Cyclic nucleotides depress action potentials in cultured aortic smooth muscle cells.

The effects of cyclic nucleotide analogs and related agents on the Ca2+ dependent action potentials of cultured rat aortic smooth muscle cells (reaggregates) were examined. The action potentials were elicited by electrical stimulation in the presence of tetraethylammonium (TEA, 5-15 mM). Superfusion of the aortic cells with analogs of cyclic AMP (dibutyryl or 8-bromo-cyclic AMP, 1 mM), isoproterenol (1-10 microM) and forskolin (1-10 microM) depressed and abolished the TEA-induced action potentials. Abolition of the action potentials by these agents was reversible and was accompanied by some hyperpolarization of the membrane. Superfusion with 8-bromo-cyclic GMP (0.1-1 mM) also depressed or abolished the TEA-induced action potentials, whereas dibutyryl cyclic GMP (1 mM) and sodium nitroprusside (10 microM) had little effect. Synthetic atrial natriuretic factor (0.01-0.1 microM) had inhibitory effects in most experiments. Thus, depression of membrane excitability may be a contributing factor in the relaxation of aortic smooth muscle produced by some agents that increase intracellular levels of cyclic nucleotides.

Effect of Bay K 8644 on tetraethylammonium-induced excitability of the rabbit pulmonary artery.

The effect of Bay K 8644 on the electrical activity of the smooth muscle cells in the main pulmonary artery of the rabbit was examined. In normal physiological solution, the resting membrane potential was -56 +/- 0.6 mV, and the cells were electrically quiescent. Tetraethylammonium (5 mM) depolarized the membrane to about -45 mV, and electrical stimulation elicited action potentials. To suppress contractile responses and thereby facilitate sustained impalements, the muscle strips were bathed with a hypertonic solution containing sucrose. The mean amplitude of the tetraethylammonium-induced action potentials in the hypertonic solution was 35 +/- 0.9 mV. The action potentials were dependent upon the extracellular Ca2+ concentration and were abolished by diltiazem (10(-6) M). Spontaneous action potentials were occasionally generated in the presence of tetraethylammonium alone and could be induced by the further addition of Ba2+ (0.5 mM). The Ca2+ agonist Bay K 8644 (10(-8) to 10(-6) M) had no effect on the resting membrane potential or excitability in normal solution. However, in the hypertonic solution containing tetraethylammonium, Bay K 8644 caused a further depolarization and oscillatory potential changes, which were not prevented by tetrodotoxin. The oscillations were suppressed or abolished by diltiazem or nilvadipine. Thus, active responses can occur in the normally quiescent smooth muscle cells of the rabbit pulmonary artery when the outward K+ current(s) are suppressed.

cAMP suppresses CA2+-dependent electrical activity of airway smooth muscle induced by TEA.

Using intracellular microelectrodes, we investigated whether exogenous dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP) or forskolin influenced the electrical effects of tetraethylammonium (TEA) on canine tracheal smooth muscle. We found that 20 mM TEA depolarized airway smooth muscle cells from a resting membrane potential (Em) of -59 +/- 4 mV (mean +/- SD) to -45 +/- 2 mV and caused spontaneous action potentials (AP's) to develop, which were 33 +/- 2 mV in amplitude. These were totally abolished in 0 Ca2+ solution. DBcAMP (1 mM) suppressed the development of this TEA-induced electrical activity and the phasic contractions electrically coupled to it. DBcAMP had no significant effect on Em in the absence of TEA however. Forskolin (1 microM) produced similar effects. Our findings suggest that Ca2+ is the principal ion responsible for the inward current associated with the TEA-induced AP's in airway smooth muscle, and that adenosine 3',5'-cyclic monophosphate may suppress the electrogenesis of this current.

8-Bromo-cyclic GMP abolishes TEA-induced slow action potentials in canine trachealis muscle.

Using intracellular microelectrodes, we investigated whether 8-bromo-guanosine 3':5'-cyclic monophosphate (cGMP) influenced the electromechanical effects of tetraethylammonium (TEA) on canine tracheal smooth muscle. We found that 20 mM TEA depolarized airway smooth muscle cells from -58 +/- 3 mV (means +/- S.D.) to -44 +/- 2 mV and caused spontaneous action potentials (APs) to develop which were 31 +/- 2 mV in amplitude. These APs, and the phasic contractions electrically coupled to them, were totally abolished in buffer containing 0.1 mM cGMP. Our findings suggest that cGMP markedly affects the channels mediating TEA-induced APs in airway smooth muscle.

Effect of methylated bepridil on slow action potentials in cardiac muscle and vascular smooth muscle.

The anti-anginal agent bepridil blocks slow Ca2+ channels in a variety of tissues. Since bepridil accumulates inside cells, the possibility exists that bepridil acts, at least partially, from inside the cell. To test this possibility, we examined the effects of a quaternary ammonium analog of bepridil, methylated bepridil, which presumably would enter the cells less readily, on the Ca2+-dependent slow action potentials of guinea pig papillary muscles (in 25 mM [K+]0) and rabbit pulmonary arteries (in tetraethylammonium chloride). In cardiac muscle, methylated bepridil had little effect on the slow action potentials at low stimulation frequencies (0.5 Hz), but at higher frequencies (1.0 and 2.0 Hz) the slow action potentials were depressed and/or the muscle was unable to follow each stimulation. These effects are similar to those obtained with bepridil, but bepridil was more potent than methylated bepridil. In vascular smooth muscle cells, methylated bepridil inhibited the slow action potentials at a somewhat lower dose than bepridil. We conclude that, in cardiac muscle, bepridil probably has two sites of action, one outside the cell (presumably on or associated with the slow Ca2+ channel) and a second site inside the cell. On the other hand, in vascular smooth muscle cells, bepridil may act only on an external site.

New dihydropyridine drug, nilvadipine, blocks the calcium slow action potential in rat-cultured aortic smooth muscle cells.

The effects of the dihydropyridine analogs, nilvadipine (FR-34235) and mesudipine, on the electrical activity of rat aortic smooth muscle cells in culture (reaggregates) were compared with the calcium antagonist verapamil. Nilvadipine blocked the tetraethylammonium-induced action potentials (APs), whose inward current is carried almost exclusively by Ca2+ through voltage-dependent slow channels. The effects of nilvadipine were dose dependent, and nilvadipine had a more potent inhibitory effect on the K+-induced contraction than on the norepinephrine-induced contraction of rabbit aorta. The ED50 value for blockade of the K+-induced contracture by nilvadipine was 6.4 X 10(-8) M, and complete blockade of the Ca2+ slow channels occurred at 10(-8) M. Mesudipine also inhibited the Ca2+ slow channels in cultured vascular smooth muscle cells in a dose-dependent manner; elevation of the [Ca]O from 1.8 to 5.4 mM partially restored the slow APs. The order of the inhibitory action on the Ca2+-dependent slow APs was: nilvadipine greater than mesudipine greater than verapamil. The inhibition of Ca2+ influx during excitation by the drugs can account for their vasodilatory properties.