Neuropharmacology and toxicity of synthetic cannabinoid legal highs (novel psychoactive substances) tested in rat brain slices and neural stem cells

Background: Legal highs or new/novel psychoactive substances (NPS) are increasingly being abused in the UK and elsewhere. There are now more than 500 NPS, most of which can be easily bought using the Internet. The largest class of NPS are the synthetic cannabinoids and anecdotal evidence suggests that these are more potent at the cannabinoid CB1 receptor than THC, there have also been suggestions that these drugs might have activity at glutamate NMDA and 5-HT2A receptors, which contribute to their psychoactive effects. Here we describe some of the neuropharmacology of 4 new synthetic cannabinoids: STS-135, 5f-AKB48, BB22 and 5F-PB22. In addition, as new evidence suggest a neurotoxic effect of synthetic cannabinoid NPSon neural stem cells (NSC),the present in vitro toxicity study aimed to investigate the effects of the above drugs in NSC proliferation (BrdU), mitochondrial metabolic activity (MTT) and cell death (LDH). Finally, we evaluated the effect of these synthetic cannabinoids on tumour necrosis factor (TNF) release in NSC.

Methods: To examine the neuropharmacology of these synthetic cannabinoids we used quantitative autoradiography in male rat brain sections representing prefrontal cortex and striatum; we tested the ability of the drugs (0.1 – 30 μM) to compete with radioligands for the glutamate NMDA and 5-HT2A receptors, [3H]MK-801 and [3H]ketanserin, respectively.

To test the addictive potential of these drugs we examined their effects (1 and 10 μM)on dopamine efflux in 400μm rat accumbens brain slices superfused with artificial cerebrospinal fluid. We electrically evoked dopamine efflux using 900ms (10 pulses at 10 Hz) stimulations and recorded dopamine using fast cyclic voltammetry at carbon fibre microelectrodes.

The above drugs were also tested for toxicity at a range of concentrations (10nM, 100nM, 1μM and 10μM),t=24h,in NSC from wild type mice (WT, C57BL6J).

Results: i) The above synthetic cannabinoids did not decrease the binding of [3H]MK801 or [3H]ketanserin suggesting that they had no effects on the NMDA or 5-HT2A receptors, respectively. ii) All synthetic cannabinoids but 5fAKB48, decreased dopamine efflux by about 30-40% at 10 μM suggesting that these drugs do not have an addictive profile. iii) All the tested synthetic cannabinoids increase cell death (LDH release) in NSC (P<0.001)iv) All the tested synthetic cannabinoids, except 5FPB22 (20% increase) decrease mitochondrial metabolic activity (MTT) in NSC (BB22:40% decrease,5FAKB48: 10% decrease and STS135: 10% decrease approx.). v) All the above tested drugs increased TNF release in NSC (BB22: P<0.001; 5FPB22: P<0.001; 5FAKB48: P<0.05; STS135: P<0.001).

Conclusion As the synthetic cannabinoids tested here did not bind to the NMDA or 5-HT2A receptors, their psychoactive effects are likely to be mediated through the CB1 receptor. These drugs tended to decrease evoked dopamine efflux, and therefore they may not be highly addictive. Their neurotoxicity induced in NSC deserves further attention.