Circadian and sleep/rest disturbances in older mice undergoing chronic ethanol treatment

1. Circadian and sleep/rest disturbances in older mice undergoing chronic ethanol treatment A.J. Brager1, R.A. Prosser2, and J.D. Glass1. 1Dept Biological Sciences, Kent State University, Kent, OH, 2Dept Biochemistry and Cellular and Molecular Biology, University of Tennessee, Knoxville, TN. INTRODUCTION Chronic exposure to ethanol is associated with various sleep and circadian pathologies; it has disruptive effects on sleep architecture and circadian rhythmicity as well as broad physiological effects on hormonal secretion (e.g. melatonin and cortisol). In alcoholics, disruptions of sleep homeostasis and circadian timing exacerbate fragmented sleep and wake episodes, and subsequent disruptions in abstinent alcoholics increase the risk of relapse. In addition, sensitivity to the locomotor incoordinating and anxiogenic properties of ethanol increases with age. Our experimental goals are three-fold: 1) to assess an ethanol-induced impairment of photic entrainment capacity 2) to quantify an ethanol-induced unconsolidation of circadian locomotor activity, and 3) to illustrate that theses ethanol-induced circadian disturbances are independent of the amount of ethanol consumption. METHODS Ethanol Treatment. Mice (22wks.) were introduced to a 10% ethanol solution (n=4), or were maintained on tap water (n=4) for 1 month under forced consumption. Food was provided ad libitum. Consumption of the ethanol solution or tap water was measured to the nearest 0.25 mL daily at 1030. Photic Entrainment Capacity. All mice had previously entrained to a 1 min light pulse (25 lux) skeleton photoperiod. General circadian locomotor activity was measured using a passive infrared motion detector interfaced with a computerized data acquisition system. Rhythm period, alpha, activity onset, and activity offset relative to the 1 min light pulse were analyzed. Circadian Locomotor Activity. An activity bout was defined as a period of locomotor activity separated by at least 10 min of locomotor quiescence. Activity bouts were quantified across the active and rest periods and across the 24 hr circadian day. Durations of activity bouts were also plotted. Quantities and durations of activity bouts were averaged over the first 3 days of ethanol treatment. RESULTS Ethanol Disrupts Photic Entrainment Capacity. Ethanol-treated mice had a delayed activity onset vs. water controls (6.36 h +/- 0.62 vs. 4.60 h +/- 0.30, respectively; p<0.04). Ethanol Unconsolidates Circadian Locomotor Activity and Increases Sporadic Activity Across The Rest Period. Ethanol-treated mice had unconsolidated circadian locomotor activity across the rest period as indicated through more activity bouts vs. water controls (9.17 +/- 0.60 vs. 6.58 +/- 0.36, respectively; p<0.02). Frequencies of durations of activity bouts (bin size = 15 min) across the rest period revealed ethanol-induced incidences of sporadic activity; ethanol-treated mice had more bouts 0-15 min in duration vs, water controls (10.75 +/- 1.14 vs. 6.33 +/- 0.89, respectively; p<0.02). Amount of Ethanol Consumption Did Not Differ From Water Consumption. Figure 2: Representative actograms for an ethanol-treated mouse (right) and a water control (left). These analyses (commencing at purple arrow) are included within a more extensive protocol assessing concentration-dependent disturbances of photic entrainment capacity and circadian locomotor activity following chronic ethanol exposure, ethanol withdrawal, and ethanol reintroduction. The solid yellow line annotates the introduction of the 1 min (25 lux) skeleton photoperiod. b Figure 3: Schematics depict the frequencies of durations of activity bouts. As illustrated, mice introduced to a 10% ethanol solution had significantly more activity bouts 0-15 min in duration (bin size=15) across the rest period. In addition, ethanol-treated mice had significantly more activity bouts < 60 min in duration across the 24 hr circadian day (bin size=60), indicating that ethanol treatment markedly unconsolidates circadian locomotor activity. ETOH SCN PHARMACOKINETICS Figure 4: The pharmacokinetics of ethanol within the SCN following an acute injection (2.0 g/kg) was measured by means of microdialysis. Ethanol concentrations of 87 mM peaked 40 min post-injection. The half life of ethanol clearance was ~ 2 hr. The acute injection of ethanol is equivocal to the substantial amount of ethanol consumed at the beginning of the dark period in this forced-drinking paradigm. CONCLUSIONS 1. Chronic ethanol exposure delays activity onset. 2. Ethanol unconsolidates circadian locomotor activity across the rest period, by increasing the quantity of locomotor activity bouts and the frequency of activity bouts <60 min in duration. 3. These ethanol-induced circadian disturbances are independent of the magnitude of ethanol consumption. Figure 1:Mice introduced to a 10% ethanol solution had significantly more activity bouts across the 24 hr circadian day (black; p<0.05) and rest period (dark grey; p<0.05). The mean duration of an activity bout across these periods did not differ between treatment groups (p>0.05, in both cases). Acknowledgements: NIH grant AA015948 to RAP and JDG