The Ca-channel gating behaviour during steady and stepwise depolarization was examined in recordings of single Ca-channel activity from cell-attached membrane patches of single ventricular cells isolated enzymatically from hearts of adult guinea pigs. The single-channel recordings were performed by means of the improved patch-clamp technique (Hamill et al. 1981) with 90 mM Ba in the pipettes.
Upon step depolarization, two types of current records were regularly observed in the ensembles: (1) traces with Ca-channel activity (in the form of closely-spaced brief pulses of inward current with a unitary amplitude) of various length, and (2) blank sweeps without any detectable single-channel opening. The records with Ca-channel activity show a distinct tendency for openings to occur towards the beginning of the clamp pulse, followed by long periods of silence. The blank sweeps seem to reflect a condition or conditions where the Ca channel is unavailable for opening. The corresponding ensemble mean currentI(t) displayed a rapid rising phase to its peak followed by a slow decay.
During steady depolarization, kinetic analysis of the distributions of all open and shut lifetimes revealed a monoexponential probability density distribution function of all open times. By contrast, more than two exponential terms were required for an accurate description of the frequency distribution of all shut lifetimes. Corresponding to the two well-separated fast closed time components, individual Ca-channel openings were grouped into bursts of openings. The bursting behaviour reflected fast gating transitions and was related to the fluctuations of the Ca channel between two short-lived closed states and one open state. This fast gating was terminated by the entrance of the Ca channel into at least one long-lived closed state, exit from which was slow in comparison to the rapid cycling. As consequence, bursts of openings were further grouped together in clusters of bursts, the cluster behaviour being related to slow gating transitions in the kinetics of the Ca channel.
The biphasic frequency distribution of the first latencies (resulting from the transit through the two short-lived shut states, before the open state is entered) superimposed on the first time derivative of the rising phase of the ensemble mean current,I(t), upon step depolarization. The time constant of the monoexponential distribution function of all cluster lifetimes matched the declining phase ofI(t) during maintained depolarization. Thus, the decrease of the probability of channel opening and the resulting decline ofI(t) seemed to be due to a transition of the Ca channel into the long-lived third class of shut state(s).
The responsiveness of the Ca channel in a series of trials decreased at positive holding potentials with a sigmoidal dependence on the potential of the conditioning depolarization due to an increasing number of blank single-channel current records within the ensembles. Traces with channel openings as well as blank sweeps tended to form sequences of consecutive single-channel current records upon conditioning depolarization. The Ca channel was only activatable, if a command pulse was applied during the occupancy of either the open state or the two short-lived shut states. If the long-lived shut state(s) was already occupied at the conditioning potential preceding the step depolarization, the Ca channel was unavailable for opening and a blank sweep was observed upon the voltage pulse.
A quantitative patch-to-patch variability in Ca-channel gating behaviour was detected. It was interpreted as the statistical deviation from the average kinetic behaviour of a single population of Ca channels.
The total time course of the ensemble mean currentI(t) was reconstructed by a convolution of the first latency and cluster lifetime distribution functions. The peak amplitude ofI(t) was mainly determined by the steady-state occupancy of the activatable states of the Ca channel.
Under comparable experimental conditions (90 mM external Ba), the pooled average behaviour of individual Ca channels in different membrane patches was the same as the bulk behaviour of all the Ca channels in the cardiac cell.
The fast and slow Ca-channel gating transitions are discussed in terms of a channel-state model where, according to lifetimes and transition rates, the respective channel states are divided into two subsets.
