Iloprost inhibits inositol-1,4,5-trisphosphate-mediated calcium mobilization stimulated by angiotensin II in cultured preglomerular vascular smooth muscle cells.
In a previous study of cultured preglomerular vascular smooth muscle cells, it was demonstrated that, although the stable prostacyclin analog iloprost alone had no effect on the intracellular calcium concentration ([Ca2+](i)), it did significantly attenuate the increase in [Ca2+](i) stimulated by angiotensin II (AngII). In this study, the mechanisms by which iloprost interacts with calcium signaling pathways stimulated by AngII were examined. [Ca2+](i) was assessed using the calcium-sensitive fluorescent dye fura-2. Initial studies identified two major components of the [Ca2+](i) response to AngII in this homogeneous preparation of vascular smooth muscle cells from renal resistance vessels. Mobilization of internal stores was evident as an immediate TMB-8-sensitive peak increase in [Ca2+](i) (52 +/- 6 to 297 +/- 26 nM, P: < 0.001) in a calcium-free medium. After [Ca2+](i) had returned to baseline levels during continued AngII stimulation, a nifedipine-sensitive entry pathway was revealed by the sustained stimulatory effect of added external calcium, which increased [Ca2+](i) to 112 +/- 13 nM (P: < 0.001). Coadministration of iloprost with AngII attenuated both the immediate peak (154 +/- 14 nM) and sustained plateau (61 +/- 9 nM) phases. Increases in endogenous levels of cAMP induced by the phosphodiesterase inhibitor milrinone mirrored the actions of iloprost, suggesting that the prostacyclin analog exerted its actions via cAMP activation. Blockade of cAMP-dependent protein kinase with KT 5720 reversed the effects of both iloprost and milrinone. When iloprost or milrinone was introduced after the initial mobilization peak had dissipated, the plateau phase of calcium entry was unchanged (92 +/- 9 nM). The concept that iloprost does not directly modulate calcium entry was further supported by data showing that the activation of L-type calcium channels by BAY-K 8644 was unchanged during iloprost treatment. On the basis of the observation that iloprost did not alter thapsigargin stimulation of Ca(2+)-ATPase activity, it is concluded that the actions of cAMP are distinct from increasing calcium uptake into the sarcoplasmic reticulum. This study provides new information on the ability of iloprost to primarily attenuate inositol-1,4,5-triphosphate-mediated calcium mobilization via cAMP, with secondary inhibition of L-type calcium entry channels. These data clarify the mechanism by which prostaglandins buffer AngII constriction in resistance arterioles.
Purdy
Kit E
KE
Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
Arendshorst
William J
WJ
Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002227:Carbazoles; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D016285:Iloprost / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000098:blood supply*; D020105:Milrinone / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D011758:Pyrroles / Q000494:pharmacology; D051381:Rats; D019284:Thapsigargin / Q000494:pharmacology
D020746:Calcium Channels, L-Type; D002227:Carbazoles; D007211:Indoles; D010726:Phosphodiesterase Inhibitors; D011758:Pyrroles; D000804:Angiotensin II; C057416:KT 5720; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D000252:Calcium-Transporting ATPases; D016285:Iloprost; D020105:Milrinone; D002118:Calcium