Cortical interneurons in schizophrenia - cause or effect?

GABAergic cortical interneurons are important components of cortical microcircuits. Their alterations are associated with a number of neurological and psychiatric disorders, and are thought to be especially important in the pathogenesis of schizophrenia. Here, we reviewed neuroanatomical and histological studies that analyzed different populations of cortical interneurons in postmortem human tissue from patients with schizophrenia and adequately matched controls. The data strongly suggests that in schizophrenia only selective interneuron populations are affected, with alterations of somatostatin and parvalbumin neurons being the most convincing. The most prominent changes are found in the prefrontal cortex, which is consistent with the impairment of higher cognitive functions characteristic of schizophrenia. In contrast, calretinin neurons, the most numerous interneuron population in primates, seem to be largely unaffected. The selective alterations of cortical interneurons are in line with the neurodevelopmental model and the multiple-hit hypothesis of schizophrenia. Nevertheless, a large number of data on interneurons in schizophrenia is still inconclusive, with different studies yielding opposing findings. Furthermore, no studies found a clear link between interneuron alterations and clinical outcomes. Future research should focus on the causes of changes in the cortical microcircuitry in order to identify potential therapeutic targets.

BMP3 Affects Cortical and Trabecular Long Bone Development in Mice.

Bone morphogenetic proteins (BMPs) have a major role in tissue development. BMP3 is synthesized in osteocytes and mature osteoblasts and has an antagonistic effect on other BMPs in bone tissue. The main aim of this study was to fully characterize cortical bone and trabecular bone of long bones in both male and female Bmp3-/- mice. To investigate the effect of Bmp3 from birth to maturity, we compared Bmp3-/- mice with wild-type littermates at the following stages of postnatal development: 1 day (P0), 2 weeks (P14), 8 weeks and 16 weeks of age. Bmp3 deletion was confirmed using X-gal staining in P0 animals. Cartilage and bone tissue were examined in P14 animals using Alcian Blue/Alizarin Red staining. Detailed long bone analysis was performed in 8-week-old and 16-week-old animals using micro-CT. The Bmp3 reporter signal was localized in bone tissue, hair follicles, and lungs. Bone mineralization at 2 weeks of age was increased in long bones of Bmp3-/- mice. Bmp3 deletion was shown to affect the skeleton until adulthood, where increased cortical and trabecular bone parameters were found in young and adult mice of both sexes, while delayed mineralization of the epiphyseal growth plate was found in adult Bmp3-/- mice.

The anatomy lesson of the SARS-CoV-2 pandemic: irreplaceable tradition (cadaver work) and new didactics of digital technology.

AIM: To compare the efficacy of different components of online and contact anatomy classes as perceived by medical students. METHODS: An anonymous course evaluation survey was conducted at the end of the academic year 2019/2020. The organization of classes due to the SARS-CoV-2 pandemic provided our students with a unique opportunity to compare online and contact classes. Students' responses were analyzed according to the type of obtained data (ratio, ordinal, and categorical). RESULTS: The response rate was 95.58%. Approximately 90% of students found anatomical dissection and practical work in general to be the most important aspect of teaching, which could not be replaced by online learning. During online classes, students missed the most the interaction with other students, followed by the interaction with student teaching assistants and teaching staff. Very few students found contact lectures useful, with most students reporting that they could be replaced with recorded video lectures. In contrast, recorded video lectures were perceived as extremely helpful for studying. Regular weekly quizzes were essential during online classes as they gave students adequate feedback and guided their learning process. Students greatly benefitted from additional course materials and interactive lessons, which were made easily available via e-learning platform. CONCLUSIONS: Anatomical dissection and interaction during contact classes remain the most important aspects of teaching anatomy. However, online teaching increases learning efficiency by allowing alternative learning strategies and by substituting certain components of contact classes, thus freeing up more time for practical work.

