Abstract

Cortical axons contain a diverse range of voltage-activated ion channels, including Ca²⁺ currents. Interestingly, Ca²⁺ channels are not only located at presynaptic terminals, but also in the axon initial segment (AIS), suggesting a potentially important role in the regulation of action potential generation and neuronal excitability. Here, using two-photon microscopy and whole-cell patch-clamp recording, we examined the properties and role of calcium channels located in the AIS and presynaptic terminals of ferret layer 5 prefrontal cortical pyramidal cells in vitro. Subthreshold depolarization of the soma resulted in an increase in baseline and spike-triggered calcium concentration in both the AIS and nearby synaptic terminals. The increase in baseline calcium concentration rose with depolarization and fell with hyperpolarization with a time constant of approximately 1 s and was blocked by removal of Ca²⁺ from the bathing medium. The increases in calcium concentration at the AIS evoked by subthreshold or suprathreshold depolarization of the soma were blocked by the P/Q-channel antagonist ω-agatoxin IVA or the N-channel antagonist ω-conotoxin GVIA or both. The presence of these channels in the AIS pyramidal cells was confirmed with immunochemistry. Block of these channels slowed axonal action potential repolarization, apparently from reduction of the activation of a Ca²⁺-activated K⁺ current, and increased neuronal excitability. These results demonstrate novel mechanisms by which calcium currents may control the electrophysiological properties of axonal spike generation and neurotransmitter release in the neocortex.