1. Before we start: What is the question? Why is it interesting?
What is the role/contribution of the different subtypes of NMDA glutamate receptor to plasticity in the brain?
Why is it interesting?
NMDA receptors play an important role in many forms of synaptic plasticity.
NMDA receptors are important for cell survival and cell death (ischemia, excess glutamate)
NMDA receptor composition changes during development and it has been proposed to be one of the main regulators of the critical period.
The developmental switch between NR2A and NR2B can be modulated by experience.

2. Glutamate Receptors: NMDA-sensitive
Tetramers with the following subunits:
NR1 (always 2)
8 splice variants.
Glycine binding site.
The different splice variants differ in their conductances, their affinity for glycine, etc.
NR2 (always 2)
Glutamate binding site.
Need NR1 to form functional receptors
4 genes. (NR2A, NR2B. NR2C, NR2D)
Only NR2A and NR2B are expressed in the forebrain.
NR2A and NR2B are developmentally regulated, with NR2B starting to decrease after a few weeks.
Very little is known about the proteins that each subunit interacts with.

3. NMDAR Receptors: Adult distribution of NR2
Nakanishi, S., 1992., Science

4. Functional Properties of NR1 and NR2
Affinity for glutamate
Inhibition by Protons
Modulation by Glycine
Potentiation by polyamines
Ca2+ current
Inhibition by Zn+2
Channel Kinetics
Some kinetics properties (very unknown)
Dendritic transport, synaptic localization, trafficking of receptors
Required for release from the ER; might be important for targeting between soma and dendrites
NR2B interacts with the PDZ domain of SAP-102 and of Sec8 (an exocyst protein). NR2B can also interact with RasGRF1, actuvating the ERK pathway. Calcium inlfuc and ERK pathway activation can lead to changes in cell proliferation and survival.
PSD-95 binds to NR2A increasing its expression at the synapse, while at the same time depressing synaptic expression of NR2B. PDS-95 KO mice have enhanced LTP.
NR2-B truncated mice do not survive.
Cterminal truncationof NR2A survives, impairs synaptic but not extrasynaptic localization of NMDA receptors and reduces LTP at the CA3-CA1 synapse.
Cull-candy and leszkiewicz, Science STKE, 2004

5. Deactivation Kinetics of NR2B subunits
> Mg2+ sensitive
< Mg2+ sensitive
Cull-candy and leszkiewicz, Science STKE, 2004

6. Plasticity in the Hippocampus
Studying Long Term Potentiation (LTP) and Long Term Depression (LTD)
Baseline
Stim. Schaffer Collateral at 0.2Hz
CA1
Induction
Stim. Schaffer Collateral at 100Hz, 1sec
CA1
100Hz
Time (min)
% EPSP
100 %
200 %
Post- Induction
Stim. Schaffer Collateral at 0.2Hz
CA1

7. LTP and LTD in CA1
HFS of the Schaffer collateral fibers leads to LTP in CA1.
LFS of the Schaffer collateral fibers leads to LTD in CA1.
These changes in synaptic strength depend on:
Intracellular calcium.
NMDA receptors.
Simple Model:
Presynaptic
Postsynaptic
Ca2+
HFS
LFS
NMDA receptor
Modifed from Lisman, 1989; Bear and Malenka, 1994.

8. Support for this model Intracellular calcium levels “LTD” “LTP”
Hippocampal slices, p 11-p22, WC Recording, Photolysis of EGTA
Intracellular calcium levels
“LTD”
“LTP”
Yang S-N. et al., J. Neurophysiol.

9. Support for this model
Degree of NMDA receptor activation  Amount of calcium
Cummings et al., Neuron
Nishiyama et al., Nature

10. NR2A KO
NR2B KO

11. “Genetic enhancement of learning and memory in mice”
BUT… NR2B over-expression:
“Genetic enhancement of learning and memory in mice”
Tang et al., Nature 1999.
“NR2B expression is downregulated during the period of transition between juvenile and adult, correlating with the gradual shortening of the EPSP duration through the NMDA channel. This could decrease NMDA-mediated plasticity, and perhaps explain decreased memory performance in adult animals…”

12. Enhanced Novel Object Recognition
“Smart Mice”: Over-expression of NR2B
Enhanced Novel Object Recognition
Tang et al., Nature.

13. Selective block of NR2A
% block
current 2B 2A
In oocytes
In hippocampal slices

14. NR2A X
LTD
LTP
HFS
LFS

15. NR2B X
LTD (1XLFS)
LTP (1XHFS)
HFS
LFS

16. Conclusions:
[Ca++]
0.1uM Ca++ (resting) too low for both calcineurin & CamIIK – stable release
~1uM Ca++ (LFS=1-5 Hz) – calcineurin: LTD
>5uM Ca++ (HFS=>Hz) – calcineurin + CamIIK… CamIIK out-competes: LTP
Molecular ID of NR2
NR2A necessary for LTP, not LTD.
NR2B necessary for LTD, not LTP.
Reduction of NR2B during development could explain increased difficulty in inducing LTD.

17. who or where???
“Spillover” during HFS

18. Time, space, co-incidence detection
Induction:
association
cooperativity
specificity
Time, space, co-incidence detection
NMDAR coincidence detection: glutamate & depolarization
Glutamate release- local
AMPAR depolarization- small ampl., passive propagation: small spread
Back-propagating AP- big ampl., active propagation: wide-spread
Time
calcineurin – high affinity, on/off
CamIIK- medium affinity, persistent on
PKC- medium affinity, on/off
Space
Ca++ via NMDAR- local glutamate, wide-spread voltage unblock
Ca++ via IP3- local glutamate (mGluR)
Ca++ via VG-CC- wide-spread Ca++ AP
Spill-over (2A/2B in synapse; 2B extra-synaptic)
NO- trans-cellular, on/off, intermed. distance- retrograde transmitter!!

19. NOS
Na+ channels AP

20. Specificity - synaptic tagging