Behavioral and Molecular Effects Induced by Cannabidiol and Valproate Administration in the GASH/Sal Model of Acute Audiogenic Seizures.

Spread the love

. 2021 Jan 22;14:612624.

doi: 10.3389/fnbeh.2020.612624. eCollection 2020.

Affiliations

Item in Clipboard

Giselda Cabral-Pereira et al. Front Behav Neurosci. .

Abstract

Despite evidence that supports cannabidiol (CBD) as an anticonvulsant agent, there remains controversy over the antiseizure efficacy, possible adverse effects, and synergistic interactions with classic antiepileptics such as valproate (VPA). The genetic audiogenic seizure hamster from the University of Salamanca (GASH/Sal) is a reliable experimental model of generalized tonic-clonic seizures in response to intense sound stimulation. The present study examines the behavioral and molecular effects of acute and chronic intraperitoneal administrations of VPA (300 mg/kg) and CBD (100 mg/kg) on the GASH/Sal audiogenic seizures, as well as the coadministration of both drugs. The GASH/Sal animals were examined prior to and after the corresponding treatment at 45 min, 7 days, and 14 days for seizure severity and neuroethology, open-field behaviors, body weight variations, and various hematological and biochemical parameters. Furthermore, the brain tissue containing the inferior colliculus (so-called epileptogenic nucleus) was processed for reverse transcription-quantitative polymerase chain reaction analysis to determine the treatment effects on the gene expression of neuronal receptors associated with drug actions and ictogenesis. Our results indicated that single dose of VPA helps prevent the animals from getting convulsions, showing complete elimination of seizures, whereas 7 days of chronic VPA treatment had few effects in seizure behaviors. Acute CBD administration showed subtle attenuation of seizure behaviors, increasing seizure latency and decreasing the duration of the convulsion phase, but without entirely seizure abolition. Chronic CBD treatments had no significant effects on sound-induced seizures, although some animals slightly improved seizure severity. Acute and chronic CBD treatments have no significant adverse effects on body weight, hematological parameters, and liver function, although locomotor activity was reduced. The combination of VPA and CBD did not alter the therapeutic outcome of the VPA monotherapy, showing no apparent synergistic effects. As compared to sham animals, chronic treatments with CBD caused abnormal mRNA expression levels for Trpv1, Adora1, Slc29a1, and Cnr1 genes, whereas no differences in gene expression were found for Htr1a and Sigmar1. Our study shed light on the behavioral and molecular effects of CBD and VPA on the GASH/Sal model and constituted the basis to develop further studies on the pharmacological effects of CBD and its interactions with other anticonvulsants.

Keywords: animal models; antiepileptic drugs; cannabis; drug interactions; epilepsy; gene expression; inferior colliculus (IC); valproic acid.

Conflict of interest statement

DS-B was hired under the research funding from RiverForce Partners Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1

Figure 1

Experimental design and timeline. (A) Animal groups and drug treatments used in the study. (B) Experimental time schedules. Basal measurements were conducted prior to the initiation of the treatments. Acute effects of each drug treatment on sound-induced seizures were evaluated 45 min after the first intraperitoneal administration. Chronic effects of each drug treatment on sound-induced seizures were evaluated after 7 and 14 days of drug administration. Tissue samples for gene expression analyses in the inferior colliculus were obtained at the final time point.

Figure 2

Figure 2

Audiogenic seizure analysis in the GASH/Sal. (A) Seizure index (SI), according to Garcia-Cairasco et al. (1996) using the behavioral descriptions and categorized severity scores transformed into discreet variables for statistical purposes. (B) Dictionary of the behavioral variables analyzed and displayed in the flowcharts of the seizure behaviors. (C) Flowcharts illustrating the graphical and statistical aspects of the seizure behaviors. The frequency and time spent performing each behavior are proportional to the height and width of the rectangle, respectively. The arrow width and direction indicate the statistical intensity and preference association between two behavioral items. The behaviors associated with wild-running phase (including wild running, jump, and falls) are depicted in yellow, and the behaviors associated with the convulsion phase (including convulsions and extentension of extremities) are depicted in red (figure modified from Barrera-Bailón et al., 2013).

