Respiratory viral infection in neonatal piglets causes marked microglia activation in the hippocampus and deficits in spatial learning

Abstract

Environmental insults during sensitive periods can affect hippocampal development and function, but little is known about peripheral infection, especially in humans and other animals whose brain is gyrencephalic and experiences major perinatal growth. Using a piglet model, the present study showed that inoculation on postnatal day 7 with the porcine reproductive and respiratory syndrome virus (PRRSV) caused microglial activation within the hippocampus with 82% and 43% of isolated microglia being MHC II(+) 13 and 20 d after inoculation, respectively. In control piglets, <5% of microglia isolated from the hippocampus were MHC II(+). PRRSV piglets were febrile (p < 0.0001), anorectic (p < 0.0001), and weighed less at the end of the study (p = 0.002) compared with control piglets. Increased inflammatory gene expression (e.g., IL-1β, IL-6, TNF-α, and IFN-γ) was seen across multiple brain regions, including the hippocampus, whereas reductions in CD200, NGF, and MBP were evident. In a test of spatial learning, PRRSV piglets took longer to acquire the task, had a longer latency to choice, and had a higher total distance moved. Overall, these data demonstrate that viral respiratory infection is associated with a marked increase in activated microglia in the hippocampus, neuroinflammation, and impaired performance in a spatial cognitive task. As respiratory infections are common in human neonates and infants, approaches to regulate microglial cell activity are likely to be important.

Keywords: PRRSV; brain; cognition; inflammation; sickness behavior.

Figure 1.

Experimental timeline.

Figure 2.

Daily recorded sickness measures (A, body weight; B, rectal temperature; C, feeding score) for control (13 and 20 d after inoculation combined, n = 14 total) and PRRSV (13 and 20 d after inoculation combined, n = 19 total) piglets throughout the course of the study. Body weight was measured from the start of the experiment, and rectal temperature and feeding score were measured beginning 1 d before inoculation. Data are LSM ± SEM. Contrasts were used to separate treatment means on each day *p < 0.05. ^#p < 0.10.

Figure 3.

Performance of control (13 and 20 d after inoculation combined, n = 14 total) and PRRSV (13 and 20 d after inoculation combined, n = 19 total) piglets during the acquisition phase (A1–A8) and reversal phase (R1–R3) of cognitive testing in a spatial T-maze task (A, proportion of correct reward arm choices/total trials) and locomotor readouts (B, latency to choice; C, total distance moved). Data are LSM ± SEM.

Figure 4.

A representative two-color dot plot for a control (A) and a PRRSV (B) piglet is shown; cells were incubated with antibodies for receptors used to identify microglia cells (CD11b and CD45, data not shown), as well as a marker of cell activation (MHC II, shown). The proportion of activated microglia cells at the end of the study in piglets killed 13 d after inoculation (control 1: n = 6; PRRSV 1: n = 8) and 20 d after inoculation (control 2: n = 8; PRRSV 2: n = 11; C). In addition, the correlation of this measure to the average rectal temperature (D), the average feeding score (E), and performance in the cognitive behavioral task (F) for piglets killed at 13 d after inoculation are shown. A–C, Data are LSM ± SEM. Contrasts were used to separate treatment means within each time point (control vs PRRSV at 13 d after inoculation; control vs PRRSV at 20 d after inoculation; ^φp < 0.05) and between cohorts (control 13 d vs control 20 d after inoculation; PRRSV 13 d vs PRRSV 20 d after inoculation; ^†p < 0.05).

Figure 5.

Relative abundance of mRNA in peripheral tissues (A, lung; B, tracheobronchial lymph node) and brain tissues (C, hippocampus; D, prefrontal cortex; E, striatum) for control (13 and 20 d after inoculation combined, n = 14 total) and PRRSV (13 and 20 d after inoculation combined, n = 19 total) piglets. Only significant treatment effects are shown. Data are LSM ± SEM. The gray line indicates the standardized Relative Quantification (RQ) baseline (RQ = 1); all data are expressed as relative fold change to the baseline. Contrasts were used to separate treatment means for each gene. *p < 0.05. ^#p < 0.10.