Abstract Synaptic vesicle (SV) recycling is critical for sustaining neurotransmission. Although FCHO1, a protein containing both an F-BAR domain and a μ-homology (μ-HD) domain, is recognized as a nucleator of clathrin-mediated endocytosis in non-neuronal systems, its physiological role at synapses remains unclear. Here, we investigated the function of FCHO1 in SV endocytosis at central synapses using a combination of shRNA-mediated knockdown and pHluorin-based live imaging. Within defined stimulation paradigms (25–300 action potentials at 10 Hz), depletion of FCHO1 markedly slowed endocytic kinetics across all stimulation intensities and was fully rescued by re-expression of an shRNA-resistant construct. Domain-specific functional analyses revealed stimulation-strength-dependent functional requirements. The F-BAR domain was sufficient to support vesicle retrieval under low stimulation conditions, whereas the μ-homology domain (μ-HD) became essential as stimulation strength increased. These findings support a model in which FCHO1 operates as a demand-sensitive scaffold within the endocytic pathway, with distinct structural domains differentially required as neural activity and consequently endocytic load escalates. Our results establish FCHO1 as a critical regulator of SV endocytosis and suggest that multidomain endocytic proteins may scale their functional contributions according to the magnitude of neuronal activation.
Lee et al. (Mon,) studied this question.