Effects of Chronic Secondhand Smoke (SHS) Exposure on Cognitive Performance and Metabolic Pathways in the Hippocampus of Wild-Type and Human Tau Mice

Abstract

Background: Exposure to secondhand smoke (SHS) is a risk factor for developing sporadic forms of sporadic dementia. A human tau (htau) mouse model is available that exhibits age-dependent tau dysregulation, neurofibrillary tangles, neuronal loss, neuroinflammation, and oxidative stress starting at an early age (3-4 months) and in which tau dysregulation and neuronal loss correlate with synaptic dysfunction and cognitive decline.

Objective: The goal of this study was to assess the effects of chronic SHS exposure (10 months' exposure to $\sim 30{\mathrm{mg}/m}^3$) on behavioral and cognitive function, metabolism, and neuropathology in mice.

Methods: Wild-type (WT) and htau female and male mice were exposed to SHS (90% side stream, 10% main stream) using the SCIREQ® inExpose™ system or air control for 168 min per day, for 312 d, 7 d per week. The exposures continued during the days of behavioral and cognitive testing. In addition to behavioral and cognitive performance and neuropathology, the lungs of mice were examined for pathology and alterations in gene expression.

Results: Mice exposed to chronic SHS exposure showed the following genotype-dependent responses: a) lower body weights in WT, but not htau, mice; b) less spontaneous alternation in WT, but not htau, mice in the Y maze; c) faster swim speeds of WT, but not htau, mice in the water maze; d) lower activity levels of WT and htau mice in the open field; e) lower expression of brain PHF1, TTCM1, $\text{IGF}1\beta$, and HSP90 protein levels in WT male, but not female, mice; and f) more profound effects on hippocampal metabolic pathways in WT male than female mice and more profound effects in WT than htau mice.

Discussion: The brain of WT mice, in particular WT male mice, might be especially susceptible to the effects of chronic SHS exposure. In WT males, independent pathways involving ascorbate, flavin adenine dinucleotide, or palmitoleic acid might contribute to the hippocampal injury following chronic SHS exposure. https://doi.org/10.1289/EHP8428.

Figure 1.

Gravimetric analysis of the SHS and plasma cotinine at 4- and 8-month, body weight in WT and htau mice, and cumulative food intake in mice exposed or sham exposed to SHS for 312 d. All data are shown as $\text{mean}\pm \text{SEM}$. (A) Mass of SHS to which male and female mice were exposed as determined by gravimetric analysis. The genotypes were combined for analysis of plasma cotinine levels and cotinine metabolites. (B) Plasma cotinine levels at the 4-month time point. (C) Plasma cotinine levels at the 8-month time point. (D) Plasma levels of cotinine-N-oxide in male and female mice at the 8-month time point. *$p=0.016$, 2-tailed t-test. (E) Plasma levels of trans-3′-hydroxy-cotinine in male and female mice at the 8-month time point. **$p=0.0034$, 2-tailed t-test. (F) Body weight in male WT mice over the course of SHS or sham exposure. **$p=0.0016$, repeated-measures ANOVA. (G) Body weight in male htau mice over the course of SHS or sham exposure. (H) Body weight in female WT mice over the course of SHS or sham exposure. [$F\left(\mathrm{1,14}\right)=3.249$, $p=0.0931$, repeated-measures ANOVA]. (I) Body weight in female htau mice over the course of SHS or sham exposure. (J) Cumulative food intake in mice exposed to SHS or air controls over the course of the exposure period. *$p=0.0112$, 2-tailed t-test. Note: ANOVA, analysis of variance; SHS, secondhand smoke; WT, wild type.

Figure 2.

