In most mammalian species, an increase in stimulation frequency (ISF) produces an increase in contractility (treppe phenomenon), which results from larger Ca2+ transients at higher frequencies, due to an increase in sarcoplasmic reticulum Ca2+ load and release. The present study attempts to elucidate the contribution of the Na+-Ca2+ exchanger (NCX) to this phenomenon. Isolated cat ventricular myocytes, loaded with [Ca2+]i- and [Na+]i-sensitive probes, were used to determine whether the contribution of the NCX to the positive inotropic effect of ISF is due to an increase in Ca2+ influx (reverse mode) and/or a decrease in Ca2+ efflux (forward mode) via the NCX, due to frequency-induced [Na+]i elevation, or whether it was due to the reduced time for the NCX to extrude Ca2+. The results showed that the positive intropic effect produced by ISF was temporally dissociated from the increase in [Na+]i and was not modified by KB-R7943 (1 or 5 μM), a specific blocker of the reverse mode of the NCX. Whereas the ISF from 10 to 30 beats min-1 (bpm) did not affect the forward mode of the NCX (assessed by the time to half-relaxation of the caffeine-induced Ca2+ transient), the ISF to 50 bpm produced a significant reduction of the activity of the forward mode of the NCX, which occurred in association with an increase in [Na+]i (from 4.33 ± 0.40 to 7.25 ± 0.50 mM). However, both changes became significant well after the maximal positive inotropic effect had been reached. In contrast, the positive inotropic effect produced by ISF from 10 to 50 bpm was associated with an increase in diastolic [Ca2+]i, which occurred in spite of a significant increase in the relaxation rate and at a time at which no increases in [Na+]i were detected. The contribution of the NCX to stimulus frequency inotropy would therefore depend on a decrease in NCX-mediated Ca2+ efflux due to the reduced diastolic interval between beats and not on [Na+]i-dependent mechanisms.Facultad de Ciencias MédicasCentro de Investigaciones Cardiovasculares