Projects

In the past year, funding agencies in the US have asked that we researchers add a section to our CV called "Contribution to Science".  While many of us (myself included) grumbled about this change, I think it is fair to say that we also were able to use this as a tool to summarize and focus the type of work that we have done and are currently doing in our research programs. I've provided a recent update of this section here to give you a glimpse into some of these contributions.  

 

Hormonal contributions to neuronal processing and behavioral changes to noxious stimuli

During PhD studies with Karen Berkley at Florida State University I used behavioral and electrophysiological methods to answer questions about how hormones drive changes in these parameters. These data showed that changes in the hormonal milieu drive changes in neuronal and behavioral responses to noxious visceral stimuli; however, the molecular mechanism for these modifications were unknown.  I have continued to build open this work to understand some of the molecular mechanisms of how hormones play a role in lipid signaling and have shown that levels of endogenous cannabinoids in the brain change across the hormonal cycle, are different in males and females, and are regulated during mating. 

    

1)     Bradshaw, HB, Temple, JL, Wood, E, Berkley KJ.  Estrous variations in behavioral responses to vaginal and uterine distention in the rat.  Pain. 1999 Aug;82(2):187-97.

2)     Bradshaw HB, Berkley KJ. Estrous changes in responses of rat gracile nucleus neurons to stimulation of skin and pelvic viscera.  J Neurosci. 2000 Oct 15;20(20):7722-7727.

3)     Bradshaw HB, Rimmerman N, Krey JF, Walker JM. Sex and hormonal cycle differences in brain levels of pain-related cannabimimetic lipid mediators. Am J Physiol Regul Integr Comp Physiol. 2006 Aug;291(2):R349-58. Epub 2006 Mar 23

4)     Stuart JM, Paris J, Frye C, Bradshaw HB.  Brain levels of prostaglandins, endocannabinoids and related lipids are affected by mating strategies. 2013 International Journal of Endocrinology Volume 2013, Article ID 436252, 14 pages, http://dx.doi.org/10.1155/2013/436252

 

Analysis of endogenous cannabinoids in a wide variety of model systems

Post-doctoral training in the lab of J Michael Walker centered on the development of lipidomics mass spectrometric techniques for the endogenous cannabinoids.  I have become a world expert on measuring these endogenous cannabinoids in all tissue types.  These lipids are routinely measured in my lab from small volumes of brain tissue, retina, bone, cartilage, lung, liver, kidney, uterus, placenta, skin, and plasma.  The ability to modify extraction and analysis techniques has allowed me to work with colleagues from all over the world and in different model systems.  This provides both a unique perspective as well as a growing data set with which to mine for systems biology.   

 

1)   Guindon J, Lai Y, Takacs SM, Bradshaw HB, Hohmann AG.  Alterations in endocannabinoid tone following chemotherapy-induced peripheral neuropathy: Effects of endocannabinoid deactivation inhibitors targeting fatty-acid amide hydrolase and monoacylglycerol lipase in comparison to reference analgesics following cisplatin treatment.  Pharmacol Res. 2013 Jan;67(1):94-109. doi: 10.1016/j.phrs.2012.10.013.

2)   Cha J, Bartos A, Egashira M, Haraguchi H, Saito-Fujita T, Leishman E, Bradshaw H, Dey SK, Hirota Y. Combinatory approaches prevent preterm birth profoundly exacerbated by gene-environment interactions. 2013 J Clin Invest. Sep 3;123(9):4063-75. doi: 10.1172/JCI70098. Epub 2013 Aug 27

3)   Crowe MS, Leishman E, Banks ML, Gujjar R, Mahadevan A, Bradshaw HB, Kinsey SG.  Dual inhibition of monoacylglycerol lipase and cyclooxygenases synergistically reduces neuropathic pain in mice. Br J Pharmacol. 2014 Nov 13. doi: 10.1111/bph.13012. PMID: 25393148

