Retrosplenial Cortex Effects Contextual Fear Formation Relying on Dysgranular Constituent in Rats

Abstract

Animal contextual fear conditioning (CFC) models are the most-studied forms used to explore the neural substances of posttraumatic stress disorder (PTSD). In addition to the well-recognized hippocampal-amygdalar system, the retrosplenial cortex (RSC) is getting more and more attention due to substantial involvement in CFC, but with a poor understanding of the specific roles of its two major constituents-dysgranular (RSCd) and granular (RSCg). The current study sought to identify their roles and underlying brain network mechanisms during the encoding processing of the rat CFC model. Rats with pharmacologically inactivated RSCd, RSCg, and corresponding controls underwent contextual fear conditioning. [¹⁸F]-fluorodeoxyglucose positron emission tomography/computed tomography (¹⁸F-FDG PET/CT) scanning was performed for each animal. The 5-h and 24-h retrieval were followed to test the formation of contextual memory. Graph theoretic tools were used to identify the brain metabolic network involved in encoding phase, and changes of nodal (brain region) properties linked, respectively, to disturbed RSCd and RSCg were analyzed. Impaired retrieval occurred in disturbed RSCd animals, not in RSCg ones. The RSC, hippocampus (Hip), amygdala (Amy), piriform cortex (Pir), and visual cortex (VC) are hub nodes of the brain-wide network for contextual fear memory encoding in rats. Nodal degree and efficiency of hippocampus and its connectivity with amygdala, Pir, and VC were decreased in rats with disturbed RSCd, while not in those with suppressed RSCg. The RSC plays its role in contextual fear memory encoding mainly relying on its RSCd part, whose condition would influence the activity of the hippocampal-amygdalar system.

Keywords: brain metabolic network; contextual fear formation; graph theory; hippocampal-amygdalar system; retrosplenial cortex.

FIGURE 1

Effects of suppressed RSCd/g on contextual fear memory formation. (A) Diagram for experimental procedures. (B) The learning curves of four group rats (RSCd vehicle/inhibitor group and RSCg vehicle/inhibitor group) show no significant difference; RSCd vehicle group n = 24, inhibitor group n = 22; RSCg vehicle group n = 35, inhibitor group n = 32. (C) Memory retrieval during the 5-h and 24-h post-learning tests are significantly impaired in the RSCd inhibitor independent groups compared to the RSCd vehicle independent groups; 5 h, RSCd vehicle group n = 12, inhibitor group n = 11; 24 h, RSCd vehicle group n = 12, inhibitor group n = 11. (D) There is no significant difference in memory retrieval between the RSCg inhibitor independent groups during the 5-h and 24-h post-learning tests compared to the RSCg vehicle independent groups; 5 h, RSCg vehicle group n = 18, inhibitor group n = 16; 24 h, RSCg vehicle group n = 17, inhibitor group n = 16. (E) Memory retrieval during the 5-h and 24-h post-learning is significantly impaired in the RSCd inhibitor groups compared to the RSCg inhibitor groups; 5 h, RSCd inhibitor group n = 11, RSCg inhibitor group n = 16; 24 h, RSCd inhibitor group n = 11, RSCg inhibitor group n = 16. ITI, intertrial interval; i.p., intraperitoneal injection. *p < 0.05, **p < 0.01, two-way ANOVA followed by Bonferroni’s posttests.

FIGURE 2

Effects of suppressed RSCd on hub nodes of the brain-wide metabolic network during CFC. Brain-wide metabolic network of (A) RSCd vehicle group and (C) inhibitor group during CFC. Distribution of nodal degree Z-scores in network of (B) RSCd vehicle group and (D) inhibitor group. (E) Decrease in nodal degree of Pir, RSCd, RSCg, and hippocampus in rats with suppressed RSCd. The error bar indicates the 95% confidence intervals obtained from 1,000-iteration bootstrapping procedures. RSCd vehicle group n = 24, RSCd inhibitor group n = 22. *p < 0.05, ***p < 0.001, permutation test.

FIGURE 3

Effects of suppressed RSCg on hub nodes of the brain-wide metabolic network during CFC. Brain-wide metabolic network of (A) RSCg vehicle group and (C) inhibitor group during CFC. Distribution of nodal degree Z-scores in network of (B) RSCg vehicle and (D) inhibitor group. (E) Decrease in nodal degree of RSCg and hippocampus in rats with suppressed RSCg. The error bar indicates the 95% confidence intervals obtained from 1,000-iteration bootstrapping procedures. RSCg vehicle group n = 35, RSCg inhibitor group n = 32. *p < 0.05, permutation test.

FIGURE 4

Effects of suppressed RSCd on its core metabolic networks during CFC. (A) Core metabolic network of RSCd vehicle group and inhibitor groups during CFC. (B) Degree centrality and (C) nodal efficiency of the hippocampus significantly decrease in rats with suppressed RSCd. (D) 10,000 permutation tests reveal a significant decrease in metabolic correlations between the hippocampus and multiple nodes in rats with suppressed RSCd. (E) The links that changed significantly in metabolic connectivity between nodes of rat brain with suppressed RSCd. MC, metabolic connectivity. RSCd vehicle group n = 24, RSCd inhibitor group n = 22. p < 0.05 was considered statistically significant.

FIGURE 5

Effects of suppressed RSCg on its core metabolic networks during CFC. (A) Core metabolic network of RSCg vehicle and inhibitor groups during CFC. (B) Degree centrality and (C) nodal efficiency of nodes in RSCg core network do not alter significantly in rats with suppressed RSCg. (D) 10,000 permutation tests reveal that the metabolic correlations of the hippocampus with other nodes also do not alter significantly in rats with suppressed RSCg. (E) The links that changed significantly in metabolic connectivity between nodes of rat brain with suppressed RSCd. MC, metabolic connectivity. RSCg vehicle group n = 35, RSCg inhibitor group n = 32. p < 0.05 was considered statistically significant.