Sarcoplasmic reticulum is a specialized membrane system in muscle involved in the energized uptake, storage, and release of Ca2+. The sulfhydryl content of normal and reconstituted sarcoplasmic reticulum was measured using Ellman reagent. For both preparations, we find 17 and 26 mol sulfhydryls per mole calcium pump protein assayed in the absence and presence of sodium dodecyl sulfate. The release of Ca2+ from sarcoplasmic reticulum, which triggers muscle contraction, likely involves the regulation of a channel. This report deals with an experimental approach to studying the Ca2+ release in isolated sarcoplasmic reticulum. We find that sulfhydryl agents of which water-soluble mercurials were most effective induce Ca2+ release. Chlorpromazine acts synergistically with the sulfhydryl reagents. Ca2+ release under optimal conditions is very rapid compared with calcium leakage from preloaded but untreated sarcoplasmic reticulum. The imposed rapid release of Ca2+ is suggestive of the opening of a channel. Ca2+ release by mercurials is retained in reconstituted sarcoplasmic reticulum membrane vesicles.