Transient Receptor Potential Ankyrin 1 (TRPA1) Mediates Hydrogen Sulfide-induced Ca(2+) Entry and Nitric Oxide Production in Human Cerebrovascular Endothelium.

INTRODUCTION: The gasotransmitter hydrogen sulfide (H(2)S) modulates various brain functions, including neuron excitability, synaptic plasticity, and Ca(2+) dynamics. Furthermore, H(2)S may stimulate nitric oxide (NO) release from cerebrovascular endothelial cells, thereby regulating NO-dependent endothelial functions, such as angiogenesis, vasorelaxation, and cerebral blood flow (CBF). However, the signaling pathway by which H(2)S induces NO release from cerebrovascular endothelial cells is still unclear. METHODS: Herein, we exploited single-cell imaging of intracellular Ca(2+), H(2)S, and NO levels to assess how H(2)S induces Ca(2+)-dependent NO release from the human cerebrovascular endothelial cell line, hCMEC/D3. RESULTS: Administration of the H(2)S donor, sodium hydrosulfide (NaHS), induced a dose-dependent increase in (Ca(2+))i only in the presence of extracellular Ca(2+). NaHS-induced extracellular Ca(2+) entry was mediated by the Ca(2+)-permeable TRPA1 channel, as shown by pharmacological and genetic manipulation of the TRPA1 protein. Furthermore, NaHS-dependent TRPA1 activation led to NO release that was abolished by buffering the concomitant increase in (Ca(2+))(i) and inhibiting eNOS. Furthermore, the endothelial agonist, adenosine trisphosphate (ATP), caused a long-lasting elevation in (Ca(2+))(i) that was driven by cystathionine γ-lyase (CSE)-dependent H2S production and by TRPA1 activation. Consistent with this, ATP-induced NO release was strongly reduced either by blocking CSE or by inhibiting TRPA1. CONCLUSION: These findings demonstrate for the time that H(2)S stimulates TRPA1 to induce NO production in human brain microvascular endothelial cells. Additionally, they show that this signaling pathway can be recruited by an endothelial agonist to modulate NO-dependent events at the human neurovascular unit.