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Aldrich RW, Corey DP, Stevens CF (1983) A reinterpretation of mammalian sodium channel gating based on single channel recording. Nature (Lond) 306:436–441
Bean BP, Nowycky MC, Tsien RW (1983) Electrical estimates of Ca channel density in heart cell membranes. Biophys J 41:295a
Bean BP, Nowycky MC, Tsien RW (1984) β-Adrenergic modulation of calcium channels in frog ventricular heart cells. Nature (Lond) 307:371–375
Brown AM, Morimoto K, Tsuda Y, Wilson DL (1981) Calcium current-dependent and voltage-dependent inactivation of calcium channels inHelix aspersa. J Physiol (Lond) 320:193–218
Brown AM, Camerer H, Kunze DL, Lux HD (1982) Similarity of unitary Ca2+ currents in three different species. Nature (Lond) 299:156–158
Brown AM, Tsuda Y, Wilson DL (1983) A description of activation and conduction in calcium channels based on tail and turn-on current measurements in the snail. J Physiol (Lond) 344:549–583
Brown AM, Lux HD, Wilson DL (1984) Activation and inactivation of single calcium channels in snail neurons. J Gen Physiol 83:751–769
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 studies with the patch clamp. Pflügers Arch 401:111–118
Cachelin AB, de Peyer JE, Kokubun S, Reuter H (1983) Ca2+ channel modulation by 8-bromocyclic AMP in cultured heart cells. Nature (Lond) 304:462–464
Carbone E, Lux HD (1984) A low voltage-activated, fully inactivating Ca channel in vertebrate sensory neurons. Nature (Lond) 310:501–502
Cavalié A, Ochi R, Pelzer D, Trautwein W (1983) Elementary currents through Ca2+ channels in guinea pig myocytes. Pflügers Arch 398:284–297
Cavalié A, McDonald TF, Pelzer D, Trautwein W (1985a) Temperature-induced transitory and steady-state changes in the calcium current of guinea pig ventricular myocytes. Pflügers Arch 405:294–296
Cavalié A, Pelzer D, Trautwein W (1985b) Does the inactivation of cardiac calcium channels depend on divalent cation entry? Pflügers Arch 403:R48
Colquhoun D, Hawkes AG (1982) On the stochastic properties of bursts of single ion channel openings and of clusters of bursts. Phil Trans R Soc Lond B 300:1–59
Colquhoun D, Sakmann B (1983) Bursts of openings in transmitter-activated ion channels. In: Sakmann B, Neher E (eds) Single-channel recording. Plenum Press, New York London, pp 345–364
Colquhoun D, Sigworth FJ (1983) Fitting and statistical analysis of single-channel records. In: Sakmann B, Neher E (eds) Single-channel recording. Plenum Press, New York London, pp 191–264
Eckert R, Chad JE (1984) Inactivation of Ca channels. Prog Biophys Mol Biol 44:215–267
Feller W (1968) An introduction to probability theory and its applications, 3rd edn, vol I. Wiley, New York
Fenwick EM, Marty A, Neher E (1982) Sodium and calcium channels in bovine chromaffin cells. J Physiol (Lond) 331:599–635
Fox AP, Hess P, Lansman JB, Nowycky MC, Tsien RW (1984) Slow variations in the gating properties of single calcium channels in guinea-pig heart cells, chick neurones and neuroblastoma cells. J Physiol (Lond) 353:75P
Hagiwara S, Byerly L (1981) Calcium channel. Annu Rev Neurosci 4:69–125
Hagiwara S, Ohmori H (1983) Studies of single calcium channel currents in rat clonal pituitary cells. J Physiol (Lond) 336:649–661
Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100
Hess P, Tsien RW (1984) Mechanism of ion permeation through calcium channels. Nature (Lond) 309:453–456
Hess P, Lansman JB, Tsien RW (1984) Different modes of Ca channel gating behaviour favoured by dihydropyridine Ca agonists and antagonists. Nature (Lond) 311:538–544
Horn R, Vandenberg CA, Lange K (1984) Statistical analysis of single sodium channels. Effects of N-bromoacetamide. Biophys J 45:323–335
Isenberg G, Klöckner U (1982) Calcium currents of isolated bovine ventricular myocytes are fast and of large amplitude. Pflügers Arch 395:30–41
Jackson MB, Wong BS, Morris CE, Lecar H, Christian CN (1983) Successive openings of the same acetylcholine receptor channel are correlated in open time. Biophys J 42:109–114
Josephson IR, Sanchez-Chapula J, Brown AM (1984) A comparison of calcium currents in rat and guinea pig single ventricular cells. Circ Res 54:144–156
Kass RS, Sanguinetti MC (1984) Inactivation of calcium channel current in the calf cardiac Purkinje fiber. Evidence for voltage-and calcium-mediated mechanisms. J Gen Physiol 84:705–726
Lee KS, Tsien RW (1982) Reversal of current through calcium channels in dialysed single heart cells. Nature (Lond) 297:498–501
