Differentiation of oligodendrocyte precursor cells (OPCs) into mature oligodendrocytes (OLs) is a key event for axonal myelination in the brain; this process fails during demyelinating pathologies. Adenosine is emerging as an important player in oligodendrogliogenesis, by activating its metabotropic receptors (A₁R, A_2AR, A_2BR, and A₃R). We previously demonstrated that the Gs‐coupled A_2BR reduced differentiation of primary OPC cultures by inhibiting delayed rectifier (I_K) as well as transient (I_A) outward K⁺ currents. To deepen the unclear role of this receptor subtype in neuron‐OL interplay and in myelination process, we tested the effects of different A_2BR ligands in a dorsal root ganglion neuron (DRGN)/OPC cocultures, a corroborated in vitro myelination assay. The A_2BR agonist, BAY60‐6583, significantly reduced myelin basic protein levels but simultaneously increased myelination index in DRGN/OPC cocultures analyzed by confocal microscopy. The last effect was prevented by the selective A_2BR antagonists, PSB‐603 and MRS1706. To clarify this unexpected data, we wondered whether A_2BRs could play a functional role on DRGNs. We first demonstrated, by immunocytochemistry, that primary DRGN monoculture expressed A_2BRs. Their selective activation by BAY60‐6583 enhanced DRGN excitability, as demonstrated by increased action potential firing, decreased rheobase and depolarized resting membrane potential and were prevented by PSB‐603. Throughout this A_2BR‐dependent enhancement of neuronal activity, DRGNs could release factors to facilitate myelination processes. Finally, silencing A_2BR in DRGNs alone prevents the increased myelination induced by BAY60‐6583 in cocultures. In conclusion, our data suggest a different role of A_2BR during oligodendrogliogenesis and myelination, depending on their activation on neurons or oligodendroglial cells.

In recent years, the use of multi-target compounds has become an increasingly pursued strategy to treat complex pathologies, including cerebral ischemia. Adenosine and its receptors (A₁AR, A_2AAR, A_2BAR, A₃AR) are known to play a crucial role in synaptic transmission either in normoxic or ischemic-like conditions. Previous data demonstrate that the selective antagonism of A_2AAR or A_2BAR delays anoxic depolarization (AD) appearance, an unequivocal sign of neuronal injury induced by a severe oxygen-glucose deprivation (OGD) insult in the hippocampus. Furthermore, the stimulation of A_2AARs or A_2BARs by respective selective agonists, CGS21680 and BAY60-6583, increases pre-synaptic neurotransmitter release, as shown by the decrease in paired-pulse facilitation (PPF) at Schaffer collateral-CA1 synapses. In the present research, we investigated the effect/s of the newly synthesized dual A_2AAR/A_2BAR antagonist, P626, in preventing A_2AAR- and/or A_2BAR-mediated effects by extracellular recordings of synaptic potentials in the CA1 rat hippocampal slices. We demonstrated that P626 prevented PPF reduction induced by CGS21680 or BAY60-6583 and delayed, in a concentration-dependent manner, AD appearance during a severe OGD. In conclusion, P626 may represent a putative neuroprotective compound for stroke treatment with the possible translational advantage of reducing side effects and bypassing differences in pharmacokinetics due to combined treatment.

Demyelinating diseases are characterized by the progressive loss of myelin in the central nervous system (CNS). Myelin protection can be achieved by fostering the differentiation of oligodendrocyte progenitor cells to mature oligodendrocytes, the only myelinating cells in the brain. Oligodendrocytes maturation is sustained, among others, by the neuromodulator adenosine through the activation of its specific receptors: A₁, A_2A, A_2B, A₃, all expressed in the brain on neurons and glial cells. The role of A_2B receptors (A2BRs) in a cuprizone-induced demyelination model in male C57BL/6 mice was investigated by administering the selective A2BR agonist BAY60-6583 or antagonist PSB 603 during the last 2 weeks of a 5-week cuprizone-based diet. We performed body weight evaluation, behavioural tests and immunofluorescence analysis. Cuprizone-fed mice showed a significant decrease in body weight gain, a motor impairment and a reduced spontaneous mobility. These effects were associated with a decrease in myelin levels, a reactive astrogliosis and microgliosis in corpus callosum medialis, striatum and motor cortex. PSB 603, was able to prevent cuprizone effects on glia, whereas both compounds promoted a significant recovery in motor deficits. The antagonism of A2BRs might represent an attractive strategy to alleviate myelin damage and glial activation in the CNS under conditions of demyelination.