Metabotropic glutamate receptors (mGluRs) play important roles in the modulation of nociception. The group I mGluRs (mGlu1 and mGlu5) modulate nociceptive plasticity via activation of extracellular signal-regulated kinase (ERK) signaling. We reported recently that the K ϩ channel Kv4.2 subunit underlies A-type K ϩ currents in the spinal cord dorsal horn and is modulated by the ERK signaling pathway. Kv4.2-mediated A-type currents are important determinants of dorsal horn neuronal excitability and central sensitization that underlies hypersensitivity after tissue injury. In the present study, we demonstrate that ERK-mediated phosphorylation of Kv4.2 is downstream of mGlu5 activation in spinal cord dorsal horn neurons. Activation of group I mGluRs inhibited Kv4.2-mediated A-type K ϩ currents and increased neuronal excitability in dorsal horn neurons. These effects were mediated by activation of mGlu5, but not mGlu1, and were dependent on ERK activation. Analysis of Kv4.2 phosphorylation site mutants clearly identified S616 as the residue responsible for mGlu5-ERK-dependent modulation of A-type currents and excitability. Furthermore, nociceptive behavior induced by activation of spinal group I mGluRs was impaired in Kv4.2 knock-out mice, demonstrating that, in vivo, modulation of Kv4.2 is downstream of mGlu5 activation. Altogether, our results indicate that activation of mGlu5 leads to ERK-mediated phosphorylation and modulation of Kv4.2-containing potassium channels in dorsal horn neurons. This modulation may contribute to nociceptive plasticity and central sensitization associated with chronic inflammatory pain conditions.

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The extracellular signal-regulated kinase (ERK) isoforms, ERK1 and ERK2, are believed to be key signaling molecules in nociception and nociceptive sensitization. Studies using inhibitors targeting the shared ERK1/2 upstream activator, mitogen-activated protein kinase kinase (MEK), and transgenic mice expressing a dominant-negative form of MEK have established the importance of ERK1/2 signaling. However, these techniques do not discriminate between ERK1 and ERK2. To dissect the function of each isoform in pain, we used mice with a targeted genetic deletion of ERK1 [ERK1 knock-out (KO)] to test the hypothesis that ERK1 is required for behavioral sensitization in rodent pain models. Despite activation (phosphorylation) of ERK1 after acute noxious stimulation and in models of chronic pain, we found that ERK1 was not required for formalin-induced spontaneous behaviors, complete Freund's adjuvant-induced heat and mechanical hypersensitivity, and spared nerve injury-induced mechanical hypersensitivity. However, ERK1 deletion did delay formalin-induced long-term heat hypersensitivity, without affecting formalin-induced mechanical hypersensitivity, suggesting that ERK1 partially shapes long-term responses to formalin. Interestingly, ERK1 deletion resulted in elevated basal ERK2 phosphorylation. However, this did not appear to influence nociceptive processing, since inflammation-induced ERK2 phosphorylation and pERK1/2 immunoreactivity in spinal cord were not elevated in ERK1 KO mice. Additionally, systemic MEK inhibition with SL327 (␣-[amino[(4-aminophenyl)thio]methylene]-2-(trifluoromethyl)benzeneacetonitrile) attenuated formalin-induced spontaneous behaviors similarly in wild-type and ERK1 KO mice, indicating that unrelated signaling pathways do not functionally compensate for the loss of ERK1. Together, these results suggest that ERK1 plays a limited role in nociceptive sensitization and support a predominant role for ERK2 in these processes.

The pre-synaptic protein, a-synuclein, has been associated with the pathogenesis of Parkinson's disease. The present study indicates that a-synuclein, but not its mutants (A53T, A30P), can protect CNS dopaminergic cells from the parkinsonism-inducing drug 1-methyl-4-phenylpyridinium (MPP + ), whereas it cannot protect from the dopaminergic toxin, 6-hydroxydopamine, hydrogen-peroxide, or the b-amyloid peptide, A-b. Protection from MPP + was directly correlated with the preservation of mitochondrial function. Specifically, a-synuclein rescued cells from MPP + mediated decreases in mitochondrial dehydrogenase activity and loss of ATP levels by utilizing ketosis. It also prevented toxin-induced activation of the creatine kinase/creatine phosphate system. Similarly, a-synuclein protected cells from the complex I inhibitor rotenone and 3-nitroproprionic acid, a complex II inhibitor. Wildtype a-synuclein-mediated neuroprotection and subsequent alterations in energy were not found in dbcAMP-differentiated cells. These results suggest that the normal physiological role for a-synuclein may change during development.

Background and purpose: Inhibitors of fatty acid amide hydrolase (FAAH), the enzyme responsible for the metabolism of the endogenous cannabinoid (CB) receptor ligand anandamide (AEA), are effective in a number of animal models of pain. Here, we investigated a series of isoflavones with respect to their abilities to inhibit FAAH. Experimental approach: In vitro assays of FAAH activity and affinity for CB receptors were used to characterize key compounds. In vivo assays used were biochemical responses to formalin in anaesthetized mice and the 'tetrad' test for central CB receptor activation. Key results: Of the compounds tested, biochanin A was adjudged to be the most promising. Biochanin A inhibited the hydrolysis of 0.5 mM AEA by mouse, rat and human FAAH with IC50 values of 1.8, 1.4 and 2.4 mM respectively. The compound did not interact to any major extent with CB1 or CB2 receptors, nor with FAAH-2. In anaesthetized mice, URB597 (30 mg i.pl.) and biochanin A (100 mg i.pl.) both inhibited the spinal phosphorylation of extracellular signal-regulated kinase produced by the intraplantar injection of formalin. The effects of both compounds were significantly reduced by the CB1 receptor antagonist/inverse agonist AM251 (30 mg i.pl.). Biochanin A (15 mg·kg -1 i.v.) did not increase brain AEA concentrations, but produced a modest potentiation of the effects of 10 mg·kg -1 i.v. AEA in the tetrad test. Conclusions and implications: It is concluded that biochanin A, in addition to its other biochemical properties, inhibits FAAH both in vitro and peripherally in vivo.