Adenosine-A2a Receptor Down-Regulates Cerebral Smooth Muscle L-Type Ca2+ Channel Activity via Protein Tyrosine Phosphatase, Not cAMP-Dependent Protein Kinase

• Published in: Molecular pharmacology 2003-09, Vol.64 (3), p.640-649
• Main Authors: Murphy, Katrina, Gerzanich, Volodymyr, Zhou, Hui, Ivanova, Svetlana, Dong, Yafeng, Hoffman, Gloria, West, G Alexander, Winn, H Richard, Simard, J Marc
• Format: Article
• Language: English
• Subjects: Animals; Basilar Artery - drug effects; Basilar Artery - metabolism; Calcium Channels, L-Type - metabolism; Cerebrovascular Circulation - drug effects; Cerebrovascular Circulation - physiology; Cyclic AMP-Dependent Protein Kinases - antagonists & inhibitors; Cyclic AMP-Dependent Protein Kinases - metabolism; Down-Regulation - drug effects; Down-Regulation - physiology; Enzyme Inhibitors - pharmacology; Female; Muscle, Smooth, Vascular - drug effects; Muscle, Smooth, Vascular - metabolism; Protein Tyrosine Phosphatases - metabolism; Rats; Rats, Wistar; Receptor, Adenosine A2A; Receptors, Purinergic P1 - physiology
• ISSN: 0026-895X; 1521-0111
• DOI: 10.1124/mol.64.3.640

Adenosine acting via A2a receptors (A2aR) is a potent cerebral vasodilator that relaxes vascular smooth muscle cells (VSMCs) by a mechanism attributed to activation of cAMP-dependent protein kinase (cAK). We examined effects of adenosine and its mechanism of action on L-type Ca 2 + channels in native VSMCs from rat basilar artery. Reverse transcription-polymerase chain reaction and immunofluorescence imaging confirmed transcription and expression of A2aR, and in situ hybridization confirmed presence of mRNA for L-type Ca v1.2b channels. In patch-clamp experiments, adenosine down-regulated Ca 2 + channel currents in a concentration-dependent manner, with receptor-subtype-specific antagonists [4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo-[2,3- a ][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM-241385) versus 1,3-dipropyl-8-cyclopentyl-1,3-dipropylxanthine] showing that this was caused by action of A2aR. Down-regulation of channel currents was mimicked by stimulation of cGMP-dependent protein kinase (cGK; 8-Br-cGMP) and by inhibition of tyrosine kinase (AG-18) but not by stimulation of cAK [forskolin and 8-bromo-cAMP (8-Br-AMP)]. Down-regulation of currents by the A2aR agonist 2-[p-(2-carboxyeth yl)phenylethylamino]-5′- N -ethyolcarboxamidoadenosine (CGS-21680) was blocked by inhibiting protein tyrosine phosphatase (PTP; orthovanodate and dephostatin), but not by inhibiting cGK (KT-5823 and H-7). Western blots of lysate or of immunoisolated Ca 2 + channels from arterial segments incubated with CGS-21680 showed 1) increased phosphorylation of vasodilator-stimulated phosphoprotein that was blocked by inhibiting cAK (KT-5720), consistent with activation of cAK by A2aR; and 2) decreased tyrosine phosphorylation of immunoisolated α1c subunit of the Ca 2 + channel. Our data show that cAK, although activated, was not germane to down-regulation of Ca 2 + channel activity by A2aR, and they delineate a novel signaling mechanism involving reduced tyrosine phosphorylation of Ca 2 + channels by A2aR probably caused by PTP activation.