As one might imagine, this could be a serious challenge to calibrating voltage signals in small dendrites or dendritic spines, although researchers can use, and have used, the neuron’s own electrical signals, such as back-propagating action potentials, as internal standards for calibration (Nuriya et al., 2006). Finally, the relatively high speed of the electrical responses of mammalian neurons also generates a serious challenge for voltage measurements. While infinite temporal resolution would be welcome, in

practice most questions can be addressed with one Ixazomib supplier millisecond resolution. As we will discuss in the next section, there are a variety of chromophores with different response times; but unfortunately, the fastest ones normally provide the smallest signals,

which has been a long-standing problem in voltage imaging (Waggoner, 1979). The reader can appreciate from the previous list of problems that for effective voltage imaging one needs to solve some nontrivial challenges. At the same time, as mentioned, the electric field at the plasma membrane is very strong and can easily alter the physical, chemical, environmental, and spectral properties of any molecule located within it. This creates the potential to tap into a rich toolbox of different physicochemical principles

and harness them to measure changes in the electric field. As we will see, there is a great diversity of approaches Selleck Buparlisib that have achieved meaningful optical voltage measurements, a tribute to the determination and ingenuity of the scientists involved ( Cohen, 1989 and Cohen and Lesher, 1986). Most of the successful experiments with voltage imaging so far have been accomplished using single photon excitation with visible light, where the absorption cross-sections of the indicators are large. Also, some light sources (arc lamps, or now LEDs) can have very low noise, making it relatively easy to detect minute changes in signal, with ratiometric measurements at multiple absorption or emission wavelengths providing additional noise immunity and sensitivity ( Yuste et al., 1997 and Zhang Terminal deoxynucleotidyl transferase et al., 1998). With typical light sources, wide field excitation is possible, and many photons can be collected from spatially extended areas, such as a section of dendrite, the entire soma, or many cells and their processes, increasing the integrated signal. But all of the typical problems of single-photon excitation apply—there is low penetration into scattering media like intact vertebrate brain tissue, and no native sectioning capability, requiring the use of confocal microscopes to afford cellular resolution.