It has recently been shown that significant amounts of Ca²⁺ are stored in the neonatal rabbit heart SR. Here we use the perforated patch-clamp technique to determine developmental changes in SR Ca²⁺ loading mechanisms and Ca²⁺ pump efficacy in rabbit ventricular myocytes at 3, 10, 20 and 56 days (d) of age. SR Ca²⁺ loading (loadSR) and SR Ca²⁺ pump Ca²⁺ affinity were greater in younger age groups. Inhibition of the L-type calcium current (ICa) with 15 µM nifedipine dramatically reduced loadSR in older but not in younger age groups (78% in 56d vs. 7% in 3d). In contrast, subsequent inhibition of the Na⁺ / Ca²⁺ exchanger (NCX) with 10 µM KB-R 7943 strongly reduced loadSR in the younger but not the older age groups (49% in 3d vs. 1% in 56d). Accordingly, the time integral of the inward NCX current (tail INCX) elicited upon repolarization was highly sensitive to nifedipine in the older age groups (94% in 56d vs. 15% in 3d ) and sensitive to KB-R 7943 in the younger groups (85% in 3d vs. 6% in 56d). We conclude that the SR loading capacity at the earliest postnatal stages is at least as large as that of adult myocytes. However, reverse mode NCX plays a prominent role in SR Ca²⁺ loading at early postnatal stages while I_Ca is the main source of SR Ca²⁺ loading at late postnatal and adult stages.