Nitric Oxide Synthase in Mouse Brain Tissue that Exhibits Alzheimers Disease

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Nitric Oxide Synthasein Mouse Brain Tissue that
Exhibits Alzheimers Disease
Patrick McCarthy 2003-04
http//www.nsrl.ttu.edu/tmot1/mus_musc.htm
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Introduction

• Alzheimers disease (AD)
• Future impact
• Characteristics

Beta-amyloid plaque
http//www.ahaf.org/alzdis/about/AmyloidPlaques.ht
m
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Introduction

• Transgenic mouse model of Alzheimers disease
  using species, Mus musculus
• Two genetically engineered mouse types
• - Transgenic positive (Tg. )
  with AD-expressing gene
• - Transgenic negative (Tg. -)
• without AD- expressing gene

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Introduction

• Nitric oxide synthase (NOS)
• - Enzyme throughout body
• 1989, NOS in brains
• - Bredt et al.
• Early 2000s, NOS and AD relation strongly showed
  
• - de La Torre et al., Law et al.

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NOS-catalyzed production of NO via L-arginine to
L-citrulline pathway
NOS
L-arginine
L-citrulline
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Introduction

• Three NOS isoforms
• - neuronal NOS (nNOS)
• - endothelial NOS (eNOS)
• - inducible NOS (iNOS)

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Hypothesis (1)

• Luth et al. found iNOS and eNOS activity
  increased in AD
• Luth et. al and Quinn et. al found nNOS activity
  increased in AD
• First hypothesis total NOS activity increased in
  AD brains

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Hypothesis (2)

• Norris et al. found number of nNOS-containing
  neurons decreased
• Fewer neurons, less total concentration of nNOS
• Second hypothesis nNOS concentrations in AD
  brains were lower

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Procedure

• Tissue preparation
• Determining activity
• - Spectrophotometer to assay

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Results

• NOS activity significantly different (p 0.014)
• Tg. NOS activity 58 greater than Tg. -

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Procedure

• Concentrations
• - Slot Blot
• - Western Blot

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Results
Tg. Tg. -

• Intensity of band corresponds to concentrations
• Darker band, higher concentration
• This test inconclusive

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Procedure

• Concentrations
• -ELISA assay

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Results

• nNOS concentrations significantly different
  (p 0.008)
• Tg. 36 less nNOS concentration than Tg. -

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Conclusion

• (1) Total NOS was increased in Tg. brains
• (2) Concentrations of nNOS lower in Tg.
  brains

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Conclusion

• Suggest neurons initially damaged or further
  degenerated by the toxicity of the free radical
  NO
• Increase in NOS activity can most likely be
  attributed to increase in nNOS and/or eNOS
• Support the role of NOS and nNOS in AD

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Potential Future Studies

• Find concentrations of other two isoforms of NOS,
  iNOS and eNOS
• Use different-aged brains to study the role of
  NOS activity and concentration before, during,
  and after the onset of AD
• Further test certain areas of the brain (human
  and mice) to investigate role in AD

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Acknowledgements

• Dr. Ian Armitage and Dr. Bruce Martin
• Abigail Tolkheim and rest of the Armitage Lab
  team
• Ms. Lois Fruen
• Team Research

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The Role of Nitric Oxide Synthase in Alzheimers
Disease

• Patrick McCarthy 2003-04