The Journal of Neuroscience, December 14, 2005, ():
Presynaptic Homeostatic Plasticity Rescues Long-Term Depression after Chronic 
9-Tetrahydrocannabinol Exposure
J. Neurosci. Mato et al.
25: 11619
Supplemental data
Files in this Data Supplement:
- supplemental material
-
Supplementary Figure 1. Similar input-output curves were obtained when stimulating PFCx afferences and recording the excitatory postsynaptic field potential (fEPSP) evoked in accumbens MSNs from vehicle- (n = 5) and THC-treated (n = 9) mice.
- supplemental material
-
Supplementary Figure 2. Repeated in-vivo treatment with THC does not alter the protein expression levels of mGluR2/3 in the accumbens. Representative immunoblots are shown on the right. NAc punches were homogenized with a handheld tissue grinder in lysis buffer (25 mM Hepes, 150 mM NaCl, 1% Triton and protease inhibitor cocktail, Roche France), and subjected to centrifugation (10000 g, 4°C, 10 min) to remove insoluble material. Protein quantification was performed using the BCA protein assay kit (Pierce, France). Samples (15 µg) were subjected to SDS-PAGE (7.5%) using a minigel apparatus (Amersham, Life Sciences France) and transferred via semidry apparatus (Bio-Rad, France). mGluR2/3 were labeled using a rabbit anti-rat antibody purchased from Upstate Biotechnology (Lake Placid, NY, USA) reactive to a peptide sequence targeted on the C terminus at a dilution of 1:1000. Labeled proteins were detected using an HRP-conjugated anti-rabbit secondary IgG diluted 1:3000 and visualized with enhanced chemoluminescence (Amersham Life Sciences, France). Immunoreactivity levels were quantified by integrating band density X area using computer-assisted densitometry (Image J 1.29, NIH). Tubulin (anti-β-Tubulin clone tub2.1, 1:10000 Sigma, France) expression was used to normalize protein quantity. The density X area measurements were averaged over three control samples for each gel, and all bands were normalized as percentage of corresponding control values.
- supplemental material
-
Supplementary Figure 3. Presynaptic mGluR2/3 are not activated by endogenous glutamate under low frequency stimulation conditions. Bath application of the selective mGluR2/3 antagonist LY341495 (200 nM, n = 13) did not increase baseline fEPSP responses in the NAc when evoking synaptic responses at 0.05 Hz.
- supplemental material
-
Supplementary Figure 4. Presynaptic A1 receptors are not involved in activity-dependent LTD in the NAc in sham- or THC-treated mice. The selective A1 antagonist DPCPX (200 nM, n = 7) had no effect on 13 Hz-induced LTD in NAc slices from sham (n = 7) or THC-treated mice (n = 8, p > 0.05).
- supplemental material
-
Supplementary Figure 5. Localization of CB1R and mGluR2 in layers V and VI of the mouse prefrontal cortex. Confocal immunofluorescence. In layer V, many supposedly projecting neurons to the nucleus accumbens co-localize CB1R (left panel) and mGluR2 (middle panel) in their perikaryal membranes (asterisks in the merged right panel). Notice the presence of crossing fibres immunoreactive for both receptors (arrows in the merged right panel). In layer VI, observe the presence of CB1R (left panel) and mGluR2 (middle panel) immunoreactivities in the same neuronal cell bodies (asterisks in the merged right panel). Also, a mesh of axon-like fibres in the neuropil co-localizes both receptors. Scale bars: 10 µm.
