Species-specific susceptibility to cannabis-induced convulsions

Abstract

Background and purpose: Numerous claims are made for cannabis' therapeutic utility upon human seizures, but concerns persist about risks. A potential confounder is the presence of both Δ⁹ -tetrahydrocannabinol (THC), variously reported to be pro- and anticonvulsant, and cannabidiol (CBD), widely confirmed as anticonvulsant. Therefore, we investigated effects of prolonged exposure to different THC/CBD cannabis extracts on seizure activity and associated measures of endocannabinoid (eCB) system signalling.

Experimental approach: Cannabis extract effects on in vivo neurological and behavioural responses, and on bioanalyte levels, were measured in rats and dogs. Extract effects on seizure activity were measured using electroencephalography telemetry in rats. eCB signalling was also investigated using radioligand binding in cannabis extract-treated rats and treatment-naïve rat, mouse, chicken, dog and human tissue.

Key results: Prolonged exposure to cannabis extracts caused spontaneous, generalized seizures, subserved by epileptiform discharges in rats, but not dogs, and produced higher THC, but lower 11-hydroxy-THC (11-OH-THC) and CBD, plasma concentrations in rats versus dogs. In the same rats, prolonged exposure to cannabis also impaired cannabinoid type 1 receptor (CB₁ receptor)-mediated signalling. Profiling CB₁ receptor expression, basal activity, extent of activation and sensitivity to THC suggested interspecies differences in eCB signalling, being more pronounced in a species that exhibited cannabis extract-induced seizures (rat) than one that did not (dog).

Conclusions and implications: Sustained cannabis extract treatment caused differential seizure, behavioural and bioanalyte levels between rats and dogs. Supporting radioligand binding data suggest species differences in eCB signalling. Interspecies variations may have important implications for predicting cannabis-induced convulsions from animal models.

Linked articles: This article is part of a themed section on 8^th European Workshop on Cannabinoid Research. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.10/issuetoc.

Figure 1

Temporal representation of acute behaviours in rats (n = 10 per group) 10 min after daily p.o. low‐dose [1.08 mg·kg⁻¹ THC (Δ⁹‐THC) + 1 mg·kg⁻¹ CBD] or high‐dose (40.5 mg·kg⁻¹ THC + 37.5 mg·kg⁻¹ CBD) cannabis extract treatment for 13 weeks. Behavioural events associated with generalized seizures in rodents are highlighted in bold.

Figure 2

Temporal representation of persistent behaviours in rats (n = 10 per group) ~23 h after daily p.o. administration of low‐dose [1.08 mg·kg⁻¹ THC (Δ⁹‐THC)+ 1 mg·kg⁻¹ CBD] or high‐dose (40.5 mg·kg⁻¹ THC + 37.5 mg·kg⁻¹ CBD) cannabis extract treatment for 13 weeks. Behavioural events associated with generalized seizures in rodents are highlighted in bold.

Figure 3

(A) Biplot of the first two principal components (F1 and F2) derived from daily behavioural data (see Methods). Positive values of the first principal component were positively correlated with observations made in the period shortly after dosing (acute), irrespective of dose, whereas the converse applied to observations made prior to daily treatment (persistent). Positive values of the second principal component were positively correlated with observations made in animals that received high‐dose (40.5 mg·kg⁻¹ THC + 37.5 mg·kg⁻¹ CBD; n = 10) cannabis extract, irrespective of dose timing, whereas the converse applied to observations made in animals that had received low‐dose (1.08 mg·kg⁻¹ THC + 1 mg·kg⁻¹ CBD; n = 10) cannabis extract. (B) Correlation plot showing association of behaviours with the first and second principal components.

Figure 4

Epileptiform events recorded via EEG in rats treated with high‐dose (40.5 mg·kg⁻¹ THC + 37.5 mg·kg⁻¹ CBD) cannabis extract (Supporting Information Table S2). Each panel shows the EEG recording of the complete epileptiform event (top left), a shorter section of the event during the period of greatest amplitude activity represented on an extended timescale (bottom left) and a spectrographic representation of each event [right; x‐axis: time (s); y‐axis: frequency (Hz); and colour bar: power (dB·mV⁻²)]. * Indicates occurrence of a seizure during drug administration or within 10 min thereafter. ¥ Indicates an epileptiform event detected via EEG that was accompanied by a motor convulsion.

