Neurobehavioral phenotype of C57BL/6J mice prenatally and neonatally exposed to cigarette smoke

Abstract

Although maternal cigarette smoking during pregnancy is a well-documented risk factor for a variety of adverse pregnancy outcomes, how prenatal cigarette smoke exposure affects postnatal neurobehavioral/cognitive development remains poorly defined. In order to investigate the cause of an altered behavioral phenotype, mice developmentally exposed to a paradigm of 'active' maternal cigarette smoke is needed. Accordingly, cigarette smoke exposed (CSE) and air-exposed C57BL/6J mice were treated for 6h per day in paired inhalation chambers throughout gestation and lactation and were tested for neurobehavioral effects while controlling for litter effects. CSE mice exhibited less than normal anxiety in the elevated zero maze, transient hypoactivity during a 1h locomotor activity test, had longer latencies on the last day of cued Morris water maze testing, impaired hidden platform learning in the Morris water maze during acquisition, reversal, and shift trials, and impaired retention for platform location on probe trials after reversal but not after acquisition or shift. CSE mice also showed a sexually dimorphic response in central zone locomotion to a methamphetamine challenge (males under-responded and females over-responded), and showed reduced anxiety in the light-dark test by spending more time on the light side. No differences on tests of marble burying, acoustic startle response with prepulse inhibition, Cincinnati water maze, matching-to-sample Morris water maze, conditioned fear, forced swim, or MK-801-induced locomotor activation were found. Collectively, the data indicate that developmental cigarette smoke exposure induces subnormal anxiety in a novel environment, impairs spatial learning and reference memory while sparing other behaviors (route-based learning, fear conditioning, and forced swim immobility). The findings add support to mounting evidence that developmental cigarette smoke exposure has long-term adverse effects on brain function.

Figure 1

Elevated zero maze: Left, time-in-open quadrants; Middle, number of head-dips; Right, number of open quadrant entries. In all figures, data are shown as least square means ± SEM. **P <0.01, ***P <0.001, CSE vs. Control. Group sizes: total (males/females): Control = 34 (14/19), CSE = 27 (14/13).

Figure 2

Locomotor activity—central distance (cm): Mice were tested for 60 min and data analyzed in 5 min intervals. *P <0.05, ***P<0.001, ^†P <0.10, CSE vs Control. Group sizes: total (males/females): Control = 34 (14/19), CSE = 27 (15/12).

Figure 3

MWM (cued). Latency to reach goal (s). Mice were given 2 trials/day on days 2-6 with the platform and goal repositioned randomly on each trial with the platform clearly marked with an orange ball mounted on a pole affixed to the center of the platform. **P<0.01, CSE vs Control. Group sizes: total (males/females): Control = 67 (33/34), CSE = 53 (27/26).

Figure 4

Locomotor activity with methamphetamine challenge—central distance (cm). Mice were re-acclimated to the test for 30 min prior to methamphetamine administration (1.0 mg/kg s.c.). After treatment with methamphetamine they were placed back in the test apparatus for an additional 180 min. Data are least square mean ± SEM per 5 min interval. *P<0.05, ^†P <0.10, CSE vs. Control. Note *P<0.05 with bracket indicates significant difference averaged across intervals for the pre-challenge test session. Post-challenge data were analyzed using the pre-challenge data as a covariate to control for differences in baseline prior to methamphetamine treatment. Group sizes: total (males/females): Control = 33 (15/18), CSE = 27 (14/13).

Figure 5

MWM Fixed hidden platform. Mice were tested in three phases (acquisition, reversal, shift) each consisting of 6 training days (4 trials/day) followed 24 h later by a single probe trial for reference memory with the platform removed. Differences were found on acquisition for initial heading error (panel A), on reversal average heading error for females (panel B), and on shift for path length (panel C) and average heading error (panel D). Heading error is expressed in degrees. Data are averaged across days to reflect the main effect of treatment or treatment and sex (mean ± SEM). *P <0.05, **P <0.01, CSE vs Control. Group sizes: total (males/females) = Control = 33 (15/18), CSE = 27 (14/13).

Figure 6

MWM probe trials. Data are least square mean ± SEM on probe trials for acquisition-probe site crossovers (panel A), for acquisition-probe percent time and percent distance in the target quadrant (panel B), for reversal-probe for percent time in the target quadrant (panel C), and on reversal-probe initial heading error (in degrees; panel D). Probe trials were given 24 h after the last platform trial of each phase. *P <0.05, **P <0.01, ***P <0.001, CSE vs Control. Group sizes: total (males/females) = Control = 33 (15/18), CSE = 27 (14/13).