Abstract
The calcium current (I Ca) in the heart is increased by phosphorylation of a protein which is part of, or close to, the Ca channel. The phosphorylation is catalysed by cAMP-dependent protein kinase (cAMP-PK). The question whether dephosphorylated channels are available to open on depolarization was examined in ventricular myocytes of guinea pig by recording whole cellI Ca during dialysis with either regulatory (R) subunit of cAMP-PK or protein kinase inhibitor (PKI) or adenosine-5′-(γ-thio)-triphosphate (ATPγS). The following results were obtained: 1) R subunit reduced and PKI reversed the isoprenaline (ISP)-induced enhancement ofI Ca, suggesting their ability to inhibit cAMP-PK. 2) R subunit and PKI, however, reduced basal (i.e. non β-adrenergically stimulated)I Ca only by about 20%. 3) Dialysis with ATPγS resulted in a slow increase in basalI Ca, presumably due to dephosphorylation-resistant thiophosphorylation. 4) When, however, the cell was dialyzed with PKI the effect of ATPγS was almost completely suppressed, suggesting no detectable phosphorylation related to the channel activity in this condition. These results support the view that even in the dephosphorylated state Ca channels are available to open on depolarization and that phosphorylation by cAMP-PK increases the opening probability.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ashby CD, Walsh DA (1972) Characterization of the interaction of a protein inhibitor with adenosine 3′, 5′-monophosphate-dependent protein kinase. J Biol Chem 247:6637–6641
Bean BP (1985) Two kinds of Ca channels in canine atrial cells: differences in kinetics, selectivity and pharmacology. J Gen Physiol 86:1–30
Bean BP, Nowycky MC, Tsien RW (1984) β-Adrenergic modulation of calcium channels in frog ventricular heart cells. Nature 307:371–375
Beavo JA, Bechtel PJ, Krebs EG (1974) Preparation of homogeneous cyclic AMP-dependent protein kinase and its subunits from rabbit skeletal muscle. Methods Enzymol 38:299–308
Bkaily G, Sperelakis N (1984) Injection of protein kinase inhibitor into cultured heart cells blocks slow calcium channels. Am J Physiol 246:H630-H634
Brum G, Flockerzi V, Hofmann F, Osterrieder W, Trautwein W (1983) Injection of catalytic subunit of cAMP-dependent protein kinase into isolated cardiac myocytes. Pflügers Arch 398:147–154
Brum G, Osterrieder W, Trautwein W (1984) β-Adrenergic increase in the calcium conductance of cardiac myocytes studied with the patch clamp. Pflügers Arch 401:111–118
Cachelin AB, De Peyer JE, Kokubun S, Reuter H (1983) Calcium channel modulation by 8-bromo-cyclics AMP in cultured heart cells. Nature 304:462–464
Cavalié A, Pelzer D, Trautwein W (1986) Fast and slow gating behaviour of single calcium channels in cardiac cells: Relation to activation and inactivation of calcium current. Pflügers Arch 406:241–258
Corbin JD, Keely SL (1977) Characterization and regulation of heart adenosine 3′:5′-monophosphate-dependent protein kinase isozymes. J Biol Chem 252:910–918
Dills WL, Beavo JA, Bechtel PJ, Krebs EG (1975) Purification of rabbit skeletal muscle protein kinase regulatory subunit using cyclic adenosine-3′:5′-monophosphate affinity columns.Biochem Biophys Res Commun 62:70–77
Hamill OP, Marty A, Neher E, Sakann B, Sigworth J (1981) Improved patch-clamp techniques for high resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100
Hofmann F (1980) Apparent constants for the interaction of regulatory and catalytic subunit of cAMP-dependent protein kinase I and II. J Biol Chem 255:1559–1564
Hofmann F, Beavo JA, Bechtel PJ, Krebs EG (1975) Comparison of adenosine-3′:5′-monophosphate-dependent protein kinases from rabbit skeletal and bovine heart muscle. J Biol Chem 250:7795–7801
Kameyama M, Hofmann 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
Khatra BS, Printz R, Cobb LE, Corbin JD (1985) Regulatory subunit of cAMP-dependent protein kinase inhibits phospho-protein phosphatase. Biochem Biophys Res Commun 130:567–573
Krebs EG, Beavo JA (1979) Phosphorylation-dephosphorylation of enzymes. Ann Rev Biochem 48:923–959
Nargeot J, Nerbonne J, Engels J, Lester H (1983) The time course of the increase in the myocardial slow inward current after a photochemically generated concentration jump of intracellular cAMP. Proc Natl Acad Sci USA 80:2395–2399
Nelson MT, French R, Kruger BK (1984) Voltage-dependent calcium channels from brain incorporated into planar lipid bilayers. Nature 308:77–80
Nilius B, Hess P, Lansman JB, Tsien RW (1985) A novel type of cardiac calcium channel in ventricular cells. Nature 316:443–446
Noma A (1983) ATP-regulated K+ channels in cardiac muscle. Nature 305:147–148
Osterrieder W, Brum G, Hescheler J, Trautwein W, Hofmann F, Flockerzi V (1982) Injection of subunits of cyclic AMP-dependent protein kinase into cardiac myocytes modulates Ca2+ current. Nature 298:576–578
Reuter H (1979) Properties of two inward membrane currents in the heart. Ann Rev Physiol 41:413–424
Reuter H (1983) Calcium channel modulation by neurotransmitters, enzymes and drugs. Nature 301:569–574
Reuter H, Stevens CF, Tsien RW, Yellen G (1982) Properties of single calcium channels in cardiac cell cultures. Nature 297:501–504
Rubin CS, Erlichman J, Rosen OM (1972) Molecular forms and subunit composition of a cyclic adenosine 3′,5′-monophosphate-dependent protein kinase purified from bovine heart muscle. J Biol Chem 247:36–44
Sperelakis N (1985) Phosphorylation hypothesis of the myocardial slow channels and control of Ca2+ influx. In: Zipes DP, Jalife J (eds) Cardiac electrophysiology and arrhythmias. Grune and Stratton, New York, pp 123–135
Trautwein W, Taniguchi J, Noma A (1982) The effect of intracellular cyclic nucleotides and calcium on the action potential and acetylcholine response of isolated cardiac cells. Pflügers Arch 392:307–314
Trautwein W, Kameyama M (1986) The mechanism of β-adrenergic regulation of Ca channels: Intracellular injection and patch-clamp studies. In: Noble D, Powell T (eds) Electrophysiology of single cardiac cells. Academic Press, London
Trautwein W, Pelzer D (1985) Voltage-dependent gating of single calcium channels in the cardiac cell membrane and its modulation by drugs. In: Marmé D (ed) Calcium and cell physiology. Springer, Berlin Heidelberg New York, pp 53–93
Trube G, Hescheler J (1984) Inward-rectifying channels in isolated patches of the heart cell membrane: ATP-dependence and comparison with cell-attached patches. Pflügers Arch 401:178–184
Author information
Authors and Affiliations
Additional information
This work was supported by the Deutsche Forschungsgemeinschaft, SFB 38 (Membranforschung), Projekt G, and HO 579/6-2
Rights and permissions
About this article
Cite this article
Kameyama, M., Hescheler, J., Hofmann, F. et al. Modulation of Ca current during the phosphorylation cycle in the guinea pig heart. Pflugers Arch. 407, 123–128 (1986). https://doi.org/10.1007/BF00580662
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00580662