Special issue: G protein-coupled adenosine receptors: molecular aspects and beyond

Adenosine is a purine nucleoside present in all human cells where it plays many different physiological roles: From being a building block for nucleic acids to a key constituent of the biological energy currency ATP. Indeed, more than 90 years ago, Drury and Szent-Györgyi reported that adenosine produces profound hypotension and bradycardia, and until the present time, the list of physiological effects of adenosine has expanded considerably. In addition, adenosine is a well-known neuromodulator in the brain and has effects on other tissues, thus exerting its physiological actions through four different subtypes of G protein-coupled adenosine receptors (i.e., A1R, A2AR, A2BR and A3R) which, as expected, are expressed in a large variety of cells throughout the body. Consequently, ARs are potential therapeutic targets in a variety of pathophysiological conditions, including cancer, cardiovascular diseases, neurological disorders, and inflammatory and autoimmune diseases

Editorial: Purinergic Pharmacology

The purine nucleotides and nucleosides constitute important extracellular signaling molecules acting as neurotransmitters and neuromodulators. Indeed, extracellular adenosine 5′-triphosphate (ATP) and adenosine, tightly controlled by nucleotidases, ribokinases, deaminases, and transporters, signal through a rich array of purinergic receptors. These receptors, which emerged early in evolution, are among the most abundant in living organisms controlling many physiological actions, thus becoming promising therapeutic targets in a wide range of pathological conditions. Thus, while P1 receptors are selective for adenosine, a breakdown product of ATP, P2 receptors are activated by purine nucleotides, as well as P2Y receptors being activated by pyrimidine nucleotides. Interestingly, purinergic receptors, both G protein-coupled (i.e., P1 and P2Y) and ligand-gated ion channel (i.e., P2X) receptors, are involved in many neuronal and non-neuronal mechanisms, including pain, immune responses, exocrine and endocrine secretion, platelet aggregation, endothelial-mediated vasodilatation and inflammation, among others

Dopaminergic-cholinergic imbalance in movement disorders: a role for the novel striatal dopamine D2-muscarinic acetylcholine M1 receptor heteromer

The striatum is the primary input structure of the basal ganglia, which participates in motivational and goal-directed behaviors (Pisani et al., 2007). In physiological conditions, local cholinergic interneurons (ChIs) and dopaminergic afferents modulate basal ganglia output through striatal projection neurons, also called medium spiny neurons (MSNs). In general, the release of the neurotransmitters dopamine (DA) and acetylcholine (ACh) elicits contradictory effects on MSNs, which express their corresponding DA receptors (DARs) and muscarinic acetylcholine receptors (mAChRs), respectively (Ztaou and Amalric, 2019). Recently, we discovered a novel receptor-receptor interaction (i.e., heteromerization) between the dopamine D2 receptor (D2R) and the muscarinic acetylcholine M1 receptor (M1R), both expressed at striatopallidal MSNs (Crans et al., 2020). The putative striatal D2R-M1R complex coordinates a sophisticated interplay between the dopaminergic and cholinergic neurotransmission systems. Fuxe et al. (2012) foresaw that the existence of this heteromer within the striatum would mechanistically justify the use of anticholinergics in Parkinson's disease (PD) treatment, thus opening up the development of novel pharmacotherapeutic strategies for PD management. As a proof of concept, we demonstrated that an M1R-selective antagonist (i.e., VU0255035, 10 mg/kg, i.p.) potentiated the antiparkinsonian-like efficacy of an ineffective D2R-selective agonist dose (i.e., sumanirole, 3 mg/kg, i.p.) in a rodent model of experimental Parkinsonism (Crans et al., 2020). Overall, the novel D2R-M1R heteromer could serve as a specific drug target to alleviate motor deficits in PD, whereas it may avoid major adverse effects associated with traditional pharmacotherapies

G protein-coupled receptor 37 (GPR37) emerges as an important modulator of adenosinergic transmission in the striatum

G protein-coupled receptor 37 (GPR37), also known as parkin associated endothelin-like (Pael) receptor, is an orphan G protein-coupled receptor, which suffers a defective parking ubiquitination in autosomal recessive Parkinson's disease promoting its endoplasmic reticulum aggregation and stress, neurotoxicity and neuronal death (Takahashi and Imai, 2003). Interestingly, we have demonstrated previously that GPR37 heteromerizes with adenosine A2A receptor (A2AR) in the striatum (Morató et al., 2017; Sokolina et al., 2017). In addition, we also reported some functional consequences of this direct interaction, whereby GPR37 deletion enhanced striatal A2AR cell surface expression with a concomitant increase in A2AR agonist-mediated cAMP accumulation (Morató et al., 2017); accordingly, an enhancement of A2AR agonist-induced catalepsy and antagonist-induced locomotor activity was observed upon GPR37 deletion (Morató et al., 2017). Overall, it has been hypothesized that GPR37 might hold a chaperone-like activity controlling A2AR cell surface targeting and function. However, the precise physiological function of GPR37 still is unidentified. The current findings now provide additional evidence for the role of GPR37 as a repressor of A2AR function

