Mast cell extracellular trap formation underlies vascular and neural injury and hyperalgesia in sickle cell disease.

Sickle cell disease (SCD) is the most common inherited monogenetic disorder. Chronic and acute pain are hallmark features of SCD involving neural and vascular injury and inflammation. Mast cells reside in the vicinity of nerve fibers and vasculature, but how they influence these structures remains unknown. We therefore examined the mechanism of mast cell activation in a sickle microenvironment replete with cell-free heme and inflammation. Mast cells exposed to this environment showed an explosion of nuclear contents with the release of citrullinated histones, suggestive of mast cell extracellular trap (MCET) release. MCETs interacted directly with the vasculature and nerve fibers, a cause of vascular and neural injury in sickle cell mice. MCET formation was dependent upon peptidylarginine deiminase 4 (PAD4). Inhibition of PAD4 ameliorated vasoocclusion, chronic and acute hyperalgesia, and inflammation in sickle mice. PAD4 activation may also underlie neutrophil trap formation in SCD, thus providing a novel target to treat the sequelae of vascular and neural injury in SCD.

Red Blood Cells as Therapeutic Target to Treat Sickle Cell Disease.

Significance: Sickle cell disease (SCD) is the most common inherited diathesis affecting mostly underserved populations globally. SCD is characterized by chronic pain and fatigue, severe acute painful crises requiring hospitalization and opioids, strokes, multiorgan damage, and a shortened life span. Symptoms may appear shortly after birth, and, in less developed countries, most children with SCD die before attaining age 5. Hematopoietic stem cell transplant and gene therapy offer a curative therapeutic approach, but, due to many challenges, are limited in their availability and effectiveness for a majority of persons with SCD. A critical unmet need is to develop safe and effective novel targeted therapies. A wide array of drugs currently undergoing clinical investigation hold promise for an expanded pharmacological armamentarium against SCD. Recent Advances: Hydroxyurea, the most widely used intervention for SCD management, has improved the survival in the Western world and more recently, voxelotor (R-state-stabilizer), l-glutamine, and crizanlizumab (anti-P-selectin antibody) have been approved by the Food and Drug Administration (FDA) for use in SCD. The recent FDA approval emphasizes the need to revisit the advances in understanding the core pathophysiology of SCD to accelerate novel evidence-based strategies to treat SCD. The biomechanical breakdown of erythrocytesis, the core pathophysiology of SCD, is associated with intrinsic factors, including the composition of hemoglobin, membrane integrity, cellular volume, hydration, andoxidative stress. Critical Issues and Future Directions: In this context, this review focuses on advances in emerging nongenetic interventions directed toward the therapeutic targets intrinsic to sickle red blood cells (RBCs), which can prevent impaired rheology of RBCs to impede disease progression and reduce the sequelae of comorbidities, including pain, vasculopathy, and organ damage. In addition, given the intricate pathophysiology of the disease, it is unlikely that a single pharmacotherapeutic intervention will comprehensively ameliorate the multifaceted complications associated with SCD. However, the availability of multiple drug options affords the opportunity for individualized therapeutic regimens tailored to specific SCD-related complications. Furthermore, it opens avenues for combination drug therapy, capitalizing on distinct mechanisms of action and profiles of adverse effects.

Pain in Hemophilia: Unexplored Role of Oxidative Stress.

Hemophilia is the most common X-linked bleeding diathesis caused by the genetic deficiency of coagulation factors VIII or IX. Despite treatment advances and improvements in clinical management to prevent bleeding, management of acute and chronic pain remains to be established. Repeated bleeding of the joints leads to arthropathy, causing pain in hemophilia. However, mechanisms underlying the pathogenesis of pain in hemophilia remain underexamined. Herein, we describe the novel perspectives on the role for oxidative stress in the periphery and the central nervous system that may contribute to pain in hemophilia. Specifically, we cross examine preclinical and clinical studies that address the contribution of oxidative stress in hemophilia and related diseases that affect synovial tissue to induce acute and potentially chronic pain. This understanding would help provide potential treatable targets using antioxidants to ameliorate pain in hemophilia.

