EFFECTS OF CYCLOPIAZONIC ACID ON [CA2+](I) AND CONTRACTION IN RAT URINARY-BLADDER SMOOTH-MUSCLE

Cyclopiazonic acid (CPA) has been reported to inhibit the Ca2+-ATPase of the sarcoplasmic reticulum (SR) in skeletal and smooth muscle. In the present study the effect of CPA on [Ca2+](i) and force in rat urinary bladder smooth muscle was examined. The fluorescent Ca2+ indicator Fura-2 was used to monitor intracellular Ca2+, simultaneously with isometric force production. Addition of CPA to unstimulated muscles bathed in 2.5 mM Ca2+ containing Krebs solution resulted in a significant and sustained increase in [Ca2+](i) from 99 +/- 7 to 273 +/- 51 nM. This increase in [Ca2+](i) was dependent upon the presence of extracellular Ca2+ since when CPA was added to muscles in Ca2+-free media it produced only a small, transient increase in [Ca2+](i) that was not sustained. Peak force levels produced by transmural stimulation, carbachol and high KCI solution were not altered by the presence of CPA, however, the increase in [Ca2+](i) associated with these contractions was larger when CPA was present. In response to transmural stimulation, the times taken for both force and [Ca2+](i) to rise to 50% of their peak values were attenuated in the presence of CPA. Conversely, there was no effect of CPA on the times taken for force or [Ca-2+](i) to fall to 50% of their stimulated values upon the cessation of stimulation. Under control conditions both carbachol and high KCI could initiate transient increases in [Ca2+](i) and force in the absence of extracellular Ca2+. In the presence of CPA, the response to carbachol was virtually completely inhibited, however, the response to high KCI was only partially inhibited. The ability of CPA to inhibit the carbachol response in Ca2+-free media suggests that this response is due to release of Ca2+ from the SR. The incomplete inhibition of the response to KCI indicates other Ca2+ storage sites may also be mobilised by sarcolemmal depolarisation. Although the mechanism whereby CPA induces a large, sustained rise in [Ca2+](i) remains unknown, the data lend support to the suggestion that depletion of intracellular Ca2+ storage sites may activate a Ca2+ entry pathway across the sarcolemma.