Evaluate the anticonvulsant properties of phytocannabinoids

Epilepsy is the most common and serious brain disorder worldwide which afflicts people of any age, ethnic, social, and geographic category. Epilepsy’s co-morbidities include cognitive decline, depressive disorders and schizophrenia (Hermann et al., 2000, Kanner et al., 2012), all which are worsened by poorly controlled seizures (Perucca et al., 2000). There are many treatments available however all have notable side effects (Ortinski and Meador, 2004, Schachter, 2007) and 25% of the cases remain pharmaco-resistant, resulting in poorly controlled seizures (Hitiris et al., 2007). Cannabis sativa has a long history of use for the control of human seizures (Mechoulam, 1986). In two hospitals in Israel, children who are pharmaco-resistant are treated with Cannabis oils as the last resort before surgery. These oil extracts are produced from two specific strains which contain low THC and high CBD levels. However, there are at least 48 other strains in Israel with low quantities of THC that could be used as anticonvulsants. However, the long-term effects of Cannabis on brain development of these treated children has yet to be investigated. The effect of Cannabis treatment on brain development in pediatric patients is a critical question since in rodent models numerous studies have shown that chronic administration of synthetic cannabinoids (CB1 agonists) during early life periods can lead to long-term memory impairment, sensorimotor gating deficiencies, as well as alteration of hippocampal protein expression.

Currently, our epilepsy research is conducted in collaboration with Professor Uri Kremer from Ichilov hospital (Dana Children’s Hospital, Tel-Aviv), who is authorized to prescribe Cannabis extracts to treat children suffering from severe and frequent epileptic seizures. Professor Kremer currently utilizes two strains out of the 48 strains available for this treatment and together we are identifying whether these strains can be replaced by more effective ones. In this research, we focus on verifying the anticonvulsant properties of each strain and their effect on brain development and behavior.

To investigate the anticonvulsant effects of specific Cannabis extracts we apply an audiogenic model of generalized seizures in mice. This model has successfully revealed that some Cannabis extracts contain anticonvulsant properties (Hill et al., 2013). We plan to examine the anticonvulsant effects of the different strains on mice and identify the most effective strains. We will also analyze the chemical composition of these plants and verify the compounds and their ratios which are the most potent for their anticonvulsant effects. These strains will be recommended as preferred strains for treatment and will be further tested in clinical trials. Furthermore, finding the best cannabinoid combinations will enable us to design and encourage the breeding of new strains that will have optimal potency and may lead to better treatment options for epileptic patients. After identifying the most effective Cannabis strains, we will analyze the chemical composition of these plants and chemically assay the constituent cannabinoid compounds.

To identify specific cannabinoids or combinations that have anti-seizure properties, we must first purify phytocannabinoids from the plant using our state of the art MV-10 ASFE System (CO2 extractor with six solvent options and twelve fractions exits) and Dionex SemiPrep HPLC. To evaluate each cannabinoid’s anticonvulsant properties, we apply the pentylenetetrazol (PTZ) test on mice. PTZ is a validated test to identify drugs that are effective in the treatment of epilepsy in humans. After administration of a discrete amount of PTZ, mice are observed and ranked according to a seizure profile by measuring latencies to twitches and seizures (clonic/tonic, tonic, clonic). Seizure signs and profiles are assessed after 30 minutes post-PTZ administration and evaluated by the automated software SeizureScan. This software is dedicated for high throughput screenings and is specialized for the automation of detection of seizures in rodents. In addition, we screen extracts for the ability to protect against audiogenic seizures in DBA/2 mice. These types of mice exhibit convulsions in response to a loud noise and are often used as a model to screen potential compounds in the treatment of epilepsy.

To investigate the impact of different Cannabis strains extracts on brain development and behavior we will administer different Cannabis oils into juvenile mice (postnatal day 30) for ten consecutive days. In order to rank the Cannabis oils according to their long-term effect on brain development, at postnatal day 120 mice will undergo behavioral and biochemical testing as described in Gleason et al. (2012). From the results of those Cannabis oils that are shown to have the most detrimental ramifications, we will attempt to identify the phytocannabinoids with adverse effects on brain development using the methods described above. By combining these two approaches, we will identify strains, specific cannabinoids patterns and combinations that have potent anticonvulsant effects but have no or low effect on brain development and behavior.