Smith M A, Silverstein J L, Westfall D P, Buxton I L
University of Nevada School of Medicine Reno 89557-0046.
Cell Signal. 1989;1(4):357-65. doi: 10.1016/0898-6568(89)90054-5.
We have previously demonstrated that adenosine causes contraction of guinea-pig myometrium in a fashion consistent with the presence of a purinergic receptor of the A1 subtype. Incubation of guinea-pig uterine smooth muscle membranes with the stable adenosine analogue [3H]cyclohexyladenosine [( 3H]CHA) resulted in rapid, reversible association of radioligand to saturable sites. The affinity (KD) of the receptor for [3H]CHA determined from kinetic experiments (3.14 nM) is in good agreement with that determined in saturation experiments (KD = 4.5 nM). Scatchard analysis of specific [3H]CHA binding (Bmax = 79 fmol/mg protein) is consistent with a single class of binding sites for [3H]CHA. Computer analysis of competition of [3H]CHA binding by the stereoisomers of phenylisopropyl adenosine, R-PIA (KI = 5.3 nM) and S-PIA (KI = 69 nM), as well as the 5'-substituted analogue, ethylcarboxamide adenosine (NECA; KI = 4.2 nM) suggest that [3H]CHA binding occurs to a single class of receptors of the AI subtype. Contractile studies employing these agents reveal that the relative order of potency, based on ED50 values, correlates well with the relative order of competition of agonist binding, based on equilibrium binding constants. Direct assay of myometrial adenylate cyclase failed to show that adenosine receptors in this smooth muscle are coupled to adenylate cyclase. We conclude here that a smooth muscle adenosine receptor is not coupled to adenylate cyclase, yet subserves muscle contraction. These data are important in light of recent attempts to classify adenosine receptors as dual regulators of adenylate cyclase.
我们之前已经证明，腺苷可使豚鼠子宫肌层收缩，其方式与存在A1亚型嘌呤能受体一致。用稳定的腺苷类似物[3H]环己基腺苷[（3H]CHA）孵育豚鼠子宫平滑肌膜，会导致放射性配体快速、可逆地与可饱和位点结合。通过动力学实验确定的受体对[3H]CHA的亲和力（KD）为3.14 nM，与饱和实验中确定的亲和力（KD = 4.5 nM）非常一致。对特异性[3H]CHA结合的Scatchard分析（Bmax = 79 fmol/mg蛋白质）与[3H]CHA的单一结合位点类别一致。对苯基异丙基腺苷的立体异构体R-PIA（KI = 5.3 nM）和S-PIA（KI = 69 nM）以及5'-取代类似物乙基羧酰胺腺苷（NECA；KI = 4.2 nM）竞争[3H]CHA结合的计算机分析表明，[3H]CHA结合发生在AI亚型的单一受体类别上。使用这些药物的收缩研究表明，基于ED50值的效力相对顺序与基于平衡结合常数的激动剂结合竞争相对顺序密切相关。对子宫肌层腺苷酸环化酶的直接测定未能表明该平滑肌中的腺苷受体与腺苷酸环化酶偶联。我们在此得出结论，平滑肌腺苷受体不与腺苷酸环化酶偶联，但仍能促进肌肉收缩。鉴于最近将腺苷受体分类为腺苷酸环化酶双重调节剂的尝试，这些数据很重要。
