Calcium-dependent modulation of the agonist affinity of the mammalian olfactory cyclic nucleotide-gated channel by calmodulin and a novel endogenous factor
Calcium-dependent modulation of the agonist affinity of the mammalian olfactory cyclic nucleotide-gated channel by calmodulin and a novel endogenous factor
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DOI：
10.1007/s002329900081
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发表时间：
1996-07-01
影响因子：
2.4
通讯作者：
Barry, PH
中科院分区：
文献类型：
--
作者：
Balasubramanian, S;Lynch, JW;Barry, PH
在酶促分离的大鼠嗅感受器神经元上，通过记录从树突状突起上切下的由内而外的斑片的cAMP激活电流，研究了环核苷酸门控（CNG）通道对环磷酸腺苷（cAMP）亲和力的钙依赖性调节。在细胞内加入0.2 mM Ca ~（2+）（0.2 Ca）后，激活半最大电流（EC（50））所需的cAMP浓度从3 μ M可逆地增加到约30 μ M。这种Ca 2+诱导的亲和力转移对钙调素拮抗剂mastoparan不敏感，通过暴露于3 mM Mg 2 + + 2 mM EGTA（Mg + EGTA）2分钟不可逆地消除，并且通过应用钙调素（CAM）不能恢复。加入CAM加0.2 mM Ca 2+（0.2 Ca + CAM），进一步可逆地将cAMP亲和力从30 μ M移至约200 μ M。这种亲和力的变化不受Mg + EGTA暴露的影响，但被逆转的mastoparan。因此，前Ca 2 +-唯一的效果必须介导的一个未知的内源性因子，从CAM不同。去除该因子也增加了通道对CAM的亲和力。由Ca 2+诱导的亲和力转移仅在不可水解的cAMP类似物，8-溴-cAMP和磷酸酶抑制剂，微囊藻毒素-LR的存在下保持，排除了磷酸二酯酶或磷酸酶的调节。我们的研究结果表明，嗅觉CNG通道调制的一个尚未确定的因素不同CAM。
The calcium-dependent modulation of the affinity of the cyclic nucleotide-gated (CNG) channels for adenosine 3',5'-cyclic monophosphate (cAMP) was studied in enzymatically dissociated rat olfactory receptor neurons, by recording macroscopic cAMP-activated currents from inside-out patches excised from their dendritic knobs. Upon intracellular addition of 0.2 mM Ca2+ (0.2 Ca) the concentration of cAMP required for the activation of half-maximal current (EC(50)) was reversibly increased from 3 mu M to about 30 mu M. This Ca2+-induced affinity shift was insensitive to the calmodulin antagonist, mastoparan, was abolished irreversibly by a 2-min exposure to 3 mM Mg2+ + 2 mM EGTA (Mg + EGTA), and was not restored by the application of calmodulin (CAM). Addition of CAM plus 0.2 mM Ca2+ (0.2 Ca + CAM), further reversibly shifted the cAMP affinity from 30 mu M to about 200 mu M. This affinity shift was not affected by Mg + EGTA exposure, but was reversed by mastoparan. Thus, the former Ca2+-only effect must be mediated by an unknown endogenous factor, distinct from CAM. Removal of this factor also increased the affinity of the channel for CAM. The affinity shift induced by Ca2+-only was maintained in the presence of the nonhydrolyzable cAMP analogue, 8-bromo-cAMP and the phosphatase inhibitor, microcystin-LR, ruling out modulation by phosphodiesterases or phosphatases. Our results indicate that the olfactory CNG channels are modulated by an as yet unidentified factor distinct from CAM.