Axon morphology of rapid Golgi-stained pyramidal neurons in the prefrontal cortex in schizophrenia.

AIM: To analyze axon morphology on rapid Golgi impregnated pyramidal neurons in the dorsolateral prefrontal cortex in schizophrenia. METHODS: Postmortem brain tissue from five subjects diagnosed with schizophrenia and five control subjects without neuropathological findings was processed with the rapid Golgi method. Layer III and layer V pyramidal neurons from Brodmann area 9 were chosen in each brain for reconstruction with Neurolucida software. The axons and cell bodies of 136 neurons from subjects with schizophrenia and of 165 neurons from control subjects were traced. The data obtained by quantitative analysis were compared between the schizophrenia and control group with the t test. RESULTS: Axon impregnation length was consistently greater in the schizophrenia group. The axon main trunk length was significantly greater in the schizophrenia than in the control group (93.7 ± 36.6 µm vs 49.8 ± 9.9 µm, P = 0.032). Furthermore, in the schizophrenia group more axons had visibly stained collaterals (14.7% vs 5.5%). CONCLUSION: Axon rapid Golgi impregnation stops at the beginning of the myelin sheath. The increased axonal staining in the schizophrenia group could, therefore, be explained by reduced axon myelination. Such a decrease in axon myelination is in line with both the disconnection hypothesis and the two-hit model of schizophrenia as a neurodevelopmental disease. Our results support that the cortical circuitry disorganization in schizophrenia might be caused by functional alterations of two major classes of principal neurons due to altered oligodendrocyte development.

Somato-dendritic morphology and axon origin site specify von Economo neurons as a subclass of modified pyramidal neurons in the human anterior cingulate cortex.

Von Economo neurons (VENs) are modified pyramidal neurons characterized by an extremely elongated rod-shaped soma. They are abundant in layer V of the anterior cingulate cortex (ACC) and fronto-insular cortex (FI) of the human brain, and have long been described as a human-specific neuron type. Recently, VENs have been reported in the ACC of apes and the FI of macaque monkeys. The first description of the somato-dendritic morphology of VENs in the FI by Cajal in 1899 (Textura del Sistema Nervioso del Hombre y de los Vertebrados, Tomo II. Madrid: Nicolas Moya) strongly suggested that they were a unique neuron subtype with specific morphological features. It is surprising that a clarification of this extremely important observation has not yet been attempted, especially as possible misidentification of other oval or fusiform cells as VENs has become relevant in many recently published studies. Here, we analyzed sections of Brodmann area 24 (ACC) stained with rapid Golgi and Golgi-Cox in five adult human specimens, and confirmed Cajal's observations. In addition, we established a comprehensive morphological description of VENs. VENs have a distinct somato-dendritic morphology that allows their clear distinction from other modified pyramidal neurons. We established that VENs have a perpendicularly oriented, stick-shaped core part consisting of the cell body and two thick extensions - an apical and basal stem. The perpendicular length of the core part was 150-250 µm and the thickness was 10-21 µm. The core part was characterized by a lack of clear demarcation between the cell body and the two extensions. Numerous thin, spiny and horizontally oriented side dendrites arose from the cell body. The basal extension of the core part typically ended by giving numerous smaller dendrites with a brush-like branching pattern. The apical extension had a topology typical for apical dendrites of pyramidal neurons. The dendrites arising from the core part had a high dendritic spine density. The most distinct feature of VENs was the distant origin site of the axon, which arose from the ending of the basal extension, often having a common origin with a dendrite. Quantitative analysis found that VENs could be divided into two groups based on total dendritic length - small VENs with a peak total dendritic length of 1500-2500 µm and large VENs with a peak total dendritic length of 5000-6000 µm. Comparative morphological analysis of VENs and other oval and fusiform modified pyramidal neurons showed that on Nissl sections small VENs might be difficult to identify, and that oval and fusiform neurons could be misidentified as VENs. Our analysis of Golgi slides of Brodmann area 9 from a total of 32 adult human subjects revealed only one cell resembling VEN morphology. Thus, our data show that the numerous recent reports on the presence of VENs in non-primates in other layers and regions of the cortex need further confirmation by showing the dendritic and axonal morphology of these cells. In conclusion, our study provides a foundation for further comprehensive morphological and functional studies on VENs between different species.

