1) What regulates the amount of transmitter per vesicle (or quantal size), the elementary unit in synaptic transmission? We have previously identified three distinct protein families that transport neurotransmitters into secretory vesicles. Using a variety of biochemical and biophysical methods including fluorescence measurements and electrophysiology, we are characterizing the properties of these proteins, and exploring the role of other synaptic vesicle components in expression of the proton electrochemical gradient that drives vesicle filling.
2) What is the molecular basis for synaptic vesicle pools? Synaptic vesicles belong to functionally distinct pools, but the basis for these differences and the role of these pools in synaptic physiology and development remains unclear. We are now testing the possibility that different recycling pathways produce biochemically distinct synaptic vesicles.
3) Recent work from our and other labs has shown that many neurons associated with the release of one classical transmitter (such as dopamine, serotonin, acetylcholine and GABA) also release glutamate. We now wish to understand the role of glutamate corelease in physiology, development and behavior, focusing on the midbrain dopamine projection.
4) How does synaptic function contribute to neural degeneration? We originally identified the vesicular monoamine transporter by virtue of its ability to protect against a neurotoxin that reproduces the selective loss of dopamine neurons seen in Parkinson’s disease (PD), and are working now to understand how this and other presynaptic mechanisms influence degeneration. We have found that α-synuclein, a protein implicated in PD, inhibits transmitter release, and are now studying the mechanism as well as the relationship to neural degeneration.
5) In contrast to most classical transmitters, dopamine and neural peptides undergo regulated release from dendrites as well as the axon terminal, and the dendritic release of neurotrophins has been proposed to serve as a retrograde synaptic signal in development and plasticity. Through a combination of optical imaging in vitro and genetic manipulation in vivo, we hope to elucidate the physiological role of dendritic dopamine release in synaptic plasticity and determine its behavioral role in the reward pathway subverted by drug abuse.
6) How are proteins targeted for regulated release? Peptide hormones, neural peptides and many growth factors undergo regulated release in response to the appropriate physiological stimuli, but the cellular mechanisms that sort them to the large dense core vesicles (LDCVs) capable of regulated exocytosis, and indeed the mechanisms responsible for this pathway remain poorly understood. Starting with an RNAi screen in Drosophila S2 cells, we have identified a role for the adaptor protein AP-3 and a novel coat protein in the formation of LDCVs. We now wish to identify additional components required for LDCV formation, and to manipulate these in vivo to understand how regulated release contributes to the activity dependence of neural development.