Lee KS, Tsien RW (1984) High selectivity of calcium channels in single dialysed heart cells of the guinea-pig. J Physiol (Lond) 354:253–272
Lee KS, Marban E, Tsien RW (1985) Inactivation of calcium channels in mammalian heart cells: Joint dependence on membrane potential and intracellular calcium. J Physiol (Lond) 364:395–411
Lux HD, Brown AM (1984a) Patch and whole cell calcium currents recorded simultaneously in snail neurones. J Gen Physiol 83:727–750
Lux HD, Brown AM (1984b) Single channel studies on inactivation of calcium currents. Science 225:432–434
McDonald TF (1982) The slow inward calcium current in the heart. Annu Rev Physiol 44:425–434
McDonald TF, Cavalié A, Trautwein W, Pelzer D (1986) Voltage-dependent properties of macroscopic and elementary calcium channel currents in guinea pig ventricular cells. Pflügers Arch (in press)
Mitchell MR, Powell T, Terrar DA, Twist VW (1983) Characteristics of the second inward current in cells isolated from rat ventricular muscle. Proc R Soc Lond B 219:447–469
Niedergerke R, Page S (1977) Analysis of catecholamine effects in single atrial trabeculae of the frog heart. Proc R Soc Lond B 197:333–362
Ochi R, Hino N, Niimi Y (1984) Prolongation of calcium channel open time by the dihydropyridine derivative BAY K 8644 in cardiac myocytes. Proc Jpn Acad B 60:153–156
Osterrieder W, Brum G, Hescheler J, Hofmann F, Flockerzi V (1982) Injection of subunits of cyclic AMP-dependent protein kinase into cardiac myocytes modulates Ca2+ current. Nature (Lond) 298:576–578
Pelzer D, Cavalié A, Trautwein W (1985a) Cardiac Ca channel currents at the level of single cells and single channels. Basic Res Cardiol 80 (Suppl 2) 65–70
Pelzer D, Cavalié A, Trautwein W (1985b) Modulation of the gating properties of single calcium channels in cardiac cell membranes by D600. In: Mironneau J (ed) Calcium regulations in smooth muscles, vol 124. INSERM, Paris, pp 415–424
Pelzer D, Cavalié A, Trautwein W (1985c) Activation and inactivation of single calcium channels in cardiac cells. Exp Brain Res (Suppl) (in press)
Pelzer D, Hescheler J, Cavalié A, Trautwein W (1985d) Electrical estimate of calcium channel density in single ventricular cells from hearts of adult guinea pigs. Pflügers Arch 403:R48
Reuter H (1979) Properties of two inward membrane currents in the heart. Annu Rev Physiol 41:413–424
Reuter H (1983) Calcium channel modulation by neurotransmitters, enzymes and drugs. Nature (Lond) 301:569–574
Reuter H (1984) Ion channels in cardic cell membranes. Annu Rev Physiol 46:473–484
Reuter H, Stevens CF, Tsien RW, Yellen G (1982) Properties of single calcium channels in cardiac cell culture. Nature (Lond) 297:501–504
Sakmann B, Patlak J, Neher E (1980) Single acetylcholine-activated channels show burst-kinetics in presence of densensitizing concentrations of agonist. Nature (Lond) 286:71–73
Standen NB, Stanfield PR (1982) A binding-site model for calcium channel inactivation that depends on calcium entry. Proc R Soc Lond B 217:101–110
Trautwein W, Pelzer D (1985a) 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 Tokyo, pp 53–93
Trautwein W, Pelzer D (1985b) Gating of single calcium channels in the membrane of enzymatically isolated ventricular myocytes from adult mammalian hearts. In: Zipes DP, Jalife J (eds) Cardiac electrophysiology and arrhythmias. Grune and Stratton, Orlando London, pp 31–42
Tsien RW (1983) Calcium channels in excitable cell membranes. Annu Rev Physiol 45:341–358
Tsien RW, Bean BP, Hess P, Nowycky MC (1983) Calcium channels: Mechanisms of β-adrenergic modulation and ion permeation. Cold Spring Harbor Symp Quant Biol 47:201–212
Vandenberg CA, Horn R (1984) Inactivation viewed through single sodium channels. J Gen Physiol 84:535–564
Yatani A, Wilson DL, Brown AM (1983) Recovery of Ca currents from inactivation: the roles of Ca influx, membrane potential, and cellular metabolism. Cell Mol Neurobiol 3:381–395
Supported by the Deutsche Forschungsgemeinschaft (DFG), SFB 38 (Membranforschung), project G1
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Cavalié, A., Pelzer, D. & Trautwein, W. Fast and slow gating behaviour of single calcium channels in cardiac cells. Pflugers Arch. 406, 241–258 (1986). https://doi.org/10.1007/BF00640910
- Single cardiac ventricular cells
- Single Ca-channel gating behaviour
- Reconstruction of ensemble average Ca-channel currents
- Relation to whole-cell Ca-channel current