Cellular effects of BAPTA: Are they only about Ca²⁺ chelation?

Intracellular Ca²⁺ signals play a vital role in a broad range of cell biological and physiological processes in all eukaryotic cell types. Dysregulation of Ca²⁺ signaling has been implicated in numerous human diseases. Over the past four decades, the understanding of how cells use Ca²⁺ as a messenger has flourished, largely because of the development of reporters that enable visualization of Ca²⁺ signals in different cellular compartments, and tools that can modulate cellular Ca²⁺ signaling. One such tool that is frequently used is BAPTA; a fast, high-affinity Ca²⁺-chelating molecule. By making use of a cell-permeable acetoxymethyl ester (AM) variant, BAPTA can be readily loaded into the cytosol of cells (referred to as BAPTAi), where it is trapped and able to buffer changes in cytosolic Ca²⁺. Due to the ease of loading of the AM version of BAPTA, this reagent has been used in hundreds of studies to probe the role of Ca²⁺ signaling in specific processes. As such, for decades, researchers have almost universally attributed changes in biological processes caused by BAPTAi to the involvement of Ca²⁺ signaling. However, BAPTAi has often been used without any form of control, and in many cases has neither been shown to be retained in cells for the duration of experiments nor to buffer any Ca²⁺ signals. Moreover, increasing evidence points to off-target cellular effects of BAPTA that are clearly not related to Ca²⁺ chelation. Here, we briefly introduce Ca²⁺ signaling and the history of Ca²⁺ chelators and fluorescent Ca²⁺ indicators. We highlight Ca²⁺-independent effects of BAPTAi on a broad range of molecular targets and describe some of BAPTAi's impacts on cell functions that occur independently of its Ca²⁺-chelating properties. Finally, we propose strategies for determining whether Ca²⁺ chelation, the binding of other metal ions, or off-target interactions with cell components are responsible for BAPTAi's effect on a particular process and suggest some future research directions.