**4. Phytocannabinoids and epilepsy: mechanisms of action and preclinical studies**

The phytocannabinoids are a class of cannabinoids that are produced by plants of the cannabis species. The phytocannabinoids are C21 aromatic compounds consisting of an aromatic isoprenyl terpenophenolic core and resorcinyl side chain. Based on the structure of the oxygen bond between the isoprenyl and resorcinyl moieties the phytocannabinoids can be placed into 6 main families. Within each family, variations of the R-chain on the resorcinyl moiety differentiate each individual cannabinoid [23]. To date, over 140 different phytocannabinoids have been identified in *C. sativa.* While there is a high degree of structural preservation among the phytocannabinoids, they appear to display widely different effects on the mammalian central nervous system. The structural and stereochemical requirements for biological activity of the cannabinoids have been well established. Most biologically active cannabinoids (with a few exceptions) have a hydroxyl group on the C1 and an alkyl group on the C3 aromatic positions. As well, naturally occurring cannabinoids are biologically active in the trans (−) enantiomer [24]. Following the first isolation of the cannabinoids it did not take long for their anticonvulsant properties to be recognized [25]. Of the cannabinoids produced by the *C. sativa* the most comprehensively studied in the field of epilepsy are Δ<sup>9</sup> -tetrahydrocannabinol (Δ<sup>9</sup> -THC) and cannabidiol (CBD).

is achieved by the production of monoacylglycerol lipase (MAGL) by astrocytes and nerve terminals which breaks down 2-AG in the synaptic cleft. This temporal and spatial control allows for precise regulation of oscillations within neuronal networks by the endocannabi-

During an epileptic seizure there is excessive glutamate release from presynaptic excitatory neurons. In rodent models of epilepsy this has been shown to cause increased production of both 2-AG and anandamide that in turn active CB1R on the glutamatergic axon terminals to decrease the release of further excessive glutamate. This prevents further neuronal hyperexcitability which may play a role in terminating seizures. The increased anandamide is felt to

Temporal lobe epilepsy secondary to mesial temporal sclerosis (scarring of the hippocampi) is a common cause of epilepsy in adults that is often amenable to surgical resection of the mesial temporal structures. Pathological examination of surgically resected specimens has shown alterations in expression of CB1R of neurons within the hippocampi that provide insight into how disruption of the endocannabinoid system could predispose to epileptogenesis. In resected hippocampi there is a downregulation of CB1R expression on the axon terminals of excitatory (glutamatergic) neurons within the inner molecular layer of the dentate gyrus and an upregulation of CB1R expression on inhibitory (GABAergic) axon terminals within the dentate molecular layer [18]. These changes in CB1R expression result in both a loss of the normal inhibition of excessive glutamate release and increased suppression of GABAergic activity both of which result in increased neuronal hyperexcitability and subsequent seizure generation [22]. In patients with chronic epilepsy, there is also a decrease in the amount of anandamide and 2-AG released with excessive neuronal activation further contributing to a

loss of the endocannabinoid mediated inhibition of excessive neuronal activation [18].

novel anticonvulsant mechanism not provided by other anticonvulsant therapies.

**4. Phytocannabinoids and epilepsy: mechanisms of action and** 

The growing body of evidence demonstrating the role the endocannabinoid system plays in the brains' mechanisms in regulating neuronal network oscillations and preventing excessive neuronal hyperexcitability coupled with alterations in the endocannabinoid receptors seen in epileptogenic tissue make the endocannabinoid system an attractive therapeutic target in the treatment of epilepsy. Modulation of the endocannabinoid system would provide a potential

The phytocannabinoids are a class of cannabinoids that are produced by plants of the cannabis species. The phytocannabinoids are C21 aromatic compounds consisting of an aromatic isoprenyl terpenophenolic core and resorcinyl side chain. Based on the structure of the oxygen bond between the isoprenyl and resorcinyl moieties the phytocannabinoids can be placed into 6 main families. Within each family, variations of the R-chain on the resorcinyl moiety differentiate each individual cannabinoid [23]. To date, over 140 different phytocannabinoids have been identified in *C. sativa.* While there is a high degree of structural preservation among the

play a role in preventing seizure induced excitatory neurotoxic effects [18, 21].

noid system [18].

