The inhibitory effect of β-stimulation on the Na/K pump current in guinea pig ventricular myocytes is mediated by a cAMP-dependent PKA pathway (original) (raw)
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2014
Adrenergic activation of L-type Ca 2+ and various K + currents is a crucial mechanism of cardiac adaptation; however, it may carry a substantial proarrhythmic risk as well. The aim of the present work was to study the timing of activation of Ca 2+ and K + currents in isolated canine ventricular cells in response to exposure to isoproterenol (ISO). Whole cell configuration of the patch-clamp technique in either conventional voltage clamp or action potential voltage clamp modes were used to monitor I Ca , I Ks , and I Kr , while action potentials were recorded using sharp microelectrodes. ISO (10 nM) elevated the plateau potential and shortened action potential duration (APD) in subepicardial and midmyocardial cells, which effects were associated with multifold enhancement of I Ca and I Ks and moderate stimulation of I Kr . The ISO-induced plateau shift and I Ca increase developed faster than the shortening of APD and stimulation of I Ks and I Kr . Blockade of β 1 -adrenoceptors (using 300 nM CGP-20712A) converted the ISO-induced shortening of APD to lengthening, decreased its latency, and reduced the plateau shift. In contrast, blockade of β 2 -adrenoceptors (by 50 nM ICI 118,551) augmented the APD-shortening effect and increased the latency of plateau shift without altering its magnitude. All effects of ISO were prevented by simultaneous blockade of both receptor types. Inhibition of phosphodiesterases decreased the differences observed in the turn on of the ISOinduced plateau shift and APD shortening. ISO-induced activation of I Ca is turned on faster than the stimulation of I Ks and I Kr in canine ventricular cells due to the involvement of different adrenergic pathways and compartmentalization.
Isoprenaline, Ca2+ and the Na(+)-K+ pump in guinea-pig ventricular myocytes
The Journal of physiology, 1992
1. The whole-cell patch clamp technique was employed to study the effects of the beta-agonist isoprenaline (ISO) on the Na(+)=K+ pump current, Ip, in acutely isolated ventricular myocytes from guinea-pig hearts. Propranolol, a beta-adrenergic antagonist, was used to demonstrate that all of the effects of ISO, stimulatory or inhibitory, are mediated by beta-receptors. 2. Below about 150 nM [Ca2+]i, we find that ISO reduces Ip, while above this [Ca2+]i ISO increases Ip. The stimulatory and inhibitory effects of ISO on Ip are independent of either intracellular sodium ([Na+]i) or extracellular potassium ([K+]o). These results suggest that the end-effect of ISO is directly on the maximum pump turnover rate (Vmax) rather than indirectly through changes in [Na+]i or [K+]o or modulatory effects on Na+ or K+ affinity. 3. The maximum effect of ISO increases Ip by 25% when [Ca2+] is buffered at 1.4 microM. A half-maximal effect is reached at roughly 10 nM-ISO and a near-maximal effect by 0.5 ...
The Journal of Physiology, 1999
1. Guinea-pig ventricle was used in the RNase protection assays to determine which á-isoforms of the Na¤-K¤ pumps are present, and ventricular myocytes were used in whole cell patch clamp studies to investigate the actions of á-and â-adrenergic agonists on Na¤-K¤ pump current. 2. RNase protection assays showed that two isoforms of the á-subunit of the Na¤-K¤-ATPase are present in guinea-pig ventricle. The mRNA for the á1-isoform comprises 82% of the total pump message, the rest being the áµ-isoform. 3. We have previously shown that â-adrenergic agonists affect Na¤-K¤ pump current (Ip) through a protein kinase A (PKA)-dependent pathway. We now show that these â-effects are targeted to the á1-isoform of the Na¤-K¤ pumps. 4. We have also previously shown that á-adrenergic agonists increase Ip through a protein kinase C (PKC)-dependent pathway. We now show that these á-isoform effects are targeted to the áµ-isoform of the Na¤-K¤ pumps. 5. These results suggest the effects of adrenergic activation on Na¤-K¤ pump activity in the heart can be regionally specific, depending on which á-isoform of the Na¤-K¤ pump is expressed. 8785
Activation of PKC increases Na + -K + pump current in ventricular myocytes from guinea pig heart
Pfl�gers Archiv European Journal of Physiology, 1999
