Changes in phosphatidylinositol-4,5-bisphosphate 10 seconds after stimulation of washed rabbit platelets with ADP

Vickers, J D; Kinlough-Rathbone, R L; Mustard, J. F.

doi:10.1182/blood.v60.6.1247.1247

Cited by 96 publications

(30 citation statements)

References 27 publications

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“…1 previous observations , the levels of P2Plphosphatidic acid increased markedly by 15s after TRH addition and that of [32P]PtdIns did not increase until after 2 min (results not shown). The rapid decreases in [32P]PtdIns(4,5)P2 and [32PlPtdIns4P in GH3 cells after TRH stimulation are similar to recent observations in rat hepatocytes stimulated by vasopressin Thomas et al, 1983) and in horse platelets stimulated by thrombin (Billah & Lapetina, 1982;Vickers et al, 1982). In cells labelled to isotopic steady-state with [32P1P , TRH caused a decrease in Ptdlns(4,5)P2 from 33 + 2.3 pmol/106 cells to 22+ 1.lpmol at 30s (P<0.01).…”

Section: Inositol Sugar Analysissupporting

confidence: 89%

Thyroliberin stimulates rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate by a phosphodiesterase in rat mammotropic pituitary cells. Evidence for an early Ca2+-independent action

Rebecchi¹,

Gershengorn²

1983

Biochemical Journal

206

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Thyrotropin-releasing hormone (TRH; thyroliberin) stimulated rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] by a phosphodiesterase (phospholipase C) in GH3 cells, a prolactin-secreting rat pituitary tumour cell line. TRH caused a rapid decrease in the level of PtdIns(4,5)P2 to 60% of control and stimulated a marked transient increase in inositol 1,4,5-trisphosphate, the unique product of phosphodiesteratic hydrolysis of PtdIns(4,5)P2, to a peak of 410% of control at 15 s. TRH also caused decreases in phosphatidylinositol 4-monophosphate (PtdIns4P) and phosphatidylinositol (PtdIns) to 65% and 93% of control at 15 s respectively. Inositol 1,4-bisphosphate was increased to a peak of 450% at 30 s; inositol 1-monophosphate and inositol were not elevated until 30 s and 1 min respectively after TRH addition. To study whether PtdIns(4,5)P2 hydrolysis may be caused by an elevation in cytosolic Ca2+ concentration, the changes induced by TRH in the levels of inositol sugars were compared with the effects of membrane depolarization by high extracellular [K+]. The elevation in cytosolic [Ca2+] induced by K+ depolarization did not change the level of inositol 1,4,5-trisphosphate. These data suggest that phosphodiesteratic hydrolysis of PtdIns(4,5)P2 may be the initial event in TRH stimulation of inositol lipid metabolism in GH3 cells and that PtdIns(4,5)P2 hydrolysis is not stimulated by an elevation in cytosolic Ca2+ concentration. The decreases in PtdIns4P and PtdIns may be due to enhanced conversion of PtdIns into PtdIns4P into PtdIns(4,5)P2 or to their direct hydrolysis by phosphomonoesterases and/or phosphodiesterases. These results are consistent with the hypothesis that TRH-stimulated PtdIns(4,5)P2 breakdown causes Ca2+ mobilization leading to prolactin secretion.

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Section: Inositol Sugar Analysissupporting

confidence: 89%

Thyroliberin stimulates rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate by a phosphodiesterase in rat mammotropic pituitary cells. Evidence for an early Ca2+-independent action

Rebecchi¹,

Gershengorn²

1983

Biochemical Journal

206

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“…We have previously reported that stimulation of rabbit platelets with ADP caused a decrease in the chemical amount of PIP2, possibly due to reduced synthesis (Vickers et al, 1982. This decrease in PIP2 was accompanied by an increase in specific radioactivity of PIP2 in platelets labelled with [32P]phosphate and [3H]glycerol.…”

Section: Introductionmentioning

confidence: 92%

“…The silica gel containing the phospholipids was scraped into test tubes for digestion with HC104 prior to phosphate analysis (Chalvardjian & Rudnicki, 1970;Duck-Chong, 1979) and determination of radioactivity. The methods have been described in detail previously (Vickers et al, 1982).…”

