Dixon C Jane, Hall John F, Boarder Michael R
School of Molecular Sciences, The Hawthorn Building, De Montfort University, The Gateway, Leicester LE1 9BH, UK.
Br J Pharmacol. 2003 Jan;138(1):272-8. doi: 10.1038/sj.bjp.0705016.
1 Accumulation of inositol (poly)phosphates (InsP(x)) has been studied in rat hepatocytes labelled with [(3)H]inositol. Stimulation with ADP resulted in a significant increase in total [(3)H]InsP(x), whereas 2-MeSADP had only a small effect and ADPbetaS was ineffective. UTP and ITP also stimulated substantial increases in [(3)H]InsP(x). 2 The dose-response curve to ADP was largely unaltered by the presence of the P2Y(1) antagonist, adenosine-3'-phosphate-5'-phosphate (A3P5P). Similarly, inclusion of MRS 2179, a more selective P2Y(1) antagonist, had no effect on the dose-response curve to ADP. 3 The inclusion of hexokinase in the assay reduced, but did not abolish, the response to ADP. 4 HPLC analysis revealed that ADP in the medium was rapidly converted to AMP and ATP. The inclusion of hexokinase removed ATP, but exacerbated the decline in ADP concentration, leading to increased levels of AMP. 2-MeSADP was stable in the medium and ATP was largely unaffected. 5 The addition of the adenylate kinase inhibitor, diadenosine pentaphosphate (Ap(5)A) significantly reduced the ADP response. HPLC analysis conducted in parallel demonstrated that this treatment inhibited conversion of ADP to ATP and AMP. 6 Inclusion of the P1 antagonist CGS 15943 had no effect on the dose-response curve to ADP. 7 These observations indicate that hepatocytes respond to ADP with an increase in inositol (poly)phosphates following conversion to ATP. P2Y(1) activation in hepatocytes does not appear to be coupled to inositol 1,4,5-trisphosphate (Ins(1,4,5)P(3)) production.
1 已在用[³H]肌醇标记的大鼠肝细胞中研究了肌醇(多)磷酸酯(InsP(x))的积累。用ADP刺激导致总[³H]InsP(x)显著增加,而2-MeSADP只有很小的作用,ADPβS则无效。UTP和ITP也刺激[³H]InsP(x)大幅增加。2 存在P2Y(1)拮抗剂腺苷-3'-磷酸-5'-磷酸(A3P5P)时,对ADP的剂量反应曲线基本未改变。同样,加入更具选择性的P2Y(1)拮抗剂MRS 2179对ADP的剂量反应曲线也没有影响。3 在测定中加入己糖激酶可降低但并未消除对ADP的反应。4 HPLC分析显示培养基中的ADP迅速转化为AMP和ATP。加入己糖激酶可去除ATP,但加剧了ADP浓度的下降,导致AMP水平升高。2-MeSADP在培养基中稳定,ATP基本不受影响。5 加入腺苷酸激酶抑制剂二腺苷五磷酸(Ap(5)A)显著降低了ADP反应。同时进行的HPLC分析表明,这种处理抑制了ADP向ATP和AMP的转化。6 加入P1拮抗剂CGS 15943对ADP的剂量反应曲线没有影响。7 这些观察结果表明,肝细胞在ADP转化为ATP后,通过肌醇(多)磷酸酯增加来对ADP作出反应。肝细胞中的P2Y(1)激活似乎未与肌醇1,4,5-三磷酸(Ins(1,4,5)P(3))的产生偶联。