4)   Wolfson ML, Correa F, Leishman E, Vercelli C, Cymeryng C, Blanco J, Bradshaw HB, Franchi AM. Lipopolysaccharide-induced murine embryonic resorption involves changes in endocannabinoid profiling and alters progesterone secretion and inflammatory response by a CB1-mediated fashion. Mol Cell Endocrinol. 2015 Aug 15;411:214-22. doi: 10.1016/j.mce.2015.04.032. Epub 2015 May 6.  PMID: 25958042

 

Lipidomics mass spectrometric techniques as a tool for identifying novel lipids and signaling systems

Developing a variety of extraction and mass spectrometric paved the way for the discovery of other structurally similar endogenous lipids.  This growing class of lipids is derived from the conjugation of a fatty acid and an amine and is collectively called N-acyl amides or lipoamines.  To date, my group has identified and characterized over 80 of these novel lipids in the mammalian brain and other model systems such as drosophila.  I worked with collaborators at Hebrew University to discover a novel lipid; N-oleoyl serine, that plays a pivotal role in the maintenance of bone density and recently published that this lipid is present in high abundance in olive oil.  Recently, we also published the largest scale lipidomics analysis of the effects of the deletion of endogenous cannabinoid metabolic enzymes on the levels of these lipids in the brain.  These data show that endogenous cannabinoids are biochemically linked to potentially hundreds of lipids in the brain and body; therefore, the regulation of these by exogenous Cannabis compounds potentially has broader effects that ever realized. 

1)   Smoum R, Bar A, Tan B, Milman G, Attar-Namdar M, Ofek O, Stuart JM, Tam J, Kram V, O’Dell D, Walker MJ, Bradshaw HB, Bab I, Mechoulam R.  Oleoyl serine, an endogenous N-acyl amide, modulates bone remodeling and mass. Proc Natl Acad Sci U S A 2010 Oct 12

2)   Tortoriello G, Rhodes BP, Takacs SM, Stuart JM, Basnet A, Harkney T, Bradshaw HB. Targeted Lipidomics in Drosophila melanogaster Identifies Novel 2-Monoacylglycerols and N-acyl Amides. 2013 PLoS ONE 8(7): e67865. doi:10.1371/journal.pone.0067865

3)   Leishman E, Cornett B, Spork K, Straiker A, Mackie, K, and Bradshaw HB.  Broad impact of deleting endogenous cannabinoid hydrolyzing enzymes and the cannabinoid receptor CB1 on the endogenous cannabinoid-related lipidome in eight regions of the mouse brain. Pharmacol Res. 2016 Apr 22. pii: S1043-6618(16)30344-9. doi: 10.1016/j.phrs.2016.04.020. [Epub ahead of print]PMID: 27109320

4)   Leishman, E, Mackie, K, Luguet S, and Bradshaw HB.  Lipidomics profile of a NAPE-PLD KO mouse provides evidence of a broader role of this enzyme in lipid metabolism in the brain. Biochim Biophys Acta. 2016 Jun;1861(6):491-500. doi: 10.1016/j.bbalip.2016.03.003. Epub 2016 Mar 5.PMID: 26956082

 

Identification of a novel GPCR that is activated by THC as well as its endogenous ligands

Mass spec lipidomics is complemented in my lab with functional assays designed to determine the signaling role at GPCRs of these newly discovered lipids.  Through a series of functional assays including cellular migration, proliferation, MAPK, and immunohistochemistry we showed NAGly is an endogenous ligand that activates GPR18 and that this signaling system plays a role in both microglial and human endometrial migration through similar intracellular cascades.  In addition our data provided evidence that the phytocannabinoid, Cannabidiol and the endogenous lipid, N-arachidonyl serine, both act as antagonists at the GPR18 receptor.  Finally, our data also provided compelling evidence that the phytocannabinoid, ∆9 THC, is a potent ligand at GPR18, which mimics the signaling properties of NAGly.  These data have set the foundation for the identification of an additional cannabinoid receptor (GPR18) that will aid in the scientific understanding of how cannabis works in the brain and body.   

