1. Evaluation of Vascular Reactivity in Alzheimer’s Disease via Iontophoresis of Vasoactive Compounds Jonathan Maltz, Ph.D. William Jagust, M.D. Jamie Eberling, Ph.D. Thomas Budinger, M.D., Ph.D.

2. Hypothesis Altered dilatory response of brain vasculature is an important factor in the development of AD. This altered response is systemic and affects the peripheral circulation as well as the brain.

3. Hypothesis suggested by 1. Increased incidence of AD in patients with cardiovascular disease risk factors. - risk of demetia 5x in women with myocardial infarction - cerebral hypoperfusion - Apolipoprotein  4 allele predisposes to both atherosclerosis and dementia 2. Cholinergic and nitric oxide (NO) biochemistry is significantly altered in the AD brain. These agents are important mediators of vasodilation throughout the body.

4. Objective To test the hypothesis that peripheral vasoactivity is altered in AD by evaluating the cutaneous vasodilatory response to vasoactive agents.

5. ACh, MCh and SNP ACh: - Relaxes vascular smooth muscle primarily via receptor stimulation of endothelial cells, which then release NO. MCh: - Same mode of action as ACh. - More stable. Seems to act directly on smooth muscle as well as on endothelial cells. - Preferential muscarinic agonist. SNP: - Releases NO on decomposition. - Acts by directly relaxing smooth muscle.

6. Iontophoresis: quantitative drug delivery

7. Perfusion response to iontophoresis

8. Laser Doppler imaging: quantitative measurement of perfusion

9. Laser Doppler perfusion imaging

10. Laser Doppler imaging of perfusion response to iontophoresis

11. Typical response to ACh

12. Review of previous work Two studies have investigated cutaneous vasoactivity in AD: Hörnqvist et al., Gerontology 33(6), 1987 Algotsson et al., Neurobiology of Aging 16(4), 1995

13. Hörnqvist et al. (1987): Subject selection 12 AD/SDAT patients Age: 52-84, mean 71 Severe dementia, hospitalized 13 controls with various dermatoses Age: 52-82, mean 70 Nicotine and caffeine allowed

14. Hörnqvist et al.

15. Hörnqvist et al.: Results AgentActionAD vs Control Phenylephrine  1 -agonist AD slightly reduced Isoproterenol  1 -agonist AD reduced p < 0.001 MethacholineMuscarinic agonist Not significant

16. Algotsson et al. (1995): subject selection 15 AD patients Mini-mental state examination (MMSE) scores > 27 16 Age-matched controls Subjects lived at home

17. Algotsson et al.

18. Algotsson et al.: Results AgentActionAD vs Control Sodium nitroprusside NO donor to smooth muscle AD reduced (not significant) Isoproterenol  1 -agonist AD reduced p < 0.01 AcetylcholineEndothelium dependent vasodilator AD reduced p < 0.05

19. Our study: subject selection 12 AD patients, age 73 - 88, mean 80.4 3 not treated with acetylcholinesterase inhibitors (ACHEI’s) 6 on donepezil (Aricept) 5 mg 3 on donepezil 10 mg MMSE range: 2 - 28, mean 19.4 8 controls, age 71 - 84, mean 78.6 12 hour fast Lipid panel performed Subjects lived at home

20. Iontophoretic dose 110  A for 60 seconds All solutions: 0.5 – 1 mM

21. Results AgentActionAD vs Control MethacholineMuscarinic endothelium dependent vasodilator (EDV) AD increased 78% p < 0.003 AcetylcholineCholinergic EDVAD increased 68% p < 0.03 Sodium nitroprusside NO donor to smooth muscle AD increased 46% p < 0.05

22. Mean response vs. time

23. Time-integrals of perfusion responses

24. Conclusions Perfusion response to MCh, ACh and SNP significantly increased in AD under donepezil therapy. Enhanced response to cholinergic agonists is opposite of what Algotsson et al. observed. It is impossible to evaluate original hypothesis in this patient population. Results suggest MCh perfusion studies may be useful in monitoring acetylcholinesterase inhibitor therapy.

25. Proposed explanation AChEIs delay the metabolic breakdown of cholinergic agonists in cutaneous tissue. This leads to enhanced and prolonged vasodilation. Why is response to endothelium independent vasodilator SNP enhanced?

26. Proposed mechanism behind enhanced endothelium-independent vasodilation SNP relaxes smooth muscle  Flow increases  Endothelium stimulated  ACh released by endothelial cells  Donepezil preserves endogenous ACh for longer by inhibiting AChE  Prolonged and enhanced dilatory response

27. Reinterpreting the result of Algotsson et al. (2000) AD with ApoE4 AD without ApoE4 Controls On AChEI: 4 of 9 On AChEI: 1 of 8 11 (1 with ApoE4) Sodium nitroprusside Perfusion enhanced 30% (p < 0.02) Not significantly different from controls (NS) AcetylcholineNS IsoproterenolNS

28. Future plans Monitoring AChEI-therapy in AD/IVD. Study of recently diagnosed, untreated subjects. Acknowledgements We thank Matthew Darmalingum for assisting in these experiments and in the preparation of this presentation. This study was performed under NIH grant AG 05890-15 and DOE OBER contract DE-AC03-76SF0098.