The precise regulation of Ca²⁺ signals plays a crucial role in the physiological functions of neurons. Here, we investigated the rapid effect of glucocorticoids on Ca²⁺ signals in cultured hippocampal neurons from both female and male rats. In cultured hippocampal neurons, glucocorticoids inhibited the spontaneous somatic Ca²⁺ spikes generated by Kv2.1-organized Ca²⁺ microdomains. Furthermore, glucocorticoids rapidly reduced the cell surface expressions of Kv2.1 and Ca_V1.2 channels in hippocampal neurons. In HEK293 cells transfected with Kv2.1 alone, glucocorticoids significantly reduced the surface expression of Kv2.1 with little effect on K⁺ currents. In HEK293 cells transfected with Ca_V1.2 alone, glucocorticoids inhibited Ca_V1.2 currents but had no effect on the cell surface expression of Ca_V1.2. Notably, in the presence of wild-type Kv2.1, glucocorticoids caused a decrease in the surface expression of Ca_V1.2 channels in HEK293 cells. However, this effect was not observed in the presence of nonclustering Kv2.1S586A mutant channels. Live-cell imaging showed that glucocorticoids rapidly decreased Kv2.1 clusters on the plasma membrane. Correspondingly, Western blot results indicated a significant increase in the cytoplasmic level of Kv2.1, suggesting the endocytosis of Kv2.1 clusters. Glucocorticoids rapidly decreased the intracellular cAMP concentration and the phosphorylation level of PKA in hippocampal neurons. The PKA inhibitor H89 mimicked the effect of glucocorticoids on Kv2.1, while the PKA agonist forskolin abrogated the effect. In conclusion, glucocorticoids rapidly suppress Ca_V1.2-mediated Ca²⁺ signals in hippocampal neurons by promoting the endocytosis of Kv2.1 channel clusters through reducing PKA activity.

Ca²⁺ 信号的精确调节在神经元的生理功能中起着至关重要的作用。在这里，我们研究了糖皮质激素对雌性和雄性大鼠培养的海马神经元中 Ca^{2 +} 信号的快速影响。在培养的海马神经元中，糖皮质激素抑制了由 Kv2.1 组织的 Ca²⁺ 微结构域产生的自发体细胞 Ca²⁺ 尖峰。此外，糖皮质激素迅速降低了海马神经元中 Kv2.1 和 Ca_V1.2 通道的细胞表面表达。在单独转染 Kv2.1 的 HEK293 细胞中，糖皮质激素显著降低了 Kv2.1 的表面表达，而对 K⁺ 电流的影响很小。在单独转染 Ca_V1.2 的 HEK293 细胞中，糖皮质激素抑制 Ca_V1.2 电流，但对 Ca_V1.2 的细胞表面表达没有影响。值得注意的是，在野生型 Kv2.1 存在下，糖皮质激素导致 HEK293 细胞中 Ca_V1.2 通道的表面表达降低。然而，在存在非聚集性 Kv2.1S586A 突变通道的情况下未观察到这种效应。活细胞成像显示糖皮质激素迅速减少质膜上的 Kv2.1 簇。相应地，Western blot 结果显示 Kv2.1 的细胞质水平显着增加，表明 Kv2.1 簇的内吞作用。糖皮质激素迅速降低海马神经元细胞内 cAMP 浓度和 PKA 磷酸化水平。PKA 抑制剂 H89 模拟糖皮质激素对 Kv2.1 的影响，而 PKA 激动剂毛喉素消除了这种作用。总之，糖皮质激素可快速抑制 Ca_V1。通过降低 PKA 活性促进 Kv2.1 通道簇的内吞作用，在海马神经元中通过 2 介导的 Ca²⁺ 信号。