A Genetically Encoded Fluorescent Sensor Enables Rapid and Specific Detection of Dopamine in Flies, Fish, and Mice.

Dopamine (DA) is a central monoamine neurotransmitter involved in many physiological and pathological processes. A longstanding yet largely unmet goal is to measure DA changes reliably and specifically with high spatiotemporal precision, particularly in animals executing complex behaviors. Here, we report the development of genetically encoded GPCR-activation-based-DA (GRABDA) sensors that enable these measurements. In response to extracellular DA, GRABDA sensors exhibit large fluorescence increases (ΔF/F0 ∼90%) with subcellular resolution, subsecond kinetics, nanomolar to submicromolar affinities, and excellent molecular specificity. GRABDA sensors can resolve a single-electrical-stimulus-evoked DA release in mouse brain slices and detect endogenous DA release in living flies, fish, and mice. In freely behaving mice, GRABDA sensors readily report optogenetically elicited nigrostriatal DA release and depict dynamic mesoaccumbens DA signaling during Pavlovian conditioning or during sexual behaviors. Thus, GRABDA sensors enable spatiotemporally precise measurements of DA dynamics in a variety of model organisms while exhibiting complex behaviors.

Upstream surface roughness and terrain are strong drivers of contrast in tornado potential between North and South America.

Central North America is the global hotspot for tornadoes, fueled by elevated terrain of the Rockies to the west and a source of warm, moist air from equatorward oceans. This conventional wisdom argues that central South America, with the Andes to the west and Amazon basin to the north, should have a "tornado alley" at least as active as central North America. Central South America has frequent severe thunderstorms yet relatively few tornadoes. Here, we show that conventional wisdom is missing an important ingredient specific to tornadoes: a smooth, flat ocean-like upstream surface. Using global climate model experiments, we show that central South American tornado potential substantially increases if its equatorward land surface is smoothed and flattened to be ocean-like. Similarly, we show that central North American tornado potential substantially decreases if its equatorward ocean surface is roughened to values comparable to forested land. A rough upstream surface suppresses the formation of tornadic environments principally by weakening the poleward low-level winds, characterized by a weakened low-level jet east of the mountain range. Results are shown to be robust for any midlatitude landmass using idealized experiments with a simplified continent and mountain range. Our findings indicate that large-scale upstream surface roughness is likely a first-order driver of the strong contrast in tornado potential between North and South America.

Circadian regulation of developmental synaptogenesis via the hypocretinergic system.

The circadian clock orchestrates a wide variety of physiological and behavioral processes, enabling animals to adapt to daily environmental changes, particularly the day-night cycle. However, the circadian clock's role in the developmental processes remains unclear. Here, we employ the in vivo long-term time-lapse imaging of retinotectal synapses in the optic tectum of larval zebrafish and reveal that synaptogenesis, a fundamental developmental process for neural circuit formation, exhibits circadian rhythm. This rhythmicity arises primarily from the synapse formation rather than elimination and requires the hypocretinergic neural system. Disruption of this synaptogenic rhythm, by impairing either the circadian clock or the hypocretinergic system, affects the arrangement of the retinotectal synapses on axon arbors and the refinement of the postsynaptic tectal neuron's receptive field. Thus, our findings demonstrate that the developmental synaptogenesis is under hypocretin-dependent circadian regulation, suggesting an important role of the circadian clock in neural development.

Imaging volumetric dynamics at high speed in mouse and zebrafish brain with confocal light field microscopy.

A detailed understanding of the function of neural networks and how they are supported by a dynamic vascular system requires fast three-dimensional imaging in thick tissues. Here we present confocal light field microscopy, a method that enables fast volumetric imaging in the brain at depths of hundreds of micrometers. It uses a generalized confocal detection scheme that selectively collects fluorescent signals from the in-focus volume and provides optical sectioning capability to improve imaging resolution and sensitivity in thick tissues. We demonstrate recording of whole-brain calcium transients in freely swimming zebrafish larvae and observe behaviorally correlated activities in single neurons during prey capture. Furthermore, in the mouse brain, we detect neural activities at depths of up to 370 µm and track blood cells at 70 Hz over a volume of diameter 800 µm × thickness 150 µm and depth of up to 600 µm.

Dysregulation of Microglial Function Contributes to Neuronal Impairment in Mcoln1a-Deficient Zebrafish.

Type IV mucolipidosis (ML-IV) is a neurodegenerative lysosome storage disorder caused by mutations in the MCOLN1 gene. However, the cellular and molecular bases underlying the neuronal phenotypes of ML-IV disease remain elusive. Using a forward genetic screening, we identified a zebrafish mutant, biluo, that harbors a hypomorphic mutation in mcoln1a, one of the two zebrafish homologs of mammalian MCOLN1. The mcoln1a-deficient mutants display phenotypes partially recapitulating the key features of ML-IV disorder, including the accumulation of enlarged late endosomes in microglia and aberrant neuronal activities in both spontaneous and visual-evoking conditions in optic tectal neurons. We further show that the accumulation of enlarged late endosomes in microglia is caused by the impairment of late endosome and lysosome fusion and the aberrant neuronal activities can be partially rescued by the reconstitution of Mcoln1a function in microglia. Our findings suggest that dysregulation of microglial function may contribute to the development and progression of ML-IV disease.

All-optical imaging and manipulation of whole-brain neuronal activities in behaving larval zebrafish.

All-optical interrogation of population neuron activity is a promising approach to deciphering the neural circuit mechanisms supporting brain functions. However, this interrogation is currently limited to local brain areas. Here, we incorporate patterned photo-stimulation into light-sheet microscopy, allowing simultaneous targeted optogenetic manipulation and brain-wide monitoring of the neuronal activities of head-restrained behaving larval zebrafish. Using this system, we photo-stimulate arbitrarily selected neurons (regions as small as ~10-20 neurons in 3D) in zebrafish larvae with pan-neuronal expression of a spectrally separated calcium indicator, GCaMP6f, and an activity actuator, ChrimsonR, and observe downstream neural circuit activation and behavior generation. This approach allows us to dissect the causal role of neural circuits in brain functions and behavior generation.