Modulation of human T-type calcium channels by synthetic cannabinoid receptor agonists in vitro.

BACKGROUND AND PURPOSE: Consumption of Synthetic Cannabinoid Receptor agonists (SCRAs) is associated with severe adverse reactions including seizures, arrhythmias and death, but the molecular mechanisms surrounding SCRA toxicity are not yet established. These disease-like symptoms are also synonymous with altered T-type calcium channel activity which controls rhythmicity in the heart and brain. This study examined whether SCRAs alter T-type activity and whether this represents a possible mechanism of toxicity. EXPERIMENTAL APPROACH: Fluorescence-based and electrophysiology assays were used to screen 16 structurally related synthetic cannabinoids for their ability to inhibit human T-type calcium channels expressed in HEK293 cells. The most potent compounds were then further examined using patch clamp electrophysiology. KEY RESULTS: MDMB-CHMICA and AMB-CHMINACA potently blocked Cav3.2 with IC50 values of 1.5 and 0.74 µM respectively. Current inhibition increased from 47 to 80% and 45-87% respectively when the channel was in slow-inactivated state. Both SCRAs had little effect on steady state inactivation, however MDMB-CHMICA significantly shifted the half activation potential by -7mV. Neither drug produced frequency dependent block, in contrast to the phytocannabinoid Δ9-THC. CONCLUSIONS AND IMPLICATIONS: SCRAs are potent agonists of CB1 receptors and can be extremely toxic, but observed toxicity also resembles symptoms associated with altered Cav3.2 activity. Many SCRAs tested were potent modulators of Cav3.2, raising the possibility that SC toxicity may be due in part to Cav3.2 modulation. This potent T-type channel modulation suggests the possibility of SCRAs as a new drug class with potential to treat diseases associated with altered T-type channel activity. This article is part of the special issue on 'Cannabinoids'.

Effects of common anesthetic agents on [18F]flumazenil binding to the GABAA receptor.

BACKGROUND: The availability of GABAA receptor binding sites in the brain can be assessed by positron emission tomography (PET) using the radioligand, [18F]flumazenil. However, the brain uptake and binding of this PET radioligand are influenced by anesthetic drugs, which are typically needed in preclinical imaging studies and clinical imaging studies involving patient populations that do not tolerate relatively longer scan times. The objective of this study was to examine the effects of anesthesia on the binding of [18F]flumazenil to GABAA receptors in mice. METHODS: Brain and whole blood radioactivity concentrations were measured ex vivo by scintillation counting or in vivo by PET in four groups of mice following administration of [18F]flumazenil: awake mice and mice anesthetized with isoflurane, dexmedetomidine, or ketamine/dexmedetomidine. Dynamic PET recordings were obtained for 60 min in mice anesthetized by either isoflurane or ketamine/dexmedetomidine. Static PET recordings were obtained at 25 or 55 min after [18F]flumazenil injection in awake or dexmedetomidine-treated mice acutely anesthetized with isoflurane. The apparent distribution volume (VT*) was calculated for the hippocampus and frontal cortex from either the full dynamic PET scans using an image-derived input function or from a series of ex vivo experiments using whole blood as the input function. RESULTS: PET images showed persistence of high [18F]flumazenil uptake (up to 20 % ID/g) in the brains of mice scanned under isoflurane or ketamine/dexmedetomidine anesthesia, whereas uptake was almost indiscernible in late samples or static scans from awake or dexmedetomidine-treated animals. The steady-state VT* was twofold higher in hippocampus of isoflurane-treated mice and dexmedetomidine-treated mice than in awake mice. CONCLUSIONS: Anesthesia has pronounced effects on the binding and blood-brain distribution of [18F]flumazenil. Consequently, considerable caution must be exercised in the interpretation of preclinical and clinical PET studies of GABAA receptors involving the use of anesthesia.