Memory is thought to be encoded by sparsely distributed neuronal ensembles in memory‐related regions. However, it is unclear how memory‐eligible neurons react during learning to encode trace fear memory and how they retrieve a memory. We implemented a fiber‐optic confocal fluorescence endomicroscope to directly visualize calcium dynamics of hippocampal CA1 neurons in freely behaving mice subjected to trace fear conditioning. Here we report that the overall activity levels of CA1 neurons showed a right‐skewed lognormal distribution, with a small portion of highly active neurons (termed Primed Neurons) filling the long‐tail. Repetitive training induced Primed Neurons to shift from random activity to well‐tuned synchronization. The emergence of activity synchronization coincided with the appearance of mouse freezing behaviors. In recall, a partial synchronization among the same subset of Primed Neurons was induced from random dynamics, which also coincided with mouse freezing behaviors. Additionally, training‐induced synchronization facilitated robust calcium entry into Primed Neurons. In contrast, most CA1 neurons did not respond to tone and foot shock throughout the training and recall cycles. In conclusion, Primed Neurons are preferably recruited to encode trace fear memory and induction of activity synchronization among Primed Neurons out of random dynamics is critical for trace memory formation and retrieval.

Primary cilia are centriole-derived sensory organelles that are present in most mammalian cells, including astrocytes and neurons. Evidence is emerging that astrocyte and neuronal primary cilia demonstrate a dichotomy in the mature mouse brain. However, it is unknown how astrocytic and neuronal primary cilia change their morphology and ciliary proteins when exposed to reactive insults including epilepsy and traumatic brain injury. We used a double transgenic mouse strain (Arl13b-mCherry; Centrin2-GFP), in which we found spontaneous seizures, and a cortical injury model to examine the morphological changes of astrocytic and neuronal primary cilia under reactive conditions. Transgenic overexpression of Arl13b drastically increases the length of astrocytic and neuronal primary cilia in the hippocampus, as well as the cilia lengths of cultured astrocytes and neurons. Spontaneous seizures shorten Arl13b-positive astrocytic cilia and AC3-positive neuronal cilia in the hippocampus. In a cortical injury model, Arl13b is not detectable in primary cilia, but Arl13b protein relocates to the cell body and has robust expression in the proximity of injured tissues. In contrast, the number of AC3-positive cilia near injured tissues remains unchanged, but their lengths become shorter. These results on astrocytic cilia implicate Arl13b in regulating astrocyte proliferation and tissue regeneration, while the shortening of AC3-positive cilia suggests adaptive changes of neuronal primary cilia under excitotoxicity.

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Type III adenylyl cyclase (AC3, ADCY3 ) is predominantly enriched in neuronal primary cilia throughout the central nervous system (CNS). Genome-wide association studies in humans have associated ADCY3 with major depressive disorder and autistic spectrum disorder, both of which exhibit sexual dimorphism. To date, it is unclear how AC3 affects protein phosphorylation and signal networks in central neurons, and what causes the sexual dimorphism of autism. We employed a mass spectrometry (MS)-based phosphoproteomic approach to quantitatively profile differences in phosphorylation between inducible AC3 knockout (KO) and wild type (WT), male and female mice. In total, we identified 4,655 phosphopeptides from 1,756 proteins, among which 565 phosphopeptides from 322 proteins were repetitively detected in all samples. Over 46% phosphopeptides were identified in at least three out of eight biological replicas. Comparison of AC3 KO and WT datasets revealed that phosphopeptides with motifs matching proline-directed kinases' recognition sites had a lower abundance in the KO dataset than in WTs. We detected 14 phosphopeptides restricted to WT dataset (i.e., Rabl6, Spast and Ppp1r14a ) and 35 exclusively in KOs (i.e., Sptan1, Arhgap20, Arhgap44, and Pde1b ). Moreover, 95 phosphopeptides (out of 90 proteins) were identified only in female dataset and 26 only in males. Label-free MS spectrum quantification using Skyline further identified phosphopeptides that had higher abundance in each sample group. In total, 204 proteins had sex-biased phosphorylation and 167 of them had increased expression in females relative to males. Interestingly, among the 204 gender-biased phosphoproteins, 31% were found to be associated with autism, including Dlg1, Dlgap2, Syn1, Syngap1, Ctnna1, Ctnnd1, Ctnnd2, Pkp4, and Arvcf . Therefore, this study also provides the first phosphoproteomics evidence suggesting that gender-biased post-translational phosphorylation may be implicated in the sexual dimorphism of autism.

