The neuropeptide landscape of human prefrontal cortex.

Human prefrontal cortex (hPFC) is a complex brain region involved in cognitive and emotional processes and several psychiatric disorders. Here, we present an overview of the distribution of the peptidergic systems in 17 subregions of hPFC and three reference cortices obtained by microdissection and based on RNA sequencing and RNAscope methods integrated with published single-cell transcriptomics data. We detected expression of 60 neuropeptides and 60 neuropeptide receptors in at least one of the hPFC subregions. The results reveal that the peptidergic landscape in PFC consists of closely located and functionally different subregions with unique peptide/transmitter-related profiles. Neuropeptide-rich PFC subregions were identified, encompassing regions from anterior cingulate cortex/orbitofrontal gyrus. Furthermore, marked differences in gene expression exist between different PFC regions (>5-fold; cocaine and amphetamine-regulated transcript peptide) as well as between PFC regions and reference regions, for example, for somatostatin and several receptors. We suggest that the present approach allows definition of, still hypothetical, microcircuits exemplified by glutamatergic neurons expressing a peptide cotransmitter either as an agonist (hypocretin/orexin) or antagonist (galanin). Specific neuropeptide receptors have been identified as possible targets for neuronal afferents and, interestingly, peripheral blood-borne peptide hormones (leptin, adiponectin, gastric inhibitory peptide, glucagon-like peptides, and peptide YY). Together with other recent publications, our results support the view that neuropeptide systems may play an important role in hPFC and underpin the concept that neuropeptide signaling helps stabilize circuit connectivity and fine-tune/modulate PFC functions executed during health and disease.

The emerging tale of microglia in psychiatric disorders.

As the professional phagocytes of the brain, microglia orchestrate the immunological response and play an increasingly important role in maintaining homeostatic brain functions. Microglia are activated by pathological events or slight alterations in brain homeostasis. This activation is dependent on the context and type of stressor or pathology. Through secretion of cytokines, chemokines and growth factors, microglia can strongly influence the response to a stressor and can, therefore, determine the pathological outcome. Psychopathologies have repeatedly been associated with long-lasting priming and sensitization of cerebral microglia. This review focuses on the diversity of microglial phenotype and function in health and psychiatric disease. We first discuss the diverse homeostatic functions performed by microglia and then elaborate on context-specific spatial and temporal microglial heterogeneity. Subsequently, we summarize microglia involvement in psychopathologies, namely major depressive disorder, schizophrenia and bipolar disorder, with a particular focus on post-mortem studies. Finally, we postulate microglia as a promising novel therapeutic target in psychiatry through antidepressant and antipsychotic treatment.

Lesch-Nyhan disease causes impaired energy metabolism and reduced developmental potential in midbrain dopaminergic cells.

Mutations in HPRT1, a gene encoding a rate-limiting enzyme for purine salvage, cause Lesch-Nyhan disease which is characterized by self-injury and motor impairments. We leveraged stem cell and genetic engineering technologies to model the disease in isogenic and patient-derived forebrain and midbrain cell types. Dopaminergic progenitor cells deficient in HPRT showed decreased intensity of all developmental cell-fate markers measured. Metabolic analyses revealed significant loss of all purine derivatives, except hypoxanthine, and impaired glycolysis and oxidative phosphorylation. real-time glucose tracing demonstrated increased shunting to the pentose phosphate pathway for de novo purine synthesis at the expense of ATP production. Purine depletion in dopaminergic progenitor cells resulted in loss of RHEB, impairing mTORC1 activation. These data demonstrate dopaminergic-specific effects of purine salvage deficiency and unexpectedly reveal that dopaminergic progenitor cells are programmed to a high-energy state prior to higher energy demands of terminally differentiated cells.

Implication of cerebral astrocytes in major depression: A review of fine neuroanatomical evidence in humans.

