Diffusional mobility of parvalbumin in spiny dendrites of cerebellar Purkinje neurons quantified by fluorescence recovery after photobleaching

Ca2+-binding proteins (CaBPs) represent key factors for the modulation of cellular Ca2+ dynamics. Especially in thin extensions of nerve cells, Ca2+ binding and buffered diffusion of Ca2+ by CaBPs is assumed to effectively control the spatio-temporal extend of Ca2+ signals. However, no quantitative data about the mobility of specific CaBPs in the neuronal cytosol are available. We quantified the diffusion of the endogenous CaPB parvalbumin (PV) in spiny dendrites of cerebellar Purkinje neurons with two-photon fluorescence recovery after photobleaching. Fluorescently labeled PV diffused readily between spines and dendrites with a median time constant of 49 ms (37-61 ms, interquartile range). Based on published data on spine geometry, this value corresponds to an apparent diffusion coefficient of 43 mum(2) s(-1) (34-56 mum(2) s(-1)). The absence of large or immobile binding partners for PV was confirmed in PV null-mutant mice. Our data validate the common but so far unproven assumption that PV is highly mobile in neurons and will facilitate simulations of neuronal Ca2+ buffering. Our experimental approach represents a versatile tool for quantifying the mobility of proteins in neuronal dendrites