Effects of Selective Breeding, Voluntary Exercise, and Sex on Endocannabinoid Levels in the Mouse Small-Intestinal Epithelium.

The endocannabinoid (eCB) system in the gut communicates with the body and brain as part of the homeostatic mechanisms that affect energy balance. Although perhaps best known for its effects on energy intake, the eCB system also regulates voluntary locomotor behavior. Here, we examined gut eCB concentrations in relation to voluntary exercise, specifically in mice selectively bred for high voluntary wheel running behavior. We measured gut eCBs in four replicate non-selected Control (C) lines and four replicate lines of High Runner (HR) mice that had been selectively bred for 74 generations based on the average number of wheel revolutions on days 5 and 6 of a 6-day period of wheel access when young adults. On average, mice from HR lines run voluntarily on wheels ∼3-fold more than C mice on a daily basis. A recent study showed that circulating levels of primary endocannabinoids 2-arachidonoyl-sn-glycerol (2-AG) and anandamide (AEA) are altered by six days of wheel access, by acute wheel running, and differ between HR and C mice in sex-specific ways [1]. We hypothesized that eCBs in the upper small-intestinal epithelium (i.e., proximal jejunum), a region firmly implicated in eCB signaling, would differ between HR and C mice (linetype), between the sexes, between mice housed with vs. without wheels for six days, and would covary with amounts of acute running and/or home-cage activity (during the previous 30 minutes). We used the same 192 mice as in [1] , half males and half females, half HR and half C (all 8 lines), and half either given or not given access to wheels for six days. We assessed the eCBs, 2-AG and AEA, and their analogs docosahexaenoylglycerol (DHG), docosahexaenoylethanolamide (DHEA), and oleoylethanolamide (OEA). Both 2-AG and DHG showed a significant 3-way interaction of linetype, wheel access, and sex. In addition, HR mice had lower concentrations of 2-AG in the small-intestinal epithelium when compared to C mice, which may be functionally related to differences in locomotor activity or to differences in body composition and/or food consumption. Moreover, the amount of home-cage activity during the prior 30 min was a negative predictor of 2-AG and AEA concentrations in jejunum mucosa, particularly in the mice with no wheel access. Lastly, 2-AG, but not AEA, was significantly correlated with 2-AG in plasma in the same mice.

Increased 2-arachidonoyl-sn-glycerol levels normalize cortical responses to sound and improve behaviors in Fmr1 KO mice.

BACKGROUND: Individuals with Fragile X syndrome (FXS) and autism spectrum disorder (ASD) exhibit an array of symptoms, including sociability deficits, increased anxiety, hyperactivity, and sensory hyperexcitability. It is unclear how endocannabinoid (eCB) modulation can be targeted to alleviate neurophysiological abnormalities in FXS as behavioral research reveals benefits to inhibiting cannabinoid (CB) receptor activation and increasing endocannabinoid ligand levels. Here, we hypothesize that enhancement of 2-arachidonoyl-sn-glycerol (2-AG) in Fragile X mental retardation 1 gene knock-out (Fmr1 KO) mice may reduce cortical hyperexcitability and behavioral abnormalities observed in FXS. METHODS: To test whether an increase in 2-AG levels normalized cortical responses in a mouse model of FXS, animals were subjected to electroencephalography (EEG) recording and behavioral assessment following treatment with JZL-184, an irreversible inhibitor of monoacylglycerol lipase (MAGL). Assessment of 2-AG was performed using lipidomic analysis in conjunction with various doses and time points post-administration of JZL-184. Baseline electrocortical activity and evoked responses to sound stimuli were measured using a 30-channel multielectrode array (MEA) in adult male mice before, 4 h, and 1 day post-intraperitoneal injection of JZL-184 or vehicle. Behavior assessment was done using the open field and elevated plus maze 4 h post-treatment. RESULTS: Lipidomic analysis showed that 8 mg/kg JZL-184 significantly increased the levels of 2-AG in the auditory cortex of both Fmr1 KO and WT mice 4 h post-treatment compared to vehicle controls. EEG recordings revealed a reduction in the abnormally enhanced baseline gamma-band power in Fmr1 KO mice and significantly improved evoked synchronization to auditory stimuli in the gamma-band range post-JZL-184 treatment. JZL-184 treatment also ameliorated anxiety-like and hyperactivity phenotypes in Fmr1 KO mice. CONCLUSIONS: Overall, these results indicate that increasing 2-AG levels may serve as a potential therapeutic approach to normalize cortical responses and improve behavioral outcomes in FXS and possibly other ASDs.

