The relative magnitudes and functional significance of Ca extrusion by Na-Ca exchange and by an Na.-independent mechanism were investigated in monolayer cultures of chick embryo ventricular cells. Abrupt exposure of cells in 0-Na., nominally 0-Ca. solution to 20 mM caffeine produced a large contracture (3 .94 ± 0.90 um of cell shortening) that relaxed with a tl, of 8.60 ± 1 .22 s. An abrupt exposure to caffeine plus 140 mM Na resulted in a contracture that was smaller in amplitude (1 .53 ± 0.50,um) and relaxed much more rapidly (t,, = 0.77 ± 0 .09 s) . An abrupt exposure to caffeine in 0-Na. solutions produced an increase in "Ca efflux that persisted for 20 s, and a net loss of Ca content, determined by atomic absorption spectroscopy (AAS), of^-4 nmol/mg protein, within 35 s. A comparable net loss of Ca was demonstrated in the presence of 100 jM [Ca]o. The abrupt exposure of cultured cells to 0 Na. in 1 .8 mM Ca produced a Ca uptake, estimated with 45 Ca, of 3.2 nmol/mg protein -15 s, but produced no increase in cell Ca content (AAS). In cells in which a 30% increase in Nai was produced by 5 min exposure to 10 -6 M ouabain, the abrupt exposure to 0 Na. produced a Ca uptake of 6 nmol/mg protein -15 s and an increase in Ca content (AAS) of 4 nmol/mg protein. We conclude that there is an Na.-independent mechanism for Ca extrusion in these cells, presumably a Ca-ATPase Ca pump, with a limited Ca transport capacity of no more than 2 nmol/mg protein -15 s. This is five times smaller than the demonstrated maximum capacity of the Na-Ca exchanger in these cells. The relaxation of twitch tension in these cells seems to be dependent primarily on sarcoplasmic reticulum uptake of Ca, with a secondary role provided by the Na-Ca exchanger. The Ca pump appears to contribute little to beat-to-beat relaxation .