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Figure 8 | Molecular Brain

Figure 8

From: HINT1 protein cooperates with cannabinoid 1 receptor to negatively regulate glutamate NMDA receptor activity

Figure 8

MOR and CNR1 associate with the NR1 subunits via HINT1: Implications for NMDAR activity. (A) The association of GPCRs, MOR and CNR1 with the NMDAR NR1 subunits is dependent on HINT1. The GPCRs were immunoprecipitated from solubilized mouse brain cortical synaptosomes from WT and HINT1-/- mice, and the proteins that co-precipitated with the NR1 subunits were analyzed by western blot. (B) The association of CNR1 with NR1 in WT and HINT1-/- cortical cell cultures uninfected or infected with 10 μL/well of HINT1-/- lentiviral particles. The presence/absence of HINT1 was determined, and the co-precipitates of the NR1 subunits and CNR1 were then assessed by immunoprecipitation and western blot. For each determination, cells from 10 wells were pooled, and the assay was repeated twice with identical results. (C) Opioid agonists promoted a higher level of responsiveness of the NMDAR by inhibiting the association between MOR and NR1. Morphine or DAMGO was icv-injected into the mice 24 h prior to the analysis of MOR/NMDA NR1 association and CaMKII activating autophosphorylation on Thr286. At 7 days post-surgery, the mice from the CCI neuropathic pain group displayed increased pThr286 CaMKII and reduced MOR-NR1 association compared to the sham-operated controls. Changes of Mechanical Withdrawal Threshold. Following ligation, animals developed significant mechanical allodynia by day 3 that remained until day 21. Naïve control and sham – operated mice failed to exhibit mechanical allodynia. Data are mean ± SEM. *P<0.05 vs. sham-operated group, ANOVA, followed by the Student-Newman-Keuls test (SigmaStat, SPSS Science Software, Erkrath, Germany) p < 0.05.

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