- Email: [email protected]
K pump in cardiac cells
10
GATING OF SODIUM CHANNELS IN CARDIAC PURKINJE CELLS. H.A. Fozzard, D.A. Hanck, B.E. Stanley, M.F. Sheets Cardiac Electrophysiology Labs., The University of Chicago, Chicago, IL. Na current, responsible for cardiac excitation and conduction, has been poorly studied because voltage clamp is difficult in multicellular tissue. We have developed a Purkinje cell voltage clamp with intracellular perfusion, equivalent to perfused squid axon (Makielski et al, 1987). This was combined with single channel recordings to describe the kinetics of the Na channel. Compared to nerve, there is 1) frequent reopening 2) biphasic voltage dependence of open time, 3) prominent inactivation without opening. Channel voltage dependence resides in activation (-4e) with small voltage dependence of inactivation (OSe). Channel density is 2-3oO/pm2, and channel gating current confirmed the voltage dependency of activation (Hanck et al, 1988). Makielski, J.C., M.F. Sheets, D.A. Hanck, C.T. January, H.A. Fozzard, Biophys J. 52:1-11, 1987. Hanck, D.A., M.F. Sheets, H.A. Fozzard, Biophys J. 53:535a, 1988.
11
VOLTAGE DEPENDENCE OF TRANSIENT AND STEADY CURRENTS GENERATED BY THE Na/K PUMP IN CARDIAC CELLS. D.C. Gadsby, M. Nakao & A. Bahinski. Lab of Cardiac Physiol, Rockefeller Univ, New York. Na/K pump current was determined as strophanthidin-sensitive, whole-cell curtent in voltageclamped, internally-dialyzed myocytes from guinea pig ventricle. Wide-tipped pipettes, and a device for rapidly exchanging the solution inside the pipette, facilitated control of the compositions of intracellular and extracellular solutions as well as membrane potential Internal (pipette) and external solutions were designed to minimize K-channel, Ca-channel, and N&aexchange currents while sustaining various modes of Nan< pump activity. Transient pump currents could be elicited by voltage jumps in the absence of K ions but not in the absence of Na ions, indicating that Na translocation steps, but not K translocation steps, involve charge movement within the membrane field. Steady-state, forward (or reverse) Na/K pump currentvoltage relationships, from -140mV and +6OmV, at various internal [Na], and external [Na] and [K], were all monotonic with noregions of negative slope. Taken together, these results suggest that the Na/K pump cycle includes a single voltage-sensitive step, which is associated with Na translocation. Reduction of external [Na] shifted the Na/K pump current-voltage relationship along the voltage axis without changing the maximum pump current at large positive potentials, suggesting a direct influence of external [Na] on the voltage-sensitive step. This, in turn, implies that the voltage-dependent step involves the release of Na ions to the external medium.
12
CARDIAC PACEMAKER ACTIVITY IN SINGLE CELLS FROM BULLFROG SINUS VENOSUS. W. Giles, E.F. Shibata, JClark, R. Rasmussen, R. Murphy, Y. Momose Dept. of Medical Physiol. & Medicine, Univ. of Calgary, Canada. The ionic basis for pacemaker activity has been studied by combining experimental measurements (whole cell voltage clamp) and computer simulations of action potentials obtained from single cells of the sinus venosus and the atrium of the bulliiog, Rana catesbiana. The results were as follows: 1) No transient inward sodium currency 2) No inwardly rectifying background K’ current; 3) Long (5-10 set) hyperpolarizing clamp pulses did not activate any identifiable 16 4) Short (5-300 mse$ depolarizing clamp steps from a holding poydal of 60mV activated a transient inward Ca, current, I,. which was blocked py LaCla (10 M) and was enhanced by 2-or-3-fold mcreases in [Ca2’], and by isoproterenol (1c M); 5) Longer (0.5-10 set) depolarizations to the plateau range elicited a noninactivating 9and voltage-dependent outward ‘delayed rectitier’ current carrled by K+; 6) A strophanthidin (lo- M) sensitive outward current was recorded followmg 1Omm exposure to O[K+], Ringer’s; 7) Recordings consistently exhibited slowly decaying inward tails following activation of Ea. Hodgkin-Huxley descriptions of h and 1, toget$er +tith formulations for;) t% equivalent circuit of the quiescent cell, ii) $Fe ehactrogenic Na /K pump, m) the Na /Ca exchanger, and iv) intracellular buffering of Ca have been combined in an attempt to simulate the pacemaker activity potential results from an interaction of E,, I, and a background inward current.
