Defense Date


Document Type


Degree Name

Doctor of Philosophy


Pharmacology & Toxicology

First Advisor

Dr. Aron Lichtman

Second Advisor

Dr. S. Steve Negus

Third Advisor

Dr. M. Imad Damaj

Fourth Advisor

Dr. John Bigbee

Fifth Advisor

Dr. Egidio Del Fabbro


The endogenous cannabinoid system tightly regulates a myriad of physiological and pathophysiological functions. Effects of cannabinoids in ameliorating opioid withdrawal signs has been known for decades. Naloxone precipitates several withdrawal signs (e.g., diarrhea, weight loss, jumps, paw flutters, and head shakes) in opioid dependent rodents. ∆9-tetrahydrocannabinol (THC) has been shown to ameliorate opioid withdrawal in mice through the activation of cannabinoid type-1 (CB1) receptors. Inhibition of the endocannabinoid degradative enzymes also attenuates these withdrawal signs (i.e., diarrhea, weight loss, jumps, paw flutters, and head shakes) in opioid-dependent mice. Specifically, inhibition of fatty acid amide hydrolase (FAAH), the primary metabolic enzyme for the endocannabinoid N-arachidonoylethanolamide (AEA; anandamide), or inhibition of monoacylglycerol lipase (MAGL), the primary degradative metabolic enzyme for the endocannabinoid 2-arachidonoylglycerol (2-AG), reduces opioid withdrawal in mice. Notably, FAAH inhibition attenuates a subset of withdrawal signs (i.e., jumping and paw fluttering) but not diarrhea or weight loss, whereas full MAGL inhibition completely reduces withdrawal signs in mice. MAGL inhibition also produces similar acute cannabimimetic side effects as THC as well as cannabinoid dependence following repeated administration. Additionally, the combination of minimal MAGL inhibition with maximal FAAH inhibition results in a greater reduction of withdrawal signs than inhibiting each enzyme alone; however, mice trained in the drug discrimination paradigm exhibited full discrimination of a dual FAAH-MAGL inhibitor, SA-57, where the CB1 receptor agonist, CP55,940, fully substituted for SA-57 suggesting intrinsic subjective effects to a direct CB1 receptor orthosteric agonist. Thus, a selective approach targeting the CB1 receptor without cannabimimetic side effects is needed. CB1 receptor positive allosteric modulators (PAMs) target and bind to a topographically distinct site, i.e., an allosteric site, that is less conserved than the prototypical orthosteric site where THC, other synthetic cannabinoids (i.e., CP55,940), and the endocannabinoids, 2-AG and AEA, readily bind and elicit a conformational change that results in enhanced efficacy and potency of CB1 receptor orthosteric ligands when bound to CB1 receptors simultaneously. This alternative approach of enhancing CB1 receptor activation has promising therapeutic potential in rodent models by eliciting antinociceptive effects in pain models and anti-withdrawal effects in THC-dependent mice without producing acute cannabimimetic effects (i.e., impaired locomotion, catalepsy, or hypothermia), or antinociceptive tolerance or cannabinoid dependence following repeated administration. Furthermore, the CB1 receptor PAM, ZCZ011, does not substitute for the synthetic cannabinoid, CP55,940, in C57BL/6J mice or AEA in FAAH (-/-) mice in the drug discrimination paradigm. Therefore, the objectives of this dissertation are to (1) assess the efficacy of the CB1 receptor PAM, ZCZ011, to reduce withdrawal signs in opioid dependent male and female mice, (2) identify complementary dependent measures that reflect enhanced transit effects associated with the severe withdrawal sign of diarrhea, and (3) determine whether the anti-diarrheal effect of ZCZ011 ameliorates these processes of enhanced transit during opioid withdrawal through CB­1 or CB2 receptors. We found that ZCZ011 fully attenuates a subset of naloxone-precipitated withdrawal signs, i.e., diarrhea and weight loss, in oxycodone-dependent male and female mice through the activation of CB1, not CB2, receptors. Moreover, ZCZ011 partially attenuates paw flutters and head shakes, while no effect on jumping was observed. Additionally, we determined that when an oral gavage of a non-absorbable charcoal solution was administered before naloxone in oxycodone-dependent mice, there was a profound enhancement of small intestinal transit during naloxone-precipitated withdrawal. ZCZ011 inhibits transit in oxycodone-dependent mice undergoing withdrawal. Subsequent studies demonstrated that, indeed, the CB1, not CB2, receptor is required for this inhibition of naloxone-enhanced small intestinal transit. Furthermore, an unforeseen finding revealed that ZCZ011 reduces transit in non-dependent mice and merited additional experiments to (1) discern whether CB1 receptors mediate this effect and (2) whether the effect undergoes tolerance following repeated administration. These studies found that ZCZ011 inhibits small intestinal transit by activating CB1 receptors and this anti-transit effect undergoes tolerance after six days of repeated administration of ZCZ011 twice a day. These studies suggest that the CB1 receptor PAM, ZCZ011, may be an alternative treatment for withdrawal-induced diarrhea; however, it is limited due to its inhibition of small intestinal in non-dependent mice. Considering the inhibitory transit effects of ZCZ011 underwent tolerance, further research is needed to determine whether maintained anti-withdrawal effects of ZCZ011 under conditions of opioid withdrawal remain or dissipate.


© Julien Dodu

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