138 PART III INTERMOLECULAR INTERACTIONS AND PHARMACOLOGY OF CARDIAC ION CHANNELS effects appear to be derived from the fact that LTCC antagonists have a higher affinity for open or inactivated states of LTCCs. The antihypertensive and antianginal effects take advantage of the fact that the resting potential of smooth muscle is more depolarized than working cardiac myocytes. Thus, LTCC block and the associated vasodila-tion can be achieved without significant negative inotropy. The use-dependent nature of certain Ca2 + channel antago-nists (such as verapamil) makes them particularly effective in the treatment of atrial tachyarrhythmia. These topics are covered in detail in Chapter 98. • REGULATION OF L-TYPE Ca2+ CHANNELS BY PROTEIN KINASES AND PHOSPHATASES Phosphorylation of the LTCC complex by protein kinases and dephosphorylation by protein phosphatases are physi-ologically and pathologically relevant processes in cardiac myocytes. The primary physiologic regulation of LTCCs occurs via the ~-adrenergic signaling pathways through activation of PKA. Phosphorylation of the LTCC complex via this pathway causes an increase in Ca2 + influx, SR Ca2 + loading, and SR Ca2 + release and underlies the associated increase in cardi<1c contr<1ctility. Abnormalities of this sig-naling cascade are well described in cardiac diseases th<1t lead to congestive heart failure. Many clinically useful dmgs, particularly ~-adrenergic antagonists, are likely to impart a portion of their beneficial effects by influencing the phosphorylation state of the LTCC complex. Here we will briefly review the regulation of the LTCC complex via well-described signaling cascades with the understanding that dmgs that activate or block these pathways will impart at least a portion of their cardiac effects via their influence on the cardiac LTCCs. It should be noted in this regard that calmodulin-dependent kinase II (CaMKII) is an important regulator of the LTCC. However, these effects have been reviewed in Chapter 2 and will not be discussed further here. Effects of 13-Adrenergic (cyclic Adenosine Monophosphate/Protein Kinase A) Signaling Pathways on L-Type Ca2+ Channels Activation of the sympathetic nervous system is a major mechanism for controlling the rate and contractility of the normal heart. Catecholamines released from sympatlletic nerves bind to ~-adrenergic receptors on the cell surface, and this leads to phosphorylation of LTCC complex via <1ctivation of PKA (Fig. 16-4). TsienH initially proposed that the stimul<1tory effect of cyclic adenosine monophos-phate (cAMP) in heart cells was caused by PKA-mediated AMP FIGURE 16-4 • Scheme of the cAMP/PKA pathway for LTCC regUlation with (}-adrenergic system as an example. When an agonist (e,g" isoproterenol [ISO]) binds to ~-adrenergic receptors, ~,-and ~2-adrenergic receptor (P,-AR and ~2-AR), the associated stimulatory G proteins (Gs) are activated and inhibitory G~y subunits are dissociated from Gas, Activated Gas then diffuses to activate adenyl cyclase (AC) that is attached to cell membrane, Active AC catalyzes the production of cyclic adenosine monophosphate (cAMP) and therefore local cAMP concentration increases dramatically, An elevated cAMP concentration activates protein kinase A (PKA) that is believed to be anchored close to LTCC by A kinase-anchoring proteins (AKAPs) and subsequently PKA phosphorylates LTCC, This diagram shows PKA-dependent phosphorylation of the lX, subunit at the C-terminal tail (COOH). There are also PKA sites on ~2 but they are omitted for simplicity, Phosphorylated LTCC is dephosphorylated by protein phosphatase 2A (PP2A) and protein phosphatase 1 (PP1), The cAMP is cleaved by phosphodiesterases (PDE), Activation of the M2 cholinergic receptor may inhibit AC activity via G, proteins, Ach, acetylcholine,16 Pharmacology of L-Type and T-Type Calcium ChanneLs in the Heart 139 r+FTc ~ -16 -12 400 ms+1om:rL -70 mV F Test potential (mV) -80-60-40-20 0 20 40 60 80 100 ~,.-..., ..J.......+--'--'----L.:=l,--J relatively low specificity. Oilier neurotransmitters and hor-mones such as histamine, glucagons, parathyroid hormone, and serotonin can also modulate Ie.,-L in cardiomyocytes via the cAMP/PKA signal pathway. These effects are very small in comparison to those via ~-adrenergic signaling. Ser-1928 on (X.le subunit and SerA78 and/or SerA79 on the ~2a subunit are tll0ught to be the PKA sites on me LTCC complex that are phosphorylated to cause an increase in Ic,_L.37 HO'wever, this hypothesis has not been weIl established with direct measurements. In addition, the role of the C-terminus of CJ.1c> at which Ser-1928 is located, is controversial because this portion of the protein can be cleaved by proteases or truncated by alternative splicing. Interestingly, it has been shown that tlle cleaved cyto-plasmic C-tenninus of the LTCC: remains tethered to the membrane-imbedded (X.t subunit. lR The relative roles of CJ. 1 and ~2' phosphorylation in regulation of LTCCs also remain largely unestablished. "aguro and omersw found that Ser-1901 in the so-called rat brain type II CJ.1< (corre-sponding to Ser-1928 in cardiac (X.le) is responsible for the increase in Pu upon phosphorylation by PKA, and phosphorylation of other PKA sites mediated the leftward shift of voltage-dependent activation. These are important issues for cardiac Ca2+channels because they could lead to the development of calcium channel-specific drugs that specifically regulate LTCC phosphorylation. Another important issue is that PKA phosphorylation of LTCCs may require A kinase-anchoring proteins (AKAPs) to NF C o ~ A 10 pNpF ..J <Il U Test potential (mV) -80-60-40-20 0 20 40 60 80 100 l.H-~1IilT'l-+--'-----'--'--=--'--....J -16 B hll,phorylation ofLTCCs. These ideas were subsequently nhrmed in many other laboratories. PKA-dependent hosphOlylation of LTCCs causes a several-fold increase fc,l. and also shifts the voltage dependence of both til'ation and inactivation to more negative membrane tentials (Fig. 16-5, F). Because PKA-dependent phos-h()rylation prolongs single-channel open time, the whole IIlcl.L should decay more slowly. However, as discussed rlier, this effect is offset by an increase in lCl-L that romotes Ca2+-dependent inactivation. Single-channel 'periments have shown that PKA-dependent LTCC hosphorylation increases