Authors :
Luis Alfredo Marquez, PhD – Center for Research and Advanced Studies
Ernesto Griego, PhD – Albert Einstein Collegue of Medicine
Estefania Gutierrez-Castañeda, PhD – Center for Research and Advanced Studies
Christopher Martinez-Aguirre, PhD – Center for Research and Advanced Studies
Mario Alonso-Vanegas, MD – HMG Coyoacan Hospital
Emilio Galvan, PhD – Center for Research and Advanced Studies
Presenting Author: Luisa Rocha, PhD – Center for Research and Advanced Studies
Rationale: Cannabidiol (CBD) combined with other antiseizure medications has been indicated to control drug resistant seizures. This raises the question of whether CBD alone may reduce the neuronal hyperexcitability and synaptic transmission in the brain of patients with drug-resistant epilepsy. The present study focused to determine in vitro the effects of CBD on excitability and synaptic transmission in layer V pyramidal neurons of patients with drug-resistant epilepsy, using patch-clamp and extracellular recordings.
Methods: Epileptic neocortex of 8 patients with drug-resistant epilepsy (5 females and 3 males; 4 frontal, 3 temporal, and 1 parietal cortex) was evaluated ≤3 hours after surgery. Informed consent was obtained from all patients. From resected epileptic neocortex we obtained coronal slices (350 µm) using a vibratome. Electrophysiological recordings were conducted 30 minutes after the slicing process. CBD (10 µM) was dissolved in DMSO (0.5%). The effects of CBD on the action potential spike were analyzed with phase plots before and during the perfusion of the drug. Using extracellular recordings, we evaluated the field excitatory postsynaptic potentials (fEPSP) slope evoked by stimulation of axons of layer I/II of the neocortex. We also analyzed the effect of CBD in the induced spontaneous synaptic activity evoked by 4-
aminopyridine (4-AP, 200 µM). At the end of the experiment, we carried out digital reconstructions from the recorded neurons using the biocytin loaded in the patch-clamp pipettes.
Results: The digital reconstruction analysis confirmed that the recorded cells were pyramidal neurons. Under basal conditions, the pyramidal neurons presented a resting membrane potential of -70.9 ± 2.25 mV, a somatic input resistance of 223 ± 27.63 MΩ, and membrane time constant of 44.01 ± 4.6 ms. The maximal frequency of the action potential discharge was 25.4 ± 3.26 Hz. Our patch-clamp recordings showed that CBD did not modify the passive properties of pyramidal neurons, such as resting membrane potential (-70.95 ± 2.05 mV, p >0.05), somatic input resistance (236.3 ± 32.9 MΩ, p >0.05), and membrane time constant (47.13 ± 4.35 ms, p >0.05). However, CBD reduced the frequency of the action potential discharge
(20 ± 2.91 Hz, p< 0.05). Using extracellular recordings, we found that the perfusion of CBD (15 min) transiently decreased the fEPSP slope (15.7 ± 4.2% of baseline response, p< 0.05), an effect not evident when the tissue was perfused with the vehicle (DMSO, 0.5%). Finally, CBD reduced the spontaneous synaptic activity evoked with 4-AP (38.9 ± 4.76% of response induced by 4-AP alone, p< 0.05).