Two-photon Na+ imaging in spines and fine dendrites of central neurons

Dendritic spines are assumed to be the smallest units of neuronal integrati on. Because of their miniature size, however, many of their functional prop erties are still unclear. New insights in spine physiology have been provid ed by two-photon laser-scanning microscopy which allows fluorescence imagin g with high spatial resolution and minimal photodamage, For example, two-ph oton imaging has been employed successfully for the measurement of activity -induced calcium transients in individual spines. Here, we describe the fir st application of two-photon imaging to measure Na+ transients in spines an d dendrites of CA1 pyramidal neurons in hippocampal slices. Whole-cell patc h-clamped neurons were loaded with the Na+-indicator dye SBFI (sodium-bindi ng benzofuran-isophthalate). In situ calibration of SBFI fluorescence:with ionophores enabled the determination of the actual magnitude of the [Na+](i ) changes. We found that back-propagating action potentials (APs) evoked Na + transients throughout the proximal part of the dendritic tree and adjacen t spines. The action-potential-induced [Na+](i) transients reached values o f 4 mM for a train of 20 APs and monotonically decayed with a time constant of several seconds. These results represent the first demonstration of act ivity-induced Na+ accumulation in spines. Our results demonstrate that two- photon Na+ imaging represents a powerful tool for extending our knowledge o n Na+ signaling in fine cellular subcompartments.