204 Recent Advances in Cannabinoid Research

**preclinical studies**

Initial research focused on the anticonvulsant effects of Δ<sup>9</sup> -THC and other CB1R agonists such as anandamide. Through their activation of CB1R, anandamide and the synthetic cannabinoid WIN 55,212-2 were able to block the production of postsynaptic neuronal spiking and excitatory post synaptic potential production. Both compounds were also able to suppress the production of abnormal burst activity in neurons placed in Mg2 + depleted solution. Depletion of Mg2+ in solution allows activation of NMDA receptors at normal resting potentials without the usual prerequisite neuronal depolarization. This effect was abolished when CB1R antagonists were added, suggesting that the effect was secondary to activation of CB1R by these agents [26]. Δ<sup>9</sup> -THC is a major phytocannabinoid in *C. sativa*. It is a high affinity partial agonist of both CB1R and CB2R that is competitive with both anandamide and 2-AG. The direct activation of CB1R by Δ<sup>9</sup> -THC is responsible for its psychoactive effects [19]. Numerous studies have assessed the anticonvulsant activity of Δ<sup>9</sup> -THC and its metabolites with conflicting results. These studies showed that Δ<sup>9</sup> -THC and its metabolites showed both anticonvulsant and proconvulsant activity depending on the dosage, animal species and seizure model used. In Maximal Electroshock (MES) and Maximal Electroshock Threshold (MEST) mouse models which mimic generalized onset convulsive seizures both Δ9-THC and its metabolites showed anticonvulsant activity by blocking or increasing the latency to hind limb extensor seizures [27]. In other studies Δ<sup>9</sup> -THC was also shown to potentiate the effects of several anticonvulsants [28]. In models that showed an anticonvulsant effect of Δ<sup>9</sup> -THC, all three of its metabolites including 11-OH-Δ<sup>9</sup> -THC showed anticonvulsant effect. The anticonvulsant effect of 11-OH-Δ<sup>9</sup> -THC was more potent than its parent compound by almost 1 order of magnitude suggesting that much of the anticonvulsant activity attributed to Δ<sup>9</sup> -THC may in fact come from its metabolites [27].

In a rat model of electrically induced limbic seizures Δ<sup>9</sup> -THC increased the threshold of electrically induced after discharges at the site of electrode implantation in the left subiculum. However, Δ<sup>9</sup> -THC increased the duration of cortically recorded after discharges in electrodes remote from the site of stimulation. This suggested that Δ<sup>9</sup> -THC may have both anticonvulsant and proconvulsant effects in focal onset epilepsies [27]. In the cobalt model of focal epilepsy in rats Δ<sup>9</sup> -THC increased the frequency of epileptic potentials at the site of the cobalt-induced lesion. This was not seen with Δ<sup>9</sup> -THC's main metabolite 11-OH-Δ<sup>9</sup> -THC. Both Δ<sup>9</sup> -THC and 11-OH-Δ<sup>9</sup> -THC seemed to increase generalized cortical excitation as seen by the production of brief intermittent cortically recorded after discharges [27]. Similar findings were seen in a rat model using iron to induce a seizure focus. While both Δ<sup>9</sup> -THC and 11-OH-Δ<sup>9</sup> -THC both caused increased cortical excitability, only Δ<sup>9</sup> -THC provoked clinical seizures. As well, the dose of Δ<sup>9</sup> -THC required to induce seizures was much higher than that required to induce electrographic changes in keeping with cortical excitation [29]. In mice, Δ<sup>9</sup> -THC has also been shown to induce kindling of a second epileptic focus in response to both electroconvulsive therapy as well as pentylenetetrazol (PTZ) and picrotoxin induced seizures [30]. When administered to rabbits with a genetic mutation causing audiogenic seizures Δ<sup>9</sup> -THC caused signs of neurotoxicity but prevented seizures when the rabbits were stimulated with a sound stimulus above their normal seizure threshold range. Conversely, in another breed of rabbits, injection with Δ<sup>9</sup> -THC induced both neurotoxicity and behavioral seizures in a dosage dependent manner [31].

