Abstract
Endocannabinoids are important players in neural development and function. They act via receptors, whose activation inhibits cAMP production. The aim of the paper was to look for calcium- and cAMP-signaling cross-talk mediated by cannabinoid CB1 receptors (CB1R) and to assess the relevance of EF-hand CaM-like calcium sensors in this regard. Using a heterologous expression system, we demonstrated that CB1R interacts with calneuron-1 and NCS1 but not with caldendrin. Furthermore, interaction motives were identified in both calcium binding proteins and the receptor, and we showed that the first two sensors competed for binding to the receptor in a Ca2+-dependent manner. Assays in neuronal primary cultures showed that, CB1R-NCS1 complexes predominate at basal Ca2+ levels, whereas in the presence of ionomycin, a calcium ionophore, CB1R-calneuron-1 complexes were more abundant. Signaling assays following forskolin-induced intracellular cAMP levels showed in mouse striatal neurons that binding of CB1R to NCS1 is required for CB1R-mediated signaling, while the binding of CB1R to calneuron-1 completely blocked Gi-mediated signaling in response to a selective receptor agonist, arachidonyl-2-chloroethylamide. Calcium levels and interaction with calcium sensors may even lead to apparent Gs coupling after CB1R agonist challenge.
Highlights
The mechanisms involved in the cross-talk at the level of the two main second messengers, Ca2+ and cAMP, are not fully elucidated despite being seminal for neuronal function
Calcium-binding protein expression was detected by fluorescence while CB1 receptors (CB1R) fused to RLuc was detected by the use of an anti-RLuc antibody and a Cy3-conjugated secondary antibody
The potential relationships between calcium-binding proteins and G-protein-coupled receptors (GPCRs) are poorly understood despite pioneering articles showing that calmodulin is involved in hormone action (Means and Dedman, 1980), regulates GPCR kinases (Iacovelli et al, 1999) and impacts on the activity of enzymes that regulate cAMP levels after activation of dopamine and opioid receptors (Hanbauer et al, 1979)
Summary
The mechanisms involved in the cross-talk at the level of the two main second messengers, Ca2+ and cAMP, are not fully elucidated despite being seminal for neuronal function. Intracellular calcium ion is a second messenger that interacts and affects the structure of a variety of proteins. The more studied has been calmodulin (CaM), other calcium binding proteins have been identified and partially characterized (Mikhaylova et al, 2011). Upon ion binding, these calcium-binding proteins are able to participate in a myriad of relevant cellular events, inter alia proliferation, differentiation and apoptosis (Hyman and Pfenninger, 1985; Stichel et al, 1987; Forscher, 1989; Joseph et al, 1993; Orrenius and Nicotera, 1994; Orrenius et al, 2003).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
