Environmental stressors transiently activate neural systems that inhibit pain responsiveness, thereby inducing a phenomenon known as stress-induced analgesia (SIA). We have recently reported that nonopioid SIA may be mediated by the mobilization of endogenous cannabinoid lipids, such as 2-arachidonoylglycerol (2-AG), in the midbrain periaqueductal gray (PAG) [Hohmann et al., Nature 435: 1108-1112, 2005]. However, the molecular mechanisms governing 2-AG signaling in the brain under physiological conditions remain unknown. Our central hypothesis is that stress stimuli that result in nonopioid SIA activate diacylglycerol lipase (DGL) in select neurons of the PAG, causing the rapid mobilization of 2-AG. Newly- released 2-AG engages local cannabinoid receptors to induce nonopioid SIA, and is then subsequently deactivated through monoacylglycerol lipase (MGL)-mediated hydrolysis. We will test this hypothesis by developing a novel, though high-risk, methodology that unites, for the first time, in vivo virally-mediated gene transfer with targeted lipidomic analyses. Our work will identify the functional consequences of virally- mediated RNA silencing of enzymes implicated in 2-AG formation (DGL) and deactivation (MGL) using three independent complementary approaches. We will use behavioral, neuroanatomical and mass spectrometric analyses (of related lipid mediators, their precursors and metabolites) to evaluate direct and indirect consequences of DGL and MGL silencing on lipid signaling pathways and SIA. Specifically, we will determine whether virally mediated silencing of the DGL and MGL genes within the PAG (i) influences 2-AG signaling and SIA; and (ii) causes distal changes in other endocannabinoid and non-cannabinoid lipid pathways. These studies are expected to demonstrate that 1) in vivo virally-mediated RNA silencing of DGL- beta - a DGL isoform recently shown to colocalize with phospholipase C, an enzyme implicated in formation of the 2-AG precursor diacylglycerol (DAG) - suppresses 2-AG formation in the PAG and SIA, and 2) in vivo virally-mediated RNA silencing of MGL stimulates 2-AG accumulation in the PAG and SIA. Validation of our in vivo gene transfer - targeted lipidomic approach should offer a critical advantage over existing methods which rely solely on neuroanatomical measurements of mRNA or protein. The results of these studies should elucidate, for the first time, the molecular, biochemical and physiological mechanisms governing 2-AG signaling in the brain and determine the role of this endocannabinoid mediator in pain modulation. Furthermore, the results will validate targeted lipidomics as a means to quantify changes in lipid signaling that occur downstream of targeted genetic manipulations. Successful validation of our technological approach and hypotheses may facilitate development of endocannabinoid-based pharmacological and gene therapies for pain. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21DA022478-01
Application #
7189402
Study Section
Special Emphasis Panel (ZDA1-MXS-M (14))
Program Officer
Satterlee, John S
Project Start
2006-09-15
Project End
2008-08-31
Budget Start
2006-09-15
Budget End
2007-08-31
Support Year
1
Fiscal Year
2006
Total Cost
$161,862
Indirect Cost
Name
University of Georgia
Department
Psychology
Type
Schools of Arts and Sciences
DUNS #
004315578
City
Athens
State
GA
Country
United States
Zip Code
30602
Rahn, Elizabeth J; Deng, Liting; Thakur, Ganesh A et al. (2014) Prophylactic cannabinoid administration blocks the development of paclitaxel-induced neuropathic nociception during analgesic treatment and following cessation of drug delivery. Mol Pain 10:27
Gregg, Laura C; Jung, Kwang-Mook; Spradley, Jessica M et al. (2012) Activation of type 5 metabotropic glutamate receptors and diacylglycerol lipase-? initiates 2-arachidonoylglycerol formation and endocannabinoid-mediated analgesia. J Neurosci 32:9457-68
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Rahn, Elizabeth J; Zvonok, Alexander M; Makriyannis, Alexandros et al. (2010) Antinociceptive effects of racemic AM1241 and its chirally synthesized enantiomers: lack of dependence upon opioid receptor activation. AAPS J 12:147-57
Guindon, Josée; Hohmann, Andrea G (2009) The endocannabinoid system and pain. CNS Neurol Disord Drug Targets 8:403-21
Eisenstein, Sarah A; Holmes, Philip V; Hohmann, Andrea G (2009) Endocannabinoid modulation of amphetamine sensitization is disrupted in a rodent model of lesion-induced dopamine dysregulation. Synapse 63:941-50
Nyilas, Rita; Gregg, Laura C; Mackie, Ken et al. (2009) Molecular architecture of endocannabinoid signaling at nociceptive synapses mediating analgesia. Eur J Neurosci 29:1964-78
Astarita, Giuseppe; Piomelli, Daniele (2009) Lipidomic analysis of endocannabinoid metabolism in biological samples. J Chromatogr B Analyt Technol Biomed Life Sci 877:2755-67
Rahn, Elizabeth J; Hohmann, Andrea G (2009) Cannabinoids as pharmacotherapies for neuropathic pain: from the bench to the bedside. Neurotherapeutics 6:713-37
Clapper, Jason R; Mangieri, Regina A; Piomelli, Daniele (2009) The endocannabinoid system as a target for the treatment of cannabis dependence. Neuropharmacology 56 Suppl 1:235-43

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