potent anticonvulsant effect against tonic seizures its effect against clonic seizures was minimal. Consroe et al. hypothesized that this effect was due to the fact that tonic seizures are spread rap-

suppressed tetanic potentiation in isolated bullfrog ganglia [27]. This coupled with the fact that CBD is effective in preventing 3-Mercaptoproprionic acid (3-MPA) induced seizures suggested that some of the anticonvulsant effect of CBD may result from its ability to increase production

CBD correlated well with its anticonvulsant effect in several animal models. This suggests that the anticonvulsant effect of CBD is due to the parent compound and not its metabolites [27].

In summary, CBD was shown to display broad spectrum anticonvulsant activity in a wide range of animal models of epilepsy including generalized seizures caused by electroshock and GABA inhibiting drugs and focal seizures induced by placement of toxic metals on the cortex. It however had no effect on models of generalized absence seizures [38]. CBD also blocked kindling of a second epileptic focus [36]. Even at high doses it failed to cause any behavioral or cognitive side effects in test animals. This would suggest that CBD is a potent anticonvulsant with limited cognitive side effects, making it an attractive potential anticon-





207







Cannabis for Pediatric and Adult Epilepsy http://dx.doi.org/10.5772/intechopen.85719

idly throughout the brain from a focal lesion via post-tetanic stimulation. Unlike Δ<sup>9</sup>

nabidivarin (CBDV) which have been shown to have anticonvulsant effects. ∆<sup>9</sup>

However significantly fewer rats exposed to PTZ that were treated with ∆<sup>9</sup>

THCV decreased the amplitude and duration of abnormal neuronal burst activity. ∆<sup>9</sup>

non-psychoactive cannabinoid that acts as a CB1R antagonist. In a Mg2+ depleted solution ∆<sup>9</sup>

potentiated the effects on neuronal bursting of both phenobarbital and valproic acid. In a PTZ

seizures compared to those that were given PTZ alone [39]. Like CBD, CBDV is believed to exert its effects via CB1R independent mechanisms and has limited neurotoxicity [40]. CBDV has been shown to decrease the amplitude and duration of abnormal bursting in mouse and rat hippocampal slices in in both Mg2+ depleted solution and solution to which 4-aminopyridine (4-AP) has been added. CBDV also significantly decreased the number of seizures seen in in vitro MES and audiogenic seizure models in mice and PTZ induced seizures in rats. Unlike CBD, CBDV also prolonged the latency of seizure induction in a dose dependent manner. Administration of CBDV had no effect on motor performance in mice regardless of the ode administered [41]. The terpenes, which are another class of compounds found in cannabis, also possess a wide range of pharmacological activity on the mammalian nervous system at very low concentrations. Individually, these terpenes have not been assessed in patients with

The combinatorial effect of the chemical components of cannabis has been postulated wherein cannabis whole plant extracts may benefit from 'entourage' effects to yield greater effectiveness

of GABA in presynaptic GABAergic neurons [36]. Unlike Δ<sup>9</sup>

vulsant in the pediatric population [33, 37].

tial anticonvulsant activity. These include ∆<sup>9</sup>

**4.1. Other cannabinoids and terpenes**

In addition, Δ<sup>9</sup>

rat model ∆<sup>9</sup>

epilepsy [42, 43].

The results of these studies show that Δ<sup>9</sup> -THC and its metabolites display anticonvulsant activity in animal models using seizure models with rapidly evoked tonic discharges which mimics certain types of generalized onset seizures in humans. However, in models mimicking focal onset seizures, Δ<sup>9</sup> -THC and its metabolites seem to display a proconvulsant effect. This is manifested by increasing the activity at the site of the focal lesion and increasing generalized cortical activity [27]. A proconvulsant effect is also seen in models mimicking genetic based generalized epilepsies and absence seizures. Δ<sup>9</sup> -THC and its metabolites seem to induce hypersynchrony with slowly propagating epileptic discharges [32]. While Δ<sup>9</sup> -THC showed some potential as an anticonvulsant agent the potential to increase seizure activity along with its neurotoxic and psychotropic side effect profile limited its potential benefit in patients with epilepsy.

