Effect of maternal lipopolysaccharide administration on the development of dopaminergic receptors and transporter in the rat offspring

Abstract

Epidemiological evidence supports that maternal infection during gestation are notable risk factors for developmental mental illnesses including schizophrenia and autism. In prenatal lipopolysaccharide (LPS) model of immune activation in rats, the offspring exhibit significant impairments in behaviors mediated by central dopamine (DA) system. This study aimed to examine the temporal and regional pattern of postnatal DA development in the male offspring of pregnant Sprague-Dawley rats administered with 100 µg/kg LPS or saline at gestational days 15/16. Using ligand autoradiography, D1 and D2 dopamine receptors (D1R, D2R) and dopamine transporter (DAT) binding levels were measured in the prefrontal cortex (PFC) and sub cortical regions (dorsal striatum and nucleus accumbens core and shell) at pre pubertal (P35) and post pubertal ages (P60). We found a significant decrease in D2R ligand [(3)H] YM-90151-2 binding in the medial PFC (mPFC) in prenatal LPS-treated animals at P35 and P60 compared to respective saline groups. The decrease in D2R levels was not observed in the striatum or accumbens of maternal LPS-treated animals. No significant changes were observed in [(3)H] SCH23390 binding to D1R. However, the level of [(125)I] RTI-121 binding to DAT was selectively reduced in the nucleus accumbens core and shell at P35 in the prenatal LPS group. Immunohistochemical analysis showed that number of D2R immunopositive cells in infralimbic/prelimbic (IL/PL) part of mPFC was significantly reduced in the LPS group at P60. Prenatal LPS treatment did not significantly affect either the total number of mature neurons or parvalbumin (PV)-immunopositive interneurons in this region. However the number of PV and D2R co-labeled neurons was significantly reduced in the IL/PL subregion of PFC of LPS treated animals. Our data suggests D2R deficit in the PFC and PV interneurons may be relevant to understanding mechanisms of cortical dysfunctions described in prenatal infection animal models as well as schizophrenia.

Figure 1. Schematic representation of the rat brain regions analyzed for the receptor expression at the levels of mPFC (A) or striatum/Nacc (B).

Figure 2. Representative autoradiograms of ligand binding to DA receptors and DA transporter (DAT) in rat brain.

Left and right panels show binding in prenatal saline and LPS treatments respectively. (A), [³H] SCH23390 (D1-like receptor binding) in Str/NAcc; (B), [³H] YM-90151-2 (D2-like receptor binding) in mPFC; (C), [³H] YM-90151-2 in Str/Nacc; and (D), [¹²⁵I] RTI-121 (DAT binding) in Str/Nacc. D1 and D2 autoradiograms are from animals at P60 whereas DAT binding is shown for P35 rat.

Figure 3. Quantitative analysis of specific binding of D1-like receptors in the prenatally saline or LPS treated offspring at P35 and P60.

(A, B) are from striatal/accumbens and (C, D) are from prefrontal cortex (PFC) brain regions. No significant differences in the expression of D1-like receptors in the striatum, nucleus accumbens or prefrontal cortex were observed as a result of prenatal exposure to LPS. Values are expressed as mean±SEM. (Str; Dorsal striatum, Nacc core; Nucleus accumbens core, Nacc shell; Nucleus accumbens shell).

Figure 4. Quantitative analysis of specific binding of D2-like receptors in the prenatally saline or LPS treated offspring at P35 and P60.

(A, B) are from striatal/accumbens and (C, D) are from prefrontal cortex (PFC) brain regions. Specific binding to the D2 like receptors was significantly decreased in PFC of LPS treated offspring at both P35 (**p<0.01) (C) and P60 (*p<0.05) (D). Further analysis of the data revealed that while there is no subregion-specific change in D2 binding at P35 (E), D2 like binding was significantly decreased in IL/PL subregion of mPFC at P60 (**p<0.01) which indicates age-specific changes in the expression of D2 like receptors as a result of prenatal exposure to LPS (F). Values are expressed as mean±SEM. (IL/PL; Infralimbic/prelimbic corteces, Cg; cingulate gyrus, M2; secondary motor cortex, Str; Dorsal striatum, Nacc core; Nucleus accumbens core, Nacc shell; Nucleus accumbens shell).

Figure 5. Quantitative analysis of specific binding to dopamine transporter in the prenatally saline or LPS treated offspring at P35 and P60.

Due to very low level of specific [¹²⁵I] RTI-121binding in the prefrontal cortex (PFC), quantitative analysis was not possible in this brain region. The data showed that DAT binding was significantly reduced in both nucleus accumbens core and shell at P35 (*p<0.05) in maternal LPS treated rats compared to saline treated animals (A). However we did not find any difference in the expression of DAT in the same regions at post pubertal age. All values are expressed as mean±SEM. (Str; Dorsal striatum, Nacc core; Nucleus accumbens core, Nacc shell; Nucleus accumbens shell).

Figure 6. Representative immunohistochemical images of coronal brain sections of the medial prefrontal cortex (mPFC) at postpubertal age.

(A) Neu-N immunolabeling (B) D2 receptor immunolabeling, (C) Parvalbumin immunolabeling and (D) Dual immunolabeling with parvalbumin (Cy3, red) and D2 receptor (FITC, green) specific antibodies.

Figure 7. Quantitative analysis of immunolabeled cells in IL/PL subregions of medial prefrontal cortex (mPFC) at P60.

(A) Total number of mature neurons (NeuN labeling) was not changed comparing maternal LPS and saline treated offspring; (B) reduction in D2-receptor immunolabeled cells in mPFC following maternal LPS challenge in comparison to saline treatment (*p<0.05); (C) number of parvalbumin positive neurons in mPFC was not changed in maternal LPS treated animals; (D) number of parvalbumin neurons which co-express D2 receptor was significantly decreased in maternal LPS animals compared to saline treated ones. (*p<0.05). All values are expressed as mean±SEM.