Fundamental differences in excitatory pyramidal cells across cortical areas and species highlight the implausibility of extrapolation from mouse to primate neurons and cortical networks. Far less is known about comparative regional and species‐specific features of neurochemically distinct cortical inhibitory interneurons. Here, we quantified the density, laminar distribution, and somatodendritic morphology of inhibitory interneurons expressing one or more of the calcium‐binding proteins (CaBPs) (calretinin [CR], calbindin [CB], and/or parvalbumin [PV]) in mouse (Mus musculus) versus rhesus monkey (Macaca mulatta) in two functionally and cytoarchitectonically distinct regions—the primary visual and frontal cortical areas—using immunofluorescent multilabeling, stereological counting, and 3D reconstructions. There were significantly higher densities of CB+ and PV+ neurons in visual compared to frontal areas in both species. The main species difference was the significantly greater density and proportion of CR+ interneurons and lower extent of CaBP coexpression in monkey compared to mouse cortices. Cluster analyses revealed that the somatodendritic morphology of layer 2–3 inhibitory interneurons is more dependent on CaBP expression than on species and area. Only modest effects of species were observed for CB+ and PV+ interneuron morphologies, while CR+ neurons showed no difference. By contrast to pyramidal cells that show highly distinctive area‐ and species‐specific features, here we found more subtle differences in the distribution and features of interneurons across areas and species. These data yield insight into how nuanced differences in the population organization and properties of neurons may underlie specializations in cortical regions to confer species‐ and area‐specific functional capacities.

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Much is known about differences in pyramidal cells across cortical areas and species, but studies of interneurons have focused on comparisons within single cortical areas and/or species. Here we quantified the distribution and somato-dendritic morphology of interneurons expressing one or more of the calcium binding proteins (CaBPs) calretinin (CR), calbindin (CB) and/or parvalbumin (PV) in mouse (Mus musculus) versus rhesus monkey (Macaca mulatta) in two functionally and cytoarchitectonically distinct regions- the primary visual and frontal cortical areas. The density, laminar distribution and morphology of interneurons were assessed in serial brain sections using immunofluorescent multi-labeling, stereological counting and 3D reconstructions. There were significantly higher densities of CB+ and PV+ neurons in visual compared to frontal areas in both species. The main species difference was the significantly greater density and proportion of CR+ interneurons and lower extent of CaBP co-expression in monkey compared to mouse cortices. Cluster analyses revealed that the somato-dendritic morphology of layer 2-3 inhibitory interneurons is more dependent on CaBP expression than on species and area. Only modest effects of species were observed for CB+ and PV+ interneuron morphologies, while CR+ neurons showed no difference. By contrast to pyramidal cells which show highly distinctive area- and species-specific features, here we found more subtle differences in the distribution and features of interneurons across areas and species. These data yield insight into how nuanced differences in the population organization and properties of neurons may underlie specializations in cortical regions to confer species and area-specific functional capacities.

Introduction First year dental students are trained to interpret clinical presentations of injuries on brainstem or spinal cord. Therefore, it is necessary for them to understand the three major sensory and motor pathways (dorsal column/medial lemniscal system, anterolateral system, corticospinal tracts) that go through brainstem and spinal cord, including their functions and cross‐sectional anatomy. Aim The present lesson was designed for first year students at Boston University School of Dental Medicine to learn the three sensory and motor pathways by using cross‐sectional figures of brainstem and spinal cord, and help students to apply the knowledge in clinical cases. Methods Three sessions with the same contents were held on two days. Two sessions were held on the first day containing 30 students in total, and the last session was held on the fifth day containing 20 students. The lesson started with a pre‐quiz, and then students worked in groups of 4 to 5 people to order cross‐sectional figures of brainstem and spinal cord. Then, the instructor gave a lecture to explain the answer, followed by a brief review on sensory and motor pathways. In the second group activity, students were asked to label the three pathways on the cross‐sectional figures. In the third group activity, students were given the location of a lesion on brainstem, and then were asked to discuss the potential clinical presentations. Instructor explained the answers after each activity. At the end, students took a post‐quiz. Both pre‐quiz and post‐quiz had a full score of 5, containing 2 questions on clinical cases, 1 question on pathway, 2 questions on cross‐section anatomy. All questions were multiple choice. The levels of difficulty in two quizzes are similar. Results The average scores from pre‐quiz and post‐quiz were compared by conducting a t‐test. Students showed significantly improvement in the post‐quiz as compared to pre‐quiz (pre: 1.50+/−1.07; post: 3.12+/−1.05; p<0.00001). Both pre and post scores of students attending on the fifth day were significantly higher as compared to the pre and post scores of students on the first day (pre: day1=1.20; day5=1.95; p=0.01; post: day1=2.76; day5=3.65; p=0.003). A t‐test was conducted to compare group differences of the improvements (first day vs. fifth day). Improvements of students were defined as the differences between post and pre‐quiz scores, and the improvements were not significantly different based on the day of the session (p=0.69). Conclusion Students showed significant improvement in understanding cross‐sectional anatomy of three major sensory and motor pathways after group study integrating with lectures. In addition, the effectiveness of this teaching methods was not affected by the baseline scores of students.