Spatial and Temporal Regulation of Receptor Endocytosis in Neuronal Dendrites Revealed by Imaging of Single Vesicle Formation

Abstract

Endocytosis in neuronal dendrites is known to play a critical role in synaptic transmission and plasticity such as long-term depression (LTD). However, the inability to detect endocytosis directly in living neurons has hampered studies of its dynamics and regulation. Here, we visualized the formation of individual endocytic vesicles containing pHluorin-tagged receptors with high temporal resolution in the dendrites of cultured hippocampal neurons. We show that transferrin receptors (TfRs) are constitutively internalized at optically static clathrin-coated structures. These structures are slightly enriched near synapses that represent preferential sites for the endocytosis of postsynaptic AMPA-type receptors (AMPARs), but not for non-synaptic TfRs. Moreover, the frequency of AMPAR endocytosis events increases after the induction of NMDAR-dependent chemical LTD, but the activity of perisynaptic endocytic zones is not differentially regulated. We conclude that endocytosis is a highly dynamic and stereotyped process that internalizes receptors in precisely localized endocytic zones.

Specific nanoscale synaptic reshuffling and control of short-term plasticity following NMDAR- and P2XR-dependent Long-Term Depression

“…However, we cannot determine whether a long term endocytosis/exocytosis unbalance is due to the maintenance of a high endocytosis rate or to a decrease of exocytosis rate as already described after NMDA treatment by Fujii et al (46). In parallel, a large increase in the AM-PAR endocytosis rate after NMDA treatment has been reproducibly reported (43)(44)(45). This increase in AMPAR internalization rate, likely responsible for the initiation of the LTD, stays for less than 10 minutes (45).…”

“…In parallel, a large increase in the AM-PAR endocytosis rate after NMDA treatment has been reproducibly reported (43)(44)(45). This increase in AMPAR internalization rate, likely responsible for the initiation of the LTD, stays for less than 10 minutes (45). Contrary to ATP treatment, the maintenance of the depression is associated with changes of both AMPAR mobility and a decrease of the number of PSD-95 slots.…”

“…Both P2XR-and NMDAR-dependent LTD induction have been attributed to a net increase in AMPAR endocytosis rate (24,(43)(44)(45). However, the mechanisms involved in the longterm maintenance of synaptic depression are less clear, and as demonstrated with simulation ( Figure 5S2), a transient increase in endocytosis rate triggers only a transient depression and not a long term one.…”

“…The existence of various phases during NMDAR-dependent LTD has been described previously. Rosendale et al demonstrated that the endocytosis rate increases immediately after LTD induction but then goes back to its original value after approximately 10 minutes (45). Sanderson et al 2016, reported a rapid and transient increase in calcium-permeable AMPARs (49).…”

Detection and characterization of individual endocytosis of AMPA‐type glutamate receptor around postsynaptic membrane

“…However, it was difficult to detect individual endocytic events of AMPAR. Very recently, individual endocytosis of SEP-tagged AMPAR was detected using TIRFM and an extracellular pH exchange method (Rosendale et al 2017). In that study, extracellular pH (7.4) was rapidly changed to acidic pH, upon which SEP-tagged AMPAR signals on the cell surface were quenched and only intracellular SEPtagged AMPAR signals soon after internalization were recorded.…”