Abstract
To identify plasma membrane ion channels mediating calcium influx at the blood-brain barrier in response to disrupting stimuli. We examined the expression and function of candidate transient receptor potential channels using reverse transcriptase polymerase chain recation, Fura-2 calcium imaging, and permeability assays. Immortalized mouse brain microvessel endothelial cells expressed multiple transient receptor potential isoforms: transient receptor potential C1, C2, C4, and C7, M2, M3, M4, and M7, and V2 and V4. Similar profiles were observed in freshly isolated cerebral microvessels and primary cultured rat brain endothelial cells. Thrombin-stimulated calcium influx in brain endothelial cells was blocked by transient receptor potential C inhibitors. Transient receptor potential V activating stimuli also increased intracellular calcium. This increase was inhibited by a transient receptor potential V blocker or by removal of extracellular calcium. Barrier integrity was compromised by thrombin, hypo-osmolar stress, and PMA treatment. The increase in barrier permeability induced by transient receptor potential V activators was blocked by transient receptor potential V inhibition, while thrombin effects were inhibited by transient receptor potential C inhibitors. These results demonstrate that transient receptor potential C and transient receptor potential V channels mediate calcium influx at the blood-brain barrier, and as a consequence, may modulate barrier integrity.
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