Figure 5

(A) EEG recordings from a rat treated with low‐dose (1.08 mg·kg⁻¹ THC + 1 mg·kg⁻¹ CBD) cannabis extract during (panels a, b) pretreatment baseline and (panels c, d) an epileptiform event. In panel c, * and ** show areas reproduced in panels d, e respectively. (B, panel a) Spectrographic representation of [A, panel c, right; x‐axis: time (s); y‐axis: frequency (Hz); and colour bar: power (dB·mV⁻²)] and (panel b) resulting PSD. (C) Mean (black) ± SEM (red dotted) PSD of epileptiform events recorded via EEG from animals treated with high‐dose (40.5 mg·kg⁻¹ THC + 37.5 mg·kg⁻¹ CBD; n = 10) cannabis extract. Inset shows overlay of individual PSD plots per event per animal. (D) (panel a) Cumulative incidence and (panel b) temporal distribution of coded behaviours associated with seizures in animals (see Methods) treated with low‐ or high‐dose cannabis extracts. (panel c) Cumulative incidence and (panel d) temporal distribution of coded behaviours not associated with seizures in animals (see Methods) treated with low‐ or high‐dose cannabis extracts.

Figure 6

Saturation binding of [³H]‐SR141716A to cerebellar membranes from rats treated with vehicle, low‐dose (1.08 mg·kg⁻¹ THC + 1 mg·kg⁻¹ CBD) or high‐dose (40.5 mg·kg⁻¹ THC + 37.5 mg·kg⁻¹ CBD) cannabis extracts for (i) 2 days, (ii) 4, (iii) 8 and (iv) 13 weeks. (B) Temporal profile of B_max (pmol·mg⁻¹) derived from (A, panels a–d). (C) Log concentration–response best‐fit curves for stimulation of [³⁵S]‐GTPγS binding by THC and WIN55 212‐2 in cerebellar membranes for week 13 data shown in (A). Data expressed as % maximal stimulation by 10 μM WIN55,212‐2 in vehicle group membranes fitted to an operational model of ligand binding (THC/high dose lack of response prevented valid curve derivation, and a subjectively assessed non‐linear fit was employed). (panel d) Overlay of best fit curves derived from panels a–c. W, WIN55,212‐2; T, THC; C, control (vehicle); L, low dose; H, high dose. No ‘T–H” curve presented. (D) (panel a) Saturation binding of [³H]‐SR141716A to human, chicken, dog, mouse and rat cerebellar membranes. (panel b) Log concentration–response curves for stimulation of [³⁵S]‐GTPγS binding (dpm) by THC in cerebellar membranes from species indicated. (panel c) Log concentration–response curves for stimulation of [³⁵S]‐GTPγS binding (normalized) by THC in cerebellar membranes from the species indicated. (E) (panel a) Equivalent (to D, panel b) curves for THC (Δ⁹‐THC)+ CBD (1.08:1.00). (panel b) Equivalent (to D, panel c) curves for THC + CBD (1.08:1.00). Data obtained from chicken, dog and human membrane samples were not amenable to sigmoidal curve fitting; here, subjective best fits are shown, but EC₅₀ not calculated. Concentration expressed as THC. (panel c) Equivalent (to D, panel b) curves for CBD. (panel d) Equivalent (to D, panel c) curves for CBD. Data obtained were not amenable to sigmoidal curve fitting; here, subjective best fits are shown but EC₅₀ not calculated. (F) Radar plot showing CB₁ receptor (‘Expression’; D, panel a), basal G‐protein turnover (‘Basal’; D, panel b) and sensitivity to and extent of agonist stimulation (‘Sensitivity’ and ‘Extent’; D, panel c) by species based upon (A–E) scaled as percentage of the species exhibiting the highest value for a given measure. With the exception of (F), all values shown are mean ± SEM; n = 3 experiments of three technical replicates in all cases.