Nest building is a novel method for indexing severity of alcohol withdrawal in mice

Abstract

Withdrawal after chronic ethanol (EtOH) affects body temperature, goal-directed behavior and motor function in mice and increases general central nervous system excitability. Nest-building tests have been used to assay these states but to this point have not been employed as measures of EtOH withdrawal severity. We first refined nest-scoring methods using a genetically heterogeneous stock of mice (HS/Npt). Mice were then made physically dependent following three days of chronic EtOH vapor inhalation to produce average blood EtOH concentrations (BECs) of 1.89 mg/mL. EtOH withdrawal affected the progression of nest building over time when mice were tested 2–4 days after removal from three days of chronic exposure to EtOH. In a separate group of mice, chronic EtOH vapor inhalation (BECs 1.84 mg/mL) suppressed nest building over days 1–2 but not days 2–3 of withdrawal. In a following experiment, EtOH withdrawal dose-dependently slowed recovery of nest building for up to 32 h. Finally, we determined that long-lasting nest-building deficits extend to mice undergoing withdrawal from a high dose (4g/kg) of acute EtOH. Sex differences for nest building were absent following EtOH exposure. In mice naïve to EtOH treatments, male mice had lower pre-test body temperatures and increased nest scores across a two-day testing period compared to females. These results suggest that nest building can be used to assess chronic and acute EtOH withdrawal severity in mice.

Effects of acute alcohol withdrawal on nest building in mice selectively bred for alcohol withdrawal severity

“…We aimed to have 10 mice per sex and ethanol treatment group for each experiment. We previously reported that this number of animals produced significant ethanol effects on nest building [22]. Mice were euthanized via CO 2 asphyxiation followed by cervical dislocation after the completion of behavioral testing.…”

“…The schedule of nest scoring allowed for the observation of nest building deficits and tracking of recovery over time. The nest scoring methods are described in more detail elsewhere [22] and were derived from published descriptions [11,18]. Briefly, if the nesting material was untouched, or if some of it had been moved, a score of 0 or 1 was given, respectively.…”

“…Once the nest walls became cupped, scores of 3, 4, or 5 were given based on whether the nest walls were less than half, approximately half, or greater than half the height of a full nest dome, respectively. The full dome height had been established to average 9.38 ± 0.60 cm [22]. …”

“…HS/Npt/Pdx mice had increased HIC scores during withdrawal from 3 days of chronic ethanol vapor exposure, and chronic ethanol dose-dependently suppressed nest building for up to 32 h into withdrawal [22]. This suppression was also observed in mice after a single, acute injection of ethanol.…”

“…In a separate line of studies using HS/Npt/Pdx mice, we found that reduced nest building was observed up to 24 h into withdrawal from an acute 4 g/kg injection of ethanol before ethanol-withdrawn mice began to recover [22]. In earlier selected generations of WSP mice, signs of acute ethanol withdrawal as measured by HICs began to subside 12 h after injection of 4 g/kg ethanol before returning to baseline levels at 24 h [31].…”