Expression of guanylate cyclase C in human prefrontal cortex depends on sex and feeding status.

Introduction: Guanylate cyclase C (GC-C) has been detected in the rodent brain in neurons of the cerebral cortex, amygdala, midbrain, hypothalamus, and cerebellum. Methods: In this study we determined GC-C protein expression in Brodmann areas (BA) 9, BA10, BA11, and BA32 of the human prefrontal cortex involved in regulation of feeding behavior, as well as in the cerebellar cortex, arcuate nucleus of hypothalamus and substantia nigra in brain samples of human 21 male and 13 female brains by ELISA with postmortem delay < 24 h. Results: GC-C was found in all tested brain areas and it was expressed in neurons of the third cortical layer of BA9. The regulation of GC-C expression by feeding was found in male BA11 and BA10-M, where GC-C expression was in negative correlation to the volume of stomach content during autopsy. In female BA11 there was no correlation detected, while in BA10-M there was even positive correlation. This suggests sex differences in GC-C expression regulation in BA11 and BA10-M. The amount of GC-C was higher in female BA9 only when the death occurred shortly after a meal, while expression of GC-C was higher in BA10-O only when the stomach was empty. The expression of GC-C in female hypothalamus was lower when compared to male hypothalamus only when the stomach was full, suggesting possibly lower satiety effects of GC-C agonists in women. Discussion: These results point toward the possible role of GC-C in regulation of feeding behavior. Since, this is first study of GC-C regulation and its possible function in prefrontal cortex, to determine exact role of GC-C in different region of prefrontal cortex, especially in humans, need further studies.

Morphological and Molecular Characteristics of Perineuronal Nets in the Human Prefrontal Cortex-A Possible Link to Microcircuitry Specialization.

Perineuronal nets (PNNs) are a type of extracellular matrix (ECM) that play a significant role in synaptic activity and plasticity of interneurons in health and disease. We researched PNNs' regional and laminar representation and molecular composition using immunohistochemistry and transcriptome analysis of Brodmann areas (BA) 9, 14r, and 24 in 25 human postmortem brains aged 13-82 years. The numbers of VCAN- and NCAN-expressing PNNs, relative to the total number of neurons, were highest in cortical layers I and VI while WFA-binding (WFA+) PNNs were most abundant in layers III-V. The ECM glycosylation pattern was the most pronounced regional difference, shown by a significantly lower proportion of WFA+ PNNs in BA24 (3.27 ± 0.69%) compared to BA9 (6.32 ± 1.73%; P = 0.0449) and BA14 (5.64 ± 0.71%; P = 0.0278). The transcriptome of late developmental and mature stages revealed a relatively stable expression of PNN-related transcripts (log2-transformed expression values: 6.5-8.5 for VCAN and 8.0-9.5 for NCAN). Finally, we propose a classification of PNNs that envelop GABAergic neurons in the human cortex. The significant differences in PNNs' morphology, distribution, and molecular composition strongly suggest an involvement of PNNs in specifying distinct microcircuits in particular cortical regions and layers.

Dendritic Spines: Synaptogenesis and Synaptic Pruning for the Developmental Organization of Brain Circuits.

