Identification of A1 adenosine receptors in rat cochlea coupled to inhibition of adenylyl cyclase.
Identification of A1 adenosine receptors in rat cochlea coupled to inhibition of adenylyl cyclase.
复制标题
大鼠耳蜗中与腺苷酸环化酶抑制相关的 A1 腺苷受体的鉴定。
DOI：
10.1152/ajpcell.1994.267.3.c731
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发表时间：
1994
期刊：
影响因子：
--
通讯作者：
Rybak,LP
中科院分区：
文献类型：
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作者：
Ramkumar,V;Ravi,R;Wilson,MC;Gettys,TW;Whitworth,C;Rybak,LP
A1腺苷受体（A1 AR）存在于身体的许多组织中，其中它们的生理作用已被确定。在耳蜗中，既没有这些受体的存在，也没有腺苷的生理作用以前已经描述过。从大鼠耳蜗制备的膜表现出与N6-2-（4-氨基-3-[125 I]碘苯基）乙基腺苷（[125 I]APNEA）（一种A1 AR激动剂）的高亲和力和可饱和结合，最大结合容量和解离常数值分别为40.5 +/- 0.5 fmol/mg蛋白和1.28 +/- 0.03 nM。腺苷类似物竞争[125 I]APNEA结合位点，这些位点的效力特征为A1 AR。[125 I]APNEA结合显着减少百日咳毒素，表明这些受体与耳蜗膜中的Gi和/或Go蛋白偶联。用A1 AR激动剂N6-2-（4-叠氮基-3 [125 I]碘苯基）乙基腺苷对受体蛋白进行光亲和标记，显示出36 kDa受体蛋白的特异性标记。用R-苯基异丙基腺苷（R-PIA）激活A1 AR导致毛喉素刺激的腺苷酸环化酶活性的抑制。通过聚合酶链反应（使用牛A1 AR的引物）扩增来自耳蜗组织的逆转录RNA产生了770 bp的产物，该产物在Southern印迹上与A1 AR cDNA探针杂交。这些数据表明，在外周听觉系统中存在抑制性受体，这可能在调节听觉功能中发挥重要作用。
A1 adenosine receptors (A1ARs) are found in a number of tissues in the body where their physiological roles have been identified. In the cochlea, neither the existence of these receptors nor a physiological role of adenosine has been described previously. Membranes prepared from rat cochlea demonstrated high affinity and saturable binding to N6-2-(4-amino-3-[125I]iodophenyl)ethyladenosine ([125I]APNEA), an A1AR agonist, with maximum binding capacity and dissociation constant values being 40.5 +/- 0.5 fmol/mg protein and 1.28 +/- 0.03 nM, respectively. Adenosine analogues competed for [125I]APNEA binding sites with a rank order of potency characteristic of these sites being the A1AR. The [125I]APNEA binding was significantly reduced by pertussis toxin, indicating coupling of these receptors with the Gi and/or Go proteins in cochlear membranes. Photoaffinity labeling of the receptor protein with the A1AR agonist N6-2-(4-azido-3[125I]iodophenyl)ethyladenosine showed specific labeling of a 36-kDa receptor protein. Activation of the A1AR with R-phenylisopropyladenosine (R-PIA) led to inhibition of forskolin-stimulated adenylyl cyclase activity. Amplification of reverse-transcribed RNA derived from cochlear tissue by polymerase chain reaction (using primers for the bovine A1AR) yielded a 770-bp product that hybridized to an A1AR cDNA probe on Southern blots. These data indicate the presence of an inhibitory receptor in the peripheral auditory system, which may play an important role in modulating auditory functions.