Chronic pain affects approximately 20% of adults in the United States, but effective analgesics with minimal side effects are limited (Nahin et al., 2023). Indeed, the path toward developing novel analgesics that possess clinically adequate therapeutic dosing windows sufficient to produce pain relief without untoward side effects has proven an arduous challenge. Although research into the endocannabinoid system has yielded multiple therapeutic targets that demonstrated initial analgesic potential, the promise of these targets remains clinically unfulfilled (Wilkerson et al., 2021). Specifically, the endogenous cannabinoid ligand 2-arachidonoyl glycerol (2-AG) is an agonist at both the cannabinoid 1 receptor (CB₁R) and the cannabinoid 2 receptor (CB₂R) (Stella et al., 1997). Due to rapid metabolism by its degradative enzyme, monoacylglycerol lipase (MAGL), and short-lasting in vivo activity, exogenous 2-AG administration has limited clinical utility (Nomura et al., 2011). Pharmacological MAGL inhibition produces long-lasting elevated 2-AG levels, which can lead to CB₁R and CB₂R activation, similar to Δ9-tetrahydrocannabinol (THC), a partial CB₁R and CB₂R agonist. Although clinical reports suggest that THC may be a useful pain reliever, it has been documented that THC possesses side effects and limitations to long-term use such as tolerance, dependence, and use disorder (Ortiz et al., 2022). Thus, the utility of THC and perhaps other direct cannabinoid agonists as analgesics remains hazy. Among MAGL inhibitors, MJN110 stands out for its high potency and selectivity (Long et al., 2009). Previous studies demonstrated that MJN110 can produce dose-dependent and reversible antinociception in a variety of different preclinical pain studies, including acute nociception as well as neuropathic and cancer pain models. Unlike THC, in mice MJN110 does not induce undesired effects such as hypoactivity, hypothermia, or catalepsy (Ignatowska-Jankowska et al., 2015). This suggests that MAGL inhibitors like MJN110 may offer therapeutic cannabinoid receptor–mediated analgesia with fewer side effects. Repeated analgesic administration poses challenges in pharmacological pain management, as it often leads to tolerance and diminished effectiveness. In mice, a different MAGL inhibitor, JZL184, at high doses produces sustained increases in 2-AG levels, complete tolerance to antinociception, and cross-tolerance to exogenous agonists (Schlosburg et al., 2010). However, a lower JZL184 dose showed sustained antinociception in certain models, indicating dose-dependent outcomes (Kinsey et al., 2013). Similar findings were observed with MJN110 in rats; high doses led to antinociceptive tolerance in an osteoarthritis model, whereas lower doses maintained antinociception (Burston et al., 2016).

Clinically, pain is nuanced and includes affective, cognitive, and sensory dimensions. The affective dimension is comprised of the negative feelings and emotions, including depression, that many pain patients experience (Price, 2000). The cognitive dimension of pain relates to the complex way that a patient views the pain experience (Ciccone and Grzesiak, 1984). The sensory dimension of pain encompasses physical location and intensity. Many preclinical pain studies only examine the sensory dimension via “pain-stimulated” or “pain-evoked” behaviors, which assess responses like paw or tail withdrawal (Ignatowska-Jankowska et al., 2015). Novel preclinical procedures have recently been developed to evaluate pain-related behavioral depression such as decreased nesting behavior in mice induced by intraperitoneal acid injection (IP acid) (Diester et al., 2021a,b). In this issue of the Journal of Pharmacology and Experimental Therapeutics, Diester and colleagues (2024) aimed to assess antinociception resulting from repeated 1.0-mg/kg MJN110 treatment in mice subjected to repeated IP acid. Behavioral studies were complemented by examining CB₁R-mediated [35S]GTPγS binding in spinal cord and brain tissues. Regions investigated included the lumbar spinal cord, periaqueductal gray, ventral tegmental area, nucleus accumbens, amygdala, and hippocampus. These regions collectively either play roles in nociception or are associated with the mesolimbic dopamine system, which is known to modulate pain-depressed behavior (Leitl et al., 2014; Serafini et al., 2020). The authors hypothesized that repeated 1.0 mg/kg per day MJN110 would produce sustained or increased antinociception with minimal effects on CB₁R signaling and/or desensitization. These studies are important to the field, as it is critical to understand the behavioral and molecular impact of repeated administration when developing effective analgesics that possess prolonged efficacy and minimal side effects.

