Differential response of Period 1 expression within the suprachiasmatic nucleus

Abstract

The suprachiasmatic nuclei (SCNs) of the hypothalamus contain a circadian clock that exerts profound control over rhythmic physiology and behavior. The clock consists of multiple autonomous cellular pacemakers distributed throughout the rat SCN. In response to a shift in the light schedule, the SCN rapidly changes phase to achieve the appropriate phase relationship with the shifted light schedule. Through use of a transgenic rat in which rhythmicity in transcription of the Period 1 gene was measured with a luciferase reporter (Per1-luc), we have been successful in tracking the time course of molecular rhythm phase readjustments in different regions of the SCN that occur in response to a shift in the light schedule. We find that different regions of the SCN phase adjust at different rates, leading to transient internal desynchrony in Per1-luc expression among SCN regions. This desynchrony among regions is most pronounced and prolonged when the light schedule is advanced compared with light schedule delays. A similar asymmetry in the speed of phase resetting is observed with locomotor behavior, suggesting that phase shifting kinetics within the SCN may underlay the differences observed in behavioral resetting to advances or delays in the light schedule.

Figure 1.

Optical recording of luminescence in SCN slices (300 μm thick) from Per1-luc transgenic rat. a, Representative pseudocolored luminescence images at 2 h intervals from a continuous recording of the SCN slice in vitro. The images integrated the luminescence for 25 min every 30 min. Note that the dorsomedial region of the SCN phase leads other regions. b, Pseudocolored luminescence image at the peak of Per1 expression with the 15 sector template overlaid. Luminescence was analyzed in each of these 15 regions. Red and blue squares indicate the dorsomedial and ventrolateral SCNs. Each square consists of 5 × 5 pixels. The scale bar represents 200 μm, equal to 8 pixels. c, Luminescence rhythm in the dorsomedial (red circles) and ventrolateral (blue circles) SCNs shown in a. Red and blue lines plot the five-point moving average for dorsomedial and ventrolateral SCNs, respectively. The tip of the vertical arrowhead indicates the peak phase.

Figure 2.

Peak phase of Per1-luc rhythm after shifts in the light schedule. The peak phase represents the averaged luminescence data from the entire SCN. Each bar represents the light schedule before (control) and after a 6 h phase delay or phase advance. The time of darkness is indicated by the filled bar, and light time is indicated by the open bar. Mean phase ± SE is indicated to the right of the figure. Vertical arrows indicate the projected steady-state phase for a 6 h delay or advance in the light schedule.

Figure 3.

Reentrainment of Per1 expression in the SCN to 6 h phase delay in the light cycle. a, Steady-state phases of the Per1-luc rhythm. The phase of the 15 regions of the SCN are displayed as histograms with the position on the surface of the SCN indicated. For quantitative and statistical comparisons, the bold histogram represents the dorsomedial region, and the histogram surrounded by dashes indicates the ventrolateral histogram. Asterisks on the tops of histograms indicate regions that were statistically significantly different in phase from the dorsomedial region (Tukey's test; p < 0.05). The phase plane formed by the tops of the histograms is tipped upward in the top left corner, revealing that the dorsomedial region phase leads other regions. b, Phase plane on day 1 after 6 h delay in light schedule. The orientation of the plane reveals that the lateral region of the SCN has shifted more than the medial regions. c, By day 3 after the light schedule delay, the phase plane orientation is similar to the control orientation, indicating that the phase shift is essentially complete in all regions of the SCN. d, Circadian rhythms in Per1-luc expression in dorsomedial (filled circles) and ventrolateral (open circles) regions of the SCN before shift in light schedule (control). e, Per1-luc rhythms on day 1 after light cycle delay. f, Per1-luc rhythms on day 3. Controls are shown in dashed lines, dark dashes for dorsomedial region. In d-f, error bars indicate SE. ZT is time of light cycle just before recording; thus, ZT is for 6 h delayed light schedule in b and c.

Figure 4.

Reentrainment of Per1 expression in the SCN to 6 h phase advance in the light cycle. a, Control phase plane. b, Phase plane on day 1 after 6 h advance in light schedule. The orientation of the phase plane formed by the tops of the histograms reveals that the lateral region of the SCN has shifted significantly more than the medial regions. c, On day 3 after the light schedule change, the phase relationships have not returned to normal, as indicated by the lack of a phase difference between the dorsomedial and ventrolateral regions. d, By 6 d after the advance in the light schedule, the regional phase differences have been restored. e, Per1-luc rhythms in dorsomedial and ventrolateral regions of the SCN before shift in light schedule (control). f-h, Per1-luc rhythms on days 1, 3, and 6 after light cycle advance. In e-h, error bars indicate SE. The conventions are the same as in Figure 3.