Significance
The neurotransmitter dopamine controls normal behavior and dopaminergic dysfunction is prevalent in multiple brain diseases. To reach a detailed understanding of how dopamine release and signaling are regulated at the subcellular level, we developed a near infrared fluorescent dopamine nanosensor ‘paint’ (AndromeDA) to directly image dopamine release and its spatiotemporal characteristics. With AndromeDA, we can ascribe discrete DA release events to defined axonal varicosities, directly assess the heterogeneity of DA release events across such release sites, and determine the molecular components of the DA release machinery. AndromeDA thus provides a new method for gaining fundamental insights into the core mechanisms of dopamine release, which with greatly benefit our knowledge of dopamine biology and pathobiology.
Abstract
The neurotransmitter dopamine (DA) controls multiple behaviors and is perturbed in several major brain diseases. DA is released from large populations of specialized structures called axon varicosities. Determining the DA release mechanisms at such varicosities is essential for a detailed understanding of DA biology and pathobiology but has been limited by the low spatial resolution of DA detection methods. We used a near-infrared fluorescent DA nanosensor paint, adsorbed nanosensors detecting release of dopamine (AndromeDA), to detect DA secretion from cultured murine dopaminergic neurons with high spatial and temporal resolution. We found that AndromeDA detects discrete DA release events and extracellular DA diffusion and observed that DA release varies across varicosities. To systematically detect DA release hotspots, we developed a machine learning–based analysis tool. AndromeDA permitted the simultaneous visualization of DA release for up to 100 dopaminergic varicosities, showing that DA release hotspots are heterogeneous and occur at only ∼17% of all varicosities, indicating that many varicosities are functionally silent. Using AndromeDA, we determined that DA release requires Munc13-type vesicle priming proteins, validating the utility of AndromeDA as a tool to study the molecular and cellular mechanism of DA secretion.
