Elucidating the transcriptomic response of adult-derived mHypoA-2/12 mouse hypothalamic neuron cell line to cannabidiol (CBD) exposure

Abstract

Cannabidiol (CBD) is a compound found in Cannabis sativa that is known for its neuroprotective, anti-inflammatory, analgesic, and anxiolytic properties. These properties make it a promising treatment for various neurological conditions. This study aimed to examine the effects of CBD on hypothalamic neurons at the transcriptome level using the adult-derived mHypoA-2/12 mouse cell line. The cells were exposed to four different CBD concentrations (ranging from 0.325 to 3 µM) for 6 and 24 h. Apart from the transcriptome analysis, apoptosis (caspase 3/7 activity) and viability (MTT) assays were performed. The obtained results showed that CBD enhanced cell viability, especially after 24 h of treatment and at lower or intermediate concentrations, and reduced apoptosis, with significant effects at the highest concentration. CBD caused moderate transcriptome profile changes (13 to 69 genes per treatment), with more genes affected at higher concentrations and shorter exposure times, indicating a stronger initial cellular response. Further analysis revealed that CBD affects several biological processes, including: intrinsic apoptosis suppression via p53 modulation, impacting genes like Bbc3, Mdm2, Cdkn1a, and Smad3. Additionally, CBD influenced genes involved in extracellular matrix organization, including metalloproteinases (Mmp-3, Mmp-13) and their inhibitors (Timp1), as well as collagen components (Col11a1) and mitochondrial respiratory chain complexes (mt-Nd5, mt-Nd4). Genes related to serotonin and dopamine biosynthesis, as well as Aldh2, were also impacted, linking CBD's effects in hypothalamic neurons to potential benefits in managing alcohol use disorders. These findings suggest the hypothalamus is a significant target for CBD, warranting further investigation.

Keywords: Apoptosis; CBD; Extracellular matrix; Hypothalamic neurons; RNA-seq.

Fig. 1

Results of caspase 3/7 and MMT assay for cells treated with various concentrations of CBD. P-values of the Kruskal–Wallis ANOVA test are provided. *Statistically significant difference (adjp < 0.05) after correction for multiple testing for comparison of a specific CBD concentration against vehicle control

Fig. 2

Expression profile differentiation using principal components analysis (PCA) method among cells treated with different CBD concentrations and after different times of CBD treatment

Fig. 3

Results of differential expression analysis among cells treated with different concentrations of CBD for 6 h. A Number of differentially expressed genes and their regulation; B volcano plot for all analyzed genes (significantly altered genes (FDR < 0.1) are marked in red/green); C heatmaps for differentially expressed genes

Fig. 4

Results of differential expression analysis among cells treated with different concentrations of CBD for 24 h. A Number of differentially expressed genes and their regulation; B volcano plot for all analyzed genes (significantly altered genes (FDR < 0.1) are marked in red/green); C heatmaps for differentially expressed genes

Fig. 5

Venn diagrams for genes altered by different CBD treatments (concentrations) and treatment times

Fig. 6

Result of overrepresentation test in GO biological processes for all genes affected by CBD in 6-h treatment

Fig. 7

Result of overrepresentation test in GO biological processes for all genes affected by CBD in 24-h treatment