Differentiation induces up‐regulation of plasma membrane Ca2+‐ATPase and concomitant increase in Ca2+ efflux in human neuroblastoma cell line IMR‐32

Precise regulation of intracellular Ca2+ concentration ([Ca2+]i) is achieved by the coordinated function of Ca2+ channels and Ca2+ buffers. Neuronal differentiation induces up‐regulation of Ca2+ channels. However, little is known about the effects of differentiation on the expression of the plasma membrane Ca2+‐ATPase (PMCA), the principal Ca2+ extrusion mechanism in neurons. In this study, we examined the regulation of PMCA expression during differentiation of the human neuroblastoma cell line IMR‐32. [Ca2+]i was monitored in single cells using indo‐1 microfluorimetry. When the Ca2+‐ATPase of the endoplasmic reticulum was blocked by cyclopiazonic acid, [Ca2+]i recovery after small depolarization‐induced Ca2+ loads was governed primarily by PMCAs. [Ca2+]i returned to baseline by a process described by a monoexponential function in undifferentiated cells (τ = 52 ± 4 s; n = 25). After differentiation for 12–16 days, the [Ca2+]i recovery rate increased by more than threefold (τ = 17 ± 1 s; n = 31). Western blots showed a pronounced increase in expression of three major PMCA isoforms in IMR‐32 cells during differentiation, including PMCA2, PMCA3 and PMCA4. These results demonstrate up‐regulation of PMCAs on the functional and protein level during neuronal differentiation in vitro. Parallel amplification of Ca2+ influx and efflux pathways may enable differentiated neurons to precisely localize Ca2+ signals in time and space.