diceCT: A Valuable Technique to Study the Nervous System of Fish

Abstract

Contrast-enhanced X-ray imaging provides a non-destructive and flexible approach to optimizing contrast in soft tissues, especially when incorporated with Lugol’s solution (aqueous I 2 KI), a technique currently referred to as diffusible iodine-based contrast-enhanced computed tomography (diceCT). This stain exhibits high rates of penetration and results in excellent contrast between and within soft tissues, including the central nervous system. Here, we present a staining method for optimizing contrast in the brain of a cartilaginous fish, the brownbanded bamboo shark, Chiloscyllium punctatum , and a bony fish, the common goldfish, Carassius auratus , using diceCT. The aim of this optimization procedure is to provide suitable contrast between neural tissue and background tissue(s) of the head, thereby facilitating digital segmentation and volumetric analysis of the central nervous system. Both species were scanned before staining and were rescanned at time (T) intervals, either every 48 h ( C. punctatum ) or every 24 h ( C. auratus ), to assess stain penetration and contrast enhancement. To compare stain intensities, raw X-ray CT data were reconstructed using air and water calibration phantoms that were scanned under identical conditions to the samples. Optimal contrast across the brain was achieved at T = 240 h for C. punctatum and T = 96 h for C. auratus . Higher resolution scans of the whole brain were obtained at the two optimized staining times for all the corresponding specimens. The use of diceCT provides a new and valuable tool for visualizing differences in the anatomic organization of both the central and peripheral nervous systems of fish.

Morphometric analysis of the elasmobranch olfactory rosette

“…We measured internal metrics of the olfactory system, including the volume of the excurrent channel (Figures 9 and 11; Table 3). A recent study using diceCT to examine an elasmobranch nervous system reported a shrinkage of 14% for brain tissue in their specimens ( Chiloscyllium punctatum ; Camilieri‐Asch, Shaw, et al, 2020). While the diceCT protocol differed from this study, such as choice of fixative and percentage of Lugol's iodine used, their results suggest that the excurrent channel measured could be partially affected by tissue shrinkage during preservation.…”

Spike Generators and Cell Signaling in the Human Auditory Nerve: An Ultrastructural, Super-Resolution, and Gene Hybridization Study

“…We used a diffusible iodine-based technique to enhance contrast of soft tissues for diffusible iodine-based contrast-enhanced computed tomography (diceCT). Increased time penetration of Lugol's iodine (aqueous I2KI, 1% I 2 , 2% KI) offers possibilities to visualize between and within soft tissue structures (Camilieri-Asch et al, 2020 ). The temporal bone was fixed in a modified Karnovsky's fixative solution of 2.5% glutaraldehyde, 1% paraformaldehyde, 4% sucrose, and 1% dimethyl sulfoxide in 0.13 M of Sorensen's phosphate buffer.…”

A Synchrotron and Micro-CT Study of the Human Endolymphatic Duct System: Is Meniere's Disease Caused by an Acute Endolymph Backflow?

“…We used a diffusible iodine-based technique to enhance contrast of soft tissues for diffusible iodine-based contrast-enhanced computed tomography (diceCT). Increased time penetration of Lugol's iodine (aqueous I2KI, 1% I 2 , 2% KI) provided possibilities to visualize between and within soft tissue structures ( 24 ). The temporal bone was fixed in a modified Karnovsky's fixative solution of 2.5% glutaraldehyde, 1% paraformaldehyde, 4% sucrose, and 1% dimethyl sulfoxide in 0.13 M of Sorensen's phosphate buffer.…”