Duration of each injection was 0.2 s and injection volume was 0.2 ml. Each injection was followed by a 60-s timeout period, during which the chamber was dark and lever presses had no programmed consequences. One-hour sessions were conducted five days per week . All monkeys had learned to respond under the FR10 schedule for the particular training drug prior to beginning this study. After completing the previous experiments, monkeys self administered the training dose of each drug for at least five sessions until responding was stable . Self administration behavior was then extinguished by substituting vehicle for THC, anandamide, or cocaine, but maintaining the presentation of the brief-stimulus associated with each injection. Then we tested reinstatement of extinguished drug-taking behavior by priming injections of THC or URB597 in all three groups of monkeys. Reinstatement effects of each pretreatment were studied for three consecutive sessions starting after at least three days of stable vehicle extinction. Food was withheld 12 h prior to this procedure. Anaesthesia was induced and maintained with isoflurane . Monkeys were weighed, prepared with a venous line , placed on a surgery table and kept warm by heat lamps.After animals were stabilized , URB597 or its vehicle was intravenously injected 1 h prior to euthanasia and the venous line was flushed with 0.5 ml of saline. Body temperature, SPO2,vertical agriculture pulse and rate of respiration were recorded every 10 min. After 1 h, monkeys were euthanized with Euthasol . Death was confirmed by the absence of respiration and heart beat on ECG. Brains were quickly removed, the cerebella were separated and forebrains were dissected. Each hemisphere was cut into 3 parts by two coronal sections made at approximately AP +12.5 and −5.
Brain fragments were snap-frozen in isopentane , wrapped in aluminium foil, and placed in dry ice. Samples were stored in the freezer for 2 days and then shipped on dry ice to the University of California Irvine for analyses. In one set of experiments, rats were sacrificed by rapid decapitation under light anaesthesia 2 h after injection of URB597 or vehicle. In other experiments, rats were food deprived for 12 h and sacrificed by decapitation 1 h after injection of URB597 or vehicle, while maintained under isoflurane anaesthesia for the duration of the experiment. In either case, brains were rapidly removed, and the hippocampus and prefrontal cortex was dissected from the fresh tissue over ice. Brain regions were frozen in dry ice, and stored at −80°C until lipid and enzymatic analyses. Cumulative-response records were obtained during all sessions to assess within-session patterns of responding. Rates of responding during self-administration sessions are expressed as responses per second averaged over the one-hour session, with responding during time-outs not included in calculations. Injections per session represent total number of injections delivered per session. Data for dose-effect curves are expressed as mean response rates and numbers of injections per session ± SEM over the last three sessions. In addition, total intake of anandamide, THC or cocaine for each session was calculated. Reinstatement data and effects of pretreatment with URB597 on drug self-administration are expressed as mean ± SEM of total numbers of injections per session over three sessions. Statistical analysis was done using single-factor repeated measures ANOVA to assess differences between vehicle and test-drug pretreatment conditions or between different doses of anandamide, THC, cocaine or URB597 and vehicle. Significant main effects were analyzed further by subsequent paired comparisons to control values using Dunnett’s test . Bonferroni t-test was used when the number of observations did not allow for the use of the Dunnett’s test. Differences between effects of vehicle and URB597 pretreatment on lipid levels and FAAH activity were analyzed using single-factor ANOVA.
Differences were considered statistically significant when p < 0.05.We found that the selective FAAH inhibitor URB597 suppresses FAAH activity and increases anandamide levels in regions of the squirrel monkey brain that participate in motivational, cognitive and emotional functions. This effect is accompanied by a marked decrease in the levels of 2-AG, a major endocannabinoid substance in the brain, even though URB597 does not affect activities of 2-AG-metabolizing enzymes such as DGL and MGL. We further observed that URB597 does not display overt reinforcing property in monkeys over a broad range of experimental conditions. Indeed, the drug did not reinforce self-administration behavior even when its cumulative intake exceeded by several folds a fully effective dose for FAAH inhibition. Furthermore, neither previous cocaine nor THC exposure predisposed monkeys to self-administer URB597: even monkeys that had previously self-administered anandamide at very high rates failed to respond to the FAAH inhibitor. Lastly, URB597 did not affect the reinforcing effects of THC or cocaine, and did not reinstate extinguished drug seeking behavior in monkeys that had previously self-administered THC or cocaine. We interpret these results to indicate that URB597, by enhancing anandamide signaling, causes a compensatory down-regulation in 2-AG mobilization; and the potentiation of anandamide-mediated transmission produced by URB597 is insufficient per se to produce reinforcing effects. Our findings further imply that FAAH inhibitors such as URB597 – which have demonstrated analgesic, anxiolytic, antidepressant and antihypertensive properties in rodents – may be used in humans without anticipated risk of inducing abuse or provoking relapse to drug use in abstinent individuals. The pharmacological profile of URB597 is strikingly different from that of THC and other direct-acting CB1 receptor agonists. Studies in rodents have shown that URB597 does not produce THC-like effects such as catalepsy, hypothermia or hyperphagia . Further, URB597 does not mimic the discriminative-stimulus actions of THC . Even further, URB597 does not increase dopamine levels in the nucleus accumbens shell of rats,hydroponics flood table a defining neurochemical feature of reinforcing drugs . Finally, URB597 does not elicit conditioned place preferences indicative of rewarding properties in rats .
