4. Neurite Growth Research Project-JanPlan 2009
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N,N-Diethyl-meta-toulamide. The active chemical in DEET: 

Flavonoids and Terpenoids are the antioxidant elements in Ginkgo Biloba Extract:

 For many years the Tilden research lab has  focused on crustacean nervous systems, specifically the neural tissue of the fiddler crab, Uca Pugilator. Most of the resarch has focused on the effects of that melatonin has on neurite growth in neurons cultured from the x-Organ/Sinus gland complex of the fiddler crab.  This study was designed with the question of how we could prolong and/or promote the neurite growth in mind. In researching common dietary and herbal supplements used in Alzheimer's patients, we came across the antioxidant Ginkgo Biloba Extract. Ginkgo Biloba Extract has been used in Chinese medicine for centuries for nearly every type of ailment, and has relatively recently made it to the United States as a treatment/preventative measure for Alzheimer's and other neurodegenerative conditions. The flavonoid compounds found in the leaves of Ginkgo Biloba are antioxidants, which bind to free radicals in the cells. By binding to the free radicals, the flavonoids prevent the free radicals from "stealing" electrons from the growing neurite membranes. We hypothesized that this antioxidant property would result in enhanced neurite growth. Furthermore, Ginkgo Biloba extract contin terpenoid compounds, which are less studied, but are thought to enhance neural connections.

The purpose of our project was to determine the impact of oxidative stress and the effects of exogenous antioxidants on neurite growth in neurons extracted from the X-organ/sinus gland of the fiddler crab Uca pugilator.

After removing the eyestalks from anesthetized crabs, we microdissected the eyestalks and removed neural tissue from the X-organ, located directly opposite the opalescent-blue sinus gland. We enzymatically digested the tissue fragments using trypsin, separated the tissue to break loose individual neurons, and cultured the cells for a period of forty-eight hours.

Neural cells grown in a normal culture medium were used as controls. Our experimental cells were cultured in a medium treated with either 100% DEET insect repellent (at a 50% lethal dose), or with 24% standardized Ginkgo Biloba extract (10X the human dose).

After forty-eight hours, the cell cultures were examined under a light microscope at 640X magnification and photographed using AxioVision software. Neurite areas and cell body areas were calculated using Image J, and a ratio of neurite to cell body was calculated.

Deet technical fact sheet from the National Pesticide Information Center

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