CBD is a low affinity negative allosteric modulator of CB1R with no psychotropic side effects due to the fact it does not cause activation of CB1R. It modulates the influx of both Ca2+ and Na+ into neurons by binding to human T-type voltage gated Ca2+ channels, Melastatin and Vanilloid type Transient Receptor Potential membrane receptors (TRPM and TRPV) and voltage gated Na+ channels [19]. This decreases neuronal excitability in response to stimulation. CBD has also been shown to inhibit intrasynaptic re-uptake of adenosine as well as activation of neuronal Serotonin, Glycine and Vanilloid receptors [33, 34]. The anticonvulsant effect of CBD is felt to be independent of activation of the endogenous CBR pathways. While the exact mechanism of anticonvulsant activity of CBD remains uncertain it appears to have a polypharmacological effect on modulating neuronal excitability.

In the Cobalt induced focal epilepsy rat model CBD had no effect on focal discharges at the lesion site but decreased the frequency of seizures. CBD also blocked the proconvulsant effects in of Δ<sup>9</sup> -THC [27, 35]. In the limbic seizure rat model CBD decreased the frequency, duration and amplitude of electrically induced after discharges at the site of stimulation in the left subiculum but did not prevent the spread of after discharges from the site of focal stimulation to distal electrodes. It had no apparent effect on generalized cortical excitability. This suggests that in focal models of epilepsy, CBD acts directly on the site of focal seizure onset [27].

Other animal studies continued to show the anticonvulsant effect of CBD in both transcorneal electroshock, drug induced and lesional epilepsies. This anticonvulsant effect was seen when a single intraperitoneal (i.p.) dose of CBD was administered alone but like Δ<sup>9</sup> -THC it also potentiated the effects of several anticonvulsant medications [33, 36, 37]. While CBD had potent anticonvulsant effect against tonic seizures its effect against clonic seizures was minimal. Consroe et al. hypothesized that this effect was due to the fact that tonic seizures are spread rapidly throughout the brain from a focal lesion via post-tetanic stimulation. Unlike Δ<sup>9</sup> -THC, CBD suppressed tetanic potentiation in isolated bullfrog ganglia [27]. This coupled with the fact that CBD is effective in preventing 3-Mercaptoproprionic acid (3-MPA) induced seizures suggested that some of the anticonvulsant effect of CBD may result from its ability to increase production of GABA in presynaptic GABAergic neurons [36]. Unlike Δ<sup>9</sup> -THC, the brain concentrations of CBD correlated well with its anticonvulsant effect in several animal models. This suggests that the anticonvulsant effect of CBD is due to the parent compound and not its metabolites [27].

In summary, CBD was shown to display broad spectrum anticonvulsant activity in a wide range of animal models of epilepsy including generalized seizures caused by electroshock and GABA inhibiting drugs and focal seizures induced by placement of toxic metals on the cortex. It however had no effect on models of generalized absence seizures [38]. CBD also blocked kindling of a second epileptic focus [36]. Even at high doses it failed to cause any behavioral or cognitive side effects in test animals. This would suggest that CBD is a potent anticonvulsant with limited cognitive side effects, making it an attractive potential anticonvulsant in the pediatric population [33, 37].

#### **4.1. Other cannabinoids and terpenes**

dose of Δ<sup>9</sup>

bits, injection with Δ<sup>9</sup>

dependent manner [31].

206 Recent Advances in Cannabinoid Research

ing focal onset seizures, Δ<sup>9</sup>

patients with epilepsy.

