Article Figures & Data
Figures
- Figure 1.
Time-lapse imaging of SCN Per1-driven GFP rhythms. A, Individual pseudocolored fluorescence images from a time-lapse recording of Per1-driven GFP fluorescence rhythms from an SCN brain slice in vitro. B, Fluorescence rhythms in the in vitro SCN from an animal housed in a 14/10 hr LD cycle. Images were captured every 30 min. The open and filled bars indicate the previous light/dark cycle for the animal. C, Wheel-running activity, double-plotted, from an animal housed in constant darkness. Vertical marks show times when the animal was active. The filled arrow marks the time the SCN slice was prepared for imaging. D, GFP fluorescence recording of the SCN from the animal in C. Images were captured every 30 min for 2 d.
- Figure 2.
Per1-driven fluorescence intensity correlates with spike frequency in SCN neurons. Right, Fluorescence intensity, measured as percent above background, versus action potential (AP) frequency plotted for SCN neurons (n = 24, 3 slices). r = 0.91; p < 0.001. Left, Example of spike output from neighboring neurons; top arrow, low fluorescence intensity (6.9% above background, 0.03 Hz); bottom arrow, high fluorescence intensity (49.5% above background, 9.14 Hz).
- Figure 3.
Individual neuron gene expression rhythms in the SCN. A, Per1-driven GFP rhythms from two individual SCN neurons in an in vitro DD SCN obtained by confocal time-lapse imaging. B, C, Example gene expression rhythms from four individual neurons in an in vitro SCN from a LD animal. Each set of symbols represents the measured fluorescence for an individual cell during the circadian cycle. The black solid line indicates the integrated overall fluorescence rhythms of the SCN as a whole.
- Figure 4.
Neuronal phase relationships are dependent on light history. A, Histograms of individual neuron fluorescence peak times in an SCN slice from an LD animal (gray bars) and an SCN slice from a DD animal (black bars). Cell peak times were binned at 1 hr intervals; n = 26 neurons for the LD slice; n = 37 neurons for the DD slice. B, Histogram of individual neuron fluorescence peak times summed from SCN slices from five LD animals. Peak times are in 1 hr bins. The black line indicates the best fit curve showing phase group peak times of ZT5, ZT8, and ZT11. C, Histogram of individual neuron fluorescence peak times summed from SCN slices from five DD animals. The black line indicates the best fit curve showing phase group peak times of CT12, CT15, and CT18. D, Cumulative probability plot of peak times from animals housed in the LD cycle (filled circle, gray line) and peak times from DD (filled diamond, black line). The median time of cellular peaks for each animal is plotted as an open symbol (circle, LD; diamond, DD).
- Figure 5.
Rhythmic neurons by SCN region. Split cylinders shown in a three-dimensional perspective rendering represent the percentage of rhythmic neurons in the SCN mapped by SCN region. The height of each half-cylinder indicates the percentage of rhythmic SCN neurons in that region. Scale bars indicate the percentage scale for the lateral-medial axis (vertical) and the dorsal medial axis (horizontal). Total cells are n = 126 cells for LD slices and 131 cells for DD slices. A, Overall distributions of rhythmic neurons. Top row, Percentage of rhythmic SCN neurons mapped in the lateral-medial axis for LD (left) and DD (right) slices. Bottom row, Percentage of rhythmic SCN neurons mapped in the dorsal-medial axis for LD (left) and DD (right) slices. B, Distributions of rhythmic neurons by phase group. Top row, Percentage of rhythmic SCN neurons from the three LD phase groups mapped in the lateral-medial axis. Bottom row, Percentage of rhythmic SCN neurons from the three DD phase groups mapped in the lateral-medial axis.
Additional Files
- temp index page
HTML Page - index.htslp
Files in this Data Supplement:
- Supplementary Figure - No effect of different levels of GFP expression on ionic currents in N2A cells. Top panel shows the mean relative intensity\ies of GFP in low and high expressing cells (N=8 each). Middle and bottom panels show mean current-voltage curves for the cells in the top panel. Middle panel shows peak currents, whereas the bottom panel shows steady state currents. There no significant differences between groups.
- Supplementary Video - Time-lapse confocal video of neuronal Per1::GFP rhythms in an SCN slice from an LD entrained animal. Fluorescence intensity is shown in false color with warmer colors (red) indicating higher intensities. This coronal SCN slice is oriented with the ventral aspect to the bottom of the frame and the medial aspect to the right. Recording begins in the early subjective day, when Per1::GFP expression is low in a majority of neurons, and continues through 24 hours with images taken every 20 minutes. It best viewed as a continuous loop. Salient features include the �twinkling� of cells as they peak at different phases, and the presence of two pairs of adjacent neurons that cycle in approximate antiphase to one another � one pair on the ventral/lateral quadrant of the SCN (lower left) and one pair in the dorsal/medial quadrant (upper right).
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