The Journal of Neuroscience, April 26, 2006, ():
Dopamine D1 Activation Potentiates Striatal NMDA Receptors by Tyrosine Phosphorylation-Dependent Subunit Trafficking
J. Neurosci. Hallett et al.
26: 4690
Supplemental data
Files in this Data Supplement:
- supplemental material
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Supplementary Figure 1. Expression of the dopamine D1 receptor in cultured striatal neurons.
(A) Immunoreactivity of the dopamine D1 receptor in striatal neurons. Total homogenates (10 μg protein) from rat striatum (lane 1) and striatal neurons at DIV21 (lane 2) were resolved on SDS-polyacrylamide gel electrophoresis and probed with antibody against the dopamine D1 receptor.
(B) Distribution of the dopamine D1 receptor in striatal cells. Striatal neurons at DIV21 were stained for the dopamine D1 receptor. Higher magnification of individual dendritic segment is shown below the panel.
(C) Striatal neurons in culture express moderate number of dendritic spines. Striatal neurons were transfected at DIV13 with GFP and doubly stained at DIV19 for GFP and DARPP-32. Shown is the spine morphology of a striatal neuron that stained positive for the DARPP-32 protein.
(D) SKF-82958 induced increase in NR2B staining intensity in dopamine D1-expressing cells. Striatal neurons at DIV21 were treated with SKF-82958 and doubled-labeled for the dopamine D1 receptor and NR2B protein. The staining intensity for NR2B subunit was quantified in D1-expressing cells.
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Supplementary Figure 2. Developmental expression of NMDA receptors in striatal neurons.
Double-label immunofluorescence staining of striatal neurons for NR1 (a-c), NR2A (d-f) or NR2B (g-i) (green) and PSD-95 (red) at DIV7, DIV14 and DIV21 as indicated. Higher magnification panels show individual dendrite segments, in gray-scale for NMDA receptor subunits and in color for merged green and red channels. Scale bars: top panels, 80 μm; higher magnification panels, 8 μm.
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Supplementary Figure 3. Potential model for modulation of NMDA receptor trafficking at striatal dendritic spine by tyrosine phosphorylation.
Preassembled NMDA receptors are delivered by components of the exocyst (Sec 6/8) (Sans et al., 2003) and KIF17 (Setou et al., 2000). Tyrosine 1472 of NR2B is phosphorylated by Fyn kinase (Suzuki and Okumura-Noji, 1995), which is found in association with the assembled receptors and PSD-95. Receptor complexes containing tyrosine-phosphorylated NR2B are rapidly trafficked to synaptic sites. Dephosphorylation is catalyzed by tonically active tyrosine phosphatases such as STEP (Boulanger et al., 1995). Dephosphorylation of tyrosine-phosphorylated NR2B exposes a site for interaction of the clathrin adapter protein AP-2 and stimulates clathrin-dependent internalization of NMDA receptors (Prybylowski et al., 2005). A key regulator of STEP is PKA (Paul et al., 2000), D1 receptor activation, and subsequent stimulation of adenyl kinase, may result in PKA activation and STEP inhibition, thereby leading to enhanced accumulation of NR2B containing receptors at synaptic sites.
