Division of Cerebral Structure, National Institute for Physiological Sciences, Okazaki 444-8787, Japan.
J Physiol. 2013 Jan 1;591(1):219-39. doi: 10.1113/jphysiol.2012.241398. Epub 2012 Oct 15.
Establishing the spatiotemporal concentration profile of neurotransmitter following synaptic vesicular release is essential for our understanding of inter-neuronal communication. Such profile is a determinant of synaptic strength, short-term plasticity and inter-synaptic crosstalk. Synaptically released glutamate has been suggested to reach a few millimolar in concentration and last for <1 ms. The synaptic cleft is often conceived as a single concentration compartment, whereas a huge gradient likely exists. Modelling studies have attempted to describe this gradient, but two key parameters, the number of glutamate in a vesicle (N(Glu)) and its diffusion coefficient (D(Glu)) in the extracellular space, remained unresolved. To determine this profile, the rat calyx of Held synapse at postnatal day 12-16 was studied where diffusion of glutamate occurs two-dimensionally and where quantification of AMPA receptor distribution on individual postsynaptic specialization on medial nucleus of the trapezoid body principal cells is possible using SDS-digested freeze-fracture replica labelling. To assess the performance of these receptors as glutamate sensors, a kinetic model of the receptors was constructed from outside-out patch recordings. From here, we simulated synaptic responses and compared them with the EPSC recordings. Combinations of N(Glu) and D(Glu) with an optimum of 7000 and 0.3 μm(2) ms(-1) reproduced the data, suggesting slow diffusion. Further simulations showed that a single vesicle does not saturate the synaptic receptors, and that glutamate spillover does not affect the conductance amplitude at this synapse. Using the estimated profile, we also evaluated how the number of multiple vesicle releases at individual active zones affects the amplitude of postsynaptic signals.
建立突触小泡释放后神经递质的时空浓度分布对于理解神经元间的通讯至关重要。该分布决定了突触强度、短期可塑性和突触间串扰。据推测，突触释放的谷氨酸浓度可达几毫摩尔，持续时间<1ms。突触间隙通常被认为是一个单一的浓度隔室，但实际上可能存在巨大的浓度梯度。建模研究试图描述这种梯度，但两个关键参数，即囊泡中的谷氨酸数量（N(Glu)）和其在细胞外空间中的扩散系数（D(Glu)），仍未得到解决。为了确定该分布，研究了出生后第 12-16 天的大鼠内膝体浦肯野细胞突触，在该突触中，谷氨酸的扩散是二维的，并且可以使用 SDS 消化的冷冻断裂复制标记在单个突触后特化上定量 AMPA 受体的分布。为了评估这些受体作为谷氨酸传感器的性能，使用从外面到里面的膜片钳记录构建了受体的动力学模型。从这里，我们模拟了突触反应，并将其与 EPSC 记录进行了比较。N(Glu)和 D(Glu)的组合为 7000 和 0.3μm(2)ms(-1)，表现最佳，表明扩散缓慢。进一步的模拟表明，单个囊泡不会使突触受体饱和，并且谷氨酸溢出不会影响该突触的电导幅度。使用估计的分布，我们还评估了单个活性区中多个囊泡释放的数量如何影响突触后信号的幅度。
