Two-Photon Na⁺ Imaging Reports Somatically Evoked Action Potentials in Rat Olfactory Bulb Mitral and Granule Cell...

Dendrodendritic synaptic interactions are a hallmark of neuronal processing in the vertebrate olfactory bulb. Many classes of olfactory bulb neurons including the principal mitral cells (MCs) and the axonless granule cells (GCs) dispose of highly efficient propagation of action potentials (AP) within their dendrites, from where they can release transmitter onto each other. So far, backpropagation in GC dendrites has been investigated indirectly via Ca²⁺ imaging. Here, we used two-photon Na⁺ imaging to directly report opening of voltage-gated sodium channels due to AP propagation in both cell types. To this end, neurons in acute slices from juvenile rat bulbs were filled with 1 mM SBFI via whole-cell patch-clamp. Calibration of SBFI signals revealed that a change in fluorescence ΔF/F by 10% corresponded to a Δ[Na⁺]_i of ∼22 mM. We then imaged proximal axon segments of MCs during somatically evoked APs (sAP). While single sAPs were detectable in ∼50% of axons, trains of 20 sAPs at 50 Hz always resulted in substantial ΔF/F of ∼15% (∼33 mM Δ[Na⁺]_i). ΔF/F was significantly larger for 80 Hz vs. 50 Hz trains, and decayed with half-durations τ_1/2 ∼0.6 s for both frequencies. In MC lateral dendrites, AP trains yielded small ΔF/F of ∼3% (∼7 mM Δ[Na⁺]_i). In GC apical dendrites and adjacent spines, single sAPs were not detectable. Trains resulted in an average dendritic ΔF/F of 7% (16 mM Δ[Na⁺]_i) with τ_1/2 ∼1 s, similar for 50 and 80 Hz. Na⁺ transients were indistinguishable between large GC spines and their adjacent dendrites. Cell-wise analysis revealed two classes of GCs with the first showing a decrease in ΔF/F along the dendrite with distance from the soma and the second an increase. These classes clustered with morphological parameters. Simulations of Δ[Na⁺]_i replicated these behaviors via negative and positive gradients in Na⁺ current density, assuming faithful AP backpropagation. Such specializations of dendritic excitability might confer specific temporal processing capabilities to bulbar principal cell-GC subnetworks. In conclusion, we show that Na⁺ imaging provides a valuable tool for characterizing AP invasion of MC axons and GC dendrites and spines.