Synaptic overproduction and elimination is a regular developmental event in the mammalian brain. In the cerebral cortex, synaptic overproduction is almost exclusively correlated with glutamatergic synapses located on dendritic spines. Therefore, analysis of changes in spine density on different parts of the dendritic tree in identified classes of principal neurons could provide insight into developmental reorganization of specific microcircuits.The activity-dependent stabilization and selective elimination of the initially overproduced synapses is a major mechanism for generating diversity of neural connections beyond their genetic determination. The largest number of overproduced synapses was found in the monkey and human cerebral cortex. The highest (exceeding adult values by two- to threefold) and most protracted overproduction (up to third decade of life) was described for associative layer IIIC pyramidal neurons in the human dorsolateral prefrontal cortex.Therefore, the highest proportion and extraordinarily extended phase of synaptic spine overproduction is a hallmark of neural circuitry in human higher-order associative areas. This indicates that microcircuits processing the most complex human cognitive functions have the highest level of developmental plasticity. This finding is the backbone for understanding the effect of environmental impact on the development of the most complex, human-specific cognitive and emotional capacities, and on the late onset of human-specific neuropsychiatric disorders, such as autism and schizophrenia.

In the eye of the beholder - how course delivery affects anatomy education.

BACKGROUND: The COVID-19 pandemic caused major shifts in students' learning strategies as well as teaching environments that profoundly affected the delivery of anatomy courses in medical schools. The Department of Anatomy at the University of Zagreb School of Medicine had a unique experience where the anatomy course in 2019/2020 was first taught in-person before transferring to an online course delivery, while the inverse happened in 2020/2021. The core curriculum, course material and examination criteria were the same in both academic years. The aim of the study was to determine whether course delivery affected students' perceptions of the course and whether it impacted students' engagement and success. METHODS: The students' perceptions of the course were assessed via an anonymous course survey (student evaluation of teaching, SET). The questions in the SET assessed the usefulness of teaching modalities rather than students' satisfaction. Most questions were in the form of statements to which students responded with their level of agreement on a five-point Likert scale. Differences between responses in 2019/2020 and 2020/2021 were analyzed using the Mann-Whitney test. Effect size was estimated using Cliff's delta and association between responses was assessed using Spearman's r coefficient. RESULTS: Students' perceptions were significantly affected by changes in course delivery. Students' success and engagement were higher in 2019/2020 when in-person teaching preceded online teaching. Furthermore, students' views on course organization and the usefulness of continuous assessment were more positive in 2019/2020. Finally, students' perceptions of the usefulness of online materials and activities were more positive in 2019/2020. All comparisons between the two academic years were statistically significant (P ≤ 0.0001 for all comparisons, Mann-Whitney test). CONCLUSIONS: Students' perceptions of the anatomy course were dependent on the teaching environment they were exposed to at the beginning of the course. A transfer from in-person to online course delivery was more successful than vice-versa. This has important implications for structuring hybrid courses in medical education in the future.

Lasting mesothalamic dopamine imbalance and altered exploratory behavior in rats after a mild neonatal hypoxic event.

Introduction: Adversities during the perinatal period can decrease oxygen supply to the fetal brain, leading to various hypoxic brain injuries, which can compromise the regularity of brain development in different aspects. To examine the catecholaminergic contribution to the link between an early-life hypoxic insult and adolescent behavioral aberrations, we used a previously established rat model of perinatal hypoxia but altered the hypobaric to normobaric conditions. Methods: Exploratory and social behavior and learning abilities were tested in 70 rats of both sexes at adolescent age. Inherent vertical locomotion, sensory-motor functions and spatial learning abilities were explored in a subset of animals to clarify the background of altered exploratory behavior. Finally, the concentrations of dopamine (DA) and noradrenaline in midbrain and pons, and the relative expression of genes for DA receptors D1 and D2, and their down-stream targets (DA- and cAMP-regulated phosphoprotein, Mr 32 kDa, the regulatory subunit of protein kinase A, and inhibitor-5 of protein phosphatase 1) in the hippocampus and thalamus were investigated in 31 rats. Results: A lesser extent of alterations in exploratory and cognitive aspects of behavior in the present study suggests that normobaric conditions mitigate the hypoxic injury compared to the one obtained under hypobaric conditions. Increased exploratory rearing was the most prominent consequence, with impaired spatial learning in the background. In affected rats, increased midbrain/pons DA content, as well as mRNA levels for DA receptors and their down-stream elements in the thalamus, but not the hippocampus, were found. Conclusion: We can conclude that a mild hypoxic event induced long-lasting disbalances in mesothalamic DA signaling, contributing to the observed behavioral alterations. The thalamus was thereby indicated as another structure, besides the well-established striatum, involved in mediating hypoxic effects on behavior through DA signaling.