Section: Fractionation Of Platelet Phospholipids and Determination Ofmentioning

confidence: 99%

“…Finally, the platelet pellet was resuspended in Tyrode/albumin solution and lipid extraction started by the addition of chloroform/methanol (1:2, v/v) extraction solvent. The lipids were then extracted with a neutral solvent and then with an acidic solvent system as previously described (Vickers et al, 1982). At 1 h the remaining platelet suspension was freed of unincorporated label by centrifuging and resuspending the platelets twice in calcium-free Tyrode solution.…”

Section: Introductionmentioning

confidence: 99%

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The phosphoinositides exist in multiple metabolic pools in rabbit platelets

Vickers¹,

Mustard²

1986

Biochemical Journal

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The labelling of the phosphoinositides and phosphatidic acid in washed rabbit platelets incubated with [32P]phosphate or [3H]glycerol was studied in the presence of isotope and after unincorporated isotope had been removed. With both isotopes the increase in the specific radioactivity of phosphatidylinositol 4,5-bisphosphate (PIP2) lagged behind that of phosphatidylinositol 4-phosphate (PIP) but the specific radioactivity remained higher after unincorporated isotope had been removed. This result was consistent with the presence of a second pool of PIP2, which interconverted slowly with the pool of PIP2 which was in direct equilibrium with PIP, proposed to explain the increase in specific radioactivity of PIP2 which accompanies the decrease in amount of PIP2 at 10 s in ADP-stimulated platelets. In platelets labelled with [3H]glycerol, the specific radioactivity of PIP2 became higher than that of PIP and the specific radioactivity of PIP became higher than that of phosphatidylinositol (PI). These results were interpreted to indicate that there were two pools of PIP; of these the pool with the higher specific radioactivity was the precursor of PIP2. Similarly, two pools of PI were proposed. The presence of pools of the phosphoinositides with different specific radioactivities necessitates the measurement of chemical amount of these compounds when studying the effect of stimulation of the platelets, since changes in labelling may not accurately reflect changes in the amount of the phosphoinositides.

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“…[ 3 H]PC was detected by Radioscanner Isomess 3000 Radio‐TLC Analyzer (‘RITA’, Raytest) or PC mass was visualized by I 2 vapor. The PC spots were scraped, extracted from the silica [21] and aliquots taken for scintillation counting and PC mass determinations as P i with a malachite green method [22].…”

Section: Methodsmentioning

confidence: 99%

Biochemical properties of platelet microparticle membranes formed during exocytosis resemble organelles more than plasma membrane

Olas¹,

Lundell²,

Holmsen³

et al. 2002

FEBS Letters

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Studies of [ 3 H]glycerol turnover in phosphatidylcholine (PC) in platelets revealed two metabolic pools, a 'low turnover PC' in collagen-induced microparticles with speci¢c radioactivity only 10% of that found in the 'high turnover PC' of bulk platelet PC. Isolated organelle fractions of [ 3 H]glycerollabelled platelets contained [ 3 H]PC with speci¢c radioactivities about 20% of that in membrane fractions. These results together with studies on distribution of concanavalin A-FITC and GPlb, a plasma membrane receptor, indicate that microparticles formed during exocytosis are not simple vesiculations of plasma membrane, but they seem rather to originate from a relatively metabolically static membrane pool not accessible to extracellular reagents. ß

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Changes in phosphatidylinositol-4,5-bisphosphate 10 seconds after stimulation of washed rabbit platelets with ADP

Cited by 96 publications

References 27 publications

Thyroliberin stimulates rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate by a phosphodiesterase in rat mammotropic pituitary cells. Evidence for an early Ca2+-independent action

Thyroliberin stimulates rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate by a phosphodiesterase in rat mammotropic pituitary cells. Evidence for an early Ca2+-independent action

The phosphoinositides exist in multiple metabolic pools in rabbit platelets

Biochemical properties of platelet microparticle membranes formed during exocytosis resemble organelles more than plasma membrane

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