However, experiments in rodents, such as those outlined above, are insufficient to model human reward based behaviors and to predict the addictive potential of drugs. Thus the present results provide the first unequivocal demonstration that URB597 lacks THC-like reinforcing properties, and suggest that this FAAH inhibitor might be used in therapy without anticipated risk of abuse or triggering relapse to drug use. Exogenous anandamide exerts potent reinforcing effects in monkeys . Thus, it may be surprising that the ability of URB597 to potentiate brain anandamide signaling does not translate into overt rewarding properties. However, there are two plausible reasons why URB597 does not support self-administration responding. First, exogenous and endogenous anandamide might each access distinct sub-populations of CB1 receptors in the brain. In particular, systemic administration could allow anandamide to reach a receptor pool that is normally engaged by 2-AG. In this context, the observation that treatment with URB597 decreases 2-AG levels in the monkey brain suggests the existence of a compensatory mechanism aimed at reducing 2-AG signaling in the face of enhanced anandamide signaling. Such a mechanism might account, at least in part, for the inability of URB597 to serve as a reinforcer. Consistent with this idea, a recent report suggests that pharmacological or genetic disruption of FAAH activity causes a down-regulation of 2-AG production in acutely dissected rodent striatal slices, which is reportedly due to vanilloid TRPV1 receptor activation . However, we were unable to replicate this observation in live animals even when using doses of URB597 that completely suppressed FAAH activity and significantly increased anandamide levels . Another possibility is that the kinetics of CB1 receptor activation may differ between anandamide and URB597 administration, as the former is likely to produce a more rapid recruitment of CB1 receptors than the latter. It is well established that effectiveness of drug reinforcement in monkeys depends on a rapid drug distribution throughout the brain . Irrespective of the mechanism involved, the impact of 2-AG down-regulation on the broad pharmacological properties of URB597 in primates remains to be determined. In conclusion, our findings with URB597 unmask a previously unsuspected functional heterogeneity within the endocannabinoid signaling system in the brain, and suggest that FAAH inhibitors such as URB597 might be used therapeutically without risk of abuse or triggering relapse to drug abuse. Feeding America is a network of food banks, food pantries, and meal programs providing food and services in the U.S.Second Harvest Heartland, a mid-western member of Feeding America, is one of the nation’s largest food banks, and supports over 1,000 food shelves and other partner programs that distribute food to over 532,000 individuals in Minnesota and western Wisconsin annually. Our institution and Second Harvest Heartland have partnered to provide food assistance to our patients since 2010. As a result of this relationship, healthy food is available in our clinics in the form of bagged groceries, and on site through an institutional food shelf. Patients are able to receive food through clinics, at discharge from an inpatient stay, or delivered to their homes through a community paramedic program or visiting nurses. To determine eligibility for food services referrals in our institution, staff or providers screen patients for food insecurity with two questions.
This two-question screening tool is a validated, abbreviated version of the 18-item U.S. Household Food Security Scale , which is used to monitor national food security.For purposes of our referral program, we modified the screening questions to be dichotomous yes/no responses from the validated responses: often, sometimes, or never. The questions included “Within the past 12 months we worried whether our food would run out before we got money to buy more;” and “Within the past 12 months the food we bought just didn’t last and we didn’t have money to get more.” In its validation cohort, an affirmative response to either question yielded a sensitivity of 97% and specificity of 83%, compared to the gold-standard, full 18-item HFSS. If deemed food insecure by this screening process, providers will then order a “referral for food” in the EMR to connect the patient to Second Harvest Heartland for support. The patient must consent to the referral and state what specific contact information they are comfortable sharing with the partner organization. This referral provides the patient’s contact information to Second Harvest Heartland through an automated fax. The food bank staff will then assist the patient in enrolling in federal nutrition programs , in addition to locating their neighborhood food shelves and meal programs, and arranging free produce distribution that they can access on a monthly basis. Though initially intended for clinic and inpatient use, starting in 2015 this order became available for use in the ED. To advertise the referral program, focused information sessions were added to the emergency medicine resident educational conferences in late 2015, and to the new resident orientation, starting in 2016. In addition to this, semi-annual updates are integrated into the resident conferences, and the details of the referral patterns are distributed to faculty. All ED personnel were encouraged to use the referral order, including ED faculty physicians, residents, physician assistants, nursing staff, social workers, ED registration, and financial support staff. Institutional financial counselors were unforeseen allies with the program, as their workflow typically incorporated several questions that touched on financial and food security issues. From January through December 2015 , a total of 1,003 referrals were made to Second Harvest Heartland; only five were made from the ED. From January 2016 through December 2016 , there were 1,519 referrals hospital-wide, and 55 referrals were made from the ED. Table 2 outlines details of the frequency of EMR order use from all clinical sites. Of the 1,519 referrals, 1,129 were successfully contacted by Second Harvest Heartland, and 954 accepted and received assistance. Of the referred and successfully contacted households, 92% were connected with at least one new form of food assistance. This assistance included new information about geographically individualized food shelves, meal sites, and produce distribution. Of households eligible for the Supplemental Nutrition Assistance Program, 76% completed applications to the federal entitlement program. This study sought to determine whether ED referrals for food resources for patients with food insecurity would increase after the implementation of an EMR referral order, as well as to introduce provider education about this referral program. To our knowledge, we report the first experience with such an institutional EMR order for food resources in the ED.