Na+

in of Δ<sup>9</sup>

age gated Na+

The results of these studies show that Δ<sup>9</sup>

genetic based generalized epilepsies and absence seizures. Δ<sup>9</sup>

pharmacological effect on modulating neuronal excitability.






been shown to induce kindling of a second epileptic focus in response to both electroconvulsive therapy as well as pentylenetetrazol (PTZ) and picrotoxin induced seizures [30]. When

signs of neurotoxicity but prevented seizures when the rabbits were stimulated with a sound stimulus above their normal seizure threshold range. Conversely, in another breed of rab-

activity in animal models using seizure models with rapidly evoked tonic discharges which mimics certain types of generalized onset seizures in humans. However, in models mimick-

This is manifested by increasing the activity at the site of the focal lesion and increasing generalized cortical activity [27]. A proconvulsant effect is also seen in models mimicking

showed some potential as an anticonvulsant agent the potential to increase seizure activity along with its neurotoxic and psychotropic side effect profile limited its potential benefit in

CBD is a low affinity negative allosteric modulator of CB1R with no psychotropic side effects due to the fact it does not cause activation of CB1R. It modulates the influx of both Ca2+ and

CBD has also been shown to inhibit intrasynaptic re-uptake of adenosine as well as activation of neuronal Serotonin, Glycine and Vanilloid receptors [33, 34]. The anticonvulsant effect of CBD is felt to be independent of activation of the endogenous CBR pathways. While the exact mechanism of anticonvulsant activity of CBD remains uncertain it appears to have a poly-

In the Cobalt induced focal epilepsy rat model CBD had no effect on focal discharges at the lesion site but decreased the frequency of seizures. CBD also blocked the proconvulsant effects

and amplitude of electrically induced after discharges at the site of stimulation in the left subiculum but did not prevent the spread of after discharges from the site of focal stimulation to distal electrodes. It had no apparent effect on generalized cortical excitability. This suggests that in focal models of epilepsy, CBD acts directly on the site of focal seizure onset [27].

Other animal studies continued to show the anticonvulsant effect of CBD in both transcorneal electroshock, drug induced and lesional epilepsies. This anticonvulsant effect was seen

also potentiated the effects of several anticonvulsant medications [33, 36, 37]. While CBD had

when a single intraperitoneal (i.p.) dose of CBD was administered alone but like Δ<sup>9</sup>


 into neurons by binding to human T-type voltage gated Ca2+ channels, Melastatin and Vanilloid type Transient Receptor Potential membrane receptors (TRPM and TRPV) and volt-

channels [19]. This decreases neuronal excitability in response to stimulation.

to induce hypersynchrony with slowly propagating epileptic discharges [32]. While Δ<sup>9</sup>





electrographic changes in keeping with cortical excitation [29]. In mice, Δ<sup>9</sup>

administered to rabbits with a genetic mutation causing audiogenic seizures Δ<sup>9</sup>

In addition, Δ<sup>9</sup> -THC and CBD several other cannabinoids have been evaluated for the potential anticonvulsant activity. These include ∆<sup>9</sup> -tetrahydrocannibivarin (∆<sup>9</sup> -THCV) and cannabidivarin (CBDV) which have been shown to have anticonvulsant effects. ∆<sup>9</sup> -THCV is a non-psychoactive cannabinoid that acts as a CB1R antagonist. In a Mg2+ depleted solution ∆<sup>9</sup> - THCV decreased the amplitude and duration of abnormal neuronal burst activity. ∆<sup>9</sup> -THCV potentiated the effects on neuronal bursting of both phenobarbital and valproic acid. In a PTZ rat model ∆<sup>9</sup> -THCV did not decrease the severity or duration of seizures or seizure mortality. However significantly fewer rats exposed to PTZ that were treated with ∆<sup>9</sup> -THCV displayed seizures compared to those that were given PTZ alone [39]. Like CBD, CBDV is believed to exert its effects via CB1R independent mechanisms and has limited neurotoxicity [40]. CBDV has been shown to decrease the amplitude and duration of abnormal bursting in mouse and rat hippocampal slices in in both Mg2+ depleted solution and solution to which 4-aminopyridine (4-AP) has been added. CBDV also significantly decreased the number of seizures seen in in vitro MES and audiogenic seizure models in mice and PTZ induced seizures in rats. Unlike CBD, CBDV also prolonged the latency of seizure induction in a dose dependent manner. Administration of CBDV had no effect on motor performance in mice regardless of the ode administered [41]. The terpenes, which are another class of compounds found in cannabis, also possess a wide range of pharmacological activity on the mammalian nervous system at very low concentrations. Individually, these terpenes have not been assessed in patients with epilepsy [42, 43].

