N-Acylethanolamines (NAEs) are bioactive acylamides which are involved in diverse biological functions in eukaryotes. Although NAEs are ubiquitous in plants and animals, occurrence of N-arachidonoylethanolamide (anandamide, AEA, NAE20:4) is limited to mammals and early land plants. Metabolism of NAEs and their functional implications in plants are yet to be fully discovered. Unlike seed plants, bryophytes possess unique fatty acid composition that includes abundance of polyunsaturated fatty acids such as arachidonic acid (AA, 20:4) and eicosapentaenoic acid (EPA, 20:5). Moss Physcomitrella patens contains ~18.7 and 15.9 % of AA in gametophores and protonemata, respectively. Therefore, it is hypothesized that P. patens may exhibit a unique NAE metabolite profile. To this extent, we performed lipid profiling and discovered long-chain NAEs and their corresponding N-acyl-phosphatidylethanolamine (NAPE) precursors in Physcomitrella and Selaginella. In protonemal tissues, N-arachidonyl-PE and N-20:5-PE contributed to about 49 % and 30 %, respectively. Matured gametophytes on the other hand showed a 12 % increase in N-20:4-PE and 20 % decline in N-20:5-PE, relative to NAPE content in protonemata. In all haploid developmental stages analyzed, NAE20:4 levels contributed to ~ 23 % of the total NAE while NAE 20:5 remained as a minor component (5 %). Interestingly, in Selaginella moellendorffi, an early vascular plant, N-18:2-PE species was most abundant; minor amounts of N-20:3-PE, N-20:4-PE and N-20:5-PE were also present with only a traceable quantity of NAE20:4. To understand biological implications of AEA, we examined the effects of exogenously applied AEA and its corresponding fatty acid (AA) on moss protonemata growth. Both AEA and AA inhibit growth of gametophytes and protonemata in a dose dependent manner. Additionally, we identified moss ortholog for NAPE-hydrolyzing phospholipase D (NAPE-PLD) enzyme that likely generates AEA. Putative PpNAPE-PLD has been expressed in E. coli for further characterization. Our data demonstrates the occurrence of evolutionarily conserved NAE metabolic pathway in the moss, with an exclusive occurrence of AEA. However, functional and evolutionary implications of this mammalian endocannabinoid in early land plants remain elusive.

N-Acylethanolamines (NAEs) including anandamide (NAE 20:4) are fatty acid ethanolamides generated by the hydrolysis of N-acylphoshotidylethanolamine (NAPE) by phospholipase D (PLD) and degraded by fatty acid amide hydrolase (FAAH). In mammals, ligands such as NAE 20:4 act through cannabinoid receptors and regulate several physiological processes like neuroprotection, pain perception, mental depression, and appetite suppression. In plants, NAE with chain length C12 to C18 are common and affect physiological processes such as cytoskeletal organization, endomembrane trafficking, cell wall and cell shape formation, seedling growth and response to stress. However, our recent identification of NAE 20:4 in moss, Physcomitrella patens prompted us to elucidate its metabolic pathway and physiological implications. We hypothesize that unique NAE metabolites such as anandamide in moss might play a role in rendering moss its ability to tolerate temperature, dehydration, salt and osmotic stress. To address the above hypothesis, three main objectives are being pursued using P patens. 1)Biochemical and molecular characterization of NAE metabolic pathway, 2) Generation and phenotypic characterization of NAE metabolite mutants, and 3) Elucidation of the physiological role of NAEs in abscisic acid-mediated dehydration tolerance. A NAPE-PLD, known to synthesize NAE 20:4 has been identified in mammals and FAAH in several eukaryotes, including plants. Here, identification and cloning of putative NAPE-PLD and FAAH genes that are likely involved in NAE synthesis and degradation, respectively, in P patens is discussed. Our long-term objective is to understand lipid-mediated stress responses in plants.

Pain is a common symptom of many clinical syndromes and diseases. In particular, the treatment of neuropathic pain represents a serious public health issue because currently available analgesia is ineffective in many cases or it has adverse effects. Treatment of pain-related suffering requires knowledge of how pain signals are initially generated and subsequently transmitted by the nervous system. A nociceptive system plays a key role in this process of encoding and transmission of pain signals. Modulation of the nociceptive synaptic transmission in the spinal cord dorsal horn represents an important mechanism in the development and maintenance of different pathological pain states. This doctoral thesis has aimed to investigate and clarify some of the mechanisms involved in the modulation of the spinal nociceptive processing in different pain states. The main attention was paid to study the following issues: (I.) Which is the role of Transient Receptor Potential Vanilloid type 1 channels (TRPV1), Toll-Like Receptors 4 (TLR4), and phosphatidylinositol 3-kinase (PI3K) in the development of neuropathic pain induced by paclitaxel (PAC) chemotherapy in acute in vitro, and subchronic in vivo murine model of PAC-induced peripheral neuropathy (PIPN)? (II.) How is affected spinal inhibitory synaptic control...