Bright daytime light enhances circadian amplitude in a diurnal mammal

Abstract

Mammalian circadian rhythms are orchestrated by a master pacemaker in the hypothalamic suprachiasmatic nuclei (SCN), which receives information about the 24 h light-dark cycle from the retina. The accepted function of this light signal is to reset circadian phase in order to ensure appropriate synchronization with the celestial day. Here, we ask whether light also impacts another key property of the circadian oscillation, its amplitude. To this end, we measured circadian rhythms in behavioral activity, body temperature, and SCN electrophysiological activity in the diurnal murid rodent Rhabdomys pumilio following stable entrainment to 12:12 light-dark cycles at four different daytime intensities (ranging from 18 to 1,900 lx melanopic equivalent daylight illuminance). R. pumilio showed strongly diurnal activity and body temperature rhythms in all conditions, but measures of rhythm robustness were positively correlated with daytime irradiance under both entrainment and subsequent free run. Whole-cell and extracellular recordings of electrophysiological activity in ex vivo SCN revealed substantial differences in electrophysiological activity between dim and bright light conditions. At lower daytime irradiance, daytime peaks in SCN spontaneous firing rate and membrane depolarization were substantially depressed, leading to an overall marked reduction in the amplitude of circadian rhythms in spontaneous activity. Our data reveal a previously unappreciated impact of daytime light intensity on SCN physiology and the amplitude of circadian rhythms and highlight the potential importance of daytime light exposure for circadian health.

Keywords: circadian; light; retina; suprachiasmatic nucleus.

Fig. 1.

Impact of increasing daytime light intensity on circadian rhythms in R. pumilio. (A) Lighting conditions. Left shows the spectral power distribution of our light source at different irradiances and that of daylight on an overcast day. Middle shows expected spectral sensitivity profile of mammalian rod opsin and melanopsin and R. pumilio medium and short wavelength-sensitive cone opsins (MWS and SWS, respectively) (61) corrected for lens transmission. Right table shows Log₁₀ effective photon fluxes for each opsin and melanopic EDI across the different lighting stimuli used (bright, mid, dim, dimmest). (B) Double-plotted actograms for general activity, wheel running and Tb (scale below) from a representative R. pumilio under different lighting conditions, over a period of 2.5 mo. Time of light exposure is indicated in yellow, and intensity of the light is shown on the Right. Entrainment to 12:12 LD cycle at each irradiance ran for 2 wk followed by 4 d in constant darkness (DD). Note that across conditions, activity is largely restricted to daytime (light phase) coinciding with higher Tb values. However, the rhythms become less robust at low light levels. Gray columns on the left indicate the 8-d period at the end of each stage used for analysis reported in SI Appendix, Table S1 and Fig. 2. (C) Mean waveforms for the recorded biological rhythms (i: general activity; ii: sustained immobility; iii: voluntary wheel-running activity; iv: Tb) across the different lighting conditions. Mean waveforms under bright intensities are shown in the Top and the dimmest conditions in panels at the Bottom. Values are expressed as mean ± SEM (n = 12), gray areas indicate period of darkness. ZT0: ZT 0 corresponds to time of lights on.

Fig. 2.

Impact of increasing daytime light intensity on reproducibility and robustness of circadian rhythms under entrained and subsequent free-running conditions. (A) Intradaily variability and (B) day-to-day reproducibility of the sustained immobility rhythm as a function of daytime irradiance under entrained conditions. (C and D) Same as in A and B but for the Tb rhythm. (E) Representative wheel-running activity actogram for an R. pumilio over the last 4 d of entrainment under dim (Top) or bright (Bottom) irradiance, and subsequent 4 d of free run in constant darkness (note difference in slope of red lines fit to activity onsets in free run between conditions). (F) Relationship between phase angle of entrainment for wheel-running activity rhythm and daytime irradiance. (G) Free-running period and (H) robustness of the activity rhythm across all animals as a function of the prior entraining irradiance. Values are expressed as mean ± SEM (n = 12, A–D; n = 11, F–H); significant relationships identified by linear regression (*P < 0.05, **P < 0.01, and ***P < 0.001).

Fig. 3.

Impact of daytime irradiance on SCN activity. (A) Image of a brain slice of R. pumilio placed over the electrode terminals of a 60-channel array to record neuronal electrical activity in the SCN. The recording site is shown in close up on Right (a1), with dotted white lines delineating the SCN area and electrodes designated as within the SCN shown in blue. (B) Neuronal MUA recorded at the array of electrodes for the preparation shown in A as a function of projected ZT for a 24-h recording epoch. Spontaneous activity within the SCN (bounded by dotted lines) is higher than surrounding hypothalamus and plotted on a different scale (scale bars to right). (C) Time-averaged firing rate (MUA) across 24-h recording epochs from all channels covering SCN harvested from bright (n = 92 channels from four slices) versus dim (n = 99 channels from four slices) conditions. (D) Percentage of channels within the SCN classified as rhythmic for each experimental condition (bright versus dim, **P < 0.01; Fisher’s exact test, data from four slices per condition). (E and F) Multiunit firing rate as a function of projected ZT for rhythmic recording sites within the SCN from animals maintained under bright (E) and dim (F) daytime conditions (bright, n = 58; dim, n = 42 channels; from a total of four animals per condition). (G) Peak-trough amplitude in the multiunit firing rhythm was significantly lower under dimmer lights. (***P < 0.001, Mann–Whitney U test, bright, n = 58; dim, n = 42 channels). Gray background indicates the projected period of lights off (ZT12 to ZT24). Data are expressed as mean ± SEM.

Fig. 4.

Increasing daytime light intensity impacts SCN neurophysiology at the single-cell level. (A) Whole-cell recording setup showing bright-field image (4×) of a living SCN in coronal brain slice either side of the third ventricle (3V) and above the optic chiasm (OX), with a patch pipette (red arrow) targeting an SCN neuron. (Scale bar, 200 μm.) (B) Schematic diagram showing the approximate location within a representative SCN coronal slice of the recorded cells giving rise to the daytime datasets under bright (blue) or dim (red) daytime irradiance. Note the similar spatial distribution between experimental groups. (C) Representative traces for each of the spontaneous membrane excitability states encountered in R. pumilio SCN neurons (from Top): highly depolarized silent; depolarized displaying low-amplitude membrane oscillations (DLAMOs); moderate RMP and firing at high or low rate; and hyperpolarized silent. (D) Positive correlation between RMP and SFR for cells recorded from bright (blue) and dim (red) conditions. Cells resting at RMP > −40 mV were excluded. P < 0.0001, linear regression analysis. (E) Pie charts showing the percentages of cells in the different electrophysiological states during the day and at night from bright and dim conditions. (*P = 0.01, chi-squared test.) (F) Example of traces from two SCN neurons (voltage clamped at −70 mV) showing PSCs recorded during the day (i) and at night (ii). (i) Cell displaying high frequency and amplitude PSCs; (ii) cell showing low frequency and amplitude PSCs. (G) Scatter plot showing the RMP of the different cells across ZTs for bright (blue) and dim (red) conditions. Each dot represents an individual cell. Gray background indicates the projected phase of lights off. (H) Mean RMP during the day and at night for SCN neurons from bright or dim conditions. (I and J) Same as in G and H but for SFR. (K and L) Same as in G and H but for Rinput. (M and N) Same as in G and H but for synaptic activity frequency. Data in bar charts are expressed as mean ± SEM. Number of cells used is indicated between brackets in each bar. *P < 0.05, **P ≤ 0.01, and ***P ≤ 0.001, Mann–Whitney U test.