UPLC-MS/MS Method for Analysis of Endocannabinoid and Related Lipid Metabolism in Mouse Mucosal Tissue.

The endocannabinoid system is expressed in cells throughout the body and controls a variety of physiological and pathophysiological functions. We describe robust and reproducible UPLC-MS/MS-based methods for analyzing metabolism of the endocannabinoids, 2-arachidonoyl-sn-glycerol and arachidonoyl ethanolamide, and related monoacylglycerols (MAGs) and fatty acid ethanolamides (FAEs), respectively, in mouse mucosal tissues (i.e., intestine and lung). These methods are optimized for analysis of activity of the MAG biosynthetic enzyme, diacylglycerol lipase (DGL), and MAG degradative enzymes, monoacylglycerol lipase (MGL) and alpha/beta hydrolase domain containing-6 (ABHD6). Moreover, we describe a novel UPLC-MS/MS-based method for analyzing activity of the FAE degradative enzyme, fatty acid amide hydrolase (FAAH), that does not require use of radioactive substrates. In addition, we describe in vivo pharmacological methods to inhibit MAG biosynthesis selectively in the mouse small-intestinal epithelium. These methods will be useful for profiling endocannabinoid metabolism in rodent mucosal tissues in health and disease.

Impact of Circulating N-Acylethanolamine Levels with Clinical and Laboratory End Points in Hemodialysis Patients.

BACKGROUND: Patients with ESRD on maintenance hemodialysis (MHD) are particularly susceptible to dysregulation of energy metabolism, which may manifest as protein energy wasting and cachexia. In recent years, the endocannabinoid system has been shown to play an important role in energy metabolism with potential relevance in ESRD. N-acylethanolamines are a class of fatty acid amides which include the major endocannabinoid ligand, anandamide, and the endogenous peroxisome proliferator-activated receptor-α agonists, oleoylethanolamide (OEA) and palmitoylethanolamide (PEA). METHODS: Serum concentrations of OEA and PEA were measured in MHD patients and their correlations with various clinical/laboratory indices were examined. Secondarily, we evaluated the association of circulating PEA and OEA levels with 12-month all-cause mortality. RESULTS: Both serum OEA and PEA levels positively correlated with high-density lipoprotein-cholesterol levels and negatively correlated with body fat and body anthropometric measures. Serum OEA levels correlated positively with serum interleukin-6 (IL-6) (rho = 0.19; p = 0.004). Serum PEA and IL-6 showed a similar but nonsignificant trend (rho = 0.12; p = 0.07). Restricted cubic spline analyses showed that increasing serum OEA and PEA both trended toward higher mortality risk, and these associations were statistically significant for PEA (PEA ≥4.7 pmol/mL; reference: PEA <4.7 pmol/mL) after adjustments in a Cox model (hazard ratio 2.99; 95% confidence interval 1.04, 8.64). CONCLUSIONS: In MHD patients, OEA and PEA are significantly correlated with variables related to lipid metabolism and body mass. Additionally, higher serum levels of PEA are associated with mortality risk. Future studies are needed to examine the potential mechanisms responsible for these findings and their clinical implications.