The Distinct Characteristics of Somatostatin Neurons in the Human Brain.

Somatostatin cells are frequently described as a major population of GABAergic neurons in the cerebral cortex. In this study, we performed a comprehensive analysis of their molecular expression, morphological features, and laminar distribution. We provided a detailed description of somatostatin neurons in the human prefrontal cortex, including their proportion in the total neuron population, laminar distribution, neurotransmitter phenotype, as well as their molecular and morphological characteristics using immunofluorescence and RNAscope in situ hybridization. We found that somatostatin neurons comprise around 7% of neocortical neurons in the human Brodmann areas 9 and 14r, without significant difference between the two regions. Somatostatin cells were NeuN positive and synthesized vesicular GABA transporter and glutamate decarboxylase 1 and 2, confirming their neuronal nature and GABAergic phenotype. Somatostatin cells in the upper cortical layers were small, had a high expression of somatostatin mRNA, a relatively low expression of somatostatin peptide, and co-expressed calbindin. In the lower cortical layers, somatostatin cells were larger with complex somato-dendritic morphology, typically showed a lower expression of somatostatin mRNA and a high expression of somatostatin peptide, and co-expressed neuronal nitric oxide synthase (nNOS) and neuropeptide Y (NPY), but not calbindin. Somatostatin neurons in the white matter co-expressed MAP2. Based on their somato-dendritic morphology, cortical somatostatin neurons could be classified into at least five subtypes. The somatostatin neurons of the human prefrontal cortex show remarkable morphological and molecular complexity, which implies that they have equally complex and distinct functions in the human brain.

Von Economo Neurons - Primate-Specific or Commonplace in the Mammalian Brain?

The pioneering work by von Economo in 1925 on the cytoarchitectonics of the cerebral cortex revealed a specialized and unique cell type in the adult human fronto-insular (FI) and anterior cingulate cortex (ACC). In modern studies, these neurons are termed von Economo neurons (VENs). In his work, von Economo described them as stick, rod or corkscrew cells because of their extremely elongated and relatively thin cell body clearly distinguishable from common oval or spindle-shaped infragranular principal neurons. Before von Economo, in 1899 Cajal depicted the unique somato-dendritic morphology of such cells with extremely elongated soma in the FI. However, although VENs are increasingly investigated, Cajal's observation is still mainly being neglected. On Golgi staining in humans, VENs have a thick and long basal trunk with horizontally oriented terminal branching (basilar skirt) from where the axon arises. They are clearly distinguishable from a spectrum of modified pyramidal neurons found in infragranular layers, including oval or spindle-shaped principal neurons. Spindle-shaped cells with highly elongated cell body were also observed in the ACC of great apes, but despite similarities in soma shape, their dendritic and axonal morphology has still not been described in sufficient detail. Studies identifying VENs in non-human species are predominantly done on Nissl or anti-NeuN staining. In most of these studies, the dendritic and axonal morphology of the analyzed cells was not demonstrated and many of the cells found on Nissl or anti-NeuN staining had a cell body shape characteristic for common oval or spindle-shaped cells. Here we present an extensive literature overview on VENs, which demonstrates that human VENs are specialized elongated principal cells with unique somato-dendritic morphology found abundantly in the FI and ACC of the human brain. More research is needed to properly evaluate the presence of such specialized cells in other primates and non-primate species.