Background and Purpose— Ischemic stroke continues to be one of the main causes of death worldwide. Inflammation accounts for a large part of damage in this pathology. The cannabinoid type 2 receptor (CB2R) has been proposed to have neuroprotective properties in neurological diseases. Therefore, our aim was to determine the effects of the activation of CB2R on infarct outcome and on ischemia-induced brain expression of classic and alternative markers of macrophage/microglial activation. Methods— Swiss wild-type and CB2R knockout male mice were subjected to a permanent middle cerebral artery occlusion. Mice were treated with either a CB2R agonist (JWH-133), with or without a CB2R antagonist (SR144528) or vehicle. Infarct outcome was determined by measuring infarct volume and neurological outcome. An additional group of animals was used to assess mRNA and protein expression of CB2R, interleukin (IL)-1β, IL-6, tumor necrosis factor α (TNF-α), monocyte chemoattractant protein–1 (MCP-1), macrophage inflammatory peptide (MIP) –1α, RANTES, inducible nitric oxide synthase (iNOS), cyclooxygenase-2, IL-4, IL-10, transforming growth factor β (TGF-β), arginase I, and Ym1. Results— Administration of JWH-133 significantly improved infarct outcome, as shown by a reduction in brain infarction and neurological impairment. This effect was reversed by the CB2R antagonist and was absent in CB2R knockout mice. Concomitantly, administration of JWH-133 led to a lower intensity of Iba1+ microglia/macrophages and a decrease in middle cerebral artery occlusion–induced gene expression of both classic (IL-6, TNF-α, MCP-1, MIP-1α, RANTES, and iNOS) and alternative mediators/markers (IL-10, TGF-β, and Ym1) of microglial/macrophage activation after permanent middle cerebral artery occlusion. Conclusions— The inhibitory effect of CB2R on the activation of different subpopulations of microglia/macrophages may account for the protective effect of the selective CB2R agonist JWH-133 after stroke.

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Diadenosine tetraphosphate (Ap4A) and diadenosine pentaphosphate (Ap5A) have been identified in bovine adrenal medullary tissue using an HPLC method. The values obtained were 0.1 +/- 0.05 mumol/g of tissue for both compounds. The subcellular fraction where Ap4A and Ap5A were present in the highest concentration was chromaffin granules: 32 nmol/mg of protein for both compounds (approximately 6 mM intragranularly). This value was 30 times higher than in the cytosolic fraction. Enzymatic degradation of Ap4A and Ap5A, isolated from chromaffin granules, with phosphodiesterase produces AMP as the final product. The Ap4A and Ap5A obtained from this tissue were potent inhibitors of adenosine kinase. Their Ki values relative to adenosine were 0.3 and 2 microM for Ap4A and Ap5A, respectively. The cytosolic fraction also contains enzymatic activities that degrade Ap4A as well as Ap5A. These activities were measured by an HPLC method; the observed Km values were 10.5 +/- 0.5 and 13 +/- 1 microM for Ap4A and Ap5A, respectively.

At synaptic boutons, metabotropic glutamate receptor 7 (mGlu7 receptor) serves as an autoreceptor, inhibiting glutamate release. In this response, mGlu7 receptor triggers pertussis toxin-sensitive G protein activation, reducing presynaptic Ca 2؉ influx and the subsequent depolarization evoked release. Here we report that receptor coupling to signaling pathways that potentiate release can be seen following prolonged exposure of nerve terminals to the agonist L-(؉)-phosphonobutyrate, L-AP4. This novel mGlu7 receptor response involves an increase in the release induced by the Ca 2؉ ionophore ionomycin, suggesting a mechanism that is independent of Ca 2؉ channel activity, but dependent on the downstream exocytotic release machinery. The mGlu7 receptor-mediated potentiation resists exposure to pertussis toxin, but is dependent on phospholipase C, and increased phosphatidylinositol (4,5)-bisphosphate hydrolysis. Furthermore, the potentiation of release does not depend on protein kinase C, although it is blocked by the diacylglycerolbinding site antagonist calphostin C. We also found that activation of mGlu7 receptors translocate the active zone protein essential for synaptic vesicle priming, munc13-1, from soluble to particulate fractions. We propose that the mGlu7 receptor can facilitate or inhibit glutamate release through multiple pathways, thereby exerting homeostatic control of presynaptic function.Metabotropic glutamate receptors belong to the G proteincoupled receptors (GPCRs) 2 superfamily and their eight receptor subtypes (mGlu1-8 receptors) are classified into three major groups. Most group III mGlu receptors (mGlu4, -6, -7, and -8 receptors) are located within the presynaptic active zone (1) where they act as autoreceptors mediating feedback inhibition of glutamate release (2-4). The signaling mechanism initiated by mGlu7 receptors to inhibit neurotransmitter release involves the activation of G i/o proteins that inhibit Ca 2ϩ channels and adenylyl cyclase (5), and probably the release process itself (6). However, mGlu7 receptor signaling is not restricted to these pathways. Thus, transfected mGlu7 receptors expressed in cerebellar granule cells inhibits somatic Ca 2ϩ currents by a mechanism that involves the activation of phospholipase C (PLC) and the hydrolysis of phosphatidylinositol (4,5)-bisphosphate thereby generating inositol trisphosphate that releases Ca 2ϩ from intracellular stores and diacylglycerol (DAG) that activates protein kinase C (PKC) (7). However, one important question that remains to be resolved is whether endogenous mGlu7 receptors at synaptic sites also signal via PLC and if so, what effect such signaling has on release modulation.Phorbol esters, stable analogues of the endogenous product of PLC, DAG, potentiate synaptic transmission by increasing neurotransmitter release (8 -15). DAG signaling at synapses has long been thought to be mediated by PKC and both presynaptic K ϩ and Ca 2ϩ channels, as well as other proteins of the release machinery, have been identified as PKC substrates....