Chronic ethanol intake alters circadian phase shifting and free-running period in mice

Abstract

Chronic alcohol intake is associated with widespread disruptions in sleep and circadian rhythms in both human alcoholics and in experimental animals. Recent studies have demonstrated that chronic and acute ethanol treatments alter fundamental properties of the circadian pacemaker--including free-running period and responsiveness to photic and nonphotic phase-shifting stimuli--in rats and hamsters. In the present work, the authors extend these observations to the C57BL/6J mouse, an inbred strain characterized by very high levels of voluntary ethanol intake and by reliable and stable free-running circadian activity rhythms. Mice were housed individually in running-wheel cages under conditions of either voluntary or forced ethanol intake, whereas controls were maintained on plain water. Forced ethanol intake significantly attenuated photic phase delays (but not phase advances) and shortened free-running period in constant darkness, but voluntary ethanol intake failed to affect either of these parameters. Thus, high levels of chronic ethanol intake, beyond those normally achieved under voluntary drinking conditions, are required to alter fundamental circadian pacemaker properties in C57BL/6J mice. These observations may be related to the relative ethanol insensitivity displayed by this strain in several other phenotypic domains, including ethanol-induced sedation, ataxia, and withdrawal. Additional experiments will investigate chronobiological sensitivity to ethanol in a range of inbred strains showing diverse ethanol-related phenotypes.

Figure 1

Representative actogram segments showing light-induced phase shifts for forced ethanol and control animals, at ZT 15 and ZT 21, during either maintained drinking or at 24 h after ethanol replacement by plain water (acute withdrawal). Bold lines superimposed on each chart connect successive activity onsets prior to and following each light pulse; stars indicate the approximate times of light pulse delivery.

Figure 2

Mean (± SEM) light-induced phase shifts (free-choice ethanol vs. controls, top; forced ethanol vs. controls, bottom). Each animal was tested a total of 4 times: at ZT 15 and ZT 21, during maintained drinking and at 24 h following ethanol withdrawal (WTD). Asterisks indicate significant attenuation of phase shifting.

Figure 3

Representative actograms showing free-running activity rhythms under long-term DD from 1 animal in each of the 3 groups (forced ethanol, free-choice ethanol, and water-only controls). All animals were maintained on plain water for the 1st 3 weeks of the experiment, after which ethanol was continuously available in the forced and free-choice ethanol groups (horizontal line indicates beginning of ethanol treatment on day 22).

Figure 4

Mean (± SEM) free-running period (top), periodogram amplitude (middle), and daily activity (bottom) for all 3 groups in successive 3-week data samples. “B” indicates the initial water-only baseline, and “E1” through “E7” indicate successive 3-week samples in which ethanol was continuously available in the forced and free-choice ethanol groups. * = controls significantly different from both ethanol-treated groups; @ = controls significantly different from forced ethanol group; # = controls significantly different from free-choice ethanol group.

Figure 5

Mean (± SEM) fluid intakes in both experiments. (Top) Total fluid intake for forced and free-choice ethanol groups and combined water-only controls. (Middle) Water and 10% ethanol intake in free-choice ethanol groups. (Bottom) Ten percent ethanol intake in forced and free-choice ethanol groups. * = forced ethanol group significantly different from both controls and free-choice ethanol groups; # = free-choice ethanol group significantly different from controls; & = experiment 2 significantly different than experiment 1; + = free-choice ethanol group significantly different from forced ethanol group.