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SCREENING DRUGS FOR SPINAL CORD REGENERATION USING AN IMPROVED LARVAL ZEBRAFISH MODEL SYSTEM

thesis
posted on 2023-03-10, 22:34 authored by Gentry Lee AndrewsGentry Lee Andrews

In an endeavor to identify novel compounds that may aid in spinal cord regeneration following traumatic injury, compounds in an FDA-approved small molecule drug library were screened for their effects on regeneration using a regeneration-permissive vertebrate model system, the danio rerio.

A novel injury paradigm for studying upper spinal cord injuries in larval danio rerio was developed for this purpose. This injury paradigm was paired with a behavioral assay designed to test the functionality of the spinal cord under drug treated conditions compared to controls and applied to the drug screen, following an initial toxicity screen of the drug library. Each compound was tested at a 10 μM concentration. Drugs were introduced at 1 hours post injury (hpi) for 48 hours (i.e., the study’s Primary Screen). After the 48-hour drug treatment, the larvae underwent the behavioral assay (e.g., Visual Motor Response Assay using a Viewpoint Zebrabox System). Drugs that met the assay’s threshold for potentially regenerative effects, based on the established criteria, were retested at the same concentration as part of a Secondary Screen. If needed, additional concentrations were tested and once a drug was confirmed as part of the Secondary Screen, imaging studies performed to assess for axonal bridging.

As of Fall 2022, the over 650 drugs had been screened as part of the Primary Screen, representing roughly one quarter of the total drug library. Screen hits through this point were classified by mechanism of action. Trends from these classifications have indicated, a large portion of drug hits have been vasoactive drugs (e.g., DAR and 5-HT agonists; cholingeric receptor antagonists), anti-inflammatory drugs (e.g., COX-2 inhibitors), antimicrobials, calcium/sodium/potassium channel blockers, transcription-related drugs (i.e., DNA and Viral), and a couple microtubule-stabilizing drugs. Histone Deacetylase (HDAC) Inhibitors were identified as repeat hits early in the Primary Screen, and to follow up on this trend, a study specific to HDAC Inhibitors was designed and executed.

An additional study designed to assess the influence nutrition has on larval locomotor and regenerative capabilities after injury was run with the intention of modeling and demonstrating how, even in regeneration permissive species, access to proper nutrition significantly influences health outcomes.

Funding

Indiana Spinal Cord and Brain Injury Research Program

Indiana CTSI Pilot Funding for Use of Core Facilities

History

Degree Type

  • Master of Science

Department

  • Biological Sciences

Campus location

  • West Lafayette

Advisor/Supervisor/Committee Chair

Daniel Suter

Additional Committee Member 2

Yuk Fai Leung

Additional Committee Member 3

Alan Friedman