Properties and regulation of cardiac Cl− channel

J Mel

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Cell

Cd+

Cardiol

CHANNEL

24 (Supplement

REGULATION

I) (1992)

BY CYCLIC

NUCLEOTIDES

IN DEVELOPING

HEARTS

Nicholas Sperelakis, Noritsugu Tohse, and Hiroshi Masuda. Department of Physiology & Biophysics, University of Cincinnati College of Medicine, Cincinnati, OH 45267. Myocardial Ca2+ slow (L-type) channels have special properties, including dependence on metabolic energy, blockade by acidosis, and regulation by cyclic nucleotides. In early (3-day) developing embryonic chick heart cells, long-lasting openings of the L-type Ca 2+ channels (mode 2) occur naturally (Tohse & Sperelakis, 1990). Mode-2 occurs infrequently in adult cells. In whole-cell voltage clamp, ICa(L) density was 8.1 and 5.1 PA/cm2 in 3-d and 17-d cells, respectively, and slower inactivation occurred m 3-d cells. In single-channel recordings (cell-attached patch), 17-d cells showed infrequent High-P, mode-2 behavior, and the ensemble-averaged current was less (37%) and decayed faster. sweeps (PO > 0.25), reflecting mode-2, were 20.2% and 3.7% in 3-d and 17-d cells. Therefore, mode-2 behavior decreased with development. In 3-d heart cells, Bay K 8644 increased (a) peak amplitude of the ensemble-averaged current by 3.9-fold and (b) number of simultaneous openings from 2.6 to 4.4, suggesting that silent channels are activated. In 3-d cells, 8-br-cGMP inhibited single-channel activity and decreased ensemble-averaged currents from multichannel patches; 1 mM completely blocked (Tohse & Sperelakis, 1991). In 17-d ventricular myocytes, 1 mM 8-br-cGMP decreased peak current (IBa (68% at 15 min) (Wahler et al., 1990). Therefore, cGMP can inhibit basal Ca2+ current, as we1 I as CAMP-stimulated current. In myocytes from 18-d fetal rat hearts, a novel isoform of Ca2+ channel was found, in addition to the L-type Ca2+ channel. This novel Ca2+ current was blocked by Co2+, but not by L-, T- (tetramethrine), or N-type blockers (w-conotoxin). Thus, rat fetal cardiomyocytes have a unique type of Ca2+ channel (ICa(fe)), which diminishes during subsequent development.

S-06-5

DIVER!fXTY OF CARDIAC PERMEATION.

K+ CHANNELS

IN MECHANISMS

OF GATING

AND

E. Carmeliet, Lab. Physiolo y, University Leuven, Belgium. On the basis of gating, K 4 channels can be subdivided into voltage-operated and ligand-operated channels. Gating involves activation and eventually inactivation. Most voltage-operated K+ channels are activated on depolarization, the “inward rectifier” iKI responsible for the resting potential, on hyperpolarization. Ligand-operated channels are activated by an increase or decrease in local concentration of an ion or metabolite. The distinction between the two classes is not absolute since voltage-operated channels can be modulated by ligands and ligand-operated channels are sensitive to voltage. Gating of the iK1 channel is ve dependent on K+ and its activation on hyperpolarization may be caused b an increase in local K 7 concentration. Cs % ions, although not permeating, can substitute for K Y as the activating ion. Permeation of K+ channels is characterized by the existence of inward or outward rectification. Outward rectification, predicted by the constant field hypothesis, is

shown by large conductance K+ channels such as the ATPi-, Na+i- and fatty acid-sensitive channels. Inward rectification shown by the other K+ than els, has been explained on the basis of a change in single channel conductance through block by Mg f+ combined in the case of iK1 and the delayed K channel, with a decrease in open probability (deactivation or fast inactivation).

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PROPERTIES AND REGULATION OF CARDIAC Cl- CHANNEL Tsuguhisa Ehara. Hiroshi Matsuura. Department of Physiology. Saga Medical School, Saga 849, Japan. g-adrenergic stimulation induces a Cl- current (lCl) in cardiac cells. This response involves intracellular cyclic AMP-dependent protein kinase (PKA). In line with the idea that phosphorylation of the channel protein or related structure activates the Cl- channel. The macroscopic 1 shows outward rectification, and is time-independent. Thus the !Cl will construct a steady backgroun 8’ current during cell excitation. When maximally activated, the Cl conductance, as relative to capacitive area of the membrane, increases to more than 0.1 mS/pF. the value large enough to attenuate the action potential configuration, and hence other voltage-dependent membrane currents. Single Cl- channel currents can be recorded with cell-attached patch clamp when the cell is exposed to p-stimulants. or when cyclic AMP is applied intracellularly. The channel is practically voltage-independent. and its I-V relation shows outward rectification with a unit conductance of about 13 pS for outward current. The most characteristic feature of the Cl- channel is its very slow open-close kinetics, with a mean open time of 0.5 to I sec. Analysis of the single channel activities that are induced by intracellular application of cyclic AMP suggests that the cyclic AMP system creates an available state of the channel without influencing its kinetic behaviour. The latter is likely to be controlled by an intrinsic mechanism. Besides the regulation by PKA, another regulatory mechanism has recently been found for the cardiac I on the whole cell level. Purinoceptor stimulation by ATP or other adenine nucleotides such as ADP. A& and adenosine also activates IC,, Like the PKA-regulated lc , purinoceptor-stimulated It., appears to be independent of the intracellular Ca 2+ level. The subcellu I ar mechanisms involved in this type of response remains to be elucidated.

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