Student Feature | Biology and Engineering are Merged for Interdisciplinary Undergraduate Research in Sea Slugs
The California sea hare (Aplysia californica) is a large sea slug native to the Pacific Coast of California and Mexico. They can grow as large as a football and get their maroon coloration from the macroalgae they feed on in the intertidal zone. Aplysia are important biomedical models for human health contributing to our understanding of neurological processes such as behavior, learning and memory, and more recently our molecular and physiological understanding of aging and tolerance to low oxygen environments. During the past four years, the Aquatic Germplasm and Genetic Resources Center (AGGRC, aggrc.com) in collaboration with the National Aplysia Resource (NAR, University of Miami), has led the development of technology and procedures required for genetic banking of these animals. Student workers Alana Fregoso Castellanos (B.S. ’27 in Coastal and Environmental Science; left) and Dylan Linsten (B.S. ’27 Mechanical Engineering; right) have combined biology and engineering to explore a specific type of cryopreservation called vitrification. For Aplysia, the spaghetti-like egg strands are processed and cooled rapidly with liquid nitrogen to -196˚C, transforming the liquid into a glass-like state without ice crystal formation.
Two technologies have been explored so far. The first was a resin 3-D printed “baton” (bottom left) created in computer aided design software (Autodesk Fusion). Each baton has compartments to hold 0.25-centimeter-long segments of egg strand. The baton can be stored inside of standard freezing containers (French straws) that can integrate with industrial-scale processing. This can also lead to a novel open-capability kit. The second technology used acupuncture needles (bottom right) to skewer 1-centimeter strand pieces through their long axis. This method was used to reduce the mass of material around the egg strand (such as with the baton), and to increase the conductivity and effectiveness of the cooling process.
Vitrification batons and acupuncture needles used for vitrification.
So far, these methods have yielded results of around 50% survival after thawing, with survival for as long as 2 weeks. Further experimentation is needed to reach the goals of development and survival to adulthood after being vitrified and thawed, as well as being replicable. This year Dylan and Alana are exploring different chemicals (cryoprotectants) to prepare the embryonic Aplysia for freezing and new open-hardware devices to promote vitrification. These efforts will involve the continued integration of biology and engineering. Interaction of these disciplines has had an immense impact on the educational journeys for Alana and Dylan and has been integral to successful development of hardware and processes for preserving the future of biomedical genetic resources. This work is funded in part by the National Institutes of Health Office of Research Infrastructure Programs (NIH-ORIP).