OGB and sulforhodamine 101 (SR101) were injected with 150 ms pulses every 15 s for 15 min at 200 and 400 μm below the dLGN surface. A tube with a glass coverslip Cobimetinib datasheet was inserted and filled with artificial cerebrospinal fluid. OGB-loaded neurons were imaged through the tube with a two-photon microscope. For visual stimulation, chlorprothixene (1 mg/kg, intramuscular injection) was administered and isoflurane was lowered to 0.3%–0.5%. More details and visual stimulation parameters

can be found in the Supplemental Experimental Procedures. Regions of interest (ROIs) were drawn around each cell in each field of view, glia were excluded using SR101 labeling, and pixels were averaged within each ROI. Calcium signal modulations were measured as

relative change in fluorescence over time compared to a prestimulus baseline (ΔF/F). Fourier transforms were taken of the signals during the stimulus period, at the first and second harmonic frequencies of the grating to measure the response of the cell to each direction of the grating. Direction selectivity was calculated by both max-null and circular variance metrics. See Supplemental Experimental Selleck Ixazomib Procedures for more details and statistics. A full derivation of the model can be found in the Supplemental Experimental Procedures. We thank the Callaway laboratory for helpful discussions and technical assistance. We also thank D. Kleinfeld for helpful discussions and D. Dombeck for imaging advice. We acknowledge support from Amisulpride NIH grants EY010742; EY022577 (E.M.C.), 1F30DC010541-01 (A.P.K.), and EY019821 (I.N.) and the Gatsby Charitable Foundation. “
“Normal nervous system function requires the development of elaborate and precise connections among neurons and their targets. Establishing this complex wiring relies on the combined functions of a large and diverse number of axon guidance molecules that coordinate neuronal process pathfinding and target recognition (Dickson, 2002). During development, neurons extend processes that have at their extending tips highly motile structures

called growth cones. Receptors expressed on growth cones recognize multiple cues present in the surrounding extracellular environment and manifest their response through the reorganization of neuronal cytoskeletal components, including actin and microtubules (Dent et al., 2011). Although molecular mechanisms that signal cytoskeletal remodeling have been uncovered for certain classes of guidance cue receptors (Bashaw and Klein, 2010; Kolodkin and Tessier-Lavigne, 2011), we are only just beginning to understand how these signaling pathways are integrated in order to allow for discreet steering of neuronal processes; for many guidance cue receptors little is known about the in vivo signaling events they initiate following ligand engagement.