Influence of altered life conditions on the regulation of AMP-activated kinase (AMPK) in inflammatory pain

点击翻译按钮获取中文摘要

The AMP-activated kinase (AMPK) is a cellular energy sensor which is activated by an increase in intracellular AMP-concentration occurring after ATP-consuming processes. AMPK regulates a number of cellular functions including glucose and lipid metabolism, redox signaling and resolution of inflammation. Furthermore, we could show in our previous work that activation of AMPK leads to antinociceptive effects. This activation has been mostly achieved by drugs and the molecular mechanisms have only been investigated in isolated cells or, in my working group, in the central nervous system. Based on current results, it has to be assumed that AMPK-mediated antinociception is largely based on molecular mechanisms in the inflamed peripheral tissue. Therefore, one of the aims of the planned project is, based on our previous results, cell-type specific determination of molecular effects of AMPK-activation in the inflamed tissues. The different cell types will be investigated in tissue cryoslides by immunohistochemical staining for cell-specific markers or will be separated and sorted by MACS/FACS-analyses. On the one hand, this makes it possible to detect co-localization with potential AMPK target genes and on the other hand different sorted cells can be assessed for AMPK activation and regulation of AMPK target genes using qRT-PCR und Western Blot.Since modulation of life conditions can also modulate AMPK activation, we will further assess if exercise training, reduction diet or age can affect the AMPK activity in different tissues, particularly in neuronal tissue, and thereby the nociceptive response. We will investigate epigenetic modulations of the AMPK gene as well as regulations of already known AMPK-mediated signal transduction mechanisms. In the experiments, mice will be trained by running on a treadmill for several weeks or will be subjected to a reduction diet. To investigate effects of age we will use mice between 12 and 24 months of age and compare them with younger mice (6-8 weeks). Afterwards, we will determine the methylation status of the AMPK promoter as a parameter of epigenetic regulations and measure AMPK activity and regulation of AMPK targets. The nociceptive behavior will be analyzed in well established models of inflammatory pain.At the end of the project we want to make a statement if AMPK can be stimulated and epigenetically regulated without using drugs and if this activation is sufficient to inhibit nociception. Moreover, we will gain new insight in the regulation of AMPK in the inflamed tissue, which might also provide novel treatment options for pain.