Development of rat CA1 neurones in acute versus organotypic slices:: role of experience in synaptic morphology and activity
Development of rat CA1 neurones in acute versus organotypic slices:: role of experience in synaptic morphology and activity
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DOI：
10.1113/jphysiol.2003.039099
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发表时间：
2003-07-01
影响因子：
5.5
通讯作者：
Edwards, FA
中科院分区：
文献类型：
--
作者：
De Simoni, A;Griesinger, CB;Edwards, FA
尽管它们被广泛使用，器官型切片的生理相关性仍然存在争议。这种培养物在出生后5天制备。虽然某些局部回路保持完整，但它们随后在与动物隔离的情况下发展，因此由于经验而没有可塑性。在这些条件下，突触连接和形态的发展可能会与行为动物不同。为了解决这些问题，膜片钳技术和共聚焦显微镜在大鼠海马CA1区比较急性切片从出生后第三周的器官型切片的各个阶段。在出生后第14、17和21天制备的急性切片被发现在突触传递和树突形态的发育方面分别与培养1、2和3周的器官型切片发育相当。抑制性和兴奋性微小突触电流的频率平行增加。在这些相应的阶段，树突长度和初级分支以及棘密度和不同棘类型的比例在两种制剂中也相似。器官型和急性切片之间最显着的差异是在器官型切片中的GABA能（但不是GABA能）微型突触后电流的绝对频率增加了4至5倍。这可能与器官型切片中高阶树突分支的复杂性增加有关，如通过分形分析测量的，导致总突触数量增加。在培养的第一周内，兴奋性微型突触电流频率和树突复杂性都已增加。在随后的阶段中，两种制备物的复杂性水平保持不变，突触频率平行增加。因此，尽管器官型切片的连通性更大，但一旦建立，两种制剂的发育仍以非常相似的速度继续。我们的结论是，对于研究的参数，变化似乎是预先编程的5天，其随后的发展在很大程度上是独立的环境。
Despite their wide use, the physiological relevance of organotypic slices remains controversial. Such cultures are prepared at 5 days postnatal. Although some local circuitry remains intact, they develop subsequently in isolation from the animal and hence without plasticity due to experience. Development of synaptic connectivity and morphology might be expected to proceed differently under these conditions than in a behaving animal. To address these questions, patch-clamp techniques and confocal microscopy were used in the CA1 region of the rat hippocampus to compare acute slices from the third postnatal week with various stages of organotypic slices. Acute slices prepared at postnatal days (P) 14, 17 and 21 were found to be developmentally equivalent to organotypic slices cultured for 1, 2 and 3 weeks, respectively, in terms of development of synaptic transmission and dendritic morphology. The frequency of inhibitory and excitatory miniature synaptic currents increased in parallel. Development of dendritic length and primary branching as well as spine density and proportions of different spine types were also similar in both preparations, at these corresponding stages. The most notable difference between organotypic and acute slices was a four- to five-fold increase in the absolute frequency of glutamatergic (but not GABAergic) miniature postsynaptic currents in organotypic slices. This was probably related to an increase in complexity of higher order dendritic branching in organotypic slices, as measured by fractal analysis, resulting in an increased total synapse number. Both increased excitatory miniature synaptic current frequency and dendritic complexity were already established during the first week in culture. The level of complexity then stayed constant in both preparations over subsequent stages, with synaptic frequency increasing in parallel. Thus, although connectivity was greater in organotypic slices, once this was established, development continued in both preparations at a remarkably similar rate. We conclude that, for the parameters studied, changes seem to be preprogrammed by 5 days and their subsequent development is largely independent of environment.