The combinatorial effect of the chemical components of cannabis has been postulated wherein cannabis whole plant extracts may benefit from 'entourage' effects to yield greater effectiveness than treatment with a purified cannabinoid [42, 44]. This is supported by preclinical studies. In the *in vitro* oxotremorine-M mouse model of epilepsy, excessive neuronal bursting activity can be suppressed with ∆<sup>9</sup> -THC, but not CBD, while a standardized cannabis extract containing both ∆<sup>9</sup> -THC and CBD can abolish the abnormal bursting activity faster than purified ∆9 -THC alone [45]. In another study, both purified ∆<sup>9</sup> -THC and CBD can increase intracellular Ca2+ in rat hippocampal neuronal and glial cells. This effect is compounded when the two compounds are mixed together, with the greatest effect occurring with whole plant extract containing both ∆<sup>9</sup> -THC and CBD [46]. These preclinical data support the hypothesis that the 'entourage' effects between the various cannabinoids provide therapeutic benefit of cannabis whole plant extract, benefit that exceeds the activity of a single purified cannabinoid. This remains to be demonstrated in the human clinical context.

Two further studies showed no significant difference in seizure reduction with the addition of CBD as an adjunctive therapy. However, in one study patients were given CBD at a dose of 300 mg/day and their plasma CBD levels were maintained at 20–30 ng/ml. Subsequently one participant who had no difference in their seizure frequency was placed on CBD at a higher dose of up to 1200 mg/day. CBD plasma levels were higher averaging 150 ng/ml. This patient had a significant decrease in their seizure frequency suggesting that higher doses of CBD (and

Cannabis for Pediatric and Adult Epilepsy http://dx.doi.org/10.5772/intechopen.85719 209

In recent years there has been a public perception that cannabis is a potent, natural, and safe alternative therapy for epilepsy. This has driven the demand for and use of cannabis and its derived products to treat epilepsy especially in those patients whose seizures are medically intractable. Coupled with the media exposure showing examples of the apparent miraculous effects of CBD oil in select epileptic patients, treating physicians have struggled to balance the patient demand for cannabis products and the need for studies to determine their, efficacy, dosing, side-effect profile, and indication. To that end, there have been multiple studies, predominantly in children, looking into these clinical questions. Unfortunately, the overwhelming majority of these studies have been retrospective, unblinded, and uncontrolled resulting in their being hampered by various forms of bias and potential placebo effect. Despite the plethora of published research on this topic, questions still remain on the use of cannabis in

In this section, we will review the limitations of the studies, the studies using artisanal and CBD enriched cannabis herbal extracts (CHE), the studies using highly purified pharmaceuti-

The widespread use of cannabis and the effect of bias are highlighted in various published surveys. McLachlan performed a survey in London, Ontario, Canada, in which more than 60% of patients declared that cannabis was effective for their seizures and stress levels [56]. Ladina et al. reported a case series of 18 patients who all found medicinal cannabis very helpful for seizure control and improvement of mood disorder [57]. By contrast, Press had reported a significant discrepancy in reported responder rate between preexisting Colorado residents and those who moved to Colorado to obtain cannabis to treat their child's epilepsy (22 vs. 47%) suggesting there is a significant perception bias among these children's caregivers as to the therapeutic benefits of cannabis [58]. Physician bias may also play a role as a recent survey by Mathern showed contrasting opinions about CBD between neurologists and the general public. In his study, a minority of epileptologists and general neurologists said that there were sufficient data safety (34%) and efficacy data (28%) and only 48% would advise using medical cannabis and only in severe cases of epilepsy. Conversely, nearly all patients and the general public responded that there was sufficient safety (96%) and efficacy (95%) data, and

higher plasma levels) were required for seizure control [55].

**6. Recent clinical trials and experience**

cal grade CBD, and a meta-analysis of the CBD studies.

98% would recommend cannabis in cases with severe epilepsy [59].

**6.1. Limitations of the studies**

epilepsy.
