Capacitative Ca²⁺ entry (CCE) refers to the influx of Ca²⁺ through plasma membrane channels activated on depletion of endoplasmic-sarcoplasmic reticulum Ca²⁺ stores. We utilized two Ca²⁺-sensitive dyes (one monitoring cytoplasmic free Ca²⁺ and the other free Ca²⁺ within the sarcoplasmic reticulum) to determine whether adult rat ventricular myocytes exhibit CCE. Treatments with inhibitors of the sarcoplasmic endoplasmic reticulum Ca²⁺-ATPases were not efficient in releasing Ca²⁺ from stores. However, when these inhibitors were coupled with either Ca²⁺ ionophores or angiotensin II (an agonist generating inositol 1,4,5 trisphosphate), depletion of stores was observed. This depletion was accompanied by a significant influx of extracellular Ca²⁺ characteristic of CCE. CCE was also observed when stores were depleted with caffeine. This influx of Ca²⁺ was sensitive to four inhibitors of CCE (glucosamine, lanthanum, gadolinium, and SKF-96365) but not to inhibitors of L-type channels or the Na⁺/Ca²⁺ exchanger. In the whole cell configuration, an inward current of ∼0.7 pA/pF at –90 mV was activated when a Ca²⁺ chelator or inositol (1,4,5)-trisphosphate was included in the pipette or when Ca²⁺ stores were depleted with a Ca²⁺-ATPase inhibitor and ionophore. The current was maximal at hyperpolarizing voltages and inwardly rectified. The channel was relatively permeant to Ca²⁺ and Ba²⁺ but only poorly to Mg²⁺ or Mn²⁺. Taken together, these data support the existence of CCE in adult cardiomyocytes, a finding with likely implications to physiological responses to phospholipase C-generating agonists.