This document summarizes research on the role of NMDA receptors in stroke. It discusses how decreased blood flow during a stroke leads to neuronal injury and death through glutamate release and calcium influx via NMDA receptor activation (excitotoxicity). While NMDA receptors are heavily studied, evidence for their role in vivo is still lacking. The document also explores alternative mechanisms of calcium-mediated neuronal injury during ischemia, such as acid-sensing ion channels, and outlines areas for further research.

1. Stroke and NMDA Receptors
Laboratory of Timothy H. Murphy, Ph.D.
University of British ColumbiaKellen Betts

2. “Global” Stroke
1. Decreased blood supply
2. Cellular response
3. Regional effect

3. Ischemia
Activation NMDA Receptors
Neuronal Injury/Death
Glutamate Release
Ca2+ Influx/Overload
• Heavily studied mechanism for
ischemic induced neuronal injury.
• Many conflicting results in in
vitro, in vivo, and clinical trials
(Hoyte et al., 2004).
• Still no direct evidence in in
vivo models.
NMDAR
Excitotoxicity
Hoyte et al. (Curr Mol Biol, 2004); Buchan et al. (J Neursci, 1991)

4. Global Ischemia
Disruption of Dendritic Structure
Spike in Ca2+
Depolarization of Cortex
NMDARs??
MK-801

5. occlusion
Cardiac
Arrest &
Recitation
Surgical
Procedures
“Pump Head”
Transient
Ischemic
Attacks
Internet Stroke Center (Wash. Univ.)

6. Occlusion
GFP or YFP
C57/Bl6
Mice
Yang et al. (Brain Res, 1997); Internet Stroke Center (Wash. Univ.)

7. XX
Medial Cerebral ArteryAnterior Cerebral Artery
Somatosensory cortex

8. X
X
X
X
X

9. occlusion
occlusion “AC Ripple”
DC depolarization

11. 20 μm
Control
CCA occlusion
10 min reperfusion
-1.2 mm/s
-2.4 mm/s
0 mm/s
0 mm/s
-0.9 mm/s
2.4 mm/s
B
Con 0 10 10-30 30-60 >60
Murphy & Li
2-Photon Imaging InVivo

12. 2-Photon Imaging InVivo
(video file)

13. pre-stroke 4 min ischemia
10 min reperfusion 90 min reperfusion

14. occlusion
“AC Ripple”
occlusion
DC depolarization

15. occlusion
spike in Ca2+
occlusion
spike in Ca2+
reperfusion

16. 20μm
pre-stroke 4 min ischemia 20 min reperfusion

17. Intrinsic Optical Signal (IOS)
(video file)

18. IOS Stimulation Map
Pre-stroke 2h post-stroke
Murphy & Li

19. NMDA
MK-801 Positive
Control in vivo

20. Hasbani et al. (J Neurosci, 2001)
control NMDA NMDA

21. Global Ischemia
Disruption of Dendritic Structure
Spike in Ca2+
Depolarization of Cortex
NMDARs??
MK-801

22. ‣ Acid-Sensing Ion Channels
Xiong et al. (Neuron, 2004) -- “Neuroprotection in ischemia:
blocking calcium-permeable acid-sensing ion channels.”
‣ TRP Channels
Aarts et al. (Cell, 2003) -- “A key role for TRPM7 channels in
anoxic neuronal death.”
‣ Gap Junction Hemichannels
Thompson et al. (Science, 2006) -- “Ischemia opens neuronal gap
junction hemichannels.”
‣ Calcium-permeable AMPA receptors
Kwak et al. (Curr Opin Neurobiol, 2006) -- “Calcium-permeable
AMPA channels in... ischemia.”
Further Research

23. Acid-Sensing Ion Channels
• 4 genes (ASIC1-4) within
Deg/ENaC superfamily
• 4 splice variants (1a/b &
2a/b)... total of 6 subunits
• Homo- & heteromultimers
form ion channels
• Activated by [H+
]e
(e.g. ASIC1a, pH 0.5 = 6.2)
• Permeable to Na+
& Ca2+
(1a
only); pNa+
/pCa2+
= 2.5
• Expressed in CNS (1a & 2a)
Waldmann et al. (Nature, 1997); Zha et al. (PNAS, 2006)

24. ASIC1a is in dendrites
ASIC1a is not in axons
ASIC1a is in spines
Zha et al (PNAS, 2006)
Localization

25. Psalmopoeus cambridgei

26. ASIC Antagonists
Amelioride
•Non-specific... blocks all ASICs and ENaCs
•K+
-sparing diuretic
•Easy to get
PcTx1
• Specific for ASIC1a
• Protein isolated from venom of Psalmopoeus cambridgei
• Venom is a specific ASIC1A antagonist (Xiong et al., Neuron, 2004)
Waldmann et al. (Nature, 1997); Escoubas et al. (Protein Sci, 2003)

27. Laboratory of Tim Murphy, Ph.D.
•Craig Brown, Ph.D.
•Heidi Erb
•Alexander Goroshkov
•Ping Li
•Richard Liu
•Zhi Liu
•Albrect Sigler, Ph.D.
•PuMin Wang
•Charles Wong
•Ian Winship, Ph.D.
Laboratory of Lynn Raymond, PhD.
Thank You!