Inhibition of Casein Kinase 2 Protects Oligodendrocytes From Excitotoxicity by Attenuating JNK/p53 Signaling Cascade.

Oligodendrocytes are highly vulnerable to glutamate excitotoxicity, a central mechanism involved in tissue damage in Multiple Sclerosis (MS). Sustained activation of AMPA receptors in rat oligodendrocytes induces cytosolic calcium overload, mitochondrial depolarization, increase of reactive oxygen species, and activation of intracelular pathways resulting in apoptotic cell death. Although many signals driven by excitotoxicity have been identified, some of the key players are still under investigation. Casein kinase 2 (CK2) is a serine/threonine kinase, constitutively expressed in all eukaryotic tissues, involved in cell proliferation, malignant transformation and apoptosis. In this study, we identify CK2 as a critical regulator of oligodendrocytic death pathways and elucidate its role as a signal inductor following excitotoxic insults. We provide evidence that CK2 activity is up-regulated in AMPA-treated oligodendrocytes and CK2 inhibition significantly diminished AMPA receptor-induced oligodendroglial death. In addition, we analyzed mitogen-activated protein kinase (MAPK) signaling after excitotoxic insult. We observed that AMPA receptor activation induced a rapid increase in c-Jun N-terminal kinase (JNK) and p38 phosphorylation that was reduced after CK2 inhibition. Moreover, blocking their phosphorylation, we enhanced oligodendrocyte survival after excitotoxic insult. Finally, we observed that the tumor suppressor p53 is activated during AMPA receptor-induced cell death and, interestingly, down-regulated by JNK or CK2 inhibition. Together, these data indicate that the increase in CK2 activity induced by excitotoxic insults regulates MAPKs, triggers p53 activation and mediates subsequent oligodendroglial loss. Therefore, targeting CK2 may be a useful strategy to prevent oligodendrocyte death in MS and other diseases involving central nervous system (CNS) white matter.

Deregulation of the endocannabinoid system and therapeutic potential of ABHD6 blockade in the cuprizone model of demyelination.

Multiple sclerosis (MS) is a chronic demyelinating disease of unknown etiology in which tissue pathology suggests both immune-dependent attacks to oligodendroglia and primary oligodendrocyte demise. The endocannabinoid system has been crucially involved in the control of autoimmune demyelination and cannabinoid-based therapies exhibit therapeutic potential, but also limitations, in MS patients. In this context, growing evidence suggests that targeting the hydrolysis of the main endocannabinoid 2-arachidonoylglycerol (2-AG) may offer a more favorable benefit-to-risk balance in MS than existing cannabinoid medicines. Here we evaluated the modulation of endocannabinoid signaling and the therapeutic potential of targeting the 2-AG hydrolytic enzyme alpha/beta-hydrolase domain-containing 6 (ABHD6) in the cuprizone model of non-immune dependent demyelination. The concentrations of N-arachidonoylethanolamine (anandamide, AEA) and its congener N-palmitoylethanolamine (PEA) were reduced following 6 weeks of cuprizone feeding. Deregulation of AEA and PEA levels was not due to differences in the expression of the hydrolytic and biosynthetic enzymes fatty acid amide hydrolase and N-acylphosphatidylethanolamine-phospholipase D, respectively. Conversely, we measured elevated transcript levels of 2-AG hydrolytic enzymes monoacylglycerol lipase, ABHD6 and ABHD12 without changes in bulk 2-AG concentration. Upregulated CB1 and CB2 receptors expression, ascribed in part to microglia, was also detected in the brain of cuprizone-treated mice. Administration of an ABHD6 inhibitor partially attenuated myelin damage, astrogliosis and microglia/macrophage reactivity associated to cuprizone feeding. However, ABHD6 blockade was ineffective at engaging protective or differentiation promoting effects in oligodendrocyte cultures. These results show specific alterations of the endocannabinoid system and modest beneficial effects resulting from ABHD6 inactivation in a relevant model of primary demyelination.

Inwardly Rectifying K+ Currents in Cultured Oligodendrocytes from Rat Optic Nerve are Insensitive to pH.

Inwardly rectifying K+ (Kir) channel expression signals at an advanced stage of maturation during oligodendroglial differentiation. Knocking down their expression halts the generation of myelin and produces severe abnormalities in the central nervous system. Kir4.1 is the main subunit involved in the tetrameric structure of Kir channels in glial cells; however, the precise composition of Kir channels expressed in oligodendrocytes (OLs) remains partially unknown, as participation of other subunits has been proposed. Kir channels are sensitive to H+; thus, intracellular acidification produces Kir current inhibition. Since Kir subunits have differential sensitivity to H+, we studied the effect of intracellular acidification on Kir currents expressed in cultured OLs derived from optic nerves of 12-day-old rats. Unexpectedly, Kir currents in OLs (2-4 DIV) did not change within the pH range of 8.0-5.0, as observed when using standard whole-cell voltage-clamp recording or when preserving cytoplasmic components with the perforated patch-clamp technique. In contrast, low pH inhibited astrocyte Kir currents, which was consistent with the involvement of the Kir4.1 subunit. The H+-insensitivity expressed in OL Kir channels was not intrinsic because Kir cloning showed no difference in the sequence reported for the Kir4.1, Kir2.1, or Kir5.1 subunits. Moreover, when Kir channels were heterologously expressed in Xenopus oocytes they behaved as expected in their general properties and sensitivity to H+. It is therefore concluded that Kir channel H+-sensitivity in OLs is modulated through an extrinsic mechanism, probably by association with a modulatory component or by posttranslational modifications.

Axon-to-Glia Interaction Regulates GABAA Receptor Expression in Oligodendrocytes.

Myelination requires oligodendrocyte-neuron communication, and both neurotransmitters and contact interactions are essential for this process. Oligodendrocytes are endowed with neurotransmitter receptors whose expression levels and properties may change during myelination. However, only scant information is available about the extent and timing of these changes or how they are regulated by oligodendrocyte-neuron interactions. Here, we used electrophysiology to study the expression of ionotropic GABA, glutamate, and ATP receptors in oligodendrocytes derived from the optic nerve and forebrain cultured either alone or in the presence of dorsal root ganglion neurons. We observed that oligodendrocytes from both regions responded to these transmitters at 1 day in culture. After the first day in culture, however, GABA sensitivity diminished drastically to less than 10%, while that of glutamate and ATP remained constant. In contrast, the GABA response amplitude was sustained and remained stable in oligodendrocytes cocultured with dorsal root ganglion neurons. Immunochemistry and pharmacological properties of the responses indicated that they were mediated by distinctive GABAA receptors and that in coculture with neurons, the oligodendrocytes bearing the receptors were those in direct contact with axons. These results reveal that GABAA receptor regulation in oligodendrocytes is driven by axonal cues and that GABA signaling may play a role in myelination and/or during axon-glia recognition.