heart. Pflügers Arch 405:285–293
Marty A, Neher E (1983) Tight-seal whole-cell recording. In: Neher E, Sakmann B (eds) Single-channel recording. Plenum Press, New York London, pp 107–121
Matsuda JJ, Lee H, Shibata EF (1992) Enhancement of rabbit cardiac sodium channels by β-adrenergic stimulation. Circ Res 70:199–207
Matsuda JJ, Lee H, Shibata EF (1993) Acetylcholine reversal of isoproterenol-stimulated sodium currents in rabbit ventricular myocytes. Circ Res 71:517–525
Muramatsu H, Kiyosue T, Arita M (1990) Intracellular perfusion of cyclic AMP decreases the sodium current of guinea pig ventricular myocytes (abstract). J Am Col Cardiol 16:144A
Nakayama T, Fozzard HA (1988) Adrenergic modulation of the transient outward current in isolated canine Purkinje cells. Circ Res 62:162–172
Nakazawa K, Sano M, Saitoh T (1976) Subcellular distribution and properties of guanylate cyclase in rat cerebellum. Biochim Biophys Acta 444:563–570
Noda M, Shimizu S, Tanabe T, Takai T, Kayano T, Ikeda T, Takahashi H, Nakayama H, Kanaoka Y, Minamino N, Kangawa K, Matsuo H, Raftery MA, Hirose T, Inayama S, Hayashida H, Miyata T, Numa S (1984) Primary structure of electrophorus electricus sodium channel deduced from cDNA sequence. Nature 312:121–127
Ono K, Kiyosue T, Arita M (1989) Isoproterenol, DBcAMP, and forskolin inhibit cardiac sodium current. Am J Physiol 256:C 1131-C 1137
Ono K, Fozzard HA, Hanck DA (1993) Mechanism of cAMP-dependent modulation of cardiac sodium current kinetics. Circ Res 72:807–815
Reuter H (1983) Calcium channel modulation by neuro-transmitters, enzymes and drugs. Circ Res 62:162–172
Schubert B, VanDongen MJ, Kirsch GE, Brown AM (1989) β-adrenergic inhibition of cardiac sodium channels by dual G-protein pathways. Science 245:516–519
Soejima M, Noma A (1984) Mode of regulation of the ACh-sensitive K-channel by the muscarinic receptor in rabbit atrial cells. Pflügers Arch 400:424–431
Sunami A, Fan Z, Nakamura F, Naka M, Sawanobori T, Hiraoka M (1991) The catalytic subunit of cyclic AMP-dependent protein kinase directly inhibits sodium current activities in guinea-pig ventricular myocytes. Pflügers Arch 419:416–417
Tanabe T, Takeshima H, Mikami A, Flockerzi V, Takahashi H, Kangawa K, Kojima M, Matsuo H, Hirose T, Numa S (1987) Primary structure of the receptor for calcium channel blockers from skeletal muscle. Nature 328:313–318
Taniguchi J, Kokubun S, Noma A, Irisawa H (1981) Spontaneously active cells isolated from the sino-atrial and atrioventricular nodes of the rabbit heart. Jpn J Physiol 31:547–558
Tempel BL, Papazian DM, Schwartz TL, Jan YN, Jan LY (1987) Sequence of a probable potassium channel component encoded at Shaker locus of Drosophila. Science 237:770–775
Trautwein W, Cavalie A, Flockerzi V, Hoffmann F, Pelzer D (1987) Modulation of calcium channel function by phosphorylation in guinea pig ventricular cells and phospholipid bilayer membrane. Circ Res 61 (Suppl I):I 17-I 23
Tytgat J, Vereecke J, Carmeliet E (1990) A combined study of sodium current and T-type calcium current in isolated cardiac cells. Pflügers Arch 417:142–148
Windisch H, Tritthart HA (1982) Isoproterenol, norepinephrine and phosphodiesterase inhibitors are blockers of the depressed fast Na+-system in ventricular muscle fibers. J Mol Cell Cardiol 14:431–434
Whitehouse S, Walsh DA (1982) Purification of a physiological form of the inhibitor protein of the cAMP-dependent protein kinase. J Biol Chem 257:6028–6032
Yatani A, Brown AM, Schwartz A (1986) Bepridil block of cardiac calcium and sodium channels. J Pharmacol Exp Ther 237:9–17
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Muramatsu, H., Kiyosue, T., Arita, M. et al. Modification of cardiac sodium current by intracellular application of cAMP. Pflugers Arch. 426, 146–154 (1994). https://doi.org/10.1007/BF00374682
Received:
Revised:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00374682