Transient Receptor Potential Ankyrin 1 (TRPA1) Mediates Hydrogen Sulfide-induced Ca2+ Entry and Nitric Oxide Production in Human Cerebrovascular Endothelium.

Introduction: The gasotransmitter hydrogen sulfide (H₂S) modulates various brain functions, including neuron excitability, synaptic plasticity, and Ca²⁺ dynamics. Furthermore, H₂S 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 H₂S induces NO release from cerebrovascular endothelial cells is still unclear.

Methods: Herein, we exploited single-cell imaging of intracellular Ca²⁺, H₂S, and NO levels to assess how H₂S induces Ca²⁺-dependent NO release from the human cerebrovascular endothelial cell line, hCMEC/D3.

Results: Administration of the H₂S donor, sodium hydrosulfide (NaHS), induced a dose-dependent increase in (Ca²⁺)i only in the presence of extracellular Ca²⁺. NaHS-induced extracellular Ca²⁺ entry was mediated by the Ca²⁺-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 (Ca²⁺)_i and inhibiting eNOS. Furthermore, the endothelial agonist, adenosine trisphosphate (ATP), caused a long-lasting elevation in (Ca²⁺)_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 H₂S 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.