NeuroExaminer: an all-glass microfluidic device for whole-brain in vivo imaging in zebrafish

While microfluidics enables chemical stimuli application with high spatio-temporal precision, light-sheet microscopy allows rapid imaging of entire zebrafish brains with cellular resolution. Both techniques, however, have not been combined to monitor whole-brain neural activity yet. Unlike conventional microfluidics, we report here an all-glass device (NeuroExaminer) that is compatible with whole-brain in vivo imaging using light-sheet microscopy and can thus provide insights into brain function in health and disease

Emotions and motivated behavior converge on an amygdala-like structure in the zebrafish

International audienceThe brain reward circuitry plays a key role in emotional and motivational behaviors, and its dysfunction underlies neuropsychiatric disorders such as schizophrenia, depression and drug addiction. Here, we characterized the neuronal activity pattern induced by acute amphetamine administration and during drug-seeking behavior in the zebrafish, and demonstrate the existence of conserved underlying brain circuitry. Combining quantitative analyses of cfos expression with neuronal subtype-specific markers at single-cell resolution, we show that acute d-amphetamine administration leads to both increased neuronal activation and the recruitment of neurons in the medial (Dm) and the lateral (Dl) domains of the adult zebrafish pallium, which contain homologous structures to the mammalian amygdala and hippocampus, respectively. Calbindin-positive and glutamatergic neurons are recruited in Dm, and glutamatergic and γ-aminobutyric acid (GABAergic) neurons in Dl. The drug-activated neurons in Dm and Dl are born at juvenile stage rather than in the embryo or during adulthood. Furthermore, the same territory in Dm is activated during both drug-seeking approach and light avoidance behavior, while these behaviors do not elicit activation in Dl. These data identify the pallial territories involved in acute psychostimulant response and reward formation in the adult zebrafish. They further suggest an evolutionarily conserved function of amygdala-like structures in positive emotions and motivated behavior in zebrafish and mammals

A 3D tailored monolithic glass chip for stimulating and recording zebrafish neuronal activity with a commercial light sheet microscope

Microfluidic technology is unrivaled in its ability to apply soluble chemical stimuli with high spatiotemporal precision. Analogous, light–sheet microscopy is unmatched in its ability of low phototoxic but fast volumetric in vivo imaging with single cell resolution. Due to their optical translucency during the larval stages, zebrafish (Danio rerio) are an ideal model to combine both techniques; yet, thus far this required light–sheet microscopes, which were in most cases custom–built and adapted to the available softlithographic chip technology. Our aim was to use a commercial light–sheet microscope to illuminate a microfluidic chip from two opposite lateral directions and to record images with the detection objective placed orthogonally above the chip. Deep tissue penetration can be achieved by superimposing beams from opposite directions to form a single light sheet. But a microfluidic chip that allows a) targeted stimulus application in a closed microenvironment, b) interference–free incoupling of excitation light from two directions and c) outcoupling of fluorescence in the perpendicular direction through an optically perfect cover glass was not known until now. Here, we present a monolithic glass chip with the required plane-parallel sidewalls and cover slide closure at the top, constructed by advanced femtosecond laser ablation, thermal bonding and surface smoothing processes. In addition, the 3D shape of a fish fixator unit was tailored to match the body shape of a zebrafish larva to ensure stable positioning during whole–brain recording. With hydrodynamic focusing a targeted partial exposure of the larva’s head to chemical stimuli and fast position switching (in less than 10 s) was possible. With the capabilities of this unique monolithic glass chip and its up–scalable wafer–level fabrication process, the new NeuroExaminer is prone to become an excellent addition to neurobiology laboratories already equipped with high–quality commercial light sheet microscopes

A comparative analysis of Danionella cerebrum and zebrafish (Danio rerio) larval locomotor activity in a light-dark test

The genus Danionella comprises some of the smallest known vertebrate species and is evolutionary closely related to the zebrafish, Danio rerio. With its optical translucency, rich behavioral repertoire, and a brain volume of just 0.6 mm3, Danionella cerebrum (Dc) holds great promise for whole-brain in vivo imaging analyses with single cell resolution of higher cognitive functions in an adult vertebrate. Little is currently known, however, about the basic locomotor activity of adult and larval Danionella cerebrum and how it compares to the well-established zebrafish model system. Here, we provide a comparative developmental analysis of the larval locomotor activity of Dc and AB wildtype as well as crystal zebrafish in a light-dark test. We find similarities but also differences in both species, most notably a striking startle response of Dc following a sudden dark to light switch, whereas zebrafish respond most strongly to a sudden light to dark switch. We hypothesize that the different startle responses in both species may stem from their different natural habitats and could represent an opportunity to investigate how neural circuits evolve to evoke different behaviors in response to environmental stimuli