Semester of Graduation

Fall 2023

Degree

Master of Renewable Natural Resources (SRNR)

Department

Renewable Natural Resources

Document Type

Thesis

Abstract

The California sea hare (Aplysia californica) is an important biomedical model for molecular neurobiology, electrophysiology, learning, and memory due to their well-mapped and large neurons and well-characterized learning capabilities. The National Resource for Aplysia (NRA, University of Miami) maintains large stocks of live animals and relies on regular shipments of wild-caught individuals to maintain genetic diversity. This is labor and cost-intensive, and environmental changes could alter the availability of wild animals increasing the need to preserve this genetic resource. One solution is vitrification, ultra-fast cooling which produces an amorphous glass that minimizes damage to cells. Aplysia californica presents challenges in that sperm cannot be extracted practically and their early life stages are encased within a protective egg strand. Thus, the goal of this thesis was to lay the groundwork for A. californica vitrification and demonstrate the benefits of open technology. The objectives were to: 1) compare early life stage development within and outside of the egg strand; 2) develop larval behavior assays to assess the toxicity of five vitrification solutions and determine a suitable equilibration time; 3) evaluate larval recovery after removal of vitrification solutions; 4) assess vitrification and post-thaw larval survival, and 5) design open-fabricated devices to assist in outreach and scientific communication of A. californica. Embryos did not develop into veligers when released from the egg strand prior to embryonic stage four. As such, larvae were vitrified while still within the egg strand. Post-thaw behavior showed that 50% dimethyl sulfoxide (DMSO) had the best results. Three-dimensional (3-D) models demonstrated the effectiveness of tactile objects to engage people of all ages, especially children. Future research should address raising thawed veligers to maturity and observing if they can reproduce. In addition, future research should address embryonic cryopreservation within the egg strand. Vitrification of A. californica embryos could provide cryopreservation options to preserve other marine invertebrate species with larvae that develop within complex structures, such as marine snails.

Date

11-17-2023

Committee Chair

Tiersch, Terrence R.

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