The Effects of BDNF on Nitric Oxide Synthase I Using Human SH-SY5Y Cells

1. The Effects of BDNF on Nitric Oxide Synthase I Using Human SH-SY5Y Cells By Amir Abyaneh, and Dr. Terrie Rife

2. Nitric Oxide Synthase I • Nitric Oxide Synthase I (NOS I) is an enzyme that catalyzes the reaction of a free radical, NO•. • Learning and Memory • Neurodegenerative Diseases/Problems • Why do we study it?

3. Nitric Oxide Synthase I

4. Human NOS I Structure 5’2 exon 2 exon 3- 29 5’4 5’3 5’1 { >20 kB } Transcription Translation Note - Figure is not drawn to scale

5. What We Know! Nerve Growth Factor NEURON p75 Trk A NOS 1 ↑

6. Brain Derived Neurotrophic Factor

7. p75 RA Trk B RA RA SH-SY5Y Cells RA RA Cell Lines • human neuroblastoma cells SH-SY5Y • A model for neuronal differentiation • Responds to BDNF after being treated with Retinoic Acid 5 days

8. Untreated Establishing a Protocol RA w/ 10%FBS for 5 Days BDNF for No Serum for 6 days

9. Methods • Pretreatment with RA and 10% FBS for 5 Days • Treated with BDNF for 2, 4, 6, and 8 days, with a No Treatment control • RNA Harvesting • Reverse Transcription • PCR Analysis

10. Primers – PCR analysis RA 2 4 8 No DNA NT GADPH 380bp nNOS 433bp

11. 5’2 exon 2 exon 3- 29 5’4 5’3 5’1 { >20 kB } Transcription Translation FUTURE • PCR analysis of transcripts: • 5’1 • 5’2 • 5’3 • 5’4 Note - Figure is not drawn to scale

12. Thanks