ß-endorphin at the intersection of pain and cancer progression: Preclinical evidence.

We examined the association between endogenous opioid ß-endorphin, cancer progression and pain in a transgenic mouse model of breast cancer, with a rat C3(1) simian virus 40 large tumor antigen fusion gene (C3TAg). C3TAg mice develop ductal epithelial atypia at 8 weeks, progression to intra-epithelial neoplasia at 12 weeks, and invasive carcinoma with palpable tumors at 16 weeks. Consistent with invasive carcinoma at 4 months of age, C3TAg mice demonstrate a significant increase in hyperalgesia compared to younger C3TAg or control FVBN mice without tumors. Our data show that the growing tumor contributes to circulating ß-endorphin. As an endogenous ligand of mu opioid receptor, ß-endorphin has analgesic activity. Paradoxically, we observed an increase in pain in transgenic breast cancer mice with significantly high circulating and tumor-associated ß-endorphin. Increased circulating ß-endorphin correlates with increasing tumor burden. ß-endorphin induced the activation of mitogenic and survival-promoting signaling pathways, MAPK/ERK 1/2, STAT3 and Akt, observed by us in human MDA-MB-231 cells suggesting a role for ß-endorphin in breast cancer progression and associated pain.

Considerations for Cannabis Use to Treat Pain in Sickle Cell Disease.

Pain in Sickle Cell Disease (SCD) is a major comorbidity and unique with acute pain due to recurrent and episodic vaso-occlusive crises as well as chronic pain, which can span an individual's entire life. Opioids are the mainstay treatment for pain in SCD. Due to recent health crises raised by adverse effects including deaths from opioid use, pain management in SCD is adversely affected. Cannabis and its products are most widely used for pain in multiple conditions and also by patients with SCD on their own. With the availability of "Medical Cannabis" and approval to use cannabis as medicine across majority of States in the United States as well as over-the-counter preparations, cannabis products are being used increasingly for SCD. The reliability of many of these products remains questionable, which poses a major health risk to the vulnerable individuals seeking pain relief. Therefore, this review provides up to date insights into available categories of cannabis-based treatment strategies, their mechanism of action and pre-clinical and clinical outcomes in SCD. It provides evidence for the benefits and risks of cannabis use in SCD and cautions about the unreliable and unvalidated products that may be adulterated with life-threatening non-cannabis compounds.

Spatiotemporal Alterations in Gait in Humanized Transgenic Sickle Mice.

Sickle cell disease (SCD) is a hemoglobinopathy affecting multiple organs and featuring acute and chronic pain. Purkinje cell damage and hyperalgesia have been demonstrated in transgenic sickle mice. Purkinje cells are associated with movement and neural function which may influence pain. We hypothesized that Purkinje cell damage and/or chronic pain burden provoke compensatory gait changes in sickle mice. We found that Purkinje cells undergoe increased apoptosis as shown by caspase-3 activation. Using an automated gait measurement system, MouseWalker, we characterized spatiotemporal gait characteristics of humanized transgenic BERK sickle mice in comparison to control mice. Sickle mice showed alteration in stance instability and dynamic gait parameters (walking speed, stance duration, swing duration and specific swing indices). Differences in stance instability may reflect motor dysfunction due to damaged Purkinje cells. Alterations in diagonal and all stance indices indicative of hesitation during walking may originate from motor dysfunction and/or arise from fear and/or anticipation of movement-evoked pain. We also demonstrate that stance duration, diagonal swing indices and all stance indices correlate with both mechanical and deep tissue hyperalgesia, while stance instability correlates with only deep tissue hyperalgesia. Therefore, objective analysis of gait in SCD may provide insights into neurological impairment and pain states.

Maternal transfer of environmentally relevant polybrominated diphenyl ethers (PBDEs) produces a diabetic phenotype and disrupts glucoregulatory hormones and hepatic endocannabinoids in adult mouse female offspring.

