Moderate Traumatic Brain Injury Causes Acute Dendritic and Synaptic Degeneration in the Hippocampal Dentate Gyrus

Chen J (2011) Moderate Traumatic Brain Injury Causes Acute Dendritic and Synaptic Degeneration in the Hippocampal Dentate Gyrus. PLoS ONE 6(9): e24566. doi:10.1371/journal.pone.0024566 Moderate Traumatic Brain Injury Causes Acute Dendritic and Synaptic Degeneration in the Hippocampal Dentate Gyrus Xiang Gao 0 Ping Deng 0 Zao C. Xu 0 Jinhui Chen 0 Colin Combs, University of North Dakota, United States of America 0 1 Spinal Cord and Brain Injury Research Group, Department of Neurosurgery, Stark Neuroscience Research Institute, Indianapolis, Indiana, United States of America, 2 Department of Anatomy and Cell Biology, Indiana University School of Medicine , Indianapolis, Indiana , United States of America Hippocampal injury-associated learning and memory deficits are frequent hallmarks of brain trauma and are the most enduring and devastating consequences following traumatic brain injury (TBI). Several reports, including our recent paper, showed that TBI brought on by a moderate level of controlled cortical impact (CCI) induces immature newborn neuron death in the hippocampal dentate gyrus. In contrast, the majority of mature neurons are spared. Less research has been focused on these spared neurons, which may also be injured or compromised by TBI. Here we examined the dendrite morphologies, dendritic spines, and synaptic structures using a genetic approach in combination with immunohistochemistry and Golgi staining. We found that although most of the mature granular neurons were spared following TBI at a moderate level of impact, they exhibited dramatic dendritic beading and fragmentation, decreased number of dendritic branches, and a lower density of dendritic spines, particularly the mushroom-shaped mature spines. Further studies showed that the density of synapses in the molecular layer of the hippocampal dentate gyrus was significantly reduced. The electrophysiological activity of neurons was impaired as well. These results indicate that TBI not only induces cell death in immature granular neurons, it also causes significant dendritic and synaptic degeneration in pathohistology. TBI also impairs the function of the spared mature granular neurons in the hippocampal dentate gyrus. These observations point to a potential anatomic substrate to explain, in part, the development of posttraumatic memory deficits. They also indicate that dendritic damage in the hippocampal dentate gyrus may serve as a therapeutic target following TBI. - Funding: This work was supported by grants from the Indiana Spinal Cord & Brain Injury Research Fund (SCBI 200-12)(http://www.in.gov/isdh/23657.htm); the Ralph W. and Grace M. Showalter Research Award, and the Indiana University Biological Research Grant to J. Chen. Both the Showalter award and the biomedical research award are internal funding, and do not have grant numbers and specific websites. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. Traumatic brain injury (TBI) not only results in immediate CNS tissue disruption (primary injury), but also causes secondary damage among the surviving cells via complex mechanisms triggered by the primary event [1,2]. This secondary injury initiated by the primary impact leads to persistent cognitive, sensory, and motor dysfunction [3]. The hippocampus undergoes similar neuropathological changes after both human closed-head injury and experimental injury models of TBI, including controlled cortical impact (CCI) injury [2,4], fluid percussion [5], and stretch injury [6]. These common changes suggest that the hippocampus is particularly vulnerable to secondary injury following TBI [7]. The pathologies in the hippocampus play a leading role in the disturbance of learning, memory [8], and higher cognitive function [8]. Hippocampal injury-associated learning and memory deficits are frequent hallmarks of brain trauma and are the most enduring and devastating consequences following TBI [9]. Currently, there is no effective treatment that can preserve or restore such functions. Therefore, it is critically important to understand the molecular and cellular mechanisms behind learning and memory impairment following TBI in hopes of finding a way to prevent the progression of damage or encourage the recovery of learning and memory. Although TBI induces cell death in the hippocampus, most of the neurons are spared following moderate or mild TBI. Less attention has been given to determine whether these spared neurons suffer injury. Injured animals also demonstrated a diffusive axonal injury [10,11,12] and profound loss of synapses in CA1 [13] with a significant deafferentation in electroactivity [14]. Less is known about potential indirect effects of TBI on dendrites. Light microscopic studies of MAP2 revealed a prominent loss of MAP2 immunofluorescence in apical dendrites of pyramidal neurons in the cortex [15,16]. This suggests dendritic damage in the spared neurons after TBI. In this study, we assessed the dendrites, dendritic spines, synapses, and function of spared granular neurons in the hippocampal dentate gyrus (HDG). The results showed extensively reduced dendritic branches in the spared granular neurons. The remaining dendrites exhibited swelling with beading, which is a hallmark of dendritic injury [17,18,19]. The density of dendritic spines and number of synapses were decreased, and electrophysiological activities were impaired. These data suggest, in addition to cell death and diffused axonal injury, extensive dendritic damage in the spared neurons could also disrupt neurocircuits and likely significantly contribute to functional impairment following TBI. Materials and Methods Animals Mice were housed with a 12/12 light/dark cycle and had free access to food and water ad libitum according to the principles outlined in Guidelines for Care and Use of Experimental Animals. They were used in experiments at an age of 810 weeks. All procedures were approved by Indiana University IACUC. The approval number is 3332. Genetically labeled the granular neurons with Enhanced Green Fluorescent Protein (EGFP) in the HDG We took advantage of a line of transgenic mice generated by Dr. Lowell and characterized by our laboratory [20,21]. These mice allowed us to easily visualize and quantify dendrites of granular neurons in the HDG in high resolution. In this line of transgenic mice, a recombinase Cre, driven by the pro-opiomalanocortin (POMC) promoter, was expressed in the granular neurons of the HDG [20]. Based on the Cre/loxP conditional recombinase system [22], we crossed the POMC-Cre mice with Z/EG reporter mice [23] [Tg(ACTB-Bgeo/GFP021Lbe Stock#003920] to obtain POMC-Cre;EGFP double transgenic mice [21]. In these mice, EGFP is expressed after Cre-mediated deletion of loxP-flanked stop codon [23]. The granular neurons with Cre expression will s (...truncated)