. Inositol phospholipids localized to caveolae in rat heart are regulated by ␣1-adrenergic receptors and by ischemia-reperfusion. Am J Physiol Heart Circ Physiol 290: H2059 -H2065, 2006. First published December 22, 2005; doi:10.1152/ajpheart.01210.2005.-Postischemic reperfusion of rat or mouse hearts causes generation of inositol (1,4,5)trisphosphate [Ins(1,4,5)P3] and the initiation of arrhythmias. In the current study we investigated the possibility that the enhanced Ins(1,4,5)P3 generation in postischemic reperfusion was associated with an increased availability of the precursor lipid phosphatidylinositol(4,5)bisphosphate (PIP2) for ␣1-adrenergic receptor-activated phospholipase C (PLC). Isolated-perfused rat hearts were labeled with [3 H]inositol and subjected to ischemia-reperfusion or stimulation with norepinephrine under normoxic conditions. Caveolar fractions were prepared by buoyant density sucrose gradient centrifugation. [3 H]PIP2 was concentrated in caveolae, along with G␣q and PLC␤1b. Caveolae contained only 27.3 Ϯ 6.9% (means Ϯ SE, n ϭ 6) of the total ␣ 1-adrenergic receptor complement of the heart. These did not migrate to PIP2-containing caveolar fractions with norepinephrine stimulation under normoxic conditions, even though caveolar PIP2 was depleted. In contrast, [3 H]PIP2 in caveolae increased during 2 min of reperfusion, independently of norepinephrine release and thus of ␣1-adrenergic receptor activation. The increased PIP2 in the caveolar fractions where signaling proteins are concentrated may be critical for the heightened generation of Ins(1,4,5)P 3 in early reperfusion. lipid signaling; light lipid rafts; phospholipase C; Gq; caveolin BRIEF PERIODS OF ISCHEMIA-REPERFUSION in the rat and mouse heart cause norepinephrine release, activation of ␣ 1 -adrenergic receptors (␣ 1 -AR), and substantial generation of inositol (1,4,5)trisphosphate [Ins(1,4,5)P 3 ] associated with the onset of arrhythmogenesis (1, 2, 13, 24, 41). Ins(1,4,5)P 3 is generated from sarcolemmal phosphatidylinositol(4,5)bisphosphate (PIP 2 ) following activation of phospholipase C (PLC). We addressed the possibility that increased availability of PIP 2 or factors required for PLC activation contributed to the heightened generation of Ins(1,4,5)P 3 during postischemic reperfusion.PIP 2 is generated from phosphatidylinositol most commonly by phosphorylation to PI(4)P, followed by further phosphorylation to PI(4,5)P 2 (9) and to a lesser extent by sequential 5Ј and 4Ј phosphorylation (23). PIP 2 is critically involved in myocardial responses. In addition to Ins(1,4,5)P 3 , PLC cleavage of PIP 2 generates sn-1,2-diacylglycerol, an activator of PKC isoforms (31). PIP 2 is also the precursor of PIP 3 , an important factor in cardiomyocyte development and cytoprotection (43). Further to these roles as precursor to other active molecules, PIP 2 itself acts as a second messenger by activating phospholipase D activity in the heart (25). PIP 2 is critically important in stabilizing various ion channels and transporters (22, 32) an...