Polybrominated diphenyl ethers (PBDEs) are brominated flame retardant chemicals and environmental contaminants with endocrine-disrupting properties that are associated with diabetes and metabolic syndrome in humans. However, their diabetogenic actions are not completely characterized or understood. In this study, we investigated the effects of DE-71, a commercial penta-mixture of PBDEs, on glucoregulatory parameters in a perinatal exposure model using female C57Bl/6 mice. Results from in vivo glucose and insulin tolerance tests and ex vivo analyses revealed fasting hyperglycemia, glucose intolerance, reduced sensitivity and delayed glucose clearance after insulin challenge, decreased thermogenic brown adipose tissue mass, and exaggerated hepatic endocannabinoid tone in F1 offspring exposed to 0.1 mg/kg DE-71 relative to control. DE-71 effects on F0 dams were more limited indicating that indirect exposure to developing offspring is more detrimental. Other ex vivo glycemic correlates occurred more generally in exposed F0 and F1, i.e., reduced plasma insulin and altered glucoregulatory endocrines, exaggerated sympathoadrenal activity and reduced hepatic glutamate dehydrogenase enzymatic activity. Hepatic PBDE congener analysis indicated maternal transfer of BDE-28 and -153 to F1 at a collective level of 200 ng/g lipid, in range with maximum values detected in serum of human females. Given the persistent diabetogenic phenotype, especially pronounced in female offspring after developmental exposure to environmentally relevant levels of DE-71, additional animal studies should be conducted that further characterize PBDE-induced diabetic pathophysiology and identify critical developmental time windows of susceptibility. Longitudinal human studies should also be conducted to determine the risk of long-lasting metabolic consequences after maternal transfer of PBDEs during early-life development.

Cannabinoid CB1 Receptors in the Intestinal Epithelium Are Required for Acute Western-Diet Preferences in Mice.

The endocannabinoid system plays an important role in the intake of palatable food. For example, endocannabinoid signaling in the upper small-intestinal epithelium is increased (i) in rats after tasting dietary fats, which promotes intake of fats, and (ii) in a mouse model of diet-induced obesity, which promotes overeating via impaired nutrient-induced gut-brain satiation signaling. We now utilized a combination of genetic, pharmacological, and behavioral approaches to identify roles for cannabinoid CB1Rs in upper small-intestinal epithelium in preferences for a western-style diet (WD, high-fat/sucrose) versus a standard rodent diet (SD, low-fat/no sucrose). Mice were maintained on SD in automated feeding chambers. During testing, mice were given simultaneous access to SD and WD, and intakes were recorded. Mice displayed large preferences for the WD, which were inhibited by systemic pretreatment with the cannabinoid CB1R antagonist/inverse agonist, AM251, for up to 3 h. We next used our novel intestinal epithelium-specific conditional cannabinoid CB1R-deficient mice (IntCB1-/-) to investigate if intestinal CB1Rs are necessary for WD preferences. Similar to AM251 treatment, preferences for WD were largely absent in IntCB1-/- mice when compared to control mice for up to 6 h. Together, these data suggest that CB1Rs in the murine intestinal epithelium are required for acute WD preferences.

Integrative approaches to treating pain in sickle cell disease: Pre-clinical and clinical evidence.

Sickle cell disease (SCD) is a genetic disorder characterized by hemolysis, end-organ damage, inflammation, and pain. Recurrent and unpredictable episodes of acute pain due to vaso-occlusive crises are a unique feature of SCD. Many patients also develop lifelong chronic pain. Opioids are the primary method of pain treatment in SCD; however, continued use is associated with several adverse effects. Integrative approaches to treating pain in SCD are increasingly being explored to prevent the side effects associated with opioids. In this review, we highlight the mechanisms of pain in SCD and describe mechanism-based integrative approaches for treating pain.

A Balanced Approach for Cannabidiol Use in Chronic Pain.

