Noise analysis of cytosolic calcium image data

Cellular Ca²⁺ signals are often constrained to cytosolic micro- or nano-domains where stochastic openings of Ca²⁺ channels cause large fluctuations in local Ca²⁺ concentration (Ca²⁺ ‘noise’). With the advent of TIRF microscopy to image the fluorescence of Ca²⁺-sensitive probes from attoliter volumes it has become possible to directly monitor these signals, which closely track the gating of plasmalemmal and ER Ca²⁺-permeable channels. Nevertheless, it is likely that many physiologically important Ca²⁺ signals are too small to resolve as discrete events in fluorescence recordings. By analogy with noise analysis of electrophysiological data, we explore here the use of statistical approaches to detect and analyze such Ca²⁺ noise in images obtained using Ca²⁺-sensitive indicator dyes. We describe two techniques - power spectrum analysis and spatio-temporal correlation - and demonstrate that both effectively identify discrete, spatially localized calcium release events (Ca²⁺ puffs). Moreover, we show they are able to detect localized noise fluctuations in a case where discrete events cannot directly be resolved.