Shaping the synapse: The molecular architecture and dynamics of the postsynaptic endocytic zone

Abstract

Neurons communicate by sending information through the axon and receiving information through the dendrites. The site where neurons make contact and communicate with each other, is called a synapse. Here, neurons communicate using neurotransmitters, that are released from presynaptic intracellular vesicles into the synaptic cleft. Neurotransmitters are then detected by receptors at the postsynaptic site, allowing the signal to be transduced. At glutamatergic synapses, glutamate receptors mediate synaptic transmission and facilitate synaptic plasticity. While fast transmission is mediated by the ionotropic NMDA and AMPA receptors, the metabotropic glutamate receptors 1 and 5 (mGluR1/5) act on longer timescales. Apart from their functional difference, the location of the receptor classes is distinct. While the ionotropic glutamate receptors are enriched in the postsynaptic density (PSD), a densely packed network of proteins closely attached to the postsynaptic membrane, the mGluRs are enriched in the perisynaptic domain surrounding the PSD. Despite the functional and organizational differences both receptor classes undergo dynamic trafficking to mediate synaptic plasticity. Membrane trafficking of glutamate receptors can be divided into multiple processes: exocytosis, lateral diffusion, endocytosis, sorting and recycling. Because glutamatergic synapses are isolated from the main dendritic branch, local mechanisms are in place to facilitate efficient membrane trafficking. As such, endocytosis of synaptic glutamate receptors is mediated by the postsynaptic endocytic zone (EZ). The EZ is a clathrin-marked structure stably associated with the PSD, and is thought to allow local endocytosis of glutamate receptors. Receptors internalized via the EZ, enter a local recycling mechanism, that retains receptors in intracellular pools and can recycle back to the synaptic membrane. The EZ plays a vital role in synaptic plasticity as loss of the EZ results in a depletion of glutamate receptors at the synaptic membrane and thereby hampers plasticity. Even though the EZ plays a critical role in synaptic plasticity, we know little about how the EZ is built to sustain local endocytosis of synaptic receptors. Moreover, the mechanisms that control the molecular composition and organization of the EZ are poorly understood. Revealing the organization of the EZ would greatly contribute to our understanding of how the EZ could allow local endocytosis of synaptic glutamate receptors. In the current thesis, we use a multitude of fluorescence microscopy techniques to visualize the architecture and dynamics of the EZ. We primarily focus on clathrin, that thus far is described to be the main protein of the EZ. As clathrin is the key component of clathrin-mediated endocytosis (CME), we and others assume that this is the mechanism of endocytosis at the EZ. While clathrin structures and function have been studied in great depth in non-neuronal cells, what types of clathrin structures are present in neurons, let alone their dynamics, architecture and involvement in receptor trafficking and plasticity are poorly understood. In the following chapters, we reveal the architecture of the EZ and the mechanisms that allow local capture and endocytosis of glutamate receptors that is so important for synaptic functioning.