No abstract

No abstract

5-bisphosphate (PIP 2) affects profoundly several cardiac ion channels and transporters, and studies of PIP2-sensitive currents in excised patches suggest that PIP 2 can be synthesized and broken down within 30 s. To test when, and if, total phosphatidylinositol 4-phosphate (PIP) and PIP2 levels actually change in intact heart, we used a new, nonradioactive HPLC method to quantify anionic phospholipids. Total PIP and PIP2 levels (10-30 mol/kg wet weight) do not change, or even increase, with activation of G␣ q/phospholipase C (PLC)-dependent pathways by carbachol (50 M), phenylephrine (50 M), and endothelin-1 (0.3 M). Adenosine (0.2 mM) and phorbol 12-myristate 13-acetate (1M) both cause 30% reduction of PIP2 in ventricles, suggesting that diacylglycerol (DAG)-dependent mechanisms negatively regulate cardiac PIP2. PIP2, but not PIP, increases reversibly by 30% during electrical stimulation (2 Hz for 5 min) in guinea pig left atria; the increase is blocked by nickel (2 mM). Both PIP and PIP2 increase within 3 min in hypertonic solutions, roughly in proportion to osmolarity, and similar effects occur in multiple cell lines. Inhibitors of several volume-sensitive signaling mechanisms do not affect these responses, suggesting that PIP2 metabolism might be sensitive to membrane tension, per se. phosphatidylinositol 4,5-bisphosphate; phosphatidylinositol; diacylglycerol; phorbol ester; cardiac muscle; G protein-coupled receptors; phospholipase C; cell volume PHOSPHATIDYLINOSITOL 4,5-BISPHOSPHATE (PIP 2 ) is the phospholipid precursor of three second messengers, D-myo-inositol 1,4,5-trisphosphate (IP 3 ), diacylglycerol (DAG), and phosphatidylinositol 3,4,5-trisphosphate (PIP 3 ) (66). At the same time, PIP 2 serves other cellular functions. It anchors and modulates the function of numerous cell signaling proteins and cytoskeleton at the cell membrane (11,17,42,65), including at least one transcription factor that is released by phospholipase C (PLC) activation (60). In addition, PIP 2 metabolism is coupled to membrane trafficking, including some forms of exo-and endocytosis (7, 46). Finally, PIP 2 modulates the function of phospholipases (14), receptor kinases (16, 52), and ion transporters and ion channels (25). Especially, the anchoring/recruitment functions and the modulatory functions of PIP 2 beg the question as to how, and if, PIP 2 might be used as a cell signal. For cardiac physiology, an answer to this question seems especially important at this time, because sarcolemmal mechanisms that affect both cardiac contraction (e.g., Na ϩ /Ca 2ϩ exchange) and contraction frequency [e.g., G protein-coupled inwardly rectifying K ϩ (GIRK) channels] are strongly PIP 2 dependent (27).The minimum biochemical mechanisms involved in cardiac myocyte PIP 2 metabolism (39) are summarized in Fig. 1. The dominant pathway of PIP 2 synthesis, as in other cells, is probably the sequential phosphorylation in the sarcolemma of phosphatidylinositol (PI) at the 4-and then the 5-positions of inositol (66). As in other cells, PIP 2 is hydr...