Optimizing Nervous System-Specific Gene Targeting with Cre Driver Lines: Prevalence of Germline Recombination and Influencing Factors.

The Cre-loxP system is invaluable for spatial and temporal control of gene knockout, knockin, and reporter expression in the mouse nervous system. However, we report varying probabilities of unexpected germline recombination in distinct Cre driver lines designed for nervous system-specific recombination. Selective maternal or paternal germline recombination is showcased with sample Cre lines. Collated data reveal germline recombination in over half of 64 commonly used Cre driver lines, in most cases with a parental sex bias related to Cre expression in sperm or oocytes. Slight differences among Cre driver lines utilizing common transcriptional control elements affect germline recombination rates. Specific target loci demonstrated differential recombination; thus, reporters are not reliable proxies for another locus of interest. Similar principles apply to other recombinase systems and other genetically targeted organisms. We hereby draw attention to the prevalence of germline recombination and provide guidelines to inform future research for the neuroscience and broader molecular genetics communities.

Brain-wide cellular resolution imaging of Cre transgenic zebrafish lines for functional circuit-mapping.

Decoding the functional connectivity of the nervous system is facilitated by transgenic methods that express a genetically encoded reporter or effector in specific neurons; however, most transgenic lines show broad spatiotemporal and cell-type expression. Increased specificity can be achieved using intersectional genetic methods which restrict reporter expression to cells that co-express multiple drivers, such as Gal4 and Cre. To facilitate intersectional targeting in zebrafish, we have generated more than 50 new Cre lines, and co-registered brain expression images with the Zebrafish Brain Browser, a cellular resolution atlas of 264 transgenic lines. Lines labeling neurons of interest can be identified using a web-browser to perform a 3D spatial search (zbbrowser.com). This resource facilitates the design of intersectional genetic experiments and will advance a wide range of precision circuit-mapping studies.

Search strategy is regulated by somatostatin signaling and deep brain photoreceptors in zebrafish.

BACKGROUND: Animals use sensory cues to efficiently locate resources, but when sensory information is insufficient, they may rely on internally coded search strategies. Despite the importance of search behavior, there is limited understanding of the underlying neural mechanisms in vertebrates. RESULTS: Here, we report that loss of illumination initiates sophisticated light-search behavior in larval zebrafish. Using three-dimensional tracking, we show that at the onset of darkness larvae swim in a helical trajectory that is spatially restricted in the horizontal plane, before gradually transitioning to an outward movement profile. Local and outward swim patterns display characteristic features of area-restricted and roaming search strategies, differentially enhancing phototaxis to nearby and remote sources of light. Retinal signaling is only required to initiate area-restricted search, implying that photoreceptors within the brain drive the transition to the roaming search state. Supporting this, orthopediaA mutant larvae manifest impaired transition to roaming search, a phenotype which is recapitulated by loss of the non-visual opsin opn4a and somatostatin signaling. CONCLUSION: These findings define distinct neuronal pathways for area-restricted and roaming search behaviors and clarify how internal drives promote goal-directed activity.