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The effects of cannabinoids on the viability and differentiation of neurons derived from retinoic acid-induced  P19 embryonal carcinoma cells
Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology. (Jacobsson)
Umeå University, Faculty of Medicine, Department of Radiation Sciences, Oncology.
Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Cannabinoids and cannabinoid receptors play an important role in development and differentiation of the nervous system, but the mechanisms behind that role have not been fully elucidated. We have examined the effects of synthetic and endogenous cannabinoids and related polyunsaturated fatty acids upon mouse embryonal carcinoma P19 stem cell viability - before, during and after retinoic acid (RA)-induced neural differentiation. Experiments were also performed to investigate whether the cannabinoids affect the differentiation of P19-derived neurons by measuring the development and growth of neurites and intracellular acetylcholinesterase activity.

Both synthetic and endogenous cannabinoids as well as related fatty acids produced a concentration-dependent decrease in undifferentiated P19 cell viability, but induction of the neural pathway reduced the sensitivity to the cytotoxic effects, and in differentiated neurons anandamide and related fatty acids showed no cytotoxicity. However, synthetic cannabinoids such as HU 210, HU 211 and WIN 55,212-2 produced cytotoxicity in both undifferentiated and differentiated cells, but there was a right-shifted concentration-effect curve in RA-induced cells and differentiated neurons compared with the undifferentiated cells.

HU 210 produced a time- and concentration-dependent decrease in cell number, percentage of cells expressing neurites, number of neurites per cell and neurite length. Statistically significant inhibition was seen at a concentration of 1 µM to 3 µM, and this was confirmed by the measurement of intracellular acetylcholinesterase activity, an enzyme that is dramatically increased during the differentiation process, where HU 210 significantly decreased the activity after six and nine days of exposure. However, these effects of HU 210 could only be observed in the same concentration range as those affecting neuronal viability. Anandamide, on the other hand, had modest effect on measured markers of neuronal differentiation but decreased the fraction of neurite expressing cells and neurite length after nine days of exposure at a concentration ≥ 10 µM. No effect on the acetylcholinesterase activity was observed.

It is concluded that cannabinoids and related fatty acids have cytotoxic effects in undifferentiated P19 embryonal carcinoma cells, but induction of the neuronal pathway reduces the sensitivity to the cytotoxic effects. The synthetic cannabinoids are more potent than the endogenous cannabinoids and fatty acids in causing cytotoxicity in differentiated neurons, but the CB-induced decrease in neurite formation and acetylcholinesterase activity in RA-induced P19-derived neurons occurs only at concentrations that cause measurable neuronal cell death. 

Keyword [en]
Cannabinoids, P19 EC cells, cell viability, neuronal differentiation, neurite formation, acetylcholinesterase activity
National Category
Pharmacology and Toxicology
Research subject
biokemisk farmakologi
URN: urn:nbn:se:umu:diva-51555OAI: diva2:484044
Available from: 2012-01-31 Created: 2012-01-26 Last updated: 2012-02-10Bibliographically approved
In thesis
1. Cannabinoids as modulators of cancer cell viability, neuronal differentiation, and embryonal development
Open this publication in new window or tab >>Cannabinoids as modulators of cancer cell viability, neuronal differentiation, and embryonal development
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Alternative title[sv]
Effekter av cannabinoider på cancerceller, neuronal differentiering och embryonal utveckling
Abstract [en]

Cannabinoids (CBs) are compounds that activate the CB1 and CB2 receptors. CB receptors mediate many different physiological functions, and cannabinoids have been reported to decrease tumor cell viability, proliferation, migration, as well as to modulate metastasis.

In this thesis, the effects of cannabinoids on human colorectal carcinoma Caco-2 cells (Paper I) and mouse P19 embryonal carcinoma (EC) cells (Paper III) were studied.  In both cell lines, the compounds examined produced a concentration- and time-dependent decrease in cell viability. In Caco-2-cells, HU 210 and the pyrimidine antagonist 5-fluorouracil produced synergistic effects upon cell viability. The mechanisms behind the cytocidal effects of cannabinoids appear to be mediated by other than solely the CB receptor, and a common mechanism in Caco-2 and P19 EC cells was oxidative stress. However, in P19 EC cells the CB receptors contribute to the cytocidal effects possibly via ceramide production.

In paper II, the association between CB1 receptor immunoreactivity (CB1IR) and different histopathological variables and disease-specific survival of colorectal cancer (CRC) was investigated. In microsatellite stable (MSS) cases there was a significant positive association of the tumor grade with the CB1IR intensity. A high CB1IR is indicative of a poorer prognosis in MSS with stage II CRC patients.

Paper IV focused on the cytotoxic effects of cannabinoids during neuronal differentiation. HU 210 affected the cell viability, neurite formation and produced a decreased intracellular AChE activity. The effects of cannabinoids on embryonic development and survival were examined in Paper V, by repeated injection of cannabinoids in fertilized chicken eggs. After 10 days of incubation, HU 210 and cannabidiol (without CB receptor affinity), decreased the viability of chick embryos, in a manner that could be blocked by α-tocopherol (antioxidant) and attenuated by AM251 (CB1 receptor antagonist).

In conclusion, based on these studies, the cannabinoid system may provide a new target for the development of drugs to treat cancer such as CRC. However, the CBs also produce seemingly unspecific cytotoxic effects, and may have negative effects on the neuronal differentiation process. This may be responsible for, at least some of, the embryotoxic effects found in ovo, but also for the cognitive and neurotoxic effects of cannabinoids in the developing and adult nervous system.

Place, publisher, year, edition, pages
Umeå: Umeå universitet, 2012. 48 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1474
Cannabinoids, cell viability, neuronal differentiation, colorectal cancer, chick embryo
National Category
Pharmacology and Toxicology
Research subject
biokemisk farmakologi; Toxicology
urn:nbn:se:umu:diva-51560 (URN)978-91-7459-358-7 (ISBN)
Public defence
2012-02-24, Sal E04, by 6E, Norrlands Universitetssjukhus, Umeå, 09:00 (Swedish)
Available from: 2012-02-03 Created: 2012-01-26 Last updated: 2012-02-27Bibliographically approved

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