(Endo)cannabinoids mediate different Ca(2+) entry mechanisms in human bronchial epithelial cells

In human bronchial epithelial (16HBE14o) cells, CB(1) and CB(2) cannabinoid receptors are present, and their activation by the endocannabinoid virodhamine and the synthetic non-selective receptor agonist CP55,940 inhibits adenylyl cyclase and cellular interleukin-8 release. Here, we analyzed changes in intracellular calcium ([Ca(2+)](i)) evoked by Delta(9)-tetrahydrocannabinol (Delta(9)-THC), CP55,940, and virodhamine in 16HBE14o(-) cells. Delta(9)-THC induced [Ca(2+)](i) increase and a large transient [Ca(2+)](i) mobilization, the latter probably reflecting store-depletion-driven capacitative Ca(2+) entry (CCE). In contrast, CP55,940 induced a rather moderate Ca(2+) influx and a sustained [Ca(2+)](i) mobilization. CP55,940-induced Ca(2+) influx was inhibited by Ni(2+), indicating CCE, possibly mediated by transient receptor potential channel TRPC1, the mRNA of which is expressed in 16HBE14o(-) cells. CP55,940-induced calcium alterations were mimicked by virodhamine concentrations below 30 mu M. Interestingly, higher virodhamine induced an additional Ca2+ entry, insensitive to Ni(2+), but sensitive to the TRPV1 antagonist capsazepine, the TRPV1-TRPV4 inhibitor ruthenium red, and the non-CCE (NCCE) inhibitors La(3+) and Gd(3+). Such pharmacological profile is supported by the presence of TRPV1, TRPV4, and TRPC6 mRNAs as well as TRPV1 and TRPC6 proteins in 16HBE14o(-) cells. Cannabinoid receptor antagonists increased virodhamine-induced Ca(2+) entry. Virodhamine also enhanced arachidonic acid release, which was insensitive to cannabinoid receptor antagonism, but sensitive to the phospholipase A(2) inhibitor quinacrine, and to capsazepine. Arachidonic acid induced [Ca(2+)](i) increase similar to virodhamine. Collectively, these observations suggest that [Ca(2+)](i) alterations induced by Delta(9)-THC, CP55,940 and by low concentrations of virodhamine involve mobilization and subsequent CCE mechanisms, whereas such responses by high virodhamine concentrations involve NCCE pathways