Non-NMDA glutamate receptor occupancy and open probability at a rat cerebellar synapse with single and multiple release sites
Non-NMDA glutamate receptor occupancy and open probability at a rat cerebellar synapse with single and multiple release sites
复制标题
DOI：
10.1113/jphysiol.1996.sp021487
复制
发表时间：
1996-07-01
影响因子：
5.5
通讯作者：
Takahashi, T
中科院分区：
文献类型：
--
作者：
Silver, RA;CullCandy, SG;Takahashi, T
1.用全细胞电压钳记录大鼠小脑脑片颗粒细胞兴奋性突触后电流（EPSC）。EPSC从个人凌乱的纤维输入，确定了他们的所有或没有出现在一个分级的刺激。通过分析EPSC的非N-甲基-D-天冬氨酸（非NMDA）组分的峰值和衰减中的电流波动，在室温（类似于24摄氏度）和接近生理温度（类似于34摄氏度）下研究了兴奋性突触传递。在一个突触的子集中，平均EPSC幅度保持不变，因为通过提高细胞外[Mg 2 +]和降低[Ca 2 +]，递质释放的概率大大降低。这些突触被认为只有一个功能性释放位点。单位点突触具有小的EPSC（139 +/- 16 pS，n = 5，在24 ° C下），具有大的变异系数（c. v.= 0.23 +/- 0.02，n = 5），并且在五种情况中有四种情况下的振幅分布与高斯分布很好地拟合。EPSC潜伏期呈单峰分布，其标准差与温度相关，温度系数（Q（10）;范围，24-35 ℃）为2.4 +/- 0.4（n = 4）。单位点EPSC的峰标度非稳态波动分析表明，在35 ℃下，潜在的非NMDA通道的平均电导为12 +/- 2 pS（n = 4）。平均通道开放概率（P-O）的上限和下限，从EPSC峰值振幅的波动计算，分别为0.51和0.38。这些估计值，连同当被发射器束缚时通道的开放概率，表明在释放一个量子的发射器之后，只有大约50%的非NMDA通道被占用。在一些多位点突触处，EPSC在34 ℃时具有低c. v.（0.04 +/- 0.01，n = 5），并且非平稳波动分析给出了抛物线方差-平均电流关系。这表明，在EPSC的峰值时，几乎所有非NMDA受体都被谷氨酸占据。因此，在这些“饱和”多位点突触处的通道开放概率（P-o = 0.84 +/-0.03，n = 5）将等于当由发射器（P-o，P-max）约束时通道的开放概率。非稳态波动分析的EPSC从“饱和”多站点突触表明，170 +/- 40突触后非NMDA通道暴露于发射器在EPSC的峰值。突触通道的平均电导为10 +/- 2 pS（n = 5）在34摄氏度。在具有多个释放位点的突触处，当释放概率降低时（通过降低外部[Ca 2 +]/[Mg 2 +]比率），EPSC衰减时间变得更快，表明递质浓度分布依赖于释放概率。我们的研究结果表明，从一个单一的释放位点释放一包递质不会饱和突触后非NMDA受体在小脑凌乱的纤维颗粒细胞突触。然而，在多位点突触中，从相邻位点释放的递质可能重叠，从而改变突触间隙中的递质浓度分布。我们的结论是，突触后受体占用的水平可以取决于在个别多站点突触的发射机释放的概率。
1. Excitatory postsynaptic currents (EPSCs) were recorded under whole-cell voltage clamp from granule cells in slices of rat cerebellum. EPSCs from individual messy fibre inputs were identified by their all-or-none appearance in response to a graded stimulus. Excitatory synaptic transmission was investigated at room temperature (similar to 24 degrees C) and at near-physiological temperature (similar to 34 degrees C) by analysing current fluctuations in the peak and decay of the non-N-methyl-D-aspartate (non-NMDA) component of EPSCs.2. In a subset of synapses the mean EPSC amplitude remained unchanged as the probability of transmitter release was substantially lowered by raising the extracellular [Mg2+] and lowering [Ca2+]. These synapses were considered to have only one functional release site. Single-site synapses had small EPSCs (139 +/- 16 pS, n = 5, at 24 degrees C) with a large coefficient of variation (c.v. = 0.23 +/- 0.02, n = 5) and an amplitude distribution that was well fitted by a Gaussian distribution in four out of five cases. The EPSC latency had a unimodal distribution and its standard deviation had a temperature dependence with a temperature coefficient (Q(10); range, 24-35 degrees C) of 2.4 +/- 0.4 (n = 4).3. Peak-scaled non-stationary fluctuation analysis of single-site EPSCs indicated that the mean conductance of the underlying non-NMDA channels was 12 +/- 2 pS (n = 4) at 35 degrees C. Upper and lower limits for mean channel open probability (P-o), calculated from fluctuations in the EPSC peak amplitude, were 0.51 and 0.38, respectively. These estimates, together with the open probability of the channel when bound by transmitter, suggest that only about 50% of the non-NMDA channels were occupied following the release of a quantum of transmitter.4. At some multi-site synapses EPSCs had a low c.v. (0.04 +/- 0.01, n = 5) at 34 degrees C and nonstationary fluctuation analysis gave a parabolic variance-mean current relationship. This suggests that practically all of the non-NMDA receptors were occupied by glutamate at the peak of the EPSC. The channel open probability (P-o = 0.84 +/- 0.03, n = 5) at these 'saturated' multi-site synapses will therefore equal the open probability of the channel when bound by transmitter (P-o,P-max).5. Non-stationary fluctuation analysis of EPSCs from 'saturating' multi-site synapses indicated that 170 +/- 40 postsynaptic non-NMDA channels were exposed to transmitter at the peak of the EPSC. The mean conductance of the synaptic channels was 10 +/- 2 pS (n = 5) at 34 degrees C.6. At synapses with multiple release sites the EPSC decay time became faster when release probability was lowered (by reducing the external [Ca2+]/[Mg2+] ratio), indicating that the transmitter concentration profile depended on release probability No such speeding of the EPSC decay was observed at single-site synapses.7. Our results suggest that release of a packet of transmitter from a single release site does not saturate postsynaptic non-NMDA receptors at cerebellar messy fibre-granule cell synapses. However, at multi-site synapses transmitter released from neighbouring sites can overlap, changing the transmitter concentration profile in the synaptic cleft. We conclude that the level of postsynaptic receptor occupancy can depend on the probability of transmitter release at individual multi-site synapses.