Summary: A new study from Indiana University shows that acetaminophen (Tylenol, paracetamol) eases pain by lowering levels of the endocannabinoid 2‑arachidonoyl glycerol (2‑AG) rather than increasing it. The researchers report that acetaminophen inhibits the enzyme diacylglycerol lipase α (DAGLα), which produces 2‑AG and helps activate CB1 receptors. This unexpected mechanism challenges long‑standing assumptions about endocannabinoids and pain signaling and could guide the design of safer, more targeted analgesics.
The research team discovered that acetaminophen reduces production of 2‑AG by blocking DAGLα activity. Because 2‑AG normally activates CB1 receptors—the same receptor family involved in the effects of cannabis—scientists had generally assumed more endocannabinoid signaling would reduce pain. These findings suggest a more nuanced role for 2‑AG in nociception: in some settings, lowering 2‑AG appears to produce analgesia.
Key Facts:
- New mechanism: Acetaminophen inhibits DAGLα and reduces levels of the endocannabinoid 2‑AG.
- Revised model: Reduced 2‑AG can be antinociceptive in certain pain tests, offering a new perspective on CB1‑related pain pathways.
- Drug development potential: DAGLα represents a potential drug target for pain relief that could avoid acetaminophen’s liver toxicity.
Institution: Indiana University
Overview of the study
Researchers Michaela Dvorakova and Alex Straiker at Indiana University’s Gill Institute for Neuroscience led the study published in Cell Reports Medicine. Their experiments indicate that acetaminophen selectively inhibits diacylglycerol lipase α (DAGLα), reducing synthesis of 2‑AG without affecting DAGLβ. The reduction in 2‑AG correlates with decreased pain responses in animal models, flipping the conventional idea that boosting endocannabinoids is always analgesic.
Acetaminophen is the most widely used over‑the‑counter analgesic and antipyretic in the United States, but its exact mechanism of action has long been debated. The IU team’s data suggest a clear biochemical target—DAGLα—that could explain at least part of acetaminophen’s pain‑relieving effects and open the door to alternative therapies that avoid hepatic toxicity.
“There are many hypotheses, but until now we did not have a definitive molecular target,” said Dvorakova. “Our results show that in the case of 2‑AG, decreased levels may actually reduce pain.”
Co‑author Alex Straiker emphasized the translational potential: “If DAGLα is the relevant target, pharmaceutical development can focus on drugs that modulate that enzyme without acetaminophen’s liver toxicity.”
The study also included behavioral pharmacology: pharmacological inhibition of DAGL with RHC80267 produced antinociception in wild‑type mice but not in CB1 knockout animals during a hot‑plate test, supporting a DAGLα–CB1 circuit involved in certain forms of pain processing.
Acetaminophen overdoses remain a major public health concern—responsible for about 500 deaths per year in the U.S. and a leading cause of liver transplantation worldwide. Because more than 60 million Americans consume products containing acetaminophen weekly, understanding its mechanism is essential to develop safer alternatives and to inform clinical guidance.
Breaking long‑standing beliefs about acetaminophen’s action required confronting decades of research supporting CB1 activation as analgesic. “It can be difficult to overturn established dogma,” Straiker said, “but our data point to a different circuit in which inhibiting DAGLα reduces nociception.”
The team plans to evaluate other common analgesics such as ibuprofen and aspirin to determine whether they also affect endocannabinoid pathways or DAGL enzymes, which would further refine understanding of pain pharmacology and therapeutic targets.
Additional contributors to the study include Ken Mackie, Taryn Bosquez‑Berger, Jenna Billingsley, Natalia Murataeva, Taylor Woodward, Emma Leishman, Anaëlle Zimmowitch, Anne Gibson, Jim Wager‑Miller, Heather Bradshaw, Ruyi Cai, Shangxuan Cai, Yulong Li, Tim Ware and Ku‑Lung Hsu.
About this pain relief and neuroscience research news
Author: Jaleesa Elliott
Source: Indiana University
Contact: Jaleesa Elliott – Indiana University
Image: Image credited to Neuroscience News
Original Research (open access):
“Acetaminophen (Paracetamol) inhibits diacylglycerol lipase synthesis of 2‑arachidonoyl glycerol: implications for nociception” by Michaela Dvorakova et al., Cell Reports Medicine. DOI: 10.1016/j.xcrm.2025.102139
Abstract
Acetaminophen (Paracetamol) inhibits diacylglycerol lipase synthesis of 2‑arachidonoyl glycerol: implications for nociception
Acetaminophen is a widely used analgesic whose mechanism of action has not been fully resolved. Although acetaminophen causes roughly 500 deaths annually in the United States due to overdose and liver injury, safer alternatives have not yet been developed. Because endocannabinoids have been implicated in acetaminophen action, we investigated interactions between acetaminophen and the endocannabinoid system.
We report that acetaminophen inhibits the activity of diacylglycerol lipase α (DAGLα), but not DAGLβ, leading to decreased synthesis of the endocannabinoid 2‑arachidonoyl glycerol (2‑AG). This finding supports the counterintuitive hypothesis that reducing 2‑AG production through DAGLα inhibition can be antinociceptive under certain conditions.
Consistent with this idea, pharmacological DAGL inhibition using RHC80267 produced antinociceptive effects in wild‑type mice but not in CB1 knockout mice in the hot‑plate assay. We propose that DAGLα activation may worsen certain forms of nociception, and that acetaminophen’s analgesic actions include inhibition of a DAGLα/CB1 circuit that facilitates at least one form of pain signaling.