Axonal targeting of trophic receptors is critical for neuronal responses to extracellular developmental cues, yet the underlying trafficking mechanisms remain unclear. Here, we report that tropomyosin-related kinase (Trk) receptors for target-derived neurotrophins are anterogradely trafficked to axons via transcytosis in sympathetic neurons. Using compartmentalized cultures, we show that mature receptors on neuronal soma surfaces are endocytosed and remobilized via Rab11-positive recycling endosomes into axons. Inhibition of dynamin-dependent endocytosis disrupted anterograde transport and localization of TrkA receptors in axons. Anterograde TrkA delivery and exocytosis into axon growth cones is enhanced by nerve growth factor (NGF), acting locally on distal axons. Perturbing endocytic recycling attenuated NGF-dependent signaling and axon growth while enhancing recycling conferred increased neuronal sensitivity to NGF. Our results reveal regulated transcytosis as an unexpected mode of Trk trafficking that serves to rapidly mobilize ready-synthesized receptors to growth cones, thus providing a positive feedback mechanism by which limiting concentrations of target-derived neurotrophins enhance neuronal sensitivity.

Dendritic arborization of neurons is regulated by brain-derived neurotrophic factor (BDNF) together with its receptor TrkB. Endocytosis is required for dendritic branching and regulates TrkB signaling, but how post-endocytic trafficking determine the neuronal response to BDNF is not well understood. The monomeric GTPase Rab11 regulates the dynamics of recycling endosomes and local delivery of receptors to specific dendritic compartments. Our aim was to study whether Rab11-dependent trafficking of TrkB in dendrites regulates BDNF-induced dendritic branching in rat hippocampal neurons. We report that TrkB in dendrites is a cargo for Rab11 endosomes and both Rab11 and its effector MyoVb are required for BDNF/TrkB-induced dendritic branching. In turn, BDNF induces accumulation of Rab11-positive endosomes and GTP-bound Rab11 in dendrites. Moreover, the expression of a constitutively active mutant of Rab11 is sufficient to increase dendritic branching by increasing TrkB localization in dendrites and enhancing sensitization to endogenous BDNF. We propose that Rab11-dependent dendritic recycling provides a mechanism to retain TrkB in dendrites and increase local signaling to regulate arborization.

Summary Endocytic events are critical for neuronal survival in response to target-derived neurotrophic cues, but whether local axon growth is mediated by endocytosis-dependent signaling mechanisms remains unclear. Here, we report that Nerve Growth Factor (NGF) promotes endocytosis of its TrkA receptors and axon growth by calcineurin-mediated dephosphorylation of the endocytic GTPase, dynamin1. Conditional deletion of calcineurin in sympathetic neurons disrupts NGF-dependent innervation of peripheral target tissues. Calcineurin signaling is required locally in sympathetic axons to support NGF-mediated growth in a manner independent of transcription, during the initial phase of axonal outgrowth. We show that calcineurin associates with dynamin1 via a PxIxIT interaction motif found only in specific dynamin1 splice variants. PxIxIT-containing dynamin1 isoforms co-localize with surface TrkA receptors, and their phosphoregulation is selectively required for NGF-dependent TrkA internalization and axon growth in sympathetic neurons. Thus, NGF-dependent phosphoregulation of dynamin1 is a critical event coordinating neurotrophin receptor endocytosis and initial axonal growth.

During the formation of neuronal circuits, neurons respond to diffusible cues secreted by target tissues. Often, target-derived signals act on nerve terminals to influence local growth events; in other cases, they are transported long-distance back to neuronal cell bodies to influence transcriptional changes necessary for neuronal survival and differentiation. Neurotrophins provide one of the best examples of target-derived cues that elicit an astonishingly diverse array of neuronal responses. Endocytic trafficking of neurotrophins and their receptors is a fundamental feature of neurotrophin signaling, allowing neurotrophins to control neuronal survival by retrograde transport of signaling endosomes containing ligand-receptor complexes. In this review, we summarize recent findings that provide new insight into the interplay between neurotrophin signaling and trafficking.

Down syndrome is the most common chromosomal disorder affecting the nervous system in humans. To date, investigations of neural anomalies in Down syndrome have focused on the central nervous system, although dysfunction of the peripheral nervous system is a common manifestation. The molecular and cellular bases underlying peripheral abnormalities have remained undefined. Here, we report the developmental loss of sympathetic innervation in human Down syndrome organs and in a mouse model. We show that excess regulator of calcineurin 1 (RCAN1), an endogenous inhibitor of the calcineurin phosphatase that is triplicated in Down syndrome, impairs neurotrophic support of sympathetic neurons by inhibiting endocytosis of the nerve growth factor (NGF) receptor, TrkA. Genetically correcting RCAN1 levels in Down syndrome mice markedly improves NGF-dependent receptor trafficking, neuronal survival and innervation. These results uncover a critical link between calcineurin signalling, impaired neurotrophin trafficking and neurodevelopmental deficits in the peripheral nervous system in Down syndrome.