Cannabidiol (CBD), the major non-psychoactive constituent of Cannabis sativa L., has gained traction as a potential treatment for intractable chronic pain in many conditions. Clinical evidence suggests that CBD provides therapeutic benefit in certain forms of epilepsy and imparts analgesia in certain conditions, and improves quality of life. CBD continues to be Schedule I or V on the list of controlled substances of the Drug Enforcement Agency of the United States. However, preparations labeled CBD are available publicly in stores and on the streets. However, use of CBD does not always resolve pain. CBD purchased freely entails the risk of adulteration by potentially hazardous chemicals. As well, CBD use by pregnant women is rising and poses a major health-hazard for future generations. In this mini-review, we present balanced and unbiased pre-clinical and clinical findings for the beneficial effects of CBD treatment on chronic pain and its deleterious effects on prenatal development.

Bile acid composition regulates GPR119-dependent intestinal lipid sensing and food intake regulation in mice.

OBJECTIVES: Lipid mediators in the GI tract regulate satiation and satiety. Bile acids (BAs) regulate the absorption and metabolism of dietary lipid in the intestine, but their effects on lipid-regulated satiation and satiety are completely unknown. Investigating this is challenging because introducing excessive BAs or eliminating BAs strongly impacts GI functions. We used a mouse model (Cyp8b1-/- mice) with normal total BA levels, but alterations in the composition of the BA pool that impact multiple aspects of intestinal lipid metabolism. We tested two hypotheses: BAs affect food intake by (1) regulating production of the bioactive lipid oleoylethanolamide (OEA), which enhances satiety; or (2) regulating the quantity and localisation of hydrolysed fat in small intestine, which controls gastric emptying and satiation. DESIGN: We evaluated OEA levels, gastric emptying and food intake in wild-type and Cyp8b1-/- mice. We assessed the role of the fat receptor GPR119 in these effects using Gpr119-/- mice. RESULTS: Cyp8b1-/- mice on a chow diet showed mild hypophagia. Jejunal OEA production was blunted in Cyp8b1-/- mice, thus these data do not support a role for this pathway in the hypophagia of Cyp8b1-/- mice. On the other hand, Cyp8b1 deficiency decreased gastric emptying, and this was dependent on dietary fat. GPR119 deficiency normalised the gastric emptying, gut hormone levels, food intake and body weight of Cyp8b1-/- mice. CONCLUSION: BAs regulate gastric emptying and satiation by determining fat-dependent GPR119 activity in distal intestine.

Circulating Endocannabinoids and Mortality in Hemodialysis Patients.

BACKGROUND: Mortality in patients with end-stage renal disease (ESRD) on maintenance hemodialysis (MHD) remains exceptionally high. While traditional risk factors such as obesity are paradoxically associated with better survival, nontraditional risk factors including cachexia increase the likelihood of poor outcomes. There is accumulating evidence that the endocannabinoid (ECB) system plays a major role in energy preservation and storage, factors which can prevent the deleterious effects of cachexia. Hence, in this study, we evaluated the association of circulating ECB levels with mortality in MHD patients. METHODS: Serum concentrations of anandamide (AEA) and 2-arachidonoyl-sn-glycerol (2-AG), major ECB ligands, were measured in MHD patients. Their correlation with various clinical/laboratory indices and association with 12-month all-cause mortality were examined. RESULTS: Serum 2-AG levels positively correlated with body mass index, serum triglycerides and body anthropometric measures. Meanwhile, serum AEA levels correlated positively with serum interleukin-6, and negatively with serum very low-density lipoprotein levels. While increased serum 2-AG levels were associated with reduced risk of all-cause mortality (hazard ratio [HR] 0.52, 95% CI 0.28-0.98), there was no clear association between serum AEA levels and mortality (HR 0.91, 95% CI 0.48-1.72). CONCLUSIONS: In MHD patients, the circulating levels of ECB ligand, 2-AG, may play an important role in determining body mass and risk of mortality. These observations were unique to 2-AG as similar findings were not obtained with serum AEA. Future studies need to investigate the mechanisms responsible for these associations and examine the modulation of the ECB system as a potential target for therapy in ESRD.

