A study by researchers in the U.S. and U.K. has revealed a previously unknown mechanism by which cannabidiol (CBD)—a non-psychoactive component of cannabis—reduces seizures in many treatment-resistant forms of pediatric epilepsy. Led by researchers at NYU Grossman School of Medicine, the new study, in rodent models, found that CBD blocked signals carried by a molecule called lysophosphatidylinositol (LPI) in neurons. LPI is thought to amplify nerve signals as part of normal function, but can be hijacked by disease to promote seizures.

The new research confirmed a previous finding that CBD blocks the ability of LPI to amplify nerve signals in the brain’s hippocampus, and argues for the first time that LPI also weakens signals that counter seizures, further explaining the value of CBD treatment.

“Our results deepen the field’s understanding of a central seizure-inducing mechanism, with many implications for the pursuit of new treatment approaches,” said corresponding author Richard W. Tsien, PhD, chair of the Department of Physiology and Neuroscience at NYU Langone Health. “The study also clarified, not just how CBD counters seizures, but more broadly how circuits are balanced in the brain. Related imbalances are present in autism and schizophrenia, so the paper may have a broader impact.”

Tsein, together with colleagues at NYU Langone, and collaborators in the U.S. and U.K., reported on their work in Neuron, in a paper titled “Cannabidiol modulates excitatory-inhibitory ratio to counter hippocampal hyperactivity.”

When a neuron “fires” it sends an electrical pulse down an extension of itself until it reaches a synapse, the gap that connects one neuron to the next cell in a neuronal pathway. When the electrical impulse reaches the cell’s end before the synapse, it triggers the release of neurotransmitters that are travel across the gap to reach the next cell. Upon crossing this synapse, such signals either encourage the next cell to fire (excitation), or apply the brakes on firing (inhibition). Balance between the two are essential to brain function; too much excitation promotes seizures, and an imbalance between inhibition and excitation is also implicated in other disorders. “Neuronal circuits require coordination between synaptic excitation (E) and inhibition (I) for proper function,” the authors wrote, ”and disruptions in the excitatory-to-inhibitory (E:I) ratio contribute to epilepsy, autism spectrum disorders, and schizophrenia.”

But while cannabidiol has been shown to reduce seizures in multiple forms of pediatric epilepsies, the mechanism(s) of anti-seizure action remain unclear. “In preclinical models, CBD reduces spontaneous recurrent seizures and regulates the E:I ratio in acute seizures; however, the molecular signaling underlying CBD’s anti-seizure actions remains poorly defined.”

Possible therapeutic targets of CBD encompass ion channels, transporters, and transmembrane signaling proteins, the team noted. Among the proposed candidates are two GPCRs: the cannabinoid receptor CB1R and the receptor GPR55.  “In one model, CBD acts at glutamatergic axon terminals, blocking the pro-excitatory actions of the endogenous membrane phospholipid lysophosphatidylinositol, at the G-protein-coupled receptor GPR55 … However, the impact of LPI-GPR55 signaling at inhibitory synapses and in epileptogenesis remains underexplored.”

For their new study, the team looked at several rodent models to explore mechanisms behind seizures. “… we focused on the LPI-GPR55 signaling pathway as a potential modulator of E:I ratio and anti-seizure target of CBD.

To do this they used a variety of techniques that included measuring information-carrying electrical current flows with fine-tipped electrodes, or by investigating the effect of LPI by genetically removing its main signaling partner, or by measuring the release of LPI following seizures. Their collective results confirmed past findings that LPI influences nerve signals by binding to GPR55, on neuron cell surfaces. This LPI-GPR55 presynaptic interaction was found to cause the release of calcium ions within the cell, which encouraged cells to release glutamate, the main excitatory neurotransmitter.

Further, when LPI activated GPR55 on the other side of the synapse, it weakened inhibition, by decreasing the supply and proper arrangement of proteins necessary for inhibition. Collectively, this creates a “dangerous” two-pronged mechanism to increase excitability, say the authors. “We found that LPI triggers aGPR55-dependent dual mechanism to elevate network excitability: a transient elevation in presynaptic excitatory release probability, complemented by a slower sustained reduction of inhibitory synaptic strength,” the investigators stated.

The research team found that LPI-mediated effects on both excitatory and inhibitory synaptic transmission could be blocked either by genetically engineering mice to lack GPR55, or by treating mice with plant-derived CBD prior to seizure-inducing stimuli. While prior studies had implicated GPR55 as a seizure-reducing target of CBD, the current work provided a more detailed, proposed mechanism of action.

The authors propose that CBD blocks a positive feedback loop in which seizures increase LPI-GPR55 signaling, which likely encourages more seizures, which in turn increases levels of both LPI and GPR55. “Our observations suggest a positive feedback loop whereby LPI promotes hyperexcitability, which in turn increases the expression of LPI and GPR55,” they further explained. “We tentatively propose that CBD can extinguish a potentially regenerative loop in which hyperactivity enhances LPI-GPR55 signaling, further shifting the E:I ratio … Our experiments provide new insights on synaptic mechanisms of CBD’s anti-seizure effects.”

The team concluded that CBD could represent a potential therapeutic agent in treatment-resistant epilepsy patients who don’t respond to current drugs, such as benzodiazepines, possibly due to LPI-GPR55 effects.

The proposed vicious cycle also represents one process that could explain repeated epileptic seizures, although future studies are needed to confirm this. Further, the current study examined the plant-based cannabinoid CBD, but the authors note that LPI is part of signaling network that includes endocannabinoids such as 2-Arachidonoylglycerol (2-AG) that occur naturally in human tissues. LPI and 2-AG target receptors also regulated by CBD, but have different actions at the synapse. While LPI amplifies incoming electrical signals, endocannabinoids such as 2-AG respond to increases in brain activity by dialing down the release of neurotransmitters from nerve cells. Interestingly, LPI and 2-AG can be converted into each other through actions of enzymes.

“Theoretically, the brain could control activity by toggling between pro-excitatory LPI and the restorative actions of 2-AG,” said first study author Evan Rosenberg, PhD, a postdoctoral scholar in the Tsein’s lab. “Drug designers could inhibit the enzymes that underpin LPI production or promote its conversion to 2-AG, as an additional approach to control seizures. LPI could also serve as a biomarker of seizures or predictor of clinical responsiveness to CBD, providing an area of future research.”

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