Focal sarcoplasmic reticulum calcium stores and diffuse inositol 1,4,5-trisphosphate and ryanodine receptors in human...

Intracellular calcium stores of human uterine myocytes in primary and second passage cell culture were visualized using the low-affinity calcium-sensitive fluorescent dye, fluo-3FF. The calcium stores appeared as numerous small (0.2-0.5 microm diameter) focal fluorescences. The stores were not depleted by exposing the cells to oxytocin or ryanodine under standard conditions. The stores were rapidly depleted by oxytocin or ryanodine exposure when sarcoplasmic reticulum (SR) calcium re-uptake was inhibited by pretreatment with thapsigargin. Immunofluorescence experiments indicated that both ryanodine and inositol 1,4,5-trisphosphate (IP(3)) receptors were smoothly distributed throughout the SR, and neither receptor co-localized with the calcium stores. Since IP(3) and ryanodine calcium channels are tightly associated with their receptor, these results suggest that SR calcium release occurs via second messenger channels that are remote from the SR calcium stores. These observations are consistent only with a mechanism for release of calcium stores where the SR serves three functions: (1) as site of calcium storage, (2) as the structure that contains the IP(3)- and ryanodine receptors and their associated release channels, and (3) as a conduit between the calcium stores and the release channels.