光能诱导 cAMP 快速增加,并激活蛙视网膜视杆细胞外节的 PKA。

IF 3.3 2区 医学 Q1 PHYSIOLOGY
Journal of General Physiology Pub Date : 2024-11-04 Epub Date: 2024-10-22 DOI:10.1085/jgp.202313530
Olga Chernyshkova, Natalia Erofeeva, Darya Meshalkina, Anna Balykina, Stepan Gambaryan, Michael Belyakov, Michael Firsov
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引用次数: 0

摘要

光传导级联使光感受器能够在很宽的强度范围内检测光而不会饱和。该级联的主要第二信使是 cGMP,主要调节机制是钙反馈。不过,一些实验数据表明,cAMP 也可能在调节光传导级联中发挥作用,但这需要 cAMP 在几秒钟的时间尺度内发生变化。目前,还缺乏有关细胞内 cAMP 水平在这一时间范围内动态变化的数据。这主要是由于光感受器的感觉模式具有特异性,因此实际上无法使用基于荧光方法的传统实验方法。在这项研究中,我们采用了在光刺激后快速冷冻固定视网膜样本并随后分离外节制备物的方法。研究采用了高灵敏度的代谢组学方法来测量 cAMP 的水平。此外,还利用 Western 印迹法测定了样本中的 PKA 活性。结果表明,当暴露在接近饱和但仍然适度的光照下时,cAMP 水平会在第一秒内短暂上升,然后恢复到刺激前的水平。cAMP 的增加激活了 PKA,导致蛙视网膜外节中 PKA 特异性底物的磷酸化。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Light induces a rapid increase in cAMP and activates PKA in rod outer segments of the frog retina.

The phototransduction cascade enables the photoreceptor to detect light over a wide range of intensities without saturation. The main second messenger of the cascade is cGMP and the primary regulatory mechanism is calcium feedback. However, some experimental data suggest that cAMP may also play a role in regulating the phototransduction cascade, but this would require changes in cAMP on a time scale of seconds. Currently, there is a lack of data on the dynamics of changes in intracellular cAMP levels on this timescale. This is largely due to the specificity of the sensory modality of photoreceptors, which makes it practically impossible to use conventional experimental approaches based on fluorescence methods. In this study, we employed the method of rapid cryofixation of retinal samples after light stimulation and subsequent isolation of outer segment preparations. The study employed highly sensitive metabolomics approaches to measure levels of cAMP. Additionally, PKA activity was measured in the samples using a western blot. The results indicate that when exposed to near-saturating but still moderate light, cAMP levels increase transiently within the first second and then return to pre-stimulus levels. The increase in cAMP activates PKA, resulting in the phosphorylation of PKA-specific substrates in frog retinal outer segments.

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来源期刊
CiteScore
6.00
自引率
10.50%
发文量
88
审稿时长
6-12 weeks
期刊介绍: General physiology is the study of biological mechanisms through analytical investigations, which decipher the molecular and cellular mechanisms underlying biological function at all levels of organization. The mission of Journal of General Physiology (JGP) is to publish mechanistic and quantitative molecular and cellular physiology of the highest quality, to provide a best-in-class author experience, and to nurture future generations of independent researchers. The major emphasis is on physiological problems at the cellular and molecular level.
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