GATING OF SODIUM CHANNELS IN CARDIAC PURKINJE CELLS. H.A. Fozzard, D.A. Hanck, B.E. Stanley, M.F. Sheets Cardiac Electrophysiology Labs., The University of Chicago, Chicago, IL. Na current, responsible for cardiac excitation and conduction, has been poorly studied because voltage clamp is difficult in multicellular tissue. We have developed a Purkinje cell voltage clamp with intracellular perfusion, equivalent to perfused squid axon (Makielski et al, 1987). This was combined with single channel recordings to describe the kinetics of the Na channel. Compared to nerve, there is 1) frequent reopening 2) biphasic voltage dependence of open time, 3) prominent inactivation without opening. Channel voltage dependence resides in activation (-4e) with small voltage dependence of inactivation (OSe). Channel density is 2-3oO/pm2, and channel gating current confirmed the voltage dependency of activation (Hanck et al, 1988). Makielski, J.C., M.F. Sheets, D.A. Hanck, C.T. January, H.A. Fozzard, Biophys J. 52:1-11, 1987. Hanck, D.A., M.F. Sheets, H.A. Fozzard, Biophys J. 53:535a, 1988.
11
VOLTAGE DEPENDENCE OF TRANSIENT AND STEADY CURRENTS GENERATED BY THE Na/K PUMP IN CARDIAC CELLS. D.C. Gadsby, M. Nakao & A. Bahinski. Lab of Cardiac Physiol, Rockefeller Univ, New York. Na/K pump current was determined as strophanthidin-sensitive, whole-cell curtent in voltageclamped, internally-dialyzed myocytes from guinea pig ventricle. Wide-tipped pipettes, and a device for rapidly exchanging the solution inside the pipette, facilitated control of the compositions of intracellular and extracellular solutions as well as membrane potential Internal (pipette) and external solutions were designed to minimize K-channel, Ca-channel, and N&aexchange currents while sustaining various modes of Nan< pump activity. Transient pump currents could be elicited by voltage jumps in the absence of K ions but not in the absence of Na ions, indicating that Na translocation steps, but not K translocation steps, involve charge movement within the membrane field. Steady-state, forward (or reverse) Na/K pump currentvoltage relationships, from -140mV and +6OmV, at various internal [Na], and external [Na] and [K], were all monotonic with noregions of negative slope. Taken together, these results suggest that the Na/K pump cycle includes a single voltage-sensitive step, which is associated with Na translocation. Reduction of external [Na] shifted the Na/K pump current-voltage relationship along the voltage axis without changing the maximum pump current at large positive potentials, suggesting a direct influence of external [Na] on the voltage-sensitive step. This, in turn, implies that the voltage-dependent step involves the release of Na ions to the external medium.
12
CARDIAC PACEMAKER ACTIVITY IN SINGLE CELLS FROM BULLFROG SINUS VENOSUS. W. Giles, E.F. Shibata, JClark, R. Rasmussen, R. Murphy, Y. Momose Dept. of Medical Physiol. & Medicine, Univ. of Calgary, Canada. The ionic basis for pacemaker activity has been studied by combining experimental measurements (whole cell voltage clamp) and computer simulations of action potentials obtained from single cells of the sinus venosus and the atrium of the bulliiog, Rana catesbiana. The results were as follows: 1) No transient inward sodium currency 2) No inwardly rectifying background K’ current; 3) Long (5-10 set) hyperpolarizing clamp pulses did not activate any identifiable 16 4) Short (5-300 mse$ depolarizing clamp steps from a holding poydal of 60mV activated a transient inward Ca, current, I,. which was blocked py LaCla (10 M) and was enhanced by 2-or-3-fold mcreases in [Ca2’], and by isoproterenol (1c M); 5) Longer (0.5-10 set) depolarizations to the plateau range elicited a noninactivating 9and voltage-dependent outward ‘delayed rectitier’ current carrled by K+; 6) A strophanthidin (lo- M) sensitive outward current was recorded followmg 1Omm exposure to O[K+], Ringer’s; 7) Recordings consistently exhibited slowly decaying inward tails following activation of Ea. Hodgkin-Huxley descriptions of h and 1, toget$er +tith formulations for;) t% equivalent circuit of the quiescent cell, ii) $Fe ehactrogenic Na /K pump, m) the Na /Ca exchanger, and iv) intracellular buffering of Ca have been combined in an attempt to simulate the pacemaker activity potential results from an interaction of E,, I, and a background inward current.