Different Subtypes of α1-Adrenoceptor Modulate Different K+ Currents via Different Signaling Pathways in Canine Ventricular Myocytes

Abstract

Multiple subtypes (␣ 1A , ␣ 1B , and ␣ 1D ) of ␣ 1 -adrenoreceptors (␣ 1 ARs) co-exist in the heart and mediate a variety of cellular functions. We studied ␣ 1 AR modulation of inward rectifier (I K1 ) and transient outward (I to ) K ؉ currents in canine ventricular myocytes. Phenylephrine at 10 M depressed only I to without affecting I K1 and at 100 M inhibited both I to and I K1 . The effect of phenylephrine on I to was abolished by (؉)niguldipine (10 nM Over the past decade, evidence from pharmacological studies and molecular cloning has been accumulating indicating that ␣ 1 -adrenoreceptors (␣ 1 ARs) 1 are actually a heterogeneous group of distinct but related protein subsets. Many cellular responses to ␣ 1 ARs are mediated by multiple subtypes (␣ 1A , ␣ 1B , and ␣ 1D ) (1-4). In the heart, whereas the ␣ 1A and ␣ 1B subtypes have been well characterized, the presence of ␣ 1D AdR was indicated only recently (5-7). Moreover, although the pathophysiological roles of ␣ 1A and ␣ 1B receptors have been well appreciated, those of ␣ 1D subtype in the heart remain to be determined. Enhanced ␣ 1 AR activity has been implicated in various types of arrhythmias, particularly those in the pathogenesis of myocardial ischemia, ischemia-reperfusion and preconditioning, cardiac hypertrophy, etc. (1, 3). Drug intervention with ␣ 1 ARs has thus become an attractive issue for developing new compounds for potential therapy. A significant mechanism underlying ␣ 1 AR-induced alteration of cardiac electrical activity is attributable to the ability of ␣ 1 ARs to modulate ion channels. To date, no less than seven cardiac ionic currents are on the list of ␣ 1 AR modulation, including inward rectifier K ϩ current (I K1 ), transient outward K ϩ current (I to ), delayed rectifier K ϩ current (I K ), ultrarapid delayed rectifier K ϩ current (I Kur ), acetylcholine-induced K ϩ current (I KACh ), calcium current (I Ca ), and chloride current (1, 3, 8 -11). However, it is not known whether the effects are the results from participation of all three different subtypes of ␣ 1 ARs or of a particular individual subtype, although evidence is accumulating that different subtypes may have different roles in regulating cardiac contraction and electrical activities (12-16). Moreover, recent studies also demonstrated subtype differences in the signal transduction (17-19). In light of these studies, we speculated that different subtypes of ␣ 1 ARs may have distinct effects on ion channels. Understanding subtype specificity of ␣ 1 ARs in ion channel regulation is of theoretical and practical importance. K ϩ currents play critical roles in determining cardiac electrical activities. Besides stabilizing resting potential, I K1 in cardiac cells also plays an important role in modulating cellular excitability and regulating membrane repolarization, therefore an important determinant of action potential initiation. Another important cardiac K ϩ current is transient outward K ϩ current (I to ), which is known to be critical for initiating cardiac repolarization...

α 1 -Adrenoceptor stimulation potentiates L-type Ca 2+ current through Ca 2+ /calmodulin-dependent PK II (CaMKII) activation in rat ventricular myocytes

“…Western immunoblot analysis confirmed an increase in active CaMKII (see also ref. 13) in parallel with the potentiation of I Ca,L . Also, we showed that PKC, which is activated by the Gq-phospholipase C-diacylglycerol pathway (2), is involved in the activation of CaMKII during ␣ 1 ARS, based on the similar effects of CaMKII and PKC inhibition on I Ca,L in the presence of phenylephrine.…”

“…KN-93 and KN-92 were obtained from Calbiochem, and 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid (BAPTA)-acetoxymethyl ester was obtained from Molecular Probes. Antiphospho-CaMKII (anti-active CaMKII, rabbit polyclonal IgG raised against threonine phosphorylated peptide corresponding to the phosphorylated Thr-256 region of the mammalian CaMKII) was obtained from Promega (12,13). Anti-total CaMKII (mouse monoclonal IgG raised against amino acids 303-478 of CaMKII of mouse origin) was obtained from Santa Cruz Biotechnology (12).…”

“…The amount of CaMKII is low compared with other PKs; consequently, it is difficult to determine CaMKII activity directly in cardiac muscle (25). To overcome this difficulty, we used an antibody against the autophosphorylation site of CaMKII, Thr-286, and an enhanced chemiluminescence system to measure CaMKII activity in whole-cell lysates in response to ␣ 1 ARS (12,13). Active CaMKII increased significantly at concentrations of Ն1 M phenylephrine (Fig.…”

α1-Adrenoceptors stimulate a Gαs protein and reduce the transient outward K+ current via a cAMP/PKA-mediated pathway in the rat heart

“…Accordingly, in the present study, neither PLC nor PKC inhibition caused a significant reduction in the effect of PE on rat I to . On the other hand, it has been demonstrated that ␣ 1 -adrenoceptor stimulation reduces the amplitude of the cardiac I to through PKC activation in the dog (45). This difference could be due to the different channels that underlie I to in the different species.…”

Effects of bisindolylmaleimide PKC inhibitors on p90RSK activity in vitro and in adult ventricular myocytes

“…The use of these inhibitors, particularly GF109203X, in myocardial tissue and cells has implicated PKC-mediated signalling events in the regulation of physiological functions, such as contraction (Pi & Walker, 2000;Szokodi et al, 2002;von Lewinski et al, 2003) and protein synthesis (Ponicke et al, 1999), as well as a variety of pathophysiological processes, such as myocyte hypertrophy (Mullan et al, 1997;Ponicke et al, 1999;Ruf et al, 2002) and ischaemic cell death (Kitakaze et al, 1996;Chen et al, 1999;Inagaki et al, 2000;Fryer et al, 2001). In addition, GF109203X and Ro31-8220 have been used to investigate the roles of PKC in the regulation of sarcolemmal ion-transporting proteins, such as K þ (Hu et al, 1996;Wang et al, 2001a), Ca 2 þ (Woo & Lee, 1999;Hu et al, 2000) and Cl À (Middleton & Harvey, 1998;Duan et al, 1999) channels and the Na þ /K þ pump (Jo et al, 2000). These inhibitors have also been utilised by this and other laboratories to explore the involvement of PKC isoforms in the stimulation of the sarcolemmal Na þ /H þ exchanger (NHE1) by diverse stimuli, such as adrenergic (Puce´at et al, 1993;Snabaitis et al, 2000), thrombin (Yasutake et al, 1996), angiotensin (Gunasegaram et al, 1999) and opioid (Bian et al, 2000) receptor agonists, anaesthetic agents (Kanaya et al, 2001) and oxidative stress (Snabaitis et al, 2002).…”