Novel tools for simultaneous optogenetic manipulation and calcium imaging in the zebrafish nervous system

The large number of neurons and neural interconnection makes the nervous system a highly dense and complex network. Understanding the functionality of such a network requires not only high-throughput recording of neural activities with cellular resolution but also a non-invasive and precisely defined interaction with the neurons. This work demonstrates two pivotal steps towards these aims:
(i) Developing a fluorescence microscope based on Bessel light-sheet illumination to record neural activities by means of calcium imaging. Chapter II describes the successful development and construction of an in vivo light-sheet fluorescence microscope (in vivo LSFM) based on conventional digitally laser scanning light-sheet microscopy (DSLM) with interchangeable Gaussian and Bessel illumination modalities to compare the performance of both methods. The Bessel illumination modality in comparison to the Gaussian one reveals not only a two-fold improvement in axial resolution but also a reduction by the factor of 4 in the shadowing artifact and consequently, 35 times improvement in detecting the correct neural activities from the calcium signals.
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In future, crossing this line with an already established line expressing a calcium indicator protein will provide the essential animal model for simultaneous optogenetic manipulation and high-fidelity neural activity recording in the intact nervous system.