Developmental exposure to concentrated ambient ultrafine particulate matter air pollution in mice results in persistent and sex-dependent behavioral neurotoxicity and glial activation

Abstract

The brain appears to be a target of air pollution. This study aimed to further ascertain behavioral and neurobiological mechanisms of our previously observed preference for immediate reward (Allen, J. L., Conrad, K., Oberdorster, G., Johnston, C. J., Sleezer, B., and Cory-Slechta, D. A. (2013). Developmental exposure to concentrated ambient particles and preference for immediate reward in mice. Environ. Health Perspect. 121, 32-38), a phenotype consistent with impulsivity, in mice developmentally exposed to inhaled ultrafine particles. It examined the impact of postnatal and/or adult concentrated ambient ultrafine particles (CAPS) or filtered air on another behavior thought to reflect impulsivity, Fixed interval (FI) schedule-controlled performance, and extended the assessment to learning/memory (novel object recognition (NOR)), and locomotor activity to assist in understanding behavioral mechanisms of action. In addition, levels of brain monoamines and amino acids, and markers of glial presence and activation (GFAP, IBA-1) were assessed in mesocorticolimbic brain regions mediating these cognitive functions. This design produced four treatment groups/sex of postnatal/adult exposure: Air/Air, Air/CAPS, CAPS/Air, and CAPS/CAPS. FI performance was adversely influenced by CAPS/Air in males, but by Air/CAPS in females, effects that appeared to reflect corresponding changes in brain mesocorticolimbic dopamine/glutamate systems that mediate FI performance. Both sexes showed impaired short-term memory on the NOR. Mechanistically, cortical and hippocampal changes in amino acids raised the potential for excitotoxicity, and persistent glial activation was seen in frontal cortex and corpus callosum of both sexes. Collectively, neurodevelopment and/or adulthood CAPS can produce enduring and sex-dependent neurotoxicity. Although mechanisms of these effects remain to be fully elucidated, findings suggest that neurodevelopment and/or adulthood air pollution exposure may represent a significant underexplored risk factor for central nervous system diseases/disorders and thus a significant public health threat even beyond current appreciation.

Fig. 1.

Experimental design. Mice were exposed in the postnatal period from PNDs 4–7 and 10–13, with adult re-exposure occurring at PNDs 57–59. Spontaneous locomotor (SLA) activity was collected every other day for three sessions from PNDs 71–78. Starting on PND93, animals were autoshaped and subsequently placed under the FI schedule of reinforcement until PND240. Novel object recognition (NOR) was carried out at ∼6 months of age. Blood was collected (represented by arrows) on PND60, 6 months of age, and upon sacrifice at approximately PND270 .

Fig. 2.

Ambient outdoor UFP and CAPS characterization. Daily outdoor ambient UFP and CAPS counts across all exposure days (left) are reported as number of particles × 10⁵/cm³ ± SDs. Concentration factor (left) was calculated as [CAPS]/[UFP]. Mean particle mass (right) is reported as μg/m³ ± SDs. Daily diameter of CAPS (right) is reported in nm ± SDs.

Fig. 3.

Overall rates, run rates, postreinforcement pause times, and interresponse times for male and female mice performing under fixed-interval schedule controlled behavior (mean ± SE). Data binned into five sessions per bin for each of FI60I, FI120, and FI60II. PN, or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.

Fig. 4.

Novel object recognition in males and females exposed to postnatal Air or CAPS, adult Air or CAPS, and their combination. Change in time spent with novel object (top) and change in number of approaches to the novel object (middle), and change in time per approach to the novel object (bottom) are reported as percent change ± SEM from trial 1 to trial 2 using the object on the same side as the novel object as baseline. The graphic offers a visual depiction of experimental set up. Position of the novel object was counterbalanced against sex and treatment group to preclude potential side bias. PN or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.

Fig. 5.

Ambulatory, horizontal, and vertical activity in male and female mice exposed to postnatal CAPS, adult CAPS, or their combination. Reported as total beam breaks across session ± SE. PN or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.

Fig. 6.

Serum corticosterone in male (left) and female (right) animals exposed to Air/Air, Air/CAPS, CAPS/Air, or CAPS/CAPS 24 h following cessation of adult exposure (point 1), at 6 months of age (point 2) or upon sacrifice (point 3). Reported as group mean ± SE. PN or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. Group indicates a main effect of group. Group × Age indicates a statistical interaction of group × age. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.

Fig. 7.

Monoamine neurotransmitters and metabolites in male and female Air/Air, Air/CAPS, CAPS/Air, and CAPS/CAPS animals. Data reported as group mean ± SE. DA TO calculated as [DOPAC]/[DA]. Reported in ng neurotransmitter or metabolite/mg protein. PN or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.

Fig. 8.

Cortical hippocampal glutamate, glutamine, glutamine:glutamate, and GABA in male and female Air/Air, Air/CAPS, CAPS/Air, and CAPS/CAPS mice as indicated. Data reported group mean ± SE. Reported in ng analyte/mg protein. Glutamine:glutamate ratio calculated as [glutamine]/[glutamate]. PN or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.

Fig. 9.

Selected neurochemistry and corticosterone regressed against FI measures in male mice exposed to Air/Air and CAPS/Air. Statistically significant (p < 0.05) regressions and associated r² and p values are listed in the body of the graph in dashed boxes for the CAPS/Air group and in solid lines for the Air/Air group.

Fig. 10.

Selected neurochemistry and corticosterone regressed against FI measures in Air/Air and Air/CAPS females. Statistically significant (p < 0.05) regressions and associated r² and p values are listed in the body of the graph in dashed boxes for the CAPS/Air group and in solid lines for the Air/Air group.

Fig. 11.

Cortical and corpus callosum astrocyte (GFAP) and microglial/macrophage (IBA-1) activation in CAPS-exposed males and females in indicated treatment groups with representative images of GFAP and IBA1 immunostaining. Relative quantification of GFAP and IBA-1 immunoreactivity in the corpus callosum cortex (left) and corpus callosum (right) of Air/Air, Air/CAPS, CAPS/Air, and CAPS/CAPS exposed males and females. PN or AD indicates statistical main effect of postnatal or adult treatment, respectively. PN × AD indicates statistical interaction of the treatments. * denotes statistical significance from Air/Air, ∼ denotes statistical significance from Air/CAPS, + denotes statistical significance from CAPS/Air, and ^∧ denotes statistical significance from CAPS/CAPS.