Serum Endocannabinoid Levels in Patients With End-Stage Renal Disease.

CONTEXT: Previous studies have shown that the endocannabinoid system plays a major role in energy metabolism through the actions of its main mediators, 2-arachidonoyl-sn-glycerol (2-AG) and anandamide (AEA). OBJECTIVE: We examined serum levels of major endocannabinoid mediators and their association with clinical parameters in patients with end-stage renal disease (ESRD). DESIGN AND SETTING: Serum concentrations of 2-AG and AEA were measured in patients on maintenance hemodialysis (MHD) and controls, and correlations with various clinical and laboratory indices were examined. 2-AG was also measured in age and sex-matched healthy subjects for comparison of levels in patients undergoing MHD. MAIN OUTCOME MEASURE: Serum 2-AG. RESULTS: Serum 2-AG levels were significantly elevated in patients with ESRD compared with healthy controls. Higher levels of 2-AG were found in patients on MHD compared to healthy subjects, and similar findings were seen in a second set of subjects in independent analyses. Among 96 patients on MHD, 2-AG levels correlated significantly and positively with serum triglycerides (ρ = 0.43; P < 0.0001), body mass index (ρ = 0.40; P < 0.0001), and body anthropometric measures and negatively with serum high-density lipoprotein cholesterol (ρ = -0.33; P = 0.001) following adjustment for demographic and clinical variables. CONCLUSIONS: In patients on MHD, levels of serum 2-AG, a major endocannabinoid mediator, were increased. In addition, increasing serum 2-AG levels correlated with increased serum triglycerides and markers of body mass. Future studies will need to evaluate the potential mechanisms responsible for these findings.

Cannabinoid CB1 Receptors Inhibit Gut-Brain Satiation Signaling in Diet-Induced Obesity.

Gut-brain signaling controls feeding behavior and energy homeostasis; however, the underlying molecular mechanisms and impact of diet-induced obesity (DIO) on these pathways are poorly defined. We tested the hypothesis that elevated endocannabinoid activity at cannabinoid CB1 receptor (CB1Rs) in the gut of mice rendered DIO by chronic access to a high fat and sucrose diet for 60 days inhibits nutrient-induced release of satiation peptides and promotes overeating. Immunoreactivity for CB1Rs was present in enteroendocrine cells in the mouse's upper small-intestinal epithelium that produce and secrete the satiation peptide, cholecystokinin (CCK), and expression of mRNA for CB1Rs was greater in these cells when compared to non-CCK producing cells. Oral gavage of corn oil increased levels of bioactive CCK (CCK-8) in plasma from mice fed a low fat no-sucrose diet. Pretreatment with the cannabinoid receptor agonist, WIN55,212-2, blocked this response, which was reversed by co-administration with the peripherally-restricted CB1R neutral antagonist, AM6545. Furthermore, monoacylglycerol metabolic enzyme function was dysregulated in the upper small-intestinal epithelium from DIO mice, which was met with increased levels of a variety of monoacylglycerols including the endocannabinoid, 2-arachidonoyl-sn-glycerol. Corn oil failed to affect levels of CCK in DIO mouse plasma; however, pretreatment with AM6545 restored the ability for corn oil to stimulate increases in levels of CCK, which suggests that elevated endocannabinoid signaling at small intestinal CB1Rs in DIO mice inhibits nutrient-induced CCK release. Moreover, the hypophagic effect of AM6545 in DIO mice was reversed by co-administration with the CCKA receptor antagonist, devazepide. Collectively, these results provide evidence that hyperphagia associated with DIO is driven by a mechanism that includes CB1R-mediated inhibition of gut-brain satiation signaling.