Lighting up G protein-coupled purinergic receptors with engineered fluorescent ligands

The use of G protein-coupled receptors fluorescent ligands is undergoing continuous expansion. In line with this, fluorescent agonists and antagonists of high affinity for G protein-coupled adenosine and P2Y receptors have been shown to be useful pharmacological probe compounds. Fluorescent ligands for A1R, A2AR, and A3R (adenosine receptors) and P2Y2R, P2Y4R, P2Y6R, and P2Y14R (nucleotide receptors) have been reported. Such ligands have been successfully applied to drug discovery and to GPCR characterization by flow cytometry, fluorescence correlation spectroscopy, fluorescence microscopy, fluorescence polarization, fluorescence resonance energy transfer and scanning confocal microscopy. Here we summarize recently reported and readily available representative fluorescent ligands of purinergic receptors. In addition, we pay special attention on the use of this family of fluorescent ligands revealing two main aspects of purinergic receptor biology, namely ligand binding and receptor oligomerization

Lighting up multiprotein complexes: lessons from GPCR oligomerization

Spatiotemporal characterization of protein protein interactions (PPIs) is essential in determining the molecular mechanisms of intracellular signaling processes. In this review, we discuss how new methodological strategies derived from non-invasive fluorescence- and luminescence-based approaches (FRET, BRET, BiFC and BiLC), when applied to the study of G protein-coupled receptor (GPCR) oligomerization, can be used to detect specific PPIs in live cells. These technologies alone or in concert with complementary methods (SRET, BRET or BiFC, and SNAP-tag or TR-FRET) can be extremely powerful approaches for PPI visualization, even between more than two proteins. Here we provide a comprehensive update on all the biotechnological aspects, including the strengths and weaknesses, of new fluorescence- and luminescence-based methodologies, with a specific focus on their application for studying PPIs

Editorial: “Purinergic Signaling 2020: The State-of-The-Art Commented by the Members of the Italian Purine Club”

Editorial on the Research Topic. Purinergic Signaling 2020: The State-of-The-Art Commented by the Members of the Italian Purine Club

Presynaptic adenosine receptor heteromers as key modulators of glutamatergic and dopaminergic neurotransmission in the striatum

Adenosine plays a very significant role in modulating striatal glutamatergic and dopaminergic neurotransmission. In the present essay we first review the extensive evidence that indicates this modulation is mediated by adenosine A1 and A2A receptors (A1Rs and A2ARs) differentially expressed by the components of the striatal microcircuit that include cortico-striatal glutamatergic and mesencephalic dopaminergic terminals, and the cholinergic interneuron. This microcircuit mediates the ability of striatal glutamate release to locally promote dopamine release through the intermediate activation of cholinergic interneurons. A1Rs and A2ARs are colocalized in the cortico-striatal glutamatergic terminals, where they form A1R-A2AR and A2AR-cannabinoid CB1 receptor (CB1R) heteromers. We then evaluate recent findings on the unique properties of A1R-A2AR and A2AR-CB1R heteromers, which depend on their different quaternary tetrameric structure. These properties involve different allosteric mechanisms in the two receptor heteromers that provide fine-tune modulation of adenosine and endocannabinoid-mediated striatal glutamate release. Finally, we evaluate the evidence supporting the use of different heteromers containing striatal adenosine receptors as targets for drug development for neuropsychiatric disorders, such as Parkinson's disease and restless legs syndrome, based on the ability or inability of the A2AR to demonstrate constitutive activity in the different heteromers, and the ability of some A2AR ligands to act preferentially as neutral antagonists or inverse agonists, or to have preferential affinity for a specific A2AR heteromer

On the role of G protein-coupled receptors oligomerization

The existence of a supramolecular organization of the G protein-coupled receptor (GPCR) is now being widely accepted by the scientific community. Indeed, GPCR oligomers may enhance the diversity and performance by which extracellular signals are transferred to the G proteins in the process of receptor transduction, although the mechanism that underlies this phenomenon still remains unsolved. Recently, it has been proposed that a trans-conformational switching model could be the mechanism allowing direct inhibition/activation of receptor activation/inhibition, respectively. Thus, heterotropic receptor-receptor allosteric regulations are behind the GPCR oligomeric function. In this paper we want to revise how GPCR oligomerization impinges on several important receptor functions like biosynthesis, plasma membrane diffusion or velocity, pharmacology and signaling. In particular, the rationale of receptor oligomerization might lie in the need of sensing complex whole cell extracellular signals and translating them into a simple computational model