Thymoquinone (TQ) is a bioactive component found in many medicinal herbs. In this study, we report the smooth and cardiac muscle relaxant activities of this compound. TQ concentration dependently suppressed spontaneously contracting rabbit jejunum while also relaxed high K+-(80 mM) induced contractions in jejunum and guinea-pig ileum, indicating activity at voltage-operated Ca++channels (VOCC). Further, TQ displaced Ca++concentration-response curves, obtained in a Ca++-free environment, to the right, showing blockade of VOCC. Similar activity was observed with verapamil, a standard VOCC blocker. TQ also exhibited nonadrenergic relaxation of agonist-induced contractions in guinea-pig trachea. When tested in fluo-4-loaded mouse lung slices, TQ inhibited ACh-induced airway narrowing and Ca++signalling in airway smooth muscle cells. In endothelium-intact and endothelium-denuded rat aorta, TQ inhibited high K+-induced contractions at significantly lower concentrations than phenylephrine-(PE-) (1 microM) induced contractions. Relaxation of PE-induced contractions was resistant to blockade by L-NAME and atropine. In guinea-pig atria, TQ showed noncholinergic relaxation of atrial force and rate of contractions. These data suggest smooth and cardiac muscle relaxant activity of TQ possibly mediated, in part, via blockade of VOCC. The results also justify the use of TQ containing plants in related health disorders like colic, diarrhoea, cough, and asthma.