Figure 3

Figure 3

Time course of CBD levels in serum from GASH/Sal animals treated with CBD (100 mg/kg, intraperitoneal). Notice that levels of CBD peak within 45 min after the intraperitoneal injection. Results are expressed as means ± SEM (n = 14).

Figure 4

Figure 4

Treatment effects of VPA and CBD on the audiogenic seizure behaviors of the GASH/Sal within 45 min after the first intraperitoneal administration. (A) Flowcharts show the sequences of behaviors in GASH/Sal animals in baseline conditions before starting the treatment. Notice that all GASH/Sal animals exhibited a complete audiogenic seizure, including the behavioral manifestations of the wild-running phase with turns to the left (GL) and right (GR), running (RU), jumps (JP), and atonic falls (AF) (depicted together with the yellow circle), as well as the behaviors associated with tonic–clonic convulsions (TCV, CCV1, CCV2, and CCVg), followed by a behavioral cluster of generalized clonic seizures with hyperextensions (HP1 and HP2) (depicted together with the red circle) and ended with postictal immobility (PIM) and breathing difficulties [dyspnea (DYS) and tachypnea (TCP)]. (B) Flowchart shows the sequences of behaviors in GASH/Sal animals treated with vehicle (sham), VPA, and CBD and with a combination of CBD and VPA, 45 min after the first intraperitoneal administration. Notice that behaviors associated with tonic–clonic convulsions (red circles) were blocked with VPA—or the combined CBD+VPA treatments, whereas animals treated with vehicle or single CBD administration remained similar to baseline conditions, although the elimination of the generalized clonic convulsions (CCVg) in the single CBD administration was noticeable. Furthermore, half of the GASH/Sal animals that followed VPA treatment or the combined CBD+VPA treatments showed seizures scores <2 with almost total absence of seizure behaviors or complete blockage of seizure activity. Each flowchart showed the compendium of behaviors in all the animals belonging to each experimental group. The seizure behaviors were analyzed in three time windows (presound, during sound, and postsound). See Figure 2 for detailed interpretation of the flowchart.

Figure 5

Figure 5

Treatment effects of VPA and CBD on the audiogenic seizure behaviors of the GASH/Sal within 7 and 14 days after chronic administration. (A) Flowcharts show the sequences of behaviors in GASH/Sal animals in baseline conditions before starting the treatment. Notice that all GASH/Sal animals exhibited a complete audiogenic seizure, including the behavioral manifestations of the wild-running phase with turns to the left (GL) and right (GR), running (RU), jumps (JP), and atonic falls (AF) (depicted together with the yellow circle), as well as the behaviors associated with tonic–clonic convulsions (TCV, CCV1, CCV2, and CCVg), followed by a behavioral cluster of generalized clonic seizures with hyperextensions (HP1 and HP2) (depicted together with the red circle) and ended with postictal immobility (PIM) and breathing difficulties [dyspnea (DYS) and tachypnea (TCP)]. (B) Flowchart shows the sequences of behaviors in GASH/Sal animals treated with vehicle (sham), VPA, CBD, and with a combination of CBD and VPA after 7 and 14 days of chronic drug administration. Notice that seizure behaviors in general remained similar to baseline conditions for all experimental groups, showing seizures scores ≥2. Notice slight modifications in the behavioral items associated with the wild-running phase, which can be seen in all experimental groups. Each flowchart showed the compendium of behaviors in all the animals belonging to each experimental group. The seizure behaviors were analyzed in three time windows (presound, during sound, and postsound). See Figure 2 for detailed interpretation of the flowchart.