Postmortem investigations have implicated astrocytes in many neurological and psychiatric conditions. Multiple brain regions from individuals with major depressive disorder (MDD) have lower expression levels of astrocyte markers and lower densities of astrocytes labeled for these markers, suggesting a loss of astrocytes in this mental illness. This paper reviews the general properties of human astrocytes, the methods to study them, and the postmortem evidence for astrocyte pathology in MDD. When comparing astrocyte density and morphometry studies, astrocytes are more abundant and smaller in human subcortical than cortical brain regions, and immunohistochemical labeling for the astrocyte markers glial fibrillary acidic protein (GFAP) and vimentin (VIM) reveals fewer than 15% of all astrocytes that are present in cortical and subcortical regions, as revealed using other staining techniques. By combining astrocyte densities and morphometry, a model was made to illustrate that domain organization is mostly limited to GFAP-IR astrocytes. Using these markers and others, alterations of astrocyte densities appear more widespread than those for astrocyte morphologies throughout the brain of individuals having died with MDD. This review suggests how reduced astrocyte densities may relate to the association of depressive episodes in MDD with elevated S100 beta (S100B) cerebrospinal fluid serum levels. Finally, a potassium imbalance theory is proposed that integrates the reduced astrocyte densities generated from postmortem studies with a hypothesis for the antidepressant effects of ketamine generated from rodent studies.

Widespread Decrease of Cerebral Vimentin-Immunoreactive Astrocytes in Depressed Suicides.

Post-mortem investigations have implicated cerebral astrocytes immunoreactive (-IR) for glial fibrillary acidic protein (GFAP) in the etiopathology of depression and suicide. However, it remains unclear whether astrocytic subpopulations IR for other astrocytic markers are similarly affected. Astrocytes IR to vimentin (VIM) display different regional densities than GFAP-IR astrocytes in the healthy brain, and so may be differently altered in depression and suicide. To investigate this, we compared the densities of GFAP-IR astrocytes and VIM-IR astrocytes in post-mortem brain samples from depressed suicides and matched non-psychiatric controls in three brain regions (dorsomedial prefrontal cortex, dorsal caudate nucleus and mediodorsal thalamus). A quantitative comparison of the fine morphology of VIM-IR astrocytes was also performed in the same regions and subjects. Finally, given the close association between astrocytes and blood vessels, we also assessed densities of CD31-IR blood vessels. Like for GFAP-IR astrocytes, VIM-IR astrocyte densities were found to be globally reduced in depressed suicides relative to controls. By contrast, CD31-IR blood vessel density and VIM-IR astrocyte morphometric features in these regions were similar between groups, except in prefrontal white matter, in which vascularization was increased and astrocytes displayed fewer primary processes. By revealing a widespread reduction of cerebral VIM-IR astrocytes in cases vs. controls, these findings further implicate astrocytic dysfunctions in depression and suicide.

Characterization of Vimentin-Immunoreactive Astrocytes in the Human Brain.

Astrocytes are commonly identified by their expression of the intermediate filament protein glial fibrillary acidic protein (GFAP). GFAP-immunoreactive (GFAP-IR) astrocytes exhibit regional heterogeneity in density and morphology in the mouse brain as well as morphological diversity in the human cortex. However, regional variations in astrocyte distribution and morphology remain to be assessed comprehensively. This was the overarching objective of this postmortem study, which mainly exploited the immunolabeling of vimentin (VIM), an intermediate filament protein expressed by astrocytes and endothelial cells which presents the advantage of more extensively labeling cell structures. We compared the densities of vimentin-immunoreactive (VIM-IR) and GFAP-IR astrocytes in various brain regions (prefrontal and primary visual cortex, caudate nucleus, mediodorsal thalamus) from male individuals having died suddenly in the absence of neurological or psychiatric conditions. The morphometric properties of VIM-IR in these brain regions were also assessed. We found that VIM-IR astrocytes generally express the canonical astrocytic markers Aldh1L1 and GFAP but that VIM-IR astrocytes are less abundant than GFAP-IR astrocytes in all human brain regions, particularly in the thalamus, where VIM-IR cells were nearly absent. About 20% of all VIM-IR astrocytes presented a twin cell morphology, a phenomenon rarely observed for GFAP-IR astrocytes. Furthermore VIM-IR astrocytes in the striatum were often seen to extend numerous parallel processes which seemed to give rise to large VIM-IR fiber bundles projecting over long distances. Moreover, morphometric analyses revealed that VIM-IR astrocytes were more complex than their mouse counterparts in functionally homologous brain regions, as has been previously reported for GFAP-IR astrocytes. Lastly, the density of GFAP-IR astrocytes in gray and white matter were inversely correlated with vascular density, but for VIM-IR astrocytes this was only the case in gray matter, suggesting that gliovascular interactions may especially influence the regional heterogeneity of GFAP-IR astrocytes. Taken together, these findings reveal special features displayed uniquely by human VIM-IR astrocytes and illustrate that astrocytes display important region- and marker-specific differences in the healthy human brain.

