DiI‐mediated analysis of presynaptic and postsynaptic structures in human postmortem brain tissue

Abstract

Most cognitive and psychiatric disorders are thought to be disorders of the synapse, yet the precise synapse defects remain unknown. Because synapses are highly specialized anatomical structures, defects in synapse formation and function can often be observed as changes in microscale neuroanatomy. Unfortunately, few methods are available for accurate analysis of synaptic structures in human postmortem tissues.Here, we present a methodological pipeline for assessing presynaptic and postsynaptic structures in human postmortem tissue that is accurate, rapid, and relatively inexpensive. Our method uses small tissue blocks from postmortem human brains, immersion fixation, lipophilic dye (DiI) labeling, and confocal microscopy. As proof of principle, we analyzed presynaptic and postsynaptic structures from hippocampi of 13 individuals aged 4 months to 71 years. Our results indicate that postsynaptic CA1 dendritic spine shape and density do not change in adults, while presynaptic DG mossy fiber boutons undergo significant structural rearrangements with normal aging. This suggests that mossy fiber synapses, which play a major role in learning and memory, may remain dynamic throughout life. Importantly, we find that human CA1 spine densities observed using this method on tissue that is up to 28 h postmortem is comparable to prior studies using tissue with much shorter postmortem intervals. Thus, the ease of our protocol and suitability on tissue with longer postmortem intervals should facilitate higher-powered studies of human presynaptic and postsynaptic structures in healthy and diseased states. K E Y W O R D S dendritic spine, DiI, hippocampus, mossy fiber bouton, postmortem

Severe childhood and adulthood stress associates with neocortical layer-specific reductions of mature spines in psychiatric disorders

“…In accordance, the quantification technique only allowed for the identification of spines in the same x-y plane as the dendrite. Although the densities in the present study are comparable to those using similar methods [32], these combined factors likely underlie why the density of spines in the present study is lower than that identified using more sensitive methods in both rodents [81] and humans [82]. In the interpretation of the presented data, we acknowledge that these impacts may actually under-represent spine losses in the human OFC and the effect size may indeed be larger using techniques which facilitate greater resolution of spine detection.…”

Small heat shock proteins determine synapse number and neuronal activity during development

“…Synapse regulation is a central event during nervous system development and adult life. Disruptions in the establishment of synapses is associated with morphological, cognitive and psychiatric disorders, but the precise mechanisms underlying these disorders remain unknown [42]. Changes in synapse structure and function are related to paralysis and muscular atrophy in amyotrophic lateral sclerosis (ALS) [43,44], impairment of the neuromuscular junction function and therefore, motor decline [45] or social and cognitive behaviors related to autism [46].…”

Cytoskeletal regulation of synaptogenesis in a model of human fetal brain development

“…In human neuronal disorders, the vast majority of research focuses on mature dendritic spines found in postmortem patient brain samples from adolescents and adults (Bao & Swaab, 2018; Hutsler & Zhang, 2010; Tang et al., 2014). In spite of recent techniques to minimize fixation artifacts (Das et al., 2019), postmortem brain tissue is limited to a static snapshot of synapse development. Furthermore, while rodent models capture synapse dynamics (Marrs, Green, & Dailey, 2001), most research focuses on peri‐ and postnatal periods of synaptic development.…”