Chronic Stress Reduces Nectin-1 mRNA Levels and Disrupts Dendritic Spine Plasticity in the Adult Mouse Perirhinal Cortex

Abstract

In adulthood, chronic exposure to stressful experiences disrupts synaptic plasticity and cognitive function. Previous studies have shown that perirhinal cortex-dependent object recognition memory is impaired by chronic stress. However, the stress effects on molecular expression and structural plasticity in the perirhinal cortex remain unclear. In this study, we applied the chronic social defeat stress (CSDS) paradigm and measured the mRNA levels of nectin-1, nectin-3 and neurexin-1, three synaptic cell adhesion molecules (CAMs) implicated in the adverse stress effects, in the perirhinal cortex of wild-type (WT) and conditional forebrain corticotropin-releasing hormone receptor 1 conditional knockout (CRHR1-CKO) mice. Chronic stress reduced perirhinal nectin-1 mRNA levels in WT but not CRHR1-CKO mice. In conditional forebrain corticotropin-releasing hormone conditional overexpression (CRH-COE) mice, perirhinal nectin-1 mRNA levels were also reduced, indicating that chronic stress modulates nectin-1 expression through the CRH-CRHR1 system. Moreover, chronic stress altered dendritic spine morphology in the main apical dendrites and reduced spine density in the oblique apical dendrites of perirhinal layer V pyramidal neurons. Our data suggest that chronic stress disrupts cell adhesion and dendritic spine plasticity in perirhinal neurons, which may contribute to stress-induced impairments of perirhinal cortex-dependent memory.

Keywords: chronic stress; corticotropin-releasing hormone receptor 1; dendritic spine; nectin-1; perirhinal cortex.

Figure 1

Experimental design. CRHR1-CKO, corticotropin-releasing hormone receptor 1-conditional knockout; CRH-COE, corticotropin-releasing hormone-conditional overexpression; CSDS, chronic social defeat stress; EYFP, enhanced yellow fluorescent protein; WT, wild-type.

Figure 2

Nectin-1 mRNA and protein expression patterns in the adult mouse perirhinal cortex. (A) An in situ hybridization image showing nectin-1 mRNA expression in the perirhinal cortex (the boxed region) and adjacent brain regions. Scale bar = 500 μm. (B) A representative image showing nectin-1 immunostaining in the perirhinal cortex (the boxed region) and adjacent brain regions. Scale bar = 500 μm. (C) The magnified image of the insert in (B) showing nectin-1-positive cells in layers II–VI of areas 36 and 35. Scale bar = 50 μm. (D) Analysis of nectin-1 immunoreactivity in layers I–VI of perirhinal areas 36 and 35. *p < 0.05, main effect. n = 3 mice. (E–G) Nectin-1 colocalized with rat pyramidal cell (RPC), a marker for excitatory pyramidal neurons. Arrowheads indicate representative area 35 neurons co-immunostained with nectin-1 and RPC. (H–J) Nectin-1 partially colocalized with glutamic acid decarboxylase 67 (GAD67), a marker for inhibitory interneurons. Arrowheads indicate area 35 neurons that co-express nectin-1 and GAD67. Arrows indicate representative nectin-1-immunoreactive neurons surrounded by GAD67-positive inhibitory boutons. (K–M) Nectin-1 partially colocalized with calbindin, a marker for a subtype of pyramidal neurons and interneurons. Arrowheads indicate area 35 neurons co-immunostained with nectin-1 and calbindin. Arrows indicate representative nectin-1-positive neurons without calbindin expression. Scale bars for (E–M) are 20 μm.

Figure 3

Effects of CSDS and the CRH-CRHR1 system on nectin-1 mRNA levels in the perirhinal cortex. (A) Representative in situ hybridization images showing nectin-1 mRNA expression in the perirhinal cortex (boxed regions) of WT or CRHR1-CKO mice with or without chronic stress exposure. (B) Nectin-1 mRNA levels in the perirhinal cortex were reduced by chronic stress in WT but not CRHR1-CKO mice. n = 6–8 mice per group. (C) Representative in situ hybridization images showing nectin-1 mRNA expression in the perirhinal cortex (boxed regions) of WT or CRH-COE mice. (D) Perirhinal nectin-1 mRNA levels were reduced in CRH-COE mice. n = 9 mice per group. All scale bars = 500 μm. *p < 0.05 vs. the control WT group; ^#p < 0.05 vs. the stressed WT group.

Figure 4

Effects of CSDS on dendritic spine plasticity in perirhinal layer V pyramidal neurons. (A) A transverse section at the ventral hippocampal level from a Thy1-EYFP-H mouse. Note that in the medial entorhinal cortex (MEA) and the lateral entorhinal cortex (LEA), very few neurons are labeled by EYFP. PRH, perirhinal cortex. Scale bar = 500 μm. (B) The magnified image of the insert in (A) showing EYFP-labeled layer V pyramidal neurons in the perirhinal cortex. The asterisk indicates the rhinal fissure. Scale bar = 50 μm. (C) High-resolution projected raw z-stacks of the main apical dendrite in the insert (solid line) in (B). Scale bar = 5 μm. (D) High-resolution projected raw z-stacks of the oblique apical dendrite in the inset (dashed line) in (B). Scale bar = 5 μm. (E) Representative deconvolved and projected z-stacks showing the main apical dendrites in the control and defeat mice. Scale bars = 5 μm. (F,G) In the main apical dendrites, spine density and spine volume were comparable between groups. (H,I) CSDS increased spine head diameter but reduced spine length. (J) Representative deconvolved and projected z-stacks showing the oblique apical dendrites in the control and defeat mice. Scale bars = 5 μm. (K) CSDS reduced spine density in the oblique apical dendrites of perirhinal layer V pyramidal neurons. (L–N) Spine volume, spine head diameter and spine length were similar between groups. *p < 0.05 vs. the control group. n = 4 mice per group.