With the increase of life expectancy, neurodegenerative disorders are becoming not only a health but also a social burden worldwide. However, due to the multitude of pathophysiological disease states, current treatments fail to meet the desired outcomes. Therefore, there is a need for new therapeutic strategies focusing on more integrated, personalized and effective approaches. The prospect of using neural stem cells (NSC) as regenerative therapies is very promising, however several issues still need to be addressed. In particular, the potential actions of pharmacological agents used to modulate NSC activity are highly relevant. With the ongoing discussion of cannabinoid usage for medical purposes and reports drawing attention to the effects of cannabinoids on NSC regulation, there is an enormous, and yet, uncovered potential for cannabinoids as treatment options for several neurological disorders, specifically when combined with stem cell therapy. In this manuscript, we review in detail how cannabinoids act as potent regulators of NSC biology and their potential to modulate several neurogenic features in the context of pathophysiology.

Chronic stress is a major risk factor for neuropsychiatric conditions such as depression. Adult hippocampal neurogenesis (AHN) has emerged as a promising target to counteract stress-related disorders given the ability of newborn neurons to facilitate endogenous plasticity. Recent data sheds light on the interaction between cannabinoids and neurotrophic factors underlying the regulation of AHN, with important effects on cognitive plasticity and emotional flexibility. Since physical exercise (PE) is known to enhance neurotrophic factor levels, we hypothesised that PE could engage with cannabinoids to influence AHN and that this would result in beneficial effects under stressful conditions. We therefore investigated the actions of modulating cannabinoid type 2 receptors (CB2R), which are devoid of psychotropic effects, in combination with PE in chronically stressed animals. We found that CB2R inhibition, but not CB2R activation, in combination with PE significantly ameliorated stress-evoked emotional changes and cognitive deficits. Importantly, this combined strategy critically shaped stress-induced changes in AHN dynamics, leading to a significant increase in the rates of cell proliferation and differentiation of newborn neurons, overall reduction in neuroinflammation, and increased hippocampal levels of BDNF. Together, these results show that CB2Rs are crucial regulators of the beneficial effects of PE in countering the effects of chronic stress. Our work emphasises the importance of understanding the mechanisms behind the actions of cannabinoids and PE and provides a framework for future therapeutic strategies to treat stress-related disorders that capitalise on lifestyle interventions complemented with endocannabinoid pharmacomodulation.

Postnatal neurogenesis has been shown to rely on the endocannabinoid system. Here we aimed at unravelling the role of Cannabidivarin (CBDV), a non-psychoactive cannabinoid, with high affinity for the non-classical cannabinoid receptor TRPV1, on subventricular zone (SVZ) postnatal neurogenesis. Using the neurosphere assay, SVZ-derived neural stem/progenitor cells (NSPCs) were incubated with CBDV and/or 5'-Iodoresinferotoxin (TRPV1 antagonist), and their role on cell viability, proliferation, and differentiation were dissected. CBDV was able to promote, through a TRPV1-dependent mechanism, cell survival, cell proliferation and neuronal differentiation. Furthermore, pulse-chase experiments revealed that CBDV-induced neuronal differentiation was a result of cell cycle exit of NSPCs. Regarding oligodendrocyte differentiation, CBDV inhibited oligodendrocyte differentiation and maturation. Since our data suggested that the CBDV-induced modulation of NSPCs acted via TRPV1, a sodium-calcium channel, and that intracellular calcium levels are known regulators of NSPCs fate and neuronal maturation, single cell calcium imaging was performed to evaluate the functional response of SVZ-derived cells. We observed that CBDV-responsive cells displayed a two-phase calcium influx profile, being the initial phase dependent on TRPV1 activation. Taken together, this work unveiled a novel and untapped neurogenic potential of CBDV via TRPV1 modulation. These findings pave the way to future neural stem cell biological studies and repair strategies by repurposing this non-psychoactive cannabinoid as a valuable therapeutic target.

