Figure 1 O’Malley, 2008

1. Figure 1 O’Malley, 2008 Fig. 1. Photoreceptors in anesthetized zebrafish. Images are from transgenic larvae expressing GFP-rhodopsin fusion protein. A,B: Fluorescent rod photoreceptors in ventral retina (arrowhead) are easily viewed using a fluorescence dissecting microscope. Confocal maximum projection images (C-F) show photoreceptors at higher resolution revealing banding patterns in rod outer segments (arrows in F). Arrow in (D) shows location of lens. Scale bars = 150 μm (A,B), 50 μm (C,D), 25 μm (E), and 10 μm (F). Reprinted from Perkins et al., Visual Neuroscience, 2003. [Permission given by Dowling and Perkins; and Visual Neuroscience].

2. C A B 10 μm 2 μm Figure 2 O’Malley, 2008 scanned line Fig. 2. Spinning Disk and Point Scanning confocal images of calcium sparks in atrial myocytes. (A) Spinning disk images acquired at 60 frames/sec show a ring of calcium influx at an early moment of an atrial action potential. (B) A surface plot of the image at 17 msec shows calcium peaks in the sub- sarcolemmal space. (C) Line-scans oriented along the subsarcolemmal space provide more detailed records of the temporal dynamics of calcium sparks at multiple locations. Reprinted from Kockskamper et al., 2001. [Permission given by Lothar Blatter and Biophysical Journal 5/20].

3. Figure 3 O’Malley, 2008 Fig. 3. Deep tissue imaging with two-photon microscopy. Methods used to visualize neurons in mouse cortex are shown in (A). Shown in (B) is a side (xz) view of two-photon image stack. In a transgenic mouse expressing the genetically-encoded chloride indicator Clomeleon, layer 5 (L5) pyramidal cells can be visualized as much as 700 μm deep into cortex. Reprinted from Helmchen and Denk, Nature Methods, 2005. [Permission given by Winfried Denk and Nature Methods].

4. nMLF L200 * * Figure 4 O’Malley, 2008 Fig. 4. Confocal montage (of maximum projections) of reticulospinal neurons in the brainstem of a restrained larval zebrafish. Neurons have been labeled using the labeled-lesion technique in which large numbers of neurons can be simul-taneously labeled with fluorescent dextrans and disconnected from their spinal targets (Gahtan and O’Malley, 2001). Such lesions result in novel but abnormal bending patterns due to deconstraint of the spinal neural circuits (Day et al., 2005). Image courtesy of Leslie Day, Dept. Biology, Northeastern University.

5. Figure 5 O’Malley, 2008 Fig. 5. Two-photon projection of reticulospinal neurons in the brainstem of a restrained larval zebrafish. Neurons have been labeled with Texas-red dextran (10,000 MW) and slow-scan imaged with a 20X, 0.95 NA objective to produce maximum resolution inside living animal. Contrast has been reversed (darkest cells are most fluorescent). Fine anatomical details (such as axons and dendrites) are evident that are difficult to resolve in confocal images of similarly-labeled fish. Image courtesy of Michael Orger, Adam Kampff, JH Bollmann and Florian Engert, Dept. Molec. Cell. Biology, Harvard University.

6. Figure 6 O’Malley 2008 Fig. 6. Deconvolution of confocal image stack. 50 nm gold beads were dispersed in immersion oil and imaged using a 3D-piezoelectric stage-scanning confocal microscope. 3D stacks consisted of 30 XY images that were 40 nm apart in the Z direction. The pixel size was 10 nm in the X and Y direction. (A) shows a rendered plot of the bead images. The beads are less than one-seventh of the illumination wavelength and so their rendered confocal images represent, in effect, the optical point spread function. (B) Use of a maximum-likelihood deconvolution algorithm restores the images to a more faithful representation of the actual object dimensions. Reprinted from Schrader et al., Applied Physical Letters, 1996. [Permission given by Stefan Hell; and by Applied Physical Letters].