Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation

Area of Science:

• Neuroscience
• Cell Biology
• Mitochondrial Biology

Background:

• Mitochondria regulate cellular calcium (Ca²⁺) signaling, impacting neuronal excitability and plasticity.
• The axon initial segment (AIS) is critical for action potential initiation, and its local microenvironment influences neuronal firing patterns.
• Mitochondrial Ca²⁺ buffering at the AIS is hypothesized to modulate Ca²⁺-dependent ion channels and action potential generation.

Purpose of the Study:

• To investigate the role of mitochondria located at the AIS in regulating local cytoplasmic Ca²⁺ transients.
• To determine the impact of AIS mitochondrial Ca²⁺ buffering on action potential initiation and dynamics in layer 5 pyramidal neurons.
• To explore potential non-electrical functions of AIS mitochondria.

Main Methods:

• 3D electron microscopy for ultrastructural analysis of AIS mitochondria distribution.
• Viral delivery of genetically encoded Ca²⁺ indicators for simultaneous imaging of cytoplasmic and mitochondrial Ca²⁺.
• Electrophysiological recordings combined with pharmacological inhibition of mitochondrial Ca²⁺ uptake (Ru360).

Main Results:

• Mitochondria are predominantly clustered at the proximal AIS in layer 5 pyramidal neurons.
• AIS mitochondria exhibit potent activity-dependent Ca²⁺ uptake from the cytosol.
• Inhibition of mitochondrial Ca²⁺ import affected the slow afterhyperpolarization but not action potential initiation, waveform, or high-frequency firing.

Conclusions:

• AIS mitochondria effectively buffer local cytoplasmic Ca²⁺ transients.
• Mitochondrial Ca²⁺ buffering at the AIS is dispensable for action potential generation and temporal/rate encoding.
• These findings suggest that AIS mitochondria primarily serve non-electrical roles in neuronal function.