Image-based investigation of the zebrafish developmental process using in vivo dynamic and multi-contrast optical coherence tomography

TL;DR

This study develops and applies in vivo dynamic optical coherence tomography (DOCT) with LIV and OCDS_l contrasts, integrated with PS-OCT, OCTA, and histology, to follow zebrafish development from 2 weeks to 12 months post-fertilization. The authors implement a 32-frame DOCT protocol, motion-corrected imaging, and slab projections to visualize skin stripe formation, vascular maturation, and spinal organization across four early developmental stages and into adulthood. Key findings include stage-dependent stripe-associated DOCT signals linked to pigment-cell dynamics, segregation and remodeling of intramuscular vessels, and a dorsal shift in spinal cord activity that aligns with dorsal-ventral neural patterning. The multi-contrast, label-free approach provides a powerful framework for longitudinal in vivo studies of development, offering insights into dynamic cellular processes and their anatomical correlates with potential applications in developmental biology and disease modeling.

Abstract

We demonstrate in vivo dynamic optical coherence tomography (DOCT) imaging of zebrafish development from 2 weeks to 12 months post-fertilization, integrated with polarization-sensitive OCT (PS-OCT), OCT angiography (OCTA), and histological validation. Two DOCT algorithms were utilized: logarithmic intensity variance and late OCT correlation decay speed, which characterize the occupancy of dynamic scatterers and their motion speeds, respectively. Our results show that skin stripes exhibit high DOCT signals and it varies among the pigment-cell types. Furthermore, the combination of DOCT and PS-OCT captures the maturation of these stripes. In addition, DOCT and OCTA successfully visualized the developmental progression of blood and lymphatic vessels, as well as spinal tissues.

Figures (12)

• Figure 1: Cross-sectional and en face images of a 2-wpf zebrafish. (a1)--(a3) OCT intensity, LIV, and $\text{OCDS}_{\textit{l}}$ images at a location indicated by the orange arrow pair in (c1). (b1)--(b3) OCT intensity, LIV, and $\text{OCDS}_{\textit{l}}$ images at a location indicated by the red arrow pair in (c1). (c1)--(c3) The corresponding en face view of OCT Intensity, LIV, and $\text{OCDS}_{\textit{l}}$. The en face slice is indicated by the green arrow pair in (a1). (d)--(e) H&E-stained histological sections of the trunk in transverse (d) and sagittal (e) views. Scale bar represents 0.25 mm in (a1--c3). Abbreviation: SC, spinal cord; MBV, major blood vessels; NC, notochord; ISV, superficial lymphatic vessels. Supplementary Figure S1 presents the identical images shown in the present figure, but without the overlaying labels.
• Figure 2: Cross-sectional and en face images of a 3-wpf zebrafish. (a1)--(a3) OCT intensity, LIV and OCDSl images at a location indicated by the orange arrow pair in (c1). (b1)--(b3) OCT intensity, LIV and OCDSl images at a location indicated by the red arrow pair in (c1). (c1)--(c3) The corresponding en face view of OCT Intensity, LIV and $\text{OCDS}_{\textit{l}}$. The en face slice is indicated by the green arrow pair in (a1). (d)--(e) H&E-stained histological sections of the trunk in transverse (d) and sagittal (e) views. Scale bar represents 0.5 mm in (a1--c3). Abbreviation: SC, spinal cord; MBV, major blood vessels; NC, notochord; ISV, intersegmental vessels; SLV, superficial lymphatic vessels; HM, horizontal myoseptum. Supplementary Figure S2 presents the identical images shown in the present figure, but without the overlaying labels.
• Figure 3: Cross-sectional and en face images of 4-wpf and 5-wpf zebrafish. (a1)-(a3) OCT Intensity, LIV and $\text{OCDS}_{\textit{l}}$ images from a cross-section of the posterior trunk in 4-wpf zebrafish. (b1)-(b3) The corresponding en face view of OCT Intensity, LIV and $\text{OCDS}_{\textit{l}}$ in the 4-wpf zebrafish. (c1)-(c3) OCT Intensity, LIV and OCDSl images from a cross-section of the posterior trunk in 5-wpf zebrafish. (d1)-(d3) The corresponding en face view of OCT Intensity, LIV and $\text{OCDS}_{\textit{l}}$ in the 5-wpf zebrafish. (e1)--(f2) H&E-stained histological sections of the 4- and 5-wpf zebrafish's trunk in transverse and sagittal views. Color-coded arrow pairs indicate the spatial relationship between the cross-sectional and en face images. Scale bar represents 0.5 mm in (a1--d3). Abbreviation: SC, spinal cord; MBV, major blood vessels; NC, notochord; ISV, intersegmental vessels. Supplementary Figure S3 presents the identical images shown in the present figure, but without the overlaying labels.
• Figure 4: Cross-sectional and en face images of the 12-mpf adult zebrafish trunk. (a1--a3) OCT Intensity, LIV and $\text{OCDS}_{\textit{l}}$ images from a cross-section of the posterior trunk in 12-mpf zebrafish. (b1--b3) and (c1--c3) show the en face images from a superficial and a deep plane within the 12-mpf zebrafish, respectively. Color-coded arrow markers indicate the spatial relationship between the cross-sectional and en face images. Scale bar represents 0.5 mm in (a1--c3). Abbreviation: SC, spinal cord; MBV, major blood vessels. Supplementary Figure S4 presents the identical images shown in the present figure, but without the overlaying labels.
• Figure 5: Multi-contrast slab projection images of zebrafish skin from 2-wpf to 12-mpf. Images were arranged in a grid, where each row represents a specific developmental stage (2-wpf to 12-mpf) and each column corresponds to a different imaging contrast. And (e1--e5) are corresponding photographs for anatomical reference. Supplementary Figure S5 presents the identical images shown in the present figure, but without the overlaying labels.