The effect of active serum albumin on PC12 cells: II. Intracellular Ca 2+ transients and their role in neurite retraction

Abstract

In the preceding paper it was shown that an isoform of serum albumin (ASA: active serum albumin) causes a rapid retraction of neurites and increases intracellular content of Ins1,4,5P3 in PC12 cells. Here we examined whether ASA's effects in nerve growth factor-differentiated PC12 cells were mediated through the Ins1,4,5P3/Ca 2+ second messenger pathway by monitoring intracellular Ca 2+ (Ca 2+ i) with Fura2. It was found that ASA caused a dose-dependent increase in Ca 2+ i. In Ca 2+-free medium, the increase in Ca 2+ i elicited by ASA was smaller, but the rise in Ins1,4,5P3 content was not appreciably changed. The small Ca 2+ i increase seen in Ca 2+-free medium was probably due to the release of Ca 2+ from Ins1,4,5P3-sensitive intracellular stores. In Ca 2+-containing medium the Ca 2+ transient induced by ASA was not affected by organic Ca 2+ channel blockers, but decreased when Co 2+, Mn 2+ or Zn 2+ were present in the extracellular medium. The effect of other ligands, such as carbachol and bradykinin, whose receptors are coupled to the phosphoinositide system was also investigated. Carbachol at concentrations from 2 to 200 μM, and bradykinin at a concentration of 2 μM did not cause neurite retraction, whereas 200 μM bradykinin caused an approximately 40% decrease in neurite length. Thapsigargin, a Ca 2+-ATPase inhibitor, caused a sustained elevation of Ca 2+ i and retraction of neurites, whereas depolarization of the cells by high K + gave only a transient elevation of Ca 2+ i , and no neurite retraction. Therefore, a sustained elevation in Ca 2+ i might be a sufficient trigger to induce neurite retraction in differentiated PC12 cells. Since ASA induces only a transient rise in Ca 2+ i in most cells, the Ca 2+ i transient evoked by ASA alone may not be sufficient to cause neurite retraction in PC12 cells.