Oligodendrocytes (OLs) are vital for myelin formation in the Central Nervous System, and their differentiation from precursor cells is critical for remyelination in demyelinating disorders. These cells originate either from the maturation of oligodendrocyte precursor cells (OPCs), present throughout the brain parenchyma, or from the differentiation of neural stem cells located in the subventricular zone (SVZ-NSCs). However, the molecular signals regulating their differentiation remain partially understood. In this work, we studied the role of brain-derived neurotrophic factor (BDNF) and adenosine A_2A receptor, as well as their potential interaction, in modulating oligodendrogenesis using postnatal SVZ-derived neurosphere cultures. In these cultures, BDNF tended to increase mRNA expression levels of OPC markers after 2 days in vitro (DIV), an effect blocked by the A_2A receptor antagonist ZM 241385. This early transcriptional regulation by BDNF was followed by changes in the percentage of both OPCs and mature OLs in culture at DIV 7 and 14. Interestingly, A_2A receptor blockade prevented the BDNF-induced increase in OL differentiation at DIV 14. Concerning the morphology of mature OLs, BDNF transiently reduced proximal branching at DIV 7, an effect absent at 14 DIV, when all treatments displayed similar morphologies. At the protein level, BDNF enhanced TrkB expression at DIV 14 in an A_2A receptor-dependent manner, without altering A_2A receptor expression itself, further supporting a modulatory role of A_2A receptors in BDNF receptor signalling. Overall, our results identify BDNF as a key regulator of OL differentiation and reveal an A_2A receptor-BDNF interaction modulating this differentiation process.

Under Multiple Sclerosis pathological conditions, oligodendrocyte precursor cells (OPCs) present in the brain parenchyma or derived from subventricular zone neural stem cells (SVZ-NSCs) can differentiate into oligodendrocytes (OLs), which migrate and partially remyelinate the lesioned areas. Previous data from our group demonstrated that activation of adenosine A2A receptors (A2AR) modulated SVZ-NSCs oligodendroglial differentiation, both in vitro and in vivo under physiological conditions. Hence, we aimed at understanding the role of A2AR in adult oligodendrogenesis derived from SVZ-NSCs in an in vivo mouse model of MS. For this, the Experimental Autoimmune Encephalomyelitis (EAE) mouse model of MS was developed, and behavioural tests were performed to evaluate motor function. Cellular differentiation was assessed by immunohistochemistry assays for bromodeoxyuridine (BrdU) colocalization with oligodendrocytic markers in brain regions of interest. Western blot and ELISA assays were used for myelin protein levels and inflammatory cytokine quantification. Our results for EAE model characterization showed that motor impairment is proportional to the score of the disease and cellular and molecular data showed an increase in the levels of the pro-inflammatory cytokine TNFα (n=5, p<0.01). A significant increase in NG2+BrdU+ cells in the corpus callosum (CC) of EAE mice was observed (n=3, p<0.05), hinting at the migration of OPCs from the SVZ to the CC. Ongoing studies encompass the in vivo modulation of A2AR and assessing its effect on EAE phenotype and adult oligodendrogenesis, ultimately unveiling the modulation of adult oligodendrogenesis derived from SVZ-NSCs by A2AR as a putative therapy for MS.

Oligodendrocytes (OLs), the myelinating cells of the Central Nervous System (CNS), are generated upon differentiation of oligodendrocyte precursor cells (OPCs), which possess proliferative and migratory capabilities and are present in the CNS postnatally. Under pathological conditions such as in Multiple Sclerosis (MS), there is a depletion of OLs, but the OPCs present in the brain parenchyma or derived from subventricular zone (SVZ) neural stem cells (NSCs) can differentiate, migrate and partially remyelinate the lesioned areas. Herein, we aimed at characterizing the MS mouse model, experimental autoimmune encephalomyelitis (EAE), studying animal behaviour as well as the process of adult oligodendrogenesis. For this, the EAE mouse model was developed, and behavioural tests (open field, rotarod, pole test) were performed to evaluate motor function. Cellular differentiation was assessed by immunohistochemistry assays for bromodeoxyuridine (BrdU) colocalization with oligodendrocytic markers in brain regions of interest. Western blot and ELISA assays were used for myelin protein levels and inflammatory cytokine quantification. Results for clinical scores and behavioural characterization of the EAE model suggested that motor impairment is proportional to the score of the disease. Moreover, an increase in the levels of the pro-inflammatory cytokine TNFα (n=5, p<0.01), and a tendency for increased IL-1β were observed in EAE mice. Importantly, a tendency for increased BrdU+ cells in the SVZ, corpus callosum (CC) and cerebral cortex (CT) was observed, accompanied by a significant increase in NG2+BrdU+ cells in the CC of EAE mice (n=3, p<0.05), hinting at the migration of precursor cells from the SVZ to the CC. Altogether, this work allowed the characterization of the oligodendrogenesis process and of the EAE model throughout time, supporting future studies involving the modulation of adult oligodendrogenesis as a putative therapy for MS.