Glucocorticoids rapidly modulate CaV1.2-mediated calcium signals through Kv2.1 channel clusters in hippocampal neurons

Abstract

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 hippocampal neurons. 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 CaV1.2 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. Glucocorticoids inhibited CaV1.2 currents but had no effect on the cell surface expression of CaV1.2 in HEK293 cells transfected with CaV1.2 alone. Notably, in the presence of wild-type Kv2.1, glucocorticoids caused a decrease in the surface expression of CaV1.2 channels in HEK293 cells. However, this effect was not observed in the presence of non-clustering 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 regulate CaV1.2-mediated Ca2+ signals in hippocampal neurons by promoting the endocytosis of Kv2.1 channel clusters through reducing PKA activity. Significance StatementThe rapid non-genomic effect of glucocorticoids on the central nervous system is not fully understood. CaV1.2-mediated Ca2+ signaling microdomains control somatic Ca2+ signals and regulate excitation-transcription coupling in hippocampal neurons. Here, we demonstrate that glucocorticoids rapidly inhibit CaV1.2-mediated somatic Ca2+ spikes in hippocampal neurons. Glucocorticoids reduce the surface expression of Kv2.1 clusters but do not affect the surface expression of non-clustering Kv2.1. Moreover, glucocorticoids induce the endocytosis of CaV1.2 channels through wild-type Kv2.1. However, glucocorticoids cannot induce the endocytosis of CaV1.2 channels through Kv2.1S586A mutant channels, which cannot form clusters. This study sheds light on the intricate interplay between glucocorticoids, Kv2.1 channels, and CaV1.2 channels, unraveling a dual mechanism that influences both overall Ca2+ signaling and the intricate organization of neural microdomains.