Brain and Gut Dissection in Surface Fish and Cavefish Populations of <em>Astyanax mexicanus</em>

Teleost fish are used to answer a wide range of research questions spanning from development to genetics and neuroscience, and more recently, to understand the complexities of the gut-brain axis, and even in a wider context, the gut-brain-liver axis. The Mexican tetra, Astyanax mexicanus, has been increasingly used in comparative and developmental evolution frameworks due to the availability of morphologically and physiologically distinct morphotypes, including river-dwelling surface fish and cave-adapted, blind cavefish. Adaptation to cave habitats is associated with changes in morphological traits, behavioral traits, and gut microbiota diversity that are also found in laboratory conditions. This has opened up the possibility to ask questions about the interactions between the microbiome, metabolic activity, neural adaptation, and complex behavioral outputs. These interactions can now be addressed through whole-brain mapping studies with cleared brains and gut microbiota sequencing to detect bacterial diversity associated with surface and cave-adapted phenotypes. However, most of these studies would benefit from intact, carefully dissected, and preserved whole organs and tissues, which allow for post-processing in downstream applications. This protocol describes a step-by-step method for brain and gut dissection, where the whole brain and abdominal organs are removed and isolated for both surface fish and cavefish morphotypes.