Behavioral performance of SHS- and air-exposed WT and htau mice. (A) Schematic illustration of the Y maze. (B) Spontaneous alternation of SHS- and air-exposed WT and htau mice. *$p=0.0316$, 2-tailed t-test; ^#$p=0.0728$, 2-tailed t-test. (C) Total arm entries, a measure of activity, of SHS- and air-exposed WT and htau mice. (D) Mice were tested for two sequential days for behavioral performance in the open field. On the third day, two identical objects were placed in the open field. On the fourth day, one familiar object was replaced by a novel one. (E) Activity levels of SHS- and air-exposed WT and htau mice on days 1 and 2 in the open field. *$p<0.05$ vs. sham, ANOVA. (F) Entries of SHS- and air-exposed WT and htau mice into the center of the open field. ⁰$p<0.05$ vs. air exposed htau, ANOVA. *$p<0.05$ vs. sham htau on day 1, 2-tailed t-test. (G) Time SHS- and air-exposed WT and htau mice spent in the center of the open field. There was a $\text{genotype}\times \text{exposure}$ interaction [$F\left(\mathrm{1,58}\right)=4.704$, $p=0.034$, ANOVA]. There was an effect of SHS in htau mice [$F\left(\mathrm{1,29}\right)=6.077$, $p=0.0199$, ANOVA]. ⁰$p<0.05$ vs. sham htau, ANOVA. **$p<0.01$ vs. sham htau on day 1, 2-tailed t-test. (H) Object recognition of SHS- and air-exposed WT and htau mice. *$p<0.05$ vs. the familiar object, 2-tailed t-test. (I) Spatial learning and memory of SHS- and air-exposed WT and htau mice in the water maze. (J) Swim speeds of SHS- and air-exposed WT and htau mice in the water maze. ***$p=0.0009$, ANOVA. (K) Learning curves of WT mice to locate the visible and hidden platform. (L) Learning curves of htau mice to locate the visible and hidden platform. (M) Spatial memory retention of WT and htau mice in the first probe trial. *$p<0.05$, Dunnett’s. (N) Spatial memory retention of WT and htau mice in the second probe trial. *$p<0.05$, Dunnett’s. (O) Cumulative distance to the platform location of WT and htau mice in the water maze probe trials. ***$p<0.0001$, ANOVA. (P) Baseline motion of SHS- and air-exposed WT and htau mice in the fear conditioning test. **$p<0.01$, 2-tailed t-test. (Q) Freezing of SHS- and air-exposed WT and htau mice between the aversive stimuli during fear learning. (R) Contextual fear memory of SHS- and air-exposed WT and htau mice. All data are shown as $\text{mean}\pm \text{SEM}$. Note: ANOVA, analysis of variance; SEM, standard error of the mean; SHS, second hand smoke; WT, wild type.

Figure 3.

Expression of common lung genes in WT male and female mice and htau male mice. The heat map illustrates the expression of molecular toxicity pathway-related genes in the lungs of the mice. The black color denotes no difference in fold-change in comparison with air controls. $n=4$ mice/genotype/sex/exposure condition. Data are expressed as fold-change in comparison with the genotype- and sex-matched control group. A change greater than $\pm 1.5\text{-fold}$ is considered significant. Note: WT, wild type.

Figure 4.

Levels of PHF1 (A–H) and TTCM1 (I–P) in female (A–D; I–L) and male (E–H; M–P) mice as assessed by dot blot analysis. ***$p<0.001$ vs. female WT mice, ANOVA. ^#$p<0.05$ vs. sham mice, ANOVA. ^###$p=0.001$ vs. sham male mice, ANOVA. *$p<0.05$ vs. WT male mice, ANOVA. WT F Air: $n=5$ mice; WT F SHS: $n=5$ mice; htau F Air: $n=5$ mice; htau F SHS: $n=5$ mice; WT M Air: $n=4$ mice; WT M SHS: $n=5$ mice; htau M Air: $n=5$ mice; htau M SHS: $n=4$ mice. All data are shown as $\text{mean}\pm \text{SEM}$. Note: ANOVA, analysis of variance; CTX, cortex; HP, hippocampus; CERB, cerebellum; F, females; M, males; SEM, standard error of the mean; SHS secondhand smoke; WT, wild type.

Figure 5.

Levels of $\text{IGF}1\mathrm{\beta }$ (A–H) and HSP90 (I–P) in female (A–D; I–L) and male (E–H; M–P) mice as assessed by dot blot analysis. *$p<0.05$ vs. WT female mice. ^#$p<0.05$ vs. sham male mice. ***$p<0.001$ vs. male WT mice, ANOVA. ^###$p=0.001$ vs. sham male mice, ANOVA. WT F Air: $n=5$ mice; WT F SHS: $n=5$ mice; htau F Air: $n=5$ mice; htau F SHS: $n=5$ mice; WT M Air: $n=4$ mice; WT M SHS: $n=5$ mice; htau M Air: $n=5$ mice; htau M SHS: $n=4$ mice. All data are shown as $\text{mean}\pm \text{SEM}$. Note: ANOVA, analysis of variance; CTX, cortex; HP, hippocampus; CERB, cerebellum; F, females; M, males; SEM, standard error of the mean; SHS, secondhand smoke; WT, wild type.

Figure 6.

Hippocampal pathways in WT male (A), WT female (B), htau male (C), and htau female (D) mice exposed to SHS. Red letters within each panel highlight the pathways most affected within each group and are listed here. For WT males (A): A. Taurine and hypotaurine metabolism; B. Alanine, aspartate and glutamate metabolism; C. Glycine, serine and threonine metabolism; and D. Arginine and proline metabolism. In WT females (B): A. Taurine and hypotaurine metabolism; B. Glycine, serine and threonine metabolism; C. Arginine and proline metabolism; and D. Arginine biosynthesis. In htau males (C): A. Arginine and proline metabolism; B. Glutathione metabolism; C. Pentose phosphate pathway; and D: beta-Alanine metabolism. In htau females (D): A. Arginine and proline metabolism; B. Phenylalanine metabolism; C. Ascorbate and aldarate metabolism; and D. beta-Alanine metabolism.