Burkhalter, 2006, Soc. Neurosci., abstract; M. Roth, F. Helmchen, and B. Kampa, 2010, Soc. Neurosci., abstract;
M. Garrett, J. Marshall, L. Nauhaus, and E. Callaway, 2010, Soc. Neurosci., abstract). While comparatively little is known about visual response properties in unanesthetized selleck compound mice (Andermann et al., 2010 and Niell and Stryker, 2010), cortical neurons in mice and other species may demonstrate visual responses of greater magnitude (Niell and Stryker, 2010), diversity (Qin et al., 2008), and context sensitivity (Pack et al., 2001) in the awake state. Therefore, to determine the degree of functional specialization in mouse higher visual areas, we developed a chronic two-photon imaging system for mapping responses in local volumes of cortical neurons across multiple areas in awake
Selleckchem Pomalidomide mice. We found striking differences in stimulus preferences across areas, demonstrating distinct functional specialization of different higher visual areas in the mouse. We characterized the functional properties of neurons in visual cortical areas of awake mice, using the following approach (see also Experimental Procedures). First, we implanted a 5 mm cranial window over visual cortex. Following recovery, mice were gradually habituated to head restraint (Andermann et al., 2010) while free to walk on a single-axis trackball (Experimental Procedures). We then performed widefield imaging of intrinsic autofluorescence signals to obtain retinotopic maps of multiple visual areas (Figure 1A and Figure S1 available online; Kalatsky and Stryker, 2003 and Schuett et al., 2002; cf. Wang and Burkhalter, 2007). We subsequently removed the cranial window under anesthesia and injected adeno-associated Sitaxentan virus AAV2/1-synapsin-1-GCaMP3 at a depth of 250 μm below the cortical surface to obtain
neuron-specific expression of the calcium indicator GCaMP3 ( Tian et al., 2009, Dombeck et al., 2010 and O’Connor et al., 2010) at approximately matched retinotopic locations in one or two visual cortical areas. Changes in cellular GCaMP3 fluorescence provide an estimate of visually driven increases in calcium influx associated with increases in neural firing rate ( Tian et al., 2009; Experimental Procedures). We measured population visual responses across cortical areas using widefield calcium imaging ( Figure 1), followed by a more detailed mapping of individual neurons using two-photon calcium imaging ( Figure 2, Figure 3, Figure 4 and Figure 5). Specifically, we assessed tuning of neurons across multiple stimulus dimensions, including spatial and temporal frequency, speed, orientation, and direction of motion. Tuning estimates in Figure 1, Figure 2, Figure 3, Figure 4 and Figure 5 included all trials, independent of whether the mouse was moving or stationary, as tuning was not strongly affected by locomotion (see Figures 6, S2, and S6, below).