The Long-Term Effects of Neonatal Inflammatory Pain on Cognitive Function and Stress Hormones Depend on the Heterogeneity of the Adolescent Period of Development in Male and Female Rats

Abstract

Exposure to stress at an early age programs the HPA axis which can lead to cognitive deficits in adults. However, it is not known whether these deficits emerge in adulthood or are expressed earlier in life. The aims of the study were to investigate (1) the immediate effects of early injury-induced stress in one-day-old (P1) and repeated stress on at P1 and P2 rat pups on plasma corticosterone levels; and (2) examine the subsequent long-term effects of this early stress on spatial learning and memory, and stress reactivity in early P26-34 and late P45-53 adolescent male and female rats. Intra-plantar injection of formalin induced prolonged and elevated levels of corticosterone in pups and impaired spatial learning and short- and long-term memory in late adolescent males and long-term memory in early adolescent females. There were sex differences in late adolescence in both learning and short-term memory. Performance on the long-term memory task was better than that on the short-term memory task for all early adolescent male and female control and stressed animals. Short-term memory was better in the late age control rats of both sexes and for formalin treated females as compared with the early age rats. These results are consistent with an impaired function of structures involved in memory (the hippocampus, amygdala, prefrontal cortex) after newborn pain. However, activation of the HPA axis by neonatal pain did not directly correlate with spatial learning and memory outcomes and the consequences of neonatal pain remain are likely multi-determined.

Keywords: adolescence; corticosterone; neonatal pain; sex differences; spatial learning; spatial memory.

FIGURE 1

Mean (±SEM) latency to find the platform in the first four training trials for 5 days and second four training trials for four training days of spatial learning in Control and Formalin male and female rats of early (P26-34) and late (P45-53) age groups. Panels (A,B) show data for male and female rats at the early age group. Panels (C,D) show data for male and female rats at the late age group. The abscissa shows the first and second training four trials (1 and 2) in each of the five training days. ⁺p < 0.05, ⁺⁺p < 0.01, ⁺⁺⁺p < 0.001 significant differences in Control rats between the first four training trials and the second four training trials each day. ^∗p < 0.05, ^∗∗p < 0.01 Formalin rats vs Control rats.

FIGURE 2

Age and Sex effects in the mean (±SEM) latency to find the platform in Control and Formalin male and female rats in the first four training trials during five training days [early P26-34 and late P45-53 age groups (A,B)] and in the second four training trials during four training days [early P26-34 and late P45-53 age groups (C,D)]. ^∗p < 0.05, ^∗∗p < 0.01 age differences in Formalin rats, ⁺p < 0.05, ⁺⁺p < 0.01 age differences in Control rats. ^#p < 0.05 sex differences in P45-P48 Formalin rats. Abscissa, training days. The number of the rats in the groups corresponds to the number of rats in Figure 1.

FIGURE 3

The index of acquisition (A) and the savings index (B) for latency to find the platform during spatial learning in male and female rats of early (P26-34) and late (P45-53) age groups. The data are: Mean (±SEM) latency differences between the first and last training trials of each of the five training days of spatial learning (A). Mean (±SEM) latency differences between the last training trial of a given day and the first training trial of the next day during the five-day spatial learning (B). Both indices illustrate a decrease of latency to find the platform with age in Control rats; neonatal formalin-induced pain leveled the age differences. ^∗p < 0.05 Formalin vs Control (A,B); ⁺p < 0.05 Control P45-49 rats vs Control P26-34 rats (A,B). The number of the rats in the groups corresponds to the number of rats in Figures 1, 2.

FIGURE 4

Mean (±SEM) latency to find the platform (A,B) for short-term (A) and long-term (B) spatial memory in the Formalin or Control male and female rats for the early (P26-34) and late (P45-53) age groups. Differences in latency between short-term and long-term memory were found in the Control male and female rats of the early age group and in females of the late age group. Formalin vs Control rats; differences between short- and long-term memory: in latency, ^∧p < 0.05, ^&⁣&p < 0.01, in Control P26-34 males and females, and ^$$p < 0.001, in Control P45-53 females. The number of the rats in the groups corresponds to the number of rats in the groups in Figures 1, 2. The graphs on the right illustrate significant results of statistical analysis. ^∗p < 0.05 significant effect of exposure.

FIGURE 5

Mean (±SEM) time in target quadrant (A,B) for short-term (A) and long-term (B) spatial memory in the Formalin or Control male and female rats for the early (P26-34) and late (P45-53) age groups. Differences in time in target quadrant between short-term and long-term memory were found in Control and Formalin males and females of early age groups. In all cases, the time in target quadrant was shorter in the short-term memory (A,B). ⁺⁺⁺p < 0.001, age differences in Control rats; ^###p < 0.001, age differences in Formalin rats. Differences between short- and long-term memory: in time in target quadrant, ^∧∧∧p < 0.001, ^&⁣&⁣&p < 0.001 in Control P26-34 males and females, ^vvvp < 0.001, ^αα p < 0.01, in Formalin P26-34 males and females; ⁰p < 0.05, sex differences in P45-53 Formalin rats. The number of the rats in the groups corresponds to the number of rats in Figures 1, 2. The graphs below illustrate significant results of statistical analysis. ^∗p < 0.05, ^∗∗p < 0.01 significant effect of exposure. Graphs (A1) and (B1) illustrate the significant outcomes of the statistical analyses.

FIGURE 6

Mean (±SEM) corticosterone levels in blood plasma in neonatal pups under basal conditions, or 30 min, 24 h and 7 days after injection of Formalin (2.5%, 0.5 μl) or Control into the pad of the left hind paw. ^∗p < 0.05, ^∗∗p < 0.01, Formalin vs saline; ⁺p < 0.05, ⁺⁺⁺p < 0.001, Formalin vs basal; ^∧∧p < 0.01, saline vs basal.

FIGURE 7

Mean (±SEM) corticosterone levels in blood plasma in response to forced swimming in Control and Formalin rats at the age of P34 and P53 after testing in the Morris Water Maze. ⁺p < 0.05, ⁺⁺p < 0.01, ⁺⁺⁺p < 0.001, Formalin vs basal; ^∧∧p < 0.01, ^∧∧∧p < 0.001 Control vs basal; ^&p < 0.05, age differences between Control rats; ^vvp < 0.01, age differences between Formalin rats. The number of the rats in the groups corresponds to the number of rats in Figures 1, 2.