The formation of a synapse: measuring vesicle cycling in growth cones and developing presynaptic terminals

大脑中的神经元在称为突触的特殊位置相互通信。在大脑发育过程中，大量神经元(约100亿个)形成数量惊人的大量突触连接(约100万亿个)，形成一个有功能的大脑。毫不奇怪，神经科学中的一个重要问题是了解突触是如何形成的，以及这些连接是如何将信息从一个神经元传递到另一个神经元的。在突触，信息的传递是通过将突触前神经元释放的神经递质包释放到突触后神经元上的特定受体来完成的。神经递质储存在膜小泡内，通过小泡与质膜的融合释放，从而将神经递质排向突触后神经元。随后从质膜中取回来自囊泡的膜，以用神经递质重新填充并再次循环使用。这种囊泡循环的一个重要方面是它受到神经元活动的严格控制。这里提出的研究将集中于了解突触前终末是如何形成和成熟的，以提供这样一个精细控制的神经递质释放系统。为此，我将使用特别设计的荧光探针，通过在每次囊泡与质膜融合时显示荧光闪烁来实时报告囊泡的循环。在神经元形成突触之前，它必须首先到达它的目标神经元。轴突寻路的过程依赖于轴突顶端的一个特殊隔室，即生长锥体，它充当导航器，引导轴突到达正确的目标。突触前终末的形成可以看作是一系列事件的过程，这些事件跨越了最初的生长锥运动，参与了轴突的寻路，然后是生长锥在靶神经元上的稳定和突触的形成。在所有这些过程中，囊泡循环也发展和成熟：从生长锥体的初始混合形式的囊泡循环，到成熟末端的纯粹依赖活动的形式。在这里，我将研究囊泡循环是如何成熟的，并导致释放神经递质的功能性突触前终末。了解这些事件将有助于揭示大脑在发育过程中如何形成连接，并帮助我们了解这一过程出错的神经疾病。

Neurons in the brain communicate with each other at specialized sites called synapses. During brain development a large number of neurons (about 10 billion) make an astonishingly large number of synaptic connections (about 100 trillion) to form a functional brain. Not surprisingly, an important question in neuroscience is to understand how synapses are formed and how these connections pass information from one neuron to the next. At the synapse, the transfer of information is carried out by the release of packets of neurotransmitter released from a presynaptic neuron onto specific receptors on a postsynaptic neuron. Neurotransmitter is stored inside membrane vesicles and is released by the fusion of vesicles with the plasma membrane, which expels the neurotransmitter towards the postsynaptic neuron. The membrane from the vesicle is subsequently retrieved from the plasma membrane to be refilled with neurotransmitter and recycled for use once again. An important aspect of this vesicle cycle is that it is tightly controlled by the activity of a neuron. The research proposed here will focus on understanding how a presynaptic terminal forms and matures to provide such an exquisitely controlled system for releasing neurotransmitter. For this purpose I will use especially designed fluorescent probes that report vesicle cycling in real time by showing flashes of fluorescence every time a vesicle fuses with the plasma membrane. Before a neuron forms a synapse it must first reach its target neuron. This process of axon pathfinding relies on a special compartment at the tip of the axon, the growth cone, which acts as a navigator, guiding the axon to its correct target. The formation of a presynaptic terminal can be seen as a progression of events spanning the initial growth cone motility involved in axon pathfinding, followed by the stabilization of the growth cone at the target neuron and the formation of a synapse. Throughout all these processes, vesicle cycling also develops and matures: from an initial mixed form of vesicle cycling at the growth cone to a purely activity-dependent form at mature terminals. Here, I will study how vesicle cycling matures and results in a functional presynaptic terminal that releases neurotransmitter. Understanding these events will help uncover how connections in the brain form during development and help us understand neurological diseases where this process has gone wrong.