Barber R, Goka T J, Butcher R W
Graduate School of Biomedical Sciences, University of Texas Health Science Center, Houston 77225.
Mol Pharmacol. 1989 Sep;36(3):459-64.
Epinephrine at concentrations approximating circulating levels in resting subjects produced significant desensitization in wild type S49 lymphoma cells after long term treatment. Desensitization by such low levels of catecholamines was measured by examining subsequent responses of the cells to higher agonist concentrations and was quantified by comparing the integral cAMP accumulations with time in naive and epinephrine-treated cells challenged with the higher epinephrine concentrations. The cells were significantly desensitized after 8 hr of treatment with 3 nM epinephrine or 3 nM terbutaline and were essentially maximally refractory after 24 hr. The 3 nM epinephrine treatment resulted in a small right shift of the EC50. Responses to epinephrine were partially restored by incubating desensitized cells for 8 hr or longer in growth medium that was free of epinephrine. The attenuation of cAMP responses was largely specific, in that the decrease in the response to prostaglandin was small and the response to forskolin was unchanged. This, together with small increases in cAMP destruction in cell-free preparations from treated cells, suggested that higher phosphodiesterase activity contributed in a minor way to the desensitization. However, the response of the adenylate cyclase system to epinephrine was dramatically attenuated, and very significant changes in the properties of the beta-adrenergic receptors were also obvious. That is, the number of binding sites for epinephrine was reduced by about 65% while the number of sites for [125I]iodocyanopindolol was unchanged. The affinity for the radioactive ligand was significantly reduced. Wild type S49 cells remained viable after several days of continuous treatment with 3 nM epinephrine or terbutaline but responded to subsequent increases in cellular cAMP levels with the expected growth arrest and cytolysis. Involvement of cAMP-dependent protein kinase in this type of desensitization was suggested by the observation that S49 kincells were not desensitized by long term incubation with 3 nM epinephrine. Further, low concentrations of dibutyryl cAMP mimicked the effect of low level epinephrine treatment. We conclude that circulating levels of epinephrine in intact animals are sufficiently high to cause desensitization in cells with sensitivities to the catecholamines in the same range as that of the S49 lymphoma cell in vitro. We would predict that cells with those characteristics would always be at least partially desensitized in vivo.
在长期处理后，接近静息受试者循环水平的肾上腺素浓度可使野生型S49淋巴瘤细胞产生显著的脱敏作用。通过检测细胞对更高激动剂浓度的后续反应来测量这种低水平儿茶酚胺引起的脱敏作用，并通过比较未处理细胞和用更高肾上腺素浓度刺激的肾上腺素处理细胞中cAMP随时间的累积积分来进行量化。用3 nM肾上腺素或3 nM特布他林处理8小时后，细胞出现显著脱敏，24小时后基本达到最大不应期。3 nM肾上腺素处理导致EC50有小的右移。将脱敏细胞在不含肾上腺素的生长培养基中孵育8小时或更长时间，对肾上腺素的反应部分恢复。cAMP反应的减弱在很大程度上具有特异性，因为对前列腺素的反应降低较小，对福斯高林的反应未改变。这一点，再加上处理细胞的无细胞制剂中cAMP破坏略有增加，表明较高的磷酸二酯酶活性对脱敏作用有较小贡献。然而，腺苷酸环化酶系统对肾上腺素的反应显著减弱，β-肾上腺素能受体的性质也有非常明显的变化。也就是说，肾上腺素的结合位点数量减少了约65%，而[125I]碘氰吲哚洛尔的位点数量未改变。对放射性配体的亲和力显著降低。用3 nM肾上腺素或特布他林连续处理数天后，野生型S49细胞仍保持活力，但对随后细胞内cAMP水平的升高会出现预期的生长停滞和细胞溶解反应。S49激酶细胞经3 nM肾上腺素长期孵育后未出现脱敏，这一观察结果提示cAMP依赖性蛋白激酶参与了这种类型的脱敏作用。此外，低浓度的二丁酰cAMP模拟了低水平肾上腺素处理的效果。我们得出结论，完整动物体内循环水平的肾上腺素足够高，足以使对儿茶酚胺敏感性与体外S49淋巴瘤细胞相同范围内的细胞产生脱敏作用。我们预测，具有这些特征的细胞在体内将始终至少部分脱敏。
