树突状突触相互作用是脊椎动物嗅球中神经元加工的标志。许多种类的嗅球神经元，包括主要的二尖瓣细胞（MC）和无轴突颗粒细胞（GC）在其树突内高效地传播动作电位（AP），从那里他们可以互相释放发射器。到目前为止,通过Ca2成像间接研究了GC树突中的反向传播。这里,我们使用双光子Na+成像直接报告由于AP在两种细胞类型中的传播而导致的电压门控钠通道的打开。为此,通过全细胞膜片钳将幼年大鼠鳞茎急性切片中的神经元填充1mMSBFI。SBFI信号的校准显示荧光ΔF/F变化10%对应于~22mM的Δ[Na+]i。然后，我们在躯体诱发AP（sAP）期间对MC的近端轴突段进行了成像。虽然在50%的轴突中可以检测到单个sAP，在50Hz下，20个sAP的列车总是导致~15%的显著ΔF/F(~33mMΔ[Na+]i)。ΔF/F明显大于80Hz与50Hz列车，并且在两个频率下都以半持续时间τ1/2~0.6s衰减。在MC侧向树突中，AP列车产生的ΔF/F小，为~3%(~7mMΔ[Na+]i)。在GC顶端树突和相邻的刺中，单个sAP检测不到。火车导致平均树突ΔF/F为7％（16mMΔ[Na]i），τ1/2〜1s，50和80Hz相似。Na瞬变在大的GC刺及其相邻的树突之间无法区分。按细胞分析显示了两类GC，第一类显示沿树突的ΔF/F随着与体细胞的距离而降低，第二类显示增加。这些类以形态学参数聚集。Δ[Na]i的模拟通过Na电流密度的负梯度和正梯度复制了这些行为，假设忠实的AP反向传播。树突状兴奋性的这种专业化可能会赋予Bulbar主要细胞GC子网络特定的时间处理能力。总之,我们表明，Na+成像为表征MC轴突和GC树突和棘的AP侵袭提供了有价值的工具。
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 τ1/2 ∼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 τ1/2 ∼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.
PDF(Sci-hub)