Figure 6

Figure 6

Effects of VPA and CBD on the seizure intensity, latency, and duration of wild-running and convulsion phases. (A) Box plot summarizing the categorized seizure-index score in each experimental group on the baseline condition (pre-treatment), after acute treatment (45 min), as well as 7 and 14 days after chronic administrations. In each box plot, the horizontal line crossing the box is the median. Asterisks indicate significant differences between experimental groups (*p < 0.05, ***p < 0.001). (B–D) Graphs showing the duration (in seconds) for the seizure latency (B), the wild-running phase (C), and the convulsion phase (D) in each experimental group on pre-treatment condition and 45 min, 7 days, and 14 days post-treatment. GASH/Sal animals with complete elimination of audiogenic seizures (seizure scores = 0) were not graphed in (B–D) as the variable time was unable to be assessed.

Figure 7

Figure 7

Effects on body weight and hematological and biochemical profiles of treated GASH/Sal animals. (A) Graphs show the time course of body weight variations monitored during the 14-day treatment in sham-, VPA–, CBD–, and CBD+VPA-treated groups. Notice that all experimental groups showed a steady and persistent body weight throughout the treatment, and no significant differences were found as compared to the pre-treatment conditions (time 0 in the horizontal axis of the graphs). Data are shown as means ± SEM. (B) Table shows hematological and biochemical profiles of GASH/Sal animals after 14 days of chronic drug treatments. The hematological assessment of blood included the following parameters: hemoglobin concentration, hematocrit (volume percentage of red blood cells in blood), and number of red blood cells, white blood cells, and platelets in a microliter (μL). The analysis of liver function included five biochemical parameters: aspartate aminotransferase (AST), alanine aminotransferase (ALT), bilirubin (BT), albumin, and total protein. Asterisks and bold values in the table denote statistical significance (**p < 0.01, ***p < 0.001) as compared to the pre-treatment conditions and the sham group.

Figure 8

Figure 8

Treatment effects of VPA and CBD on the behavior of the GASH/Sal in the open-field test. (A) Plots show representative examples of the animal’s track in each experimental group at the protocol-defined time points (pre-treatment and 7 and 14 day drug treatments). Note that levels of global locomotor activity decrease in GASH/Sal animals that received CBD alone or the combination of CBD and VPA after 7 and 14 days of treatment. Notice the trace images of GASH/Sal animals treated with CBD or CBD+VPA that spent less time moving in the periphery and longer time in the center of the arena compared to those animals treated with the vehicle or VPA alone, also compared to the pre-treatment condition. Histograms show the total distance traveled (B), time of activity (C), number of rearing (D), time spent in the center (E), and number of grooming (F) for all experimental groups at the protocol-defined time points. Behavior indicators for the locomotor pattern (total distance traveled, time of activity, and number of rearings) and for the emotionality (time in the central zone and number of self-grooming) revealed differential effects of CBD in both monotherapy and combined therapy with VPA when compared to baseline activity (pre-treatment), as well as to vehicle and VPA monotherapies. Data are shown as means ± SEM. Asterisks indicate significant differences between experimental groups (*p < 0.05, **p < 0.01).

Figure 9

Figure 9

Gene expression changes in the inferior colliculus after 14-days treatment with VPA, CBD or the combination of CBD+VPA. Histogram shows relative quantities of transcripts of Trpv1
(A), 5-Htr1a
(B), Sigmar1
(C), Adora1
(D), Slc29a1
(E), and Cnr1
(F) genes in the inferior colliculus of GASH/Sal animals treated with vehicle (sham), VPA, CBD, and CBD+VPA. Notice that disruption of mRNA expression levels was found for the Trpv1, Adora1, Slc29a1, and Cnr1 genes, whereas no differential gene expression was found for Htr1a and Sigmar1. The relative mRNA expression of each gene was normalized to β-actin. ΔCt values were normalized to the average ΔCt of the inferior colliculus of sham animals. Each bar in the histograms is an average ± SEM. Asterisks indicate significant differences between experimental groups (*p < 0.05, **p < 0.01, ***p < 0.001).

