Spatial learning and memory in brain-injured and non-injured mice: investigating the roles of diacylglycerol lipase-α and -β.

A growing body of evidence implicates the importance of the endogenous cannabinoid 2-arachidonyl glycerol (2-AG) in memory regulation. The biosynthesis of 2-AG occurs primarily through the diacylglycerol lipases (DAGL-α and -β), with 2-AG serving as a bioactive lipid to both activate cannabinoid receptors and as a rate limiting precursor for the production of arachidonic acid and subsequent pro-inflammatory mediators. Gene deletion of DAGL-α shows decrements in synaptic plasticity and hippocampal neurogenesis suggesting this biosynthetic enzyme may be important for processes of normal spatial memory. Additionally, 2-AG is elevated in response to pathogenic events such as traumatic brain injury (TBI), suggesting its regulatory role may extend to conditions of neuropathology. As such, this dissertation investigates the in vivo role of DAGL-α and -β to regulate spatial learning and memory in the healthy brain and following neuropathology (TBI). The first part of this dissertation developed a mouse model of learning and memory impairment following TBI, using hippocampal-dependent tasks of the Morris water maze (MWM). We found modest, but distinct differences in MWM performance between left and right unilateral TBI despite similar motor deficits, histological damage, and glial reactivity. These findings suggest that laterality in mouse MWM deficit might be an important consideration when modeling TBI-induced functional consequences. The second part of this dissertation work evaluated DAGL-β as a target to protect against TBI-induced learning and memory deficit given its selective expression on microglia and the role of 2-AG as a precursor for eicosanoid production. The gene deletion of DAGL-β did not protect against TBI-induced MWM or motor deficits, but unexpectedly produced a survival protective phenotype. These findings suggest that while DAGL-β does not contribute to injury-induced memory deficit, it may contribute to TBI-induced mortality. The third and final set of experiments investigated the role of DAGL-α in mouse spatial learning and memory under physiological conditions (given the predominantly neuronal expression of DAGL-α). Complementary pharmacological and genetic manipulations produced task specific impaired MWM performance, as well as impaired long-term potentiation and alterations to endocannabinoid lipid levels. These results suggest that DAGL-α may play a selective role in the integration of new spatial information in the normal mouse brain. Overall, these data point to DAGL-α, but not DAGL-β, as an important contributor to hippocampal-dependent learning and memory. In contrast, DAGL-β may contribute to TBI-induced mortality

The Endogenous Cannabinoid System: A Budding Source of Targets for Treating Inflammatory and Neuropathic Pain

A great need exists for the development of new medications to treat pain resulting from various disease states and types of injury. Given that the endogenous cannabinoid (that is, endocannabinoid) system modulates neuronal and immune cell function, both of which play key roles in pain, therapeutics targeting this system hold promise as novel analgesics. Potential therapeutic targets include the cannabinoid receptors, type 1 and 2, as well as biosynthetic and catabolic enzymes of the endocannabinoids N-arachidonoylethanolamine and 2-arachidonoylglycerol. Notably, cannabinoid receptor agonists as well as inhibitors of endocannabinoid-regulating enzymes fatty acid amide hydrolase and monoacylglycerol lipase produce reliable antinociceptive effects, and offer opioid-sparing antinociceptive effects in myriad preclinical inflammatory and neuropathic pain models. Emerging clinical studies show that ‘medicinal’ cannabis or cannabinoid-based medications relieve pain in human diseases such as cancer, multiple sclerosis, and fibromyalgia. However, clinical data have yet to demonstrate the analgesic efficacy of inhibitors of endocannabinoid-regulating enzymes. Likewise, the question of whether pharmacotherapies aimed at the endocannabinoid system promote opioid-sparing effects in the treatment of pain reflects an important area of research. Here we examine the preclinical and clinical evidence of various endocannabinoid system targets as potential therapeutic strategies for inflammatory and neuropathic pain conditions

Liposomal Delivery of Diacylglycerol Lipase-Beta Inhibitors to Macrophages Dramatically Enhances Selectivity and Efficacy <i>in Vivo</i>

Diacylglycerol lipase-beta (DAGLβ) hydrolyzes arachidonic acid (AA)-containing diacylglycerols to produce bioactive lipids including endocannabinoids and AA-derived eicosanoids involved in regulation of inflammatory signaling. Previously, we demonstrated that DAGLβ inactivation using the triazole urea inhibitor KT109 blocked macrophage inflammatory signaling and reversed allodynic responses of mice in inflammatory and neuropathic pain models. Here, we tested whether we could exploit the phagocytic capacity of macrophages to localize delivery of DAGLβ inhibitors to these cells <i>in vivo</i> using liposome encapsulated KT109. We used DAGLβ-tailored activity-based probes and chemical proteomic methods to measure potency and selectivity of liposomal KT109 in macrophages and tissues from treated mice. Surprisingly, delivery of ∼5 μg of liposomal KT109 was sufficient to achieve ∼80% inactivation of DAGLβ in macrophages with no apparent activity in other tissues <i>in vivo</i>. Our macrophage-targeted delivery resulted in a >100-fold enhancement in antinociceptive potency compared with free compound in a mouse inflammatory pain model. Our studies describe a novel anti-inflammatory strategy that is achieved by targeted <i>in vivo</i> delivery of DAGLβ inhibitors to macrophages

Proposition of conception of using micro-additions for High Performance Concrete

The modern, contemporarily used cement composite types make use of various additives. This diploma thesis is focused on designs of mortars that have been enriched with micro and nano additives, which lead to higher mechanical strength through optimized grading of mortar mixes. This thesis also evaluates the influence of material properties on resulting properties of mortars in fresh and hardened state