In their present study, Deister et al. (2024) examine the intricate interplay of behavioral antinociception and changes in CB₁R function resulting from prolonged treatment with the MAGL inhibitor MJN110 in both male and female mice. Their comprehensive analysis yielded several pivotal findings, unraveling insights into the interplay of pain-related behaviors, antinociception, and CB₁R signaling. The study first assessed the impact of repeated daily treatment with IP acid on nesting behavior, revealing a significant, sustained depression linked to pain, with a potential trend for reduced effectiveness in females. Previous research in rats and mice demonstrated that repeated IP acid administration induces pain-related behavioral depression and weight loss, and the study by Diester and colleagues (2024) extended these findings by using this model as an opportune platform to evaluate MJN110 as a potential analgesic. The significant antinociception observed on the first day of MJN110 treatment mirrored previous findings, highlighting the acute antinociceptive potential of MAGL inhibitors in pain models.

However, the study’s nuanced approach extended beyond acute effects, examining the impact of repeated MJN110 treatment over several days. Contrary to the relatively consistent antinociception observed with acute MAGL inhibitor treatment, the effects of repeated administration have previously proven more variable, contingent on the chronically administered dose. Specifically, previous studies found that although high MAGL inhibitor doses led to initial antinociception diminishing over time, lower MAGL doses produce sustained or increasing antinociception without evidence of tolerance (Schlosburg et al., 2010; Kinsey et al., 2013; Burston et al., 2016; Curry et al., 2018). Expecting sustained or increasing antinociception with the chosen 1.0-mg/kg per day MJN110 dose, the study yielded unexpected results. Although partial but significant antinociception persisted in males, females exhibited a decrease in the potency of MJN110, indicating the emergence of antinociceptive tolerance. This sex-specific divergence in antinociceptive responses aligns with previous research indicating that female mice are more susceptible to CB₁R agonist antinociceptive tolerance (Henderson-Redmond et al., 2022). Further investigations into changes in other endocannabinoids, CB₂R, and eicosanoids after repeated administration are needed to unravel the mechanisms behind the observed antinociceptive tolerance. Additionally, the current study recognized the potential impact of the estrous cycle on pain-related behaviors, urging further exploration of hormonal and anatomic determinants.

The study’s comprehensive approach also shed light on the broader effects of MJN110 beyond antinociception, revealing its role in alleviating IP acid–induced weight loss. However, disentangling these effects from direct antinociception proved challenging, as MJN110 increased body weight irrespective of the presence or absence of the noxious stimulus. Notably, the sex specificity of MJN110’s effect on body weight, significant only in females, added another layer of complexity, especially given the simultaneous development of tolerance in females to MJN110’s antinociceptive effects.

An additional layer to the current study focused on evaluating CB₁R-mediated G-protein activity after repeated MJN110 treatment with or without IP acid. The use of CP55,940, a CB₁R agonist, as a CB₁R ligand for [35S]GTPγS binding is well established, and its use is a notable strength. The results unveiled modest but significant decreases in CP55,940-induced maximal stimulation, which corresponds to the Emax value, in various central nervous system (CNS) regions of MJN110-treated mice. Males exhibited greater MJN110-induced decreases in brain CB₁R activity, especially in mesolimbic regions and the periaqueductal gray, which might have contributed to the observed decrease in MJN110 potency over time. Females showed greater decreases in lumbar spinal cord and periaqueductal gray CB₁R activity, which likely accounts for the rapid development of analgesic tolerance to MJN110. These studies suggest that different CNS regions may underlie the development of analgesic tolerance in a sex-specific manner.

This study does have a few limitations that bear mentioning. The discussed research only examines the potential of CB₁R desensitization and does not directly assess CB₁R downregulation. It will be important for future studies to address whether MNJ110 causes sex-dependent differences in CB₁R downregulation. It may also be important to investigate whether chronic MJN110 leads to cannabinoid dependence, often assessed by measuring precipitated withdrawal. Notably, the study by Diester et al. (2024) highlights a challenge in administering MJN110 at doses sufficient for alleviating episodic visceral pain without triggering compensatory antinociceptive tolerance and CB₁R desensitization. The intricate dance among drug potency, sex-specific responses, and regional variations in CB₁R adaptation underscores the complexity of developing effective analgesics. The need for nuanced investigations, considering hormonal, anatomic, and behavioral factors, emerges as a key takeaway from this multifaceted and informative study. In conclusion, this study unveils a rich tapestry of findings, ranging from the behavioral effects of repeated IP acid treatment to MJN110-induced antinociception and CB₁R tolerance. The study not only contributes to the understanding of MAGL inhibitors and their potential analgesic development but also underscores the challenges in developing analgesics that navigate the intricate landscape of pain-related behaviors, sex-specific responses, and complex signaling pathways.

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