Oliveira J C, Sebastião A M, Ribeiro J A
Laboratory of Pharmacology, Gulbenkian Institute of Science, Oeiras, Portugal.
J Neurochem. 1991 Oct;57(4):1165-71. doi: 10.1111/j.1471-4159.1991.tb08275.x.
The specific binding of L-N6-[3H]phenylisopropyladenosine (L-[3H]PIA) to solubilized receptors from rat brain membranes was studied. The interaction of these receptors with relatively low concentrations of L-[3H]PIA (0.5-12.0 nM) in the presence of Mg2+ showed the existence of two binding sites for this agonist, with respective dissociation constant (KD) values of 0.24 and 3.56 nM and respective receptor number (Bmax) values of 0.28 +/- 0.03 and 0.66 +/- 0.05 pmol/mg of protein. In the presence of GTP, the binding of L-[3H]PIA also showed two sites with KD values of 24.7 and 811.5 nM and Bmax values of 0.27 +/- 0.09 and 0.93 +/- 0.28 pmol/mg of protein for the first and the second binding site, respectively. Inhibition of specific L-[3H]PIA binding by 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) (0.1-300 nM) performed with the same preparations revealed two DPCPX binding sites with Ki values of 0.29 and 13.5 nM, respectively. [3H]DPCPX saturation binding experiments also showed two binding sites with respective KD values of 0.81 and 10.7 nM and respective Bmax values of 0.19 +/- 0.02 and 0.74 +/- 0.06 pmol/mg of protein. The results suggest that solubilized membranes from rat brain possess two adenosine receptor subtypes: one of high affinity with characteristics of the A1 subtype and another with lower affinity with characteristics of the A3 subtype of adenosine receptor.
研究了L-N6-[3H]苯基异丙基腺苷（L-[3H]PIA）与大鼠脑膜溶解受体的特异性结合。在Mg2+存在下，这些受体与相对低浓度的L-[3H]PIA（0.5 - 12.0 nM）相互作用，表明该激动剂存在两个结合位点，其解离常数（KD）值分别为0.24和3.56 nM，受体数量（Bmax）值分别为0.28±0.03和0.66±0.05 pmol/mg蛋白质。在GTP存在下，L-[3H]PIA的结合也显示出两个位点，第一个和第二个结合位点的KD值分别为24.7和811.5 nM，Bmax值分别为0.27±0.09和0.93±0.28 pmol/mg蛋白质。用相同制剂进行的1,3 - 二丙基 - 8 - 环戊基黄嘌呤（DPCPX）（0.1 - 300 nM）对特异性L-[3H]PIA结合的抑制作用揭示了两个DPCPX结合位点，其Ki值分别为0.29和13.5 nM。[3H]DPCPX饱和结合实验也显示出两个结合位点，其KD值分别为0.81和10.7 nM，Bmax值分别为0.19±0.02和0.74±0.06 pmol/mg蛋白质。结果表明，大鼠脑溶解膜具有两种腺苷受体亚型：一种具有A1亚型特征的高亲和力亚型，另一种具有腺苷受体A3亚型特征的低亲和力亚型。
