Subthreshold membrane depolarization powerfully engages intracellular calcium dynamics in the brain

Membrane voltage (Vm) regulates spike timing and intracellular signaling. While Vm is extensively modulated by behavior, it is unclear how subthreshold Vm dynamics engage intracellular signaling in the awake mammalian brain. We developed a bicistronic viral vector to express genetically encoded and color compatible voltage and calcium (Ca ²⁺ ) indicators in the same neuron, and simultaneously recorded cellular Vm and Ca ²⁺ dynamics in awake mice. We report that prolonged subthreshold Vm depolarization is closely accompanied by prominent large amplitude Ca ²⁺ elevation, whereas isolated spikes are coupled with weak Ca ²⁺ rise. Additionally, individual spikes differentially engage intracellular Ca ²⁺ dynamics depending on post-spiking Vm depolarization, consistent with a prominent role of slow Vm depolarization in regulating cellular signaling. While brief intracranial electrical stimulation consistently leads to Vm depolarization and Ca ²⁺ increase, longer stimulation disrupts Vm and Ca ²⁺ coupling, highlighting a tightly regulated cellular mechanism that relays slow Vm depolarization to intracellular signaling.

Prolonged membrane depolarization engages cytosolic calcium.