References

    1. Andersen C. L., Jensen J. L., Ørntoft T. F. (2004). Normalization of real-time quantitative reverse transcription-PCR Data: a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. Cancer Res. 64, 5245–5250. 10.1158/0008-5472.CAN-04-0496 – DOI PubMed
    1. Barrera-Bailón B., Oliveira J. A. C., López D. E., Muñoz L. J., Garcia-Cairasco N., Sancho C. (2013). Pharmacological and neuroethological studies of three antiepileptic drugs in the genetic audiogenic seizure hamster (GASH:Sal). Epilepsy Behav. 28, 413–425. 10.1016/j.yebeh.2013.05.028 – DOI PubMed
    1. Barrera-Bailón B., Oliveira J. A. C., López D. E., Muñoz L. J., Garcia-Cairasco N., Sancho C. (2017). Pharmacological and neuroethological study of the acute and chronic effects of lamotrigine in the genetic audiogenic seizure hamster (GASH:Sal). Epilepsy Behav. 71, 207–217. 10.1016/j.yebeh.2015.11.005 – DOI PubMed
    1. Bisogno T., Burston J. J., Rai R., Allarà M., Saha B., Mahadevan A., et al. . (2009). Synthesis and pharmacological activity of a potent inhibitor of the biosynthesis of the endocannabinoid 2-arachidonoylglycerol. ChemMedChem 4, 946–950. 10.1002/cmdc.200800442 – DOI PubMed
    1. Boison D. (2016). Adenosinergic signaling in epilepsy. Neuropharmacology 104, 131–139. 10.1016/j.neuropharm.2015.08.046 – DOI PMC PubMed

Schaka

Related Posts

Synthesis, characterization and stress-testing of a robust quillaja saponin stabilized oil-in-water phytocannabinoid nanoemulsion

Aurora Cannabis Faces Class-Action Lawsuit, Closing of Edmonton Facility

North Carolina Lawmakers May Postpone Vote on Medical Cannabis Legislation Until 2022

Dried matrix spots in forensic toxicology

Signez la pétition !!!

 

846 signatures

Pétition ASBL Cannabis Belgique

Pourquoi une pétition ?

Nous sommes des personnes qui en avons assez de devoir aller dans la rue et avoir affaire à des réseaux criminels sans savoir où cela va nous conduire par après.

Nous sommes des personnes ayant des maladies, qui pour certaines sont rares, et utilisant pour médication le cannabis sous diverses formes (CBD,THC,THCv,CBDa,,,) sous l'accord de notre médecin.

Nous sommes des personnes responsables et honnêtes qui avons une vie épanouie et sans problèmes de vie ou sociaux.

Nous avons également une passion pour la plante de cannabis en elle-même et la cultiver est notre bonheur. De plus, nous pouvons nous soigner avec notre médication sans avoir peur des produits ou autres additifs contenus dans une plante que l'on peut trouver autre part.

Nous souhaitons pouvoir avoir notre médicament dans les normes de la santé publique, car un cannabis sain aide à réduire les frais de santé parfois conséquents pour la collectivité et le malade lui-même.

Nous sommes également des personnes responsables avec un rôle dans la société qui en avons assez d’être considérés comme des « hippies ou autres drogués », nous avons juste choisi notre médication et celle-ci a apporté les preuves de son efficacité dans le monde.

Nous connaissons déjà les produits dérivés comme le CBD et le THC que nous maîtrisons pour nous aider dans notre maladie « Je précise que nous ne sommes pas médecin et que nous nous basons sur 20 ans d’expérience médicale du cannabis des membres de notre ASBL et l'avis du médecin de famille ».

Nous désirons simplement ne plus nous cacher, et pouvoir aider les autres personnes le souhaitant.

Nous somme soucieux des ados et de la prévention à leur égard. Effectivement, nous sommes les acteurs parfaits pour répondre aux questions qu’ils se posent vu notre expérience cannabique et, de plus, nous pourrons leur expliquer les risques qu’ils encourent en achetant du cannabis dans la rue.

Le projet complet peut être demandé via mail " info@mcb.care " et sur le site internet : " http://mcb.care "

@ASBL McB

**votre signature**

Partagez avec vos amis

Articles récents

Catégories