BK_Ca channel regulates calcium oscillations induced by alpha-2-macroglobulin in human myometrial smooth muscle cells

The large-conductance, voltage-gated, calcium (Ca²⁺)-activated potassium channel (BK_Ca) plays an important role in regulating Ca²⁺ signaling and is implicated in the maintenance of uterine quiescence during pregnancy. We used immunopurification and mass spectrometry to identify proteins that interact with BK_Ca in myometrium samples from term pregnant (≥37 wk gestation) women. From this screen, we identified alpha-2-macroglobulin (α₂M). We then used immunoprecipitation followed by immunoblot and the proximity ligation assay to confirm the interaction between BK_Ca and both α₂M and its receptor, low-density lipoprotein receptor-related protein 1(LRP1), in cultured primary human myometrial smooth muscle cells (hMSMCs). Single-channel electrophysiological recordings in the cell-attached configuration demonstrated that activated α₂M (α₂M^∗) increased the open probability of BK_Ca in an oscillatory pattern in hMSMCs. Furthermore, α₂M^∗ caused intracellular levels of Ca²⁺ to oscillate in oxytocin-primed hMSMCs. The initiation of oscillations required an interaction between α₂M^∗ and LRP1. By using Ca²⁺-free medium and inhibitors of various Ca²⁺ signaling pathways, we demonstrated that the oscillations required entry of extracellular Ca²⁺ through store-operated Ca²⁺ channels. Finally, we found that the specific BK_Ca blocker paxilline inhibited the oscillations, whereas the channel opener NS11021 increased the rate of these oscillations. These data demonstrate that α₂M^∗ and LRP1 modulate the BK_Ca channel in human myometrium and that BK_Ca and its immunomodulatory interacting partners regulate Ca²⁺ dynamics in hMSMCs during pregnancy.