Authors :
Presenting Author: Luisa Rocha, PhD – Center for Research and Advanced Studies
Christopher Martínez-Aguirre, Master in Science – Pharmacobiology – Center for Research and Advanced Studies; Mario Alonso-Vanegas, MD – International Center for Epilepsy Surgery – HMG-Coyoacán Hospital; Francia Carmona-Cruz, Ms – Pharmacobiology – Center for Research and Advanced Studies; Manola Cuellar-Herrera, PhD – Epilepsy Clinic – Hospital General de México Dr. Eduardo Liceaga
Rationale: br>Drug-resistant epilepsy (
DRE) is associated with high extracellular levels of glutamate, a condition that facilitates the excitotoxicity and neuronal damage. On the other hand, studies support the idea that
cannabinol (CBN), a non-psychoactive phytocannabinoid, regulates homeostasis of calcium, an ion involved in glutamate release. At present it is unknown if CBN modifies the glutamate over-release in the brain of patients with DRE. The aim of this
study was to investigate if CBN reduces the evoked glutamate release in cortical synaptic terminals obtained from patients with DRE.Methods:
Synaptic terminals (synaptosomes) were obtained from neocortex of patients with DRE submitted to epilepsy surgery (n=2 parietal; n=4 frontal; n=2 temporal). Immediately after resected, the tissue was immersed in saccharose (0.32 M), oxygenated by bubbling (0.5 l/h) and transported (< 45 min) from the surgery room to the laboratory. Synaptosomes were highly purified by Percoll-sucrose density gradient and resuspended in artificial cerebrospinal fluid. Synaptosome-homogenates were divided in 7 fractions and used to estimate basal glutamate release and evoked glutamate release with high KCl (33 mM) in synaptosomes preincubated with CBN (100 nM, 1
µM, 10 µM, 100 µM or 1 mM ) or vehicle, during 15 min. Then, synaptosomes were centrifugated and the supernatant was used to estimate the glutamate release by HPLC. Results:
Under basal conditions, basal glutamate release was 40.86 ± 9.9 nmol/mg of protein. The KCl-induced depolarization augmented the extrasynaptosomal glutamate concentration (422%, p=0.007 vs basal). Synaptosomes preincubated with CBN showed reduced glutamate release (100 nM, -88%, p=0.0056; 1
µM, -84%, p=0.0162; 10 µM, -88%, p=0.006 vs KCl alone). At higher concentrations, this effect was persistent in synaptosomes obtained from 5 patients (62.5%) (100 µM, -72%, p=0.001; 1 mM, -64%, p=0.0038 vs KCl alone), while it was lost in the fractions obtained from 3 patients (37.5%) (100 µM, +2.15%, p=0.99; 1 mM, +8%, p=0.998 vs KCl alone). Conclusions:
Acute exposure to low concentrations of CBN reduces glutamate release from cortical synaptic terminals obtained from patients with different etiologies of DRE. Further studies are required to determine if this effect is also evident after chronic CBN exposure, as well as if this result is dependent on specific clinical conditions.
Funding: National Council of Science and Technology (CONACyT,
scholarship