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Fig. 4 Scanning Bessel foci with variable axial lengths probes varying volumes of spinal projection neurons in vivo. Reticulospinal and vestibulospinal neurons labeled with Alexa Fluor 488 in a larval zebrafish were imaged by scanning either (a,b) Gaussian or (c-j) Bessel foci. Left panels: neuron images; Right panels: axial profiles of 2-μm-diameter beads. (a) An image acquired using Gaussian focus at the relative depth of 18 μm (114 μm from the surface) contains 38 neurons. (b) Mean intensity projection over 74 μm axial range (absolute depth from 96 μm to 170 μm), color-coded by relative depths. (c-j) Volumetric images acquired by scanning Bessel foci with different axial lengths. Longer foci revealed more structures, e.g., (c) 65 neurons; (g): 87 neurons and (j): 108 neurons. Arrowheads (same color code as in b) in each image point to example new structures (neurons or axons) compared to the previous image. Axicon 1 and Mask 1 were employed here.