We observed distinct expression patterns of the kinase subfamilies

n neurons against extended culture periods or 6-OHDA. To examine whether the agonist-induced protection likely was mediated via mitochondrial complex I inhibition, we treated the cultured DA midbrain neurons with rotenone, which, like MPP+, is a mitochondrial complex I inhibitor. While rotenone did induce a dose-dependent cell death in the TH-positive neurons, no rescue was seen in the agonist treated subpopulation. Agonist-Dopamine Transporter Interaction As both 6-OHDA and MPP+ are taken up via the DAT, it was not likely that the agonists mediated their protection against MPP+ by merely blocking the DAT. However, to further exclude the possibility that the protection against MPP+ by the agonist compound 3 is mediated by blocking MPP+ uptake, rather than signaling through GPR139, DA and noradrenaline uptake in the presence of the agonist was tested. At 10 M compound 3 inhibited DA uptake only 13% and norepinephrine uptake only -13%, suggesting that the protective effect observed with the agonist is likely not mediated by directly acting on the DAT. Ten micrometres compound 1 inhibited binding to the DAT or the norepinephrine transporter by 4% or -8%, respectively compound 2 inhibited binding to the DAT or the norepinephrine transporter 2% or -4%, respectively. We next sought to examine whether the agonists would also provide neuroprotection in vivo. However, we found that ADME properties of the compounds were not favorable to provide sufficient brain exposure and receptor occupancy, neither by oral application, nor by subcutaneous delivery via osmotic pumps. 6 Bayer Andersen et al. GPR139 Agonists in Parkinson Model DISCUSSION We demonstrate that three GPR139 agonists dose-dependently protect primary DA neurons against MPP+ toxicity. When 71939-50-9 treating cultured DA midbrain neurons with rotenone or 6-OHDA, we also observed dose-dependent cell death; however, GPR139 agonist treatment did not rescue the neurons. PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19816862 Moreover, no protection was demonstrated against prolonged culture periods, suggesting that GPR139 agonism does not enhance general cellular viability and resistance against apoptotic stimuli. Previously, not peer-reviewed work suggested that GPR139 KO mice display a deficit in motor performance. However, in pilot experiments, we could not confirm those deficits using the rotarod and the balance beam test. That discrepancy might be due to a different and variable background of the mice examined compared to the mice in the earlier study. Work by Song et al. has previously reported that the bibenzyl compound Chrysotoxine could antagonize the toxicity of MPP+, but not rotenone in SH-SY5Y cells. However, the authors explain that the MPP+ protection is, at least partly, due to inhibition of the DAT. In our study the protective effect of the agonists seen in the MPP+ treated cultures is unlikely to be due to DAT inhibition. First, 6-OHDA is also partly taken up via the DAT and no protection of the agonists was seen when cultures were treated with 6-OHDA. Next, agonist compound 1 and compound 2 did not show cross-reactivity with the DAT. It cannot be entirely ruled out that compound 3 does interfere with the DAT; however, considering the very similar effect of the three agonist on MPP+ toxicity, that mechanism is not very likely. We further examined the specificity of the agonist protection against MPP+ toxicity by showing that a GPR139 antagonist To confirm the motor deficits earlier described in Gpr139 KO mice, the mice underwent behavioral