The interplay between plasma membrane and endoplasmic reticulum Ca(2+)ATPases in agonist-induced temporal Ca(2+) dynamics.

Abstract

A change in the intracellular free Ca(2+) concentration ([Ca(2+)]i) functions as a transmitter for signal transduction and shows a broad temporal pattern. Even genetically homogeneous cell types show different Ca(2+) response patterns under permanent agonist stimulation. In Ca(2+) signaling, the dynamics of the Ca(2+) release from the Ca(2+) channels during continuous agonist stimulation and the simultaneous effect of the pumps are unclear. In this study, the dynamic interaction of the Ca(2+) ATPases in the plasma membrane (PMCA) and the endoplasmic reticulum membrane (SERCA) during continuous ACh stimulation is monitored using Fluo-3 and Fura-2 loaded HEK 293 cells. We characterize Ca(2+) release patterns at the sub-maximal and maximal stimulation doses in the absence of extracellular Ca(2+). We analyze the responses regarding their types, oscillation frequency and response times. La(3+) (PMCA blocker) do not change the frequency and time courses in sub-maximal ACh treatment, while with the maximal stimulation oscillation frequency increase as oscillations superimpose on robust release, and response time of [Ca(2+)]i is elongated. A similar effect of La(3+) is observed in quantal Ca(2+) release phenomenon. In the presence of CPA, a SERCA blocker, oscillations are completely abolished, but response time does not change. We also observe that during continuous receptor stimulation, Ca(2+) release do not cease. These data may suggest that Ca(2+) release continues during agonist stimulation, but SERCA and PMCA form a new steady state and return [Ca(2+)]i to its physiological concentration.