Laboratory of Preclinical Pharmacology, National Institute of Mental Health, Saint Elizabeths Hospital, Washington, D.C. 20032.
Proc Natl Acad Sci U S A. 1982 Jan;79(2):690-4. doi: 10.1073/pnas.79.2.690.
Opioid agonists caused a rapid dose-related elevation of the cGMP content of N4TG1 murine neuroblastoma cells. An excellent correlation was found between the rank order of potency of agonists in stimulating cGMP accumulation and in displacing [(3)H]etorphine ([(3)H]ETP) bound to intact cells. The narcotic antagonists naloxone and diprenorphine failed to increase cGMP content; moreover, in the presence of 5 muM naloxone, the EC(50) of ETP increased from approximately 9 nM to > 1 muM. N4TG1 cells that had been incubated for 20 min with 0.32 muM ETP and thoroughly washed displayed a marked loss in sensitivity to subsequent ETP challenge. This desensitization was characterized by a 40-50% decrease in maximal response and an increase in the apparent K(a) of ETP from 4 to 50 nM. Desensitization was complete after a 7-min incubation with 0.32 muM ETP (t((1/2)) approximately 1 min) and was only slowly reversible (t((1/2)) > 60 min). Naloxone (5 muM) and diprenorphine (0.1 muM) failed to elicit desensitization, but they blocked ETP-induced desensitization. Dextrophan and (+)-ethylketazocine were <1% as effective as levorphanol and (-)-ethylketazocine, respectively, in both stimulating cGMP accumulation and inducing desensitization. When the binding of [(3)H]ETP (0.2-20 nM) was examined under identical experimental conditions, cells that were completely desensitized by incubation with ETP (7 min with 0.32 muM or 20 min with 15 nM) showed no loss of high-affinity recognition sites. After longer incubation with ETP (0.32 muM for 20-60 min), the maximal binding of [(3)H]ETP was reduced 17-41%. The specific short-term desensitization of cGMP accumulation is not mediated or accompanied by a decrement in the number of agonist binding sites.
阿片样物质激动剂引起 N4TG1 鼠神经母细胞瘤细胞中环鸟苷酸含量的快速、剂量相关的升高。激动剂刺激 cGMP 积累的效力顺序与它们置换结合完整细胞的 [(3)H]埃托啡 ([(3)H]ETP) 的能力之间存在极好的相关性。麻醉拮抗剂纳洛酮和二苯哌啶不能增加环鸟苷酸的含量；此外，在存在 5 μM 纳洛酮的情况下，ETP 的 EC(50)从约 9 nM 增加到 >1 μM。用 0.32 μM ETP 孵育 20 分钟并彻底洗涤的 N4TG1 细胞对随后的 ETP 挑战显示出明显的敏感性降低。这种脱敏作用的特征是最大反应减少 40-50%，以及 ETP 的表观 K(a)从 4 增加到 50 nM。用 0.32 μM ETP 孵育 7 分钟后，脱敏完全（t((1/2))约 1 分钟），并且仅缓慢可逆（t((1/2))>60 分钟）。纳洛酮（5 μM）和二苯哌啶（0.1 μM）不能引起脱敏，但它们可以阻断 ETP 诱导的脱敏。右旋苯丙胺和 (+)-乙基酮基左啡诺分别比左啡诺和 (-)-乙基酮基左啡诺的效力低 1%左右，在刺激 cGMP 积累和诱导脱敏方面。当在相同的实验条件下检查 [(3)H]ETP 的结合时（0.2-20 nM），用 ETP 孵育完全脱敏的细胞（7 分钟用 0.32 μM 或 20 分钟用 15 nM）没有失去高亲和力的识别位点。在用 ETP 孵育更长时间（0.32 μM 20-60 分钟）后，[(3)H]ETP 的最大结合减少了 17-41%。cGMP 积累的特异性短期脱敏不是由激动剂结合位点数量的减少介导或伴随的。