Stimulation of L-type calcium channels increases tyrosine hydroxylase and dopamine in ventral midbrain cells induced from somatic cells.

Making high-quality dopamine (DA)-producing cells for basic biological or small molecule screening studies is critical for the development of novel therapeutics for disorders of the ventral midbrain. Currently, many ventral midbrain assays have low signal-to-noise ratio due to low levels of cellular DA and the rate-limiting enzyme of DA synthesis, tyrosine hydroxylase (TH), hampering discovery efforts. Using intensively characterized ventral midbrain cells derived from human skin, which demonstrate calcium pacemaking activity and classical electrophysiological properties, we show that an L-type calcium agonist can significantly increase TH protein levels and DA content and release. Live calcium imaging suggests that it is the immediate influx of calcium occurring simultaneously in all cells that drives this effect. Genome-wide expression profiling suggests that L-type calcium channel stimulation has a significant effect on specific genes related to DA synthesis and affects expression of L-type calcium receptor subunits from the CACNA1 and CACNA2D families. Together, our findings provide an advance in the ability to increase DA and TH levels to improve the accuracy of disease modeling and small molecule screening for disorders of the ventral midbrain, including Parkinson's disease.

Evidence of decreased gap junction coupling between astrocytes and oligodendrocytes in the anterior cingulate cortex of depressed suicides.

Glial dysfunction is a major pathophysiological feature of mood disorders. While altered astrocyte (AS) and oligodendrocyte-lineage (OL) functions have been associated with depression, the crosstalk between these glial cell types has never been assessed in that context. AS are potent regulators of myelination, in part through gap junction (GJ) channels formed by the heterotypic coupling of AS-specific (Cx30 and Cx43) and OL-specific (Cx32 and Cx47) connexins. This study therefore aimed at addressing the integrity of AS/OL coupling in the anterior cingulate cortex (ACC) of depressed suicides. Using immunofluorescence and confocal imaging, we characterized the distribution of Cx30 and mapped its expression onto OL somas, myelinated axons, and brain vasculature in postmortem brain samples from depressed suicides (N = 48) and matched controls (N = 23). Differential gene expression of key components of the GJ nexus was also screened through RNA-sequencing previously generated by our group, and validated by quantitative real-time PCR. We show that Cx30 expression localized onto OL cells and myelinated fibers is decreased in deep cortical layers of the ACC in male-depressed suicides. This effect was associated with decreased expression of OL-specific connexins, as well as the downregulation of major connexin-interacting proteins essential for the scaffolding, trafficking, and function of GJs. These results provide a first evidence of impaired AS/OL GJ-mediated communication in the ACC of individuals with mood disorders. These changes in glial coupling are likely to have significant impact on brain function, and may contribute to the altered OL function previously reported in this brain region.

The 10,000 PhDs project at the University of Toronto: Using employment outcome data to inform graduate education.

The purpose of the 10,000 PhDs Project was to determine the current (2016) employment status of the 10,886 individuals who graduated from the University of Toronto with a PhD in all disciplines from 2000-2015. Using internet searches, we found that about half (51%) of the PhD graduates are employed in the post-secondary education sector, 26% as tenure-track professors, with an additional 3% as adjunct professors and 2% as full-time teaching-stream professors. Over the time-period 2000-2015 there has been a near doubling in PhD graduates with the biggest increase in graduation numbers for the Physical (2.6-fold) and Life Sciences (2.2-fold). Increasingly, these graduates are finding employment in the private and public sectors providing the highly qualified personnel needed to drive an innovation economy.

Disruption of GRIN2B Impairs Differentiation in Human Neurons.

Heterozygous loss-of-function mutations in GRIN2B, a subunit of the NMDA receptor, cause intellectual disability and language impairment. We developed clonal models of GRIN2B deletion and loss-of-function mutations in a region coding for the glutamate binding domain in human cells and generated neurons from a patient harboring a missense mutation in the same domain. Transcriptome analysis revealed extensive increases in genes associated with cell proliferation and decreases in genes associated with neuron differentiation, a result supported by extensive protein analyses. Using electrophysiology and calcium imaging, we demonstrate that NMDA receptors are present on neural progenitor cells and that human mutations in GRIN2B can impair calcium influx and membrane depolarization even in a presumed undifferentiated cell state, highlighting an important role for non-synaptic NMDA receptors. It may be this function, in part, which underlies the neurological disease observed in patients with GRIN2B mutations.