Molecular mechanisms of synapse development and plasticity

通过使用基因敲除和基因敲除小鼠，我们证明BAD-Bax-caspase-3级联反应是诱导NMDA受体依赖的LTD和AMPA受体内吞，而不是诱导mGluR-LTD或突触传递的长期增强(Li等人，Cell 2010；Jiao等人，Neuron 2011)。然而，与细胞凋亡不同的是，BAD只被短暂和温和地激活，导致适度和短暂的caspase-3激活，不足以诱导细胞死亡。我们的发现揭示了LTD诱导的核心信号级联，并提供了证据表明，caspase-3激活的机制和数量上的差异对于确定其是否诱导LTD或细胞凋亡至关重要。 为了确定Bad-Bax-caspase-3级联是否调节突触传递的其他特性，我们测量了诱发和自发的兴奋性突触反应，并使用电子显微镜评估了突触的超微结构。为了确定促进LTD和AMPA受体内吞的BAD-Bax-caspase-3级联反应的下游效应因子，我们与Sandy Markey博士合作寻找在LTD中被切割的caspase底物。我们将使用电生理学来确定所鉴定的caspase底物是否参与LTD的诱导。 我们之前曾与Richard Youle博士合作研究Bax在维持线粒体形态中的功能(Norris等人，Cell Death and Differential 2010)。线粒体形态对支持突触的发育和功能具有重要意义。为了测试Bad-Bax-caspase-3级联是否调节线粒体的形态，我们研究了Bad、Bax和caspase-3基因敲除小鼠的线粒体。我们在BAD和Bax基因敲除神经元中发现了线粒体的碎片化表型和线粒体融合缺陷。因此，我们认为BAD是Bax的重要激活剂，对维持神经元线粒体形态有重要作用。

By using gene knockdown and knockout mice, we demonstrated that the BAD-BAX-caspase-3 cascade is to induce NMDA receptor-dependent LTD and AMPA receptor endocytosis, but not for mGluR-LTD or long-term potentiation of synaptic transmission (Li et al., Cell 2010; Jiao et al., Neuron 2011). Unlike in apoptosis, however, BAD is activated only transiently and mildly leading to modest and transient caspase-3 activation insufficient to induce cell death. Our findings reveal a core signaling cascade for LTD induction and provide evidence that mechanistic and quantitative differences in caspase-3 activation are critical for determining whether it induces LTD or apoptosis. To determine if the BAD-BAX-caspase-3 cascade regulates other properties of synaptic transmission, we measured evoked and spontaneous excitatory synaptic response, and using electron microscopy to assess the ultrastructure of synapse in the knockout mice. To identify downstream effectors of the BAD-BAX-caspase-3 cascade that promotes LTD and AMPA receptor endocytosis, we collaborate with Dr. Sandy Markey to search for caspase substrates that are cleaved in LTD. We will use electrophysiology to determine if the identified caspase substrate is involved in LTD induction. We have previously collaborated with Dr. Richard Youle to examine the function of BAX in the maintenance of mitochondrial morphology (Norris et al., Cell Death and Differentiation 2010). Mitochondria morphology is important for mitochondria to support the development and function of synapses. To test whether the BAD-BAX-caspase-3 cascade regulates mitochondria morphology, we have examined mitochondria in BAD, BAX and caspase-3 knockout mice. We identified a fragmented mitochondrial phenotype and mitochondrial fusion defect in BAD and BAX knockout neurons. Therefore, we propose that BAD is an important activator of BAX for maintenance of mitochondria morphology in neurons.