Channels carrying slow inward Ca current are vital to the rhythm and contraction of the healthy heart, and play an important part in arrhythmias. We will study the basic properties of cardiac Ca channels, their inhibition by """"""""Ca antagonists"""""""", and their modulation by beta-adrenergic and cholinergic neurotransmitters. We will take advantage of two new and powerful approaches: the dialyzed cell method and the patch clamp technique. As we have shown in recent papers, it is possible to use these methods to record Ca channel activity from single heart cells, or even individual channels. The methods will also allow delivery of drugs, enzymes, or putative messenger molecules to the inside surface of the cell membrane. These approaches will help answer some fundamental questions. What is the ion flux through a single open Ca channel? How many functional Ca channels does a single cell have, and does the number change under the influence of neurohormones? Does the Ca channel open by a single one-step process or a multi-step mechanism? How is Ca channel inactivation affected by intracellular Ca and membrane potential? Resolving these unsettled physiological issues will help answer some important pharmacological questions. Although """"""""Ca antagonists"""""""" have already achieved clinical prominence, their basic mechanism(s) of action remain poorly understood. Do such drugs block current flowing outward through the channel as well as inward (as lidocaine or tetrodotoxin do in the case of the Na channel)? Do they act from inside the cell? Do they really compete with Ca ions, as the term """"""""antagonist"""""""" implies? Do they block open or inactivated channels preferentially? In answering these questions, we might find important differences between verapamil, diltiazem and nifedipine, and the classes of drug they represent, while also gaining basic insights into Ca channel function. We will pursue preliminary hints that isoproterenol increases Ca entry through changes in the percentage of time individual Ca channels stay open, and perhaps also through a slowing of Ca channel inactivation. We will find out whether dimetrically opposite mechanisms are at work when acetylcholine decreases Ca current. The possible involvement of cyclic nucleotides and protein phosphorylation in mediating the actions of beta-adrenergic and cholinergic agents will be tested at the level of single Ca channels.