We have previously shown activation of α 1adrenergic receptors increases Na + -K + pump current (I p ) in guinea pig ventricular myocytes, and the increase is eliminated by blockers of phosphokinase C (PKC). In this study we examined the effect of activators of PKC on I p . Phorbol 12-myristate 13-acetate (PMA), a PKC activator, increased I P at each test potential without shifting its voltage dependence. The concentration required for a half-maximal response (K 0.5 ) was 6 µM at 15 nM cytosolic [Ca 2+ ] ([Ca 2+ ] i ) and13 nM at 314 nM [Ca 2+ ] i . The maximal increase at either [Ca 2+ ] i was about 30%. Another activator of PKC, 1,2-dioctanoylsn-glycerol (diC 8 ), increased I p similarly. The effect of PMA on I P was eliminated by the PKC inhibitor staurosporine, but not by the peptide PKI, an inhibitor of protein kinase A (PKA). PMA and α 1 -adrenergic agonist effects both were sensitive to [Ca 2+ ] i , blocked by PKC inhibitors, unaffected by PKA inhibition, and increased I p uniformly at all voltages. However, they differed in that α 1 -activation caused a maximum increase of 15% vs 30% via PMA, and α 1 -effects were less sensitive to [Ca 2+ ] i than PMA effects. These results demonstrate that activation of PKC causes an increase in I p in guinea pig ventricular myocytes. Moreover, they suggest that the coupling of α 1 -adrenergic activation to I p is entirely through PKC, however α 1 -activation may be coupled to a specific population of PKC whereas PMA is a more global agonist.
Acetylcholine reversal of isoproterenol-stimulated sodium currents in rabbit ventricular myocytes
Circulation Research, 1993
We have recently shown that J3-adrenergic agonists enhance the cardiac sodium current (INa) in rabbits through dual G-protein regulatory pathways. To determine if muscarinic cholinergic receptor stimulation can also modulate INa, we studied the effects of acetylcholine (ACh) and carbachol on INa in enzymatically dispersed rabbit ventricular myocytes. Whole-cell patch-clamp experiments done at room temperature using 20 mM [Na+L] showed that 100 nM isoproterenol increased INa and accelerated current decay as previously described. ACh (1 ,uM) or carbachol (1 ,uM) significantly reversed the stimulatory isoproterenol effects at test potentials throughout the INa activation range and at holding potentials negative to-80 mV. This effect was completely inhibited by atropine (1 uM) and was confirmed by studying singlechannel INa from cell-attached patches. When INa was stimulated by forskolin (1 ,M), carbachol (1 ,uM) significantly reversed the effect. The muscarinic-mediated inhibition Of INa was inhibited by pertussis toxin (0.1 or 1.0 gg/ml) incubation (12-15 hours), suggesting that the effect was inhibitory G-protein dependent. Further investigation of the ACh inhibitory mechanism revealed that ACh alone had no effect on INa and that when cells were dialyzed with cAMP (5 ,uM), ACh failed to inhibit INa. Furthermore, cGMP failed to inhibit the effect of isoproterenol on INa. These data suggest that ACh acts at or proximal to adenylate cyclase stimulation. Thus, rabbit cardiac Na+ channels are regulated by muscarinic agonists in a fashion similar to cardiac Ca2' channels. (Circulation Research 1993;72:517-525) KEY WORDS * sodium channels * cardiac myocytes * muscarinic receptors * 3-adrenergic receptors I soproterenol (ISO), a p-adrenergic agonist, modulates several voltage-dependent ionic currents in the heart, including the calcium current (ICa), the delayed rectifier potassium current (IK), the transient outward potassium current (Ito), and the pacemaker current (If) (see Reference 1 for review). The ISOinduced modulation of these currents contributes to increases in cardiac contractile strength (positive inotropic effect) and in the rate of the heart beat (positive chronotropic effect) (see Reference 1). For example, the ISO-induced stimulation of 'Ca leads to elevated cytoplasmic Ca2' ([Ca2"I,) levels, causing an increase in cardiac contractility. We recently discovered that ISO also increases the voltage-dependent sodium current (INa) in the rabbit heart through dual G-protein regulatory pathways.2 The physiological significance of the ISO-induced increase in INa has not been determined. However, continuous cable theory predicts that enhancements of INa should increase cardiac impulse