Abstract
The dorsal striatum is a key node for many neurobiological processes such as motor activity, cognitive functions, and affective processes. The proper functioning of striatal neurons relies critically on metabotropic receptors. Specifically, the main adenosine and endocannabinoid receptors present in the striatum, ie, adenosine A2A receptor (A2AR) and cannabinoid CB1 receptor (CB1R), are of pivotal importance in the control of neuronal excitability. Facilitatory and inhibitory functional interactions between striatal A2AR and CB1R have been reported, and evidence supports that this cross-talk may rely, at least in part, on the formation of A2AR-CB1R heteromeric complexes. However, the specific location and properties of these heteromers have remained largely unknown. Here, by using techniques that allowed a precise visualization of the heteromers in situ in combination with sophisticated genetically modified animal models, together with biochemical and pharmacological approaches, we provide a high-resolution expression map and a detailed functional characterization of A2AR-CB1R heteromers in the dorsal striatum. Specifically, our data unveil that the A2AR-CB1R heteromer (i) is essentially absent from corticostriatal projections and striatonigral neurons, and, instead, is largely present in striatopallidal neurons, (ii) displays a striking G protein-coupled signaling profile, where co-stimulation of both receptors leads to strongly reduced downstream signaling, and (iii) undergoes an unprecedented dysfunction in Huntington’s disease, an archetypal disease that affects striatal neurons. Altogether, our findings may open a new conceptual framework to understand the role of coordinated adenosine-endocannabinoid signaling in the indirect striatal pathway, which may be relevant in motor function and neurodegenerative diseases.
Highlights
The dorsal striatum is a key node for many neurobiological processes such as motor activity, cognitive functions, and affective processes
adenosine subtype 2A receptor (A2AR) and cannabinoid type 1 receptor (CB1R)-evoked signaling was essentially insensitive to pertussis toxin (PTX) or cholera toxin (CTX) (Figure 3b), indicating that these receptors do not significantly couple to Gi or Gs proteins in these cells. This notion was further supported by the observation that, in both STHdhQ7 cells (Supplementary Figure S4a) and STHdhQ111 cells (Supplementary Figure S4b), assays, we found that pretreatment of brain slices with TM5, neither the A2AR agonist nor the CB1R agonist was able to TM6 or both peptides disrupted (i) the ability affect basal or forskolin-elevated cAMP concentrations in the of the CB1R agonist WIN-55,212-2 and the CB1R antagonist SR141716 to dampen A2AR-evoked actions on extracellular signalregulated kinase (ERK) and Akt, as well as (ii) the ability of the A2AR agonist CGS21680 and the A2AR antagonist ZM241385 to dampen CB1R-evoked actions on these two signaling pathways (Figure 2c)
By using (i) mice lacking CB1R selectively in GABAergic neurons, (ii) CB1R-deficient mice in which CB1R expression is selectively rescued in GABAergic neurons, (iii) mice lacking CB1R selectively in D1 receptor (D1R)-expressing MSNs, and (iv) CB1R-floxed mice in which CB1R is selectively excised in MSNs, we cogently demonstrated that the A2AR-CB1R heteromer is present in indirect-pathway MSNs (Figure 1 and Supplementary Figure S2)
Summary
The dorsal striatum is a key node for many neurobiological processes such as motor activity, cognitive functions, and affective processes. CB1R is highly expressed in the terminals of both striatonigral and Previous studies on the A2AR-CB1R heteromer have relied essentially on energy transfer-based assays in cells ectopically expressing A2AR and CB1R, as well as co-immunolocalization and co-immunoprecipitation experiments (Carriba et al, 2007; Navarro et al, 2008; Bonaventura et al, 2014). These approaches, widely exploited and certainly striatopallidal MSNs, where it mediates endocannabinoid- valuable, possess limitations of spatial resolution (co-immudependent inhibition of GABA release, decreasing nolocalization), molecular specificity (co-immunoprecipitamotor activity (Katona and Freund, 2008; Castillo et al, tion), and biological interpretation
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