High Precision and Fast Functional Mapping of Brain Circuitry through Laser Scanning Photostimulation and Fast Dye Imaging

The development of modern neuroscience tools has a significant impact on the progress in the field of neuroscience (Kandel, 1982). New tools has greatly faciliated the neuroscience resreach, and can be critical for studies of brain circuit organization and function. Although many approaches are useful by themselves, it is desirable to combine existing powerful techniques to harness each technique's advantages and compensate for limitations. In many brain areas, neuronal circuits are segregated into anatomically discrete areas such as specific lamina and modules or compartments (Mountcastle, 1997). Functional imaging of these brain areas is particularly useful in characterizing circuit properties. Fast voltagesensitive dye (VSD) imaging, which detects neuronal membrane potential changes via shifts in the dye absorption /or fluorescence emission in response to varying membrane potentials, offers a great means of simultaneous monitoring neuronal activities from many locations with high spatial and temporal resolutions. With new dyes and modern imaging apparatus, VSD imaging has been widely used to study spatiotemporal dynamics of population neuronal activity in cortical tissue both in vivo and in vitro (Grinvald & Hildesheim, 2004). Particularly, for in vitro brain slices, fast VSD imaging is important for mapping circuit organization and response dynamics, and more recently has been used to probe functional abnormalities in models of neurological and psychiatric disorders (Ang et al., 2006; Airan et al., 2007). One major limitation of most in vitro VSD imaging studies, however, lies in that the imaged neuronal responses are either spontaneous seizure activities through pharmacological manipulations or induced by electric stimulations (Petersen & Sakmann, 2001; Huang et al., 2004; Ang et al., 2006). Significant disadvantages of electric stimulation include indiscriminate activation of axons of passage, slow and inefficient placement of multiple stimulation locations, and tissue damage. In comparison, optical stimulation including laser scanning photostimulation (LSPS) either by glutamate uncaging or direct activation of light-sensitive channels (e.g., channelrhodopsin-2) enables rapid and noninvasive photoactivation of neurons with great convenience and superior spatial resolution in practical experiments (Callaway & Katz, 1993; Boyden et al., 2005; Petreanu et al., 2009). Combining whole-cell recordings from single neurons with photostimulation of clusters of presynaptic neurons permits extensive mapping of local functional inputs to individually recorded neurons (Schubert et al., 2003; Shepherd & Svoboda, 2005; Xu & Callaway, 2009).