Hydrogen sulfide (H2S) is recognized as a physiologic mediator produced in a variety of tissues. It is produced by three enzymes, cystathionine p-synthase (CBS), cystathionine gamma-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3MST). However, the regulation of H2S production by CSE has not well been understood. Here we show that H2S producing activity of CSE is regulated by intracellular Ca2+ concentrations. In the presence of pyridoxal 5'-phosphate (PLP) CSE efficiently produces H2S at the steady-state low Ca2+ concentrations but is suppressed at high Ca2+ concentrations. In the absence of PLP H2S production maintains the suppressed levels at high Ca2+ concentrations and decreased further at low Ca2+ concentrations. These observations suggest that CSE produces H2S at the steady-state in cells and that the production is suppressed when the intracellular Ca2+ concentrations are increased. (C) 2013 Elsevier Inc. All rights reserved.