alpha-Adrenoceptor-mediated depletion of phosphatidylinositol 4, 5-bisphosphate inhibits activation of volume-regulated anion channels in mouse ventricular myocytes.
OBJECTIVE
Volume-regulated anion channels (VRACs) play an important role in cell-volume regulation. alpha(1)-Adrenoceptor stimulation by phenylephrine (PE) suppressed the hypotonic activation of VRAC current in mouse ventricular cells and regulatory volume decrease (RVD) was also absent in PE-treated cells. We examined whether the effects of alpha(1)-adrenoceptor stimuli on VRAC current were modulated by phosphatidylinositol signalling.


METHODS
Whole-cell patch-clamp method was used to record the hypotonicity-induced VRAC current in mouse ventricular cells. RVD was analyzed by videomicroscopic measurement of cell images.


RESULTS
The attenuation of VRAC current by PE was suppressed by alpha(1A)-adrenoceptor antagonists (prazosin and WB-4101), anti-G(q) protein antibody and a specific phosphoinositide-specific phospholipase C (PLC) inhibitor (U-73122), but not by antagonists for alpha(1B)-, alpha(1D)- or beta-adrenoceptor, or protein kinase C inhibitors. The inhibition of VRAC by PE was antagonized by intracellular excess phosphatidylinositol 4,5-bisphosphate (PIP(2)), while intracellular anti-PIP(2) antibody (PIP(2) Ab) inhibited the activation of VRAC currents. When cells were loaded with phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) with or without PIP(2) Ab, PE little affected the VRAC current. Extracellular m-3M3FBS (an activator of PLC) suppressed VRAC in the absence of PE, and this effect was reversed by intracellular excess PIP(2).


CONCLUSIONS
Our results indicate that the stimulation of alpha(1A)-adrenoceptors by PE inhibited the activation of cardiac VRAC current via PIP(3) depletion brought about by PLC-dependent reduction of membrane PIP(2) level.
Ichishima
K
K
Department of Physiology, Faculty of Medicine, Saga University, Saga, Japan.
Yamamoto
S
S
Iwamoto
T
T
Ehara
T
T
D058646:Adrenergic alpha-1 Receptor Agonists / Q000494:pharmacology; D000818:Animals; D000906:Antibodies; D004146:Dioxanes / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D032383:Myocytes, Cardiac / Q000378:metabolism*; D010656:Phenylephrine / Q000494:pharmacology*; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000494:pharmacology*; D011224:Prazosin / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology; D011942:Receptors, Adrenergic, alpha / Q000378:metabolism*; D050994:Voltage-Dependent Anion Channels / Q000378:metabolism*
D058646:Adrenergic alpha-1 Receptor Agonists; D000906:Antibodies; D004146:Dioxanes; D004963:Estrenes; D019269:Phosphatidylinositol 4,5-Diphosphate; D011760:Pyrrolidinones; D011942:Receptors, Adrenergic, alpha; D050994:Voltage-Dependent Anion Channels; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D010656:Phenylephrine; C010654:(2-(2',6'-dimethoxy)phenoxyethylamino)methylbenzo-1,4-dioxane; D011224:Prazosin