Partial agonists and antagonists reveal a second permeability state of human lymphocyte P2Z/P2X7 channel

被引:88
作者
Wiley, JS [1 ]
Gargett, CE
Zhang, W
Snook, MB
Jamieson, GA
机构
[1] Univ Sydney, Dept Med, Nepean Hosp, Penrith, NSW 2750, Australia
[2] Austin & Repatriat Med Ctr, Dept Haematol, Heidelberg, Vic 3084, Australia
来源
AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY | 1998年 / 275卷 / 05期
关键词
extracellular adenosine 5 '-triphosphate; chronic lymphocytic leukemia; B lymphocytes;
D O I
10.1152/ajpcell.1998.275.5.C1224
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Extracellular ATP is known to trigger apoptosis of thymocytes and lymphocytes through a P2Z receptor at which ATP is a partial agonist, giving only 70% of the maximum response of 3'-O-(4-benzoyl)benzoyl-adenosine 5'-triphosphate (BzATP), a full agonist. This cytolytic receptor and its associated ion channel are Ca2+ (and Ba2+) selective but also pass molecules up to the size of ethidium cation (314 Da). RT-PCR showed identity between lymphocyte P2Z and the hP2X(7) gene recently cloned from human monocytes. When human leukemic B lymphocytes were incubated with ATP and Ba-133(2+), immediate influx of isotope occurred. It was augmented by 45% when ATP was added 10 min before isotope. Time-resolved flow cytometry was used to examine kinetics of ethidium uptake in cells incubated with BzATP or the partial agonists ATP, 2-methylthioadenosine 5'-triphosphate, or adenosine 5'-O-(3-thiotriphosphate). Maximally effective concentrations of BzATP (50 mu M) induced immediate uptake of ethidium at a rate linear with time. In contrast, a delay was observed (30 s) before ethidium uptake commenced after addition of maximally effective ATP concentrations (500 mu M) at 37 degrees C, and the delay was longer at 24 degrees C. ATP addition 2-10 min before ethidium abolished the delay. The delay was longer with other partial agonists and inversely related to maximal flux produced by agonist. A delay was also observed for submaximal BzATP concentrations (10-20 mu M). P2Z/P2X(7) inhibitors, KN-62 and 5-(N,N-hexamethylene)-amiloride, reduced the rate of agonist-induced ethidium uptake and lengthened the delay. The results support a model in which agonists for P2Z/P2X(7) receptor mediate an immediate channel opening allowing passage of small inorganic cations, followed by a slow further permeability increase allowing passage of larger permeant cations like ethidium. The rate of the second step depends on time and temperature and the efficacy and concentration of agonist and is slowed by antagonists, suggesting it depends on the fraction of P2Z/P2X(7) channels held in the initial open state.
引用
收藏
页码:C1224 / C1231
页数:8
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