Abstract
Objective: To gain some insight on the lesser arrhythmogenic properties of PST2744 [(E,Z)-3-((2-aminoethoxy)imino)androstane-6,17-dione hydrochloride] compared with digoxin, we compared modulation of intracellular Ca2+ dynamics by the two agents. Methods: SERCA (sarcoplasmic reticulum Ca2+-ATPase) activity and Ca2+ leak rate were measured in sarcoplasmic reticulum (SR) vesicles from guinea pig ventricles. Membrane current, intracellular Ca2+, and twitch amplitude were evaluated in guinea pig ventricular myocytes with or without blockade of the Na+/Ca2+ exchanger. Results: In SR vesicles, PST2744 (30–300 nM), but not digoxin, increased SERCA activity; digoxin only (≥0.1 nM) increased SR Ca2+ leak. In myocytes with blocked Na+/Ca2+ exchanger, Ca2+ reloading of caffeine-depleted SR was enhanced by PST2744 and slightly inhibited by digoxin. In myocytes with functioning Na+/Ca2+ exchanger, both agents increased diastolic Ca2+, SR Ca2+ content, the gain of Ca2+-induced Ca2+ release, the rate of cytosolic Ca2+ decay, twitch amplitude, and relaxation rate. Consistent with the observations in SR vesicles, the effects on SR Ca2+ content and Ca2+ decay rate were significantly larger for PST2744 than for digoxin. Conclusions: In isolated SR vesicles, PST2744 and digoxin directly affected SR function in opposite ways; this could be reproduced in myocytes during Na+/Ca2+ exchanger blockade. Under physiological conditions (functioning Na+/Ca2+ exchanger), the two agents affected Ca2+ dynamics in the same direction, as expected by their Na+/K+ pump inhibition; however, differential SR modulation was still expressed by quantitative differences. Thus, the more favorable inotropy-to-toxicity ratio previously described for PST2744 appears to be associated with direct SERCA stimulation and/or lack of enhancement of Ca2+ leak.
Footnotes
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This work was funded by Istituto di Ricerche Prassis Sigma-Tau and from Grant Ministero Università e Ricerca Scientifica e Tecnologica (MURST) 2000 (to A.Z.). Collaboration with Debrecen University was supported by an Italy/Hungary cooperation program of the Ministry for Foreign Affairs.
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doi:10.1124/jpet.104.077933.
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ABBREVIATIONS: SR, sarcoplasmic reticulum; PST2744, (E,Z)-3-((2-aminoethoxy)imino)androstane-6,17-dione hydrochloride; PIPES, 1,4-piperazinediethanesulfonic acid; SERCA, sarcoplasmic reticulum Ca2+-ATPase; MOPS, 4-morpholinepropanesulfonic acid; A.U., activity units (micromoles of Pi per milligrams of protein per minute; APIII, antipyrylazo-III; CICR, Ca2+-induced Ca2+ release; DMSO, dimethyl sulfoxide.
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↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
- Received September 15, 2004.
- Accepted November 30, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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