The role of extracellular vesicles as a novel trophic signaling platform in nervous system development - Project Summary/Abstract
In the developing nervous system, neurons are overproduced and subsequently pared down through cell
death to ensure the proper connections are supported and stabilized. Neurons that do not receive sufficient
support will die and be removed from the system. In the developing peripheral nervous system, sympathetic
circuits are refined by limiting amounts of nerve growth factor (NGF) that is secreted by target organs (e.g. eye
or salivary gland). NGF binds its receptor, TrkA, at the distal axon of innervating sympathetic neurons and
internalizes onto signaling endosomes (SEs). Since the cell bodies reside in sympathetic chain ganglia, SEs
need to be retrogradely trafficked to the soma for survival signaling. Insufficient NGF/TrkA uptake and trafficking
leads to cell death. Sympathetic neurons are innervated by autonomic preganglionic ChAT-positive neurons in
the spinal cord. Their survival is also NGF-dependent, despite lacking TrkA or access to soluble NGF. How these
preganglionic neurons survive without access to NGF is unknown. We hypothesize that TrkA-containing (TrkA+)
extracellular vesicles (EVs) secreted from sympathetic neurons provide this neurotrophic support.
Our recent publication is one of the first to show that sympathetic neurons secrete EVs with characteristic
sizes and protein marker composition. We identify TrkA as a novel EV cargo, which can be present in its active,
phosphorylated state. Using a compartmentalized culture system, we show that TrkA originating from distal
axons can be packaged in EVs and secreted from the somatodendritic domain. In addition, our preliminary data
show that sympathetic EVs reduce death of preganglionic ChAT-positive neurons, suggesting they can indeed
provide survival signaling. Interestingly, pilot experiments suggest that the TrkA+ EVs mediate this survival
response. Together, our foundational data led to the hypothesis that sympathetic EVs provide neurotrophic
support to preganglionic neurons, potentially via the TrkA+ subpopulation.
In this application, I aim to discover the pathways by which these signaling-competent EVs contribute to
preganglionic neuron survival. Aim 1: I will investigate the pathways by which EVs initiate signaling in recipient
cells using mass cytometry, a powerful technique that will enable the assessment of 20 signaling states with
single-cell resolution. I will test the hypothesis that a subpopulation selectively initiates survival signaling
pathways when exposed to EVs. Aim 2: Using live imaging in a compartmentalized culture system, I will
determine the contribution of long-distance communication within the preganglionic neuron to the neurotrophic
effects of sympathetic EVs. I will test the hypothesis that local accessibility of sympathetic EVs confers specificity
in recipient response and whether these EVs and their components undergo retrograde transport. The impact of
this work would be the discovery of a novel component of long-distance trophic signaling, i.e. signaling-
competent sympathetic EVs. Understanding mechanisms of sympathetic EV responses will greatly improve our
understanding of how a neuronal target dictates the entirety of a circuit to its needs.