 

1)   McHugh D, Hu S S-J, Rimmerman N, Vogil Z., Walker JM, Bradshaw HB. N-arachidonoyl glycine, an abundant endogenous lipid, potently drives directed cellular migration through GPR18, the putative abnormal cannabidiol receptor. BMC Neurosci 2010 Mar 26;11:44

2)   McHugh D, Wager-Miller J, Page J, and Bradshaw HB. siRNA knockdown of GPR18 receptors in BV-2 microglia attenuates N-arachidonoyl glycine induced cell migration.  J Mol Signal. 2012 Jul 26;7(1):10.

3)   McHugh D, Page J, Dunn E, Bradshaw HB. Δ(9) -THC and N-arachidonoyl glycine are full agonists at GPR18 and cause migration in the human endometrial cell line, HEC-1B. Br J Pharmacol 2012 Apr;165(8):2414-24. doi: 10.1111/j.1476-5381.2011.01497.x. PMID: 21595653.

4)   McHugh D, Roskowski D, Xie S, and Bradshaw HB.  ∆9-THC and N-arachidonoyl glycine regulate BV-2 microglial morphology and cytokine release plasticity. Front Pharmacol. 2014 Jan 2;4:162. doi: 10.3389/fphar.2013.00162. eCollection 2014.PMID:24427137

Novel endogenous ligand identification of TRP receptors

An important emerging field of signaling is focused on the transient receptor potential (TRP) channels.  There are 28 known variants in mammalian species and the endogenous ligands for most are unknown.  Recently, we identified 20 lipoamines with activity at TRPV1-4 and showed that at least 8 of these were up or down regulated in an acute model of peripheral pain.  In collaboration with Sven Jordt we also showed that those that activate TRPV4 were present in lung and differentially regulated with edema.  Recently, we collaborated with Andrea Hohmann to discover that mice lacking the ability to metabolize many of these endogenous TRP activators had an elevated response to noxious stimuli.  This same enzyme metabolizes the endogenous cannabinoid, Anandamide, which illustrates how these signaling systems are interconnected.    

1)   Huang SM, Lee h, Chung M-K, Yu YY, Bradshaw HB, Coulombe PA, Walker JM, Caterina MJ. Overexpressed transient receptor potential vanilloid 3 ion channels in skin keratinocyte modulate pain sensitivity via prostaglandin E2.  J Neurosci 2008 Dec 17;28(51):13727-37

2)   Raboune S, Stuart JM, Leishman E, Takacs SM, Rhodes B, Basnet A, Jameyfield E, McHugh D, Widlanski T, Bradshaw HB.  Novel endogenous N-acyl amides activate TRPV1-4 receptors, BV-2 microglia, and are regulated in brain in an acute model of inflammation.  Front Cell Neurosci. 2014 Aug 1;8:195. doi: 10.3389/fncel.2014.00195. eCollection 2014

3)   Balakrishna S, Song W, Achanta S, Doran SF, Liu B, Kaelberer MM, Yu Z, Sui A, Cheung M, Leishman E, Eidam HS, Ye G, Willette RN, Thorneloe KS, Bradshaw HB, Matalon S, Jordt SE. TRPV4 inhibition counteracts edema and inflammation and improves pulmonary function and oxygen saturation in chemically induced acute lung injury.  Am J Physiol Lung Cell Mol Physiol. 2014 Jul 15;307(2):L158-72. doi: 10.1152/ajplung.00065.2014. Epub 2014 May 16

4)   Carry L, Slivicki R, Leishman, E, Bradshaw HB, Hohmann A.  A pro-nociceptive phenotype revealed in mice lacking the anandamide hydrolyzing enzyme fatty-acid amide hydroase.  Mol Pain. 2016 May 13;12. pii: 1744806916649192. Print 2016.

Comments