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Noradrenaline up-regulates volume-regulated chloride current by PKA-independent cAMP/exchange protein activated by cAMP pathway in human atrial myocytes.

ABSTRACT: BACKGROUND AND PURPOSE:Adrenergic regulation of cell volume-regulated chloride current (ICl.vol ) is species-dependent. The present study investigates the mechanism underlying adrenergic regulation of ICl.vol in human atrial myocytes. EXPERIMENTAL APPROACH:Conventional whole-cell patch voltage-clamp techniques were used to record membrane current in human atrial myocytes. ICl.vol was evoked by hyposmotic bath solution (0.6 times isosmotic, 0.6 T). KEY RESULTS:ICl.vol was augmented by noradrenaline (1 μM) during cell swelling in 0.6 T but not under isosmotic (1 T) conditions. Up-regulation of ICl.vol in 0.6 T was blocked by the β-adrenoceptor antagonist propranolol (2 μM), but not by the α1 -adrenoceptor antagonist prazosin (2 μM). This β-adrenergic response involved cAMP but was independent of PKA; the protein kinase inhibitor H-89 (2 μM) or PKI (10 μM in pipette solution) failed to prevent ICl.vol up-regulation by noradrenaline. Moreover, the PI3K/PKB inhibitor LY294002 (50 μM) and the PKG inhibitor KT5823 (10 μM) did not affect noradrenaline-induced increases in ICl.vol . Interestingly, the exchange protein directly activated by cAMP (Epac) agonist 8-pCPT-2'-O-Me-cAMP (50 μM) also up-regulated ICl.vol , and the noradrenaline-induced increase of ICl.vol in 0.6 T was reversed or prevented by the Epac inhibitor ESI-09 (10 μM). CONCLUSION AND IMPLICATIONS:These data show that ICl.vol evoked by cell swelling of human atrial myocytes is up-regulated by noradrenaline via a PKA-independent cAMP/Epac pathway in human atrial myocytes. cAMP/Epac-induced ICl.vol is expected to shorten action potential duration during human atrial myocytes swelling and may be involved in abnormal cardiac electrical activity during cardiac pathologies that evoke β-adrenoceptor signalling.

SUBMITTER: Xiao GS

PROVIDER: S-EPMC6057899 | biostudies-other | 2018 Aug

REPOSITORIES: biostudies-other

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Noradrenaline up-regulates volume-regulated chloride current by PKA-independent cAMP/exchange protein activated by cAMP pathway in human atrial myocytes.

Xiao Guo-Sheng GS Zhang Yan-Hui YH Wang Yan Y Sun Hai-Ying HY Baumgarten Clive M CM Li Gui-Rong GR

British journal of pharmacology 20180708 16

<h4>Background and purpose</h4>Adrenergic regulation of cell volume-regulated chloride current (I<sub>Cl.vol</sub> ) is species-dependent. The present study investigates the mechanism underlying adrenergic regulation of I<sub>Cl.vol</sub> in human atrial myocytes.<h4>Experimental approach</h4>Conventional whole-cell patch voltage-clamp techniques were used to record membrane current in human atrial myocytes. I<sub>Cl.vol</sub> was evoked by hyposmotic bath solution (0.6 times isosmotic, 0.6 T).< ...[more]

PMID: 29900525

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Project description:We previously reported in atrial myocytes that inhibition of cAMP-dependent protein kinase (PKA) by laminin (LMN)-integrin signaling activates ?2-adrenergic receptor (?2-AR) stimulation of cytosolic phospholipase A2 (cPLA2). The present study sought to determine the signaling mechanisms by which inhibition of PKA activates ?2-AR stimulation of cPLA2. We therefore determined the effects of zinterol (0.1 ?M; zint-?2-AR) to stimulate ICa,L in atrial myocytes in the absence (+PKA) and presence (-PKA) of the PKA inhibitor (1 ?M) KT5720 and compared these results with atrial myocytes attached to laminin (+LMN). Inhibition of Raf-1 (10 ?M GW5074), phospholipase C (PLC; 0.5 ?M edelfosine), PKC (4 ?M chelerythrine) or IP3 receptor (IP3R) signaling (2 ?M 2-APB) significantly inhibited zint-?2-AR stimulation of ICa,L in-PKA but not +PKA myocytes. Western blots showed that zint-?2-AR stimulation increased ERK1/2 phosphorylation in-PKA compared to +PKA myocytes. Adenoviral (Adv) expression of dominant negative (dn) -PKC?, dn-Raf-1 or an IP3 affinity trap, each inhibited zint-?2-AR stimulation of ICa,L in + LMN myocytes compared to control +LMN myocytes infected with Adv-?gal. In +LMN myocytes, zint-?2-AR stimulation of ICa,L was enhanced by adenoviral overexpression of wild-type cPLA2 and inhibited by double dn-cPLA2S505A/S515A mutant compared to control +LMN myocytes infected with Adv-?gal. In-PKA myocytes depletion of intracellular Ca2+ stores by 5 ?M thapsigargin failed to inhibit zint-?2-AR stimulation of ICa,L via cPLA2. However, disruption of caveolae formation by 10 mM methyl-?-cyclodextrin inhibited zint-?2-AR stimulation of ICa,L in-PKA myocytes significantly more than in +PKA myocytes. We conclude that inhibition of PKA removes inhibition of Raf-1 and thereby allows ?2-AR stimulation to act via PKC?/Raf-1/MEK/ERK1/2 and IP3-mediated Ca2+ signaling to stimulate cPLA2 signaling within caveolae. These findings may be relevant to the remodeling of ?-AR signaling in failing and/or aging heart, both of which exhibit decreases in adenylate cyclase activity.

Dataset Information

Noradrenaline up-regulates volume-regulated chloride current by PKA-independent cAMP/exchange protein activated by cAMP pathway in human atrial myocytes.

Publications

Noradrenaline up-regulates volume-regulated chloride current by PKA-independent cAMP/exchange protein activated by cAMP pathway in human atrial myocytes.

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