Interindividual variation in human cortical cell type abundance and expression.

Single-cell transcriptomic studies have identified a conserved set of neocortical cell types from small postmortem cohorts. We extended these efforts by assessing cell type variation across 75 adult individuals undergoing epilepsy and tumor surgeries. Nearly all nuclei map to one of 125 robust cell types identified in the middle temporal gyrus. However, we found interindividual variance in abundances and gene expression signatures, particularly in deep-layer glutamatergic neurons and microglia. A minority of donor variance is explainable by age, sex, ancestry, disease state, and cell state. Genomic variation was associated with expression of 150 to 250 genes for most cell types. This characterization of cellular variation provides a baseline for cell typing in health and disease.

Target cell-specific synaptic dynamics of excitatory to inhibitory neuron connections in supragranular layers of human neocortex.

Rodent studies have demonstrated that synaptic dynamics from excitatory to inhibitory neuron types are often dependent on the target cell type. However, these target cell-specific properties have not been well investigated in human cortex, where there are major technical challenges in reliably obtaining healthy tissue, conducting multiple patch-clamp recordings on inhibitory cell types, and identifying those cell types. Here, we take advantage of newly developed methods for human neurosurgical tissue analysis with multiple patch-clamp recordings, post-hoc fluorescent in situ hybridization (FISH), machine learning-based cell type classification and prospective GABAergic AAV-based labeling to investigate synaptic properties between pyramidal neurons and PVALB- vs. SST-positive interneurons. We find that there are robust molecular differences in synapse-associated genes between these neuron types, and that individual presynaptic pyramidal neurons evoke postsynaptic responses with heterogeneous synaptic dynamics in different postsynaptic cell types. Using molecular identification with FISH and classifiers based on transcriptomically identified PVALB neurons analyzed by Patch-seq, we find that PVALB neurons typically show depressing synaptic characteristics, whereas other interneuron types including SST-positive neurons show facilitating characteristics. Together, these data support the existence of target cell-specific synaptic properties in human cortex that are similar to rodent, thereby indicating evolutionary conservation of local circuit connectivity motifs from excitatory to inhibitory neurons and their synaptic dynamics.

Signature morpho-electric, transcriptomic, and dendritic properties of human layer 5 neocortical pyramidal neurons.

In the neocortex, subcerebral axonal projections originate largely from layer 5 (L5) extratelencephalic-projecting (ET) neurons. The unique morpho-electric properties of these neurons have been mainly described in rodents, where retrograde tracers or transgenic lines can label them. Similar labeling strategies are infeasible in the human neocortex, rendering the translational relevance of findings in rodents unclear. We leveraged the recent discovery of a transcriptomically defined L5 ET neuron type to study the properties of human L5 ET neurons in neocortical brain slices derived from neurosurgeries. Patch-seq recordings, where transcriptome, physiology, and morphology were assayed from the same cell, revealed many conserved morpho-electric properties of human and rodent L5 ET neurons. Divergent properties were often subtler than differences between L5 cell types within these two species. These data suggest a conserved function of L5 ET neurons in the neocortical hierarchy but also highlight phenotypic divergence possibly related to functional specialization of human neocortex.

Functional enhancer elements drive subclass-selective expression from mouse to primate neocortex.

Viral genetic tools that target specific brain cell types could transform basic neuroscience and targeted gene therapy. Here, we use comparative open chromatin analysis to identify thousands of human-neocortical-subclass-specific putative enhancers from across the genome to control gene expression in adeno-associated virus (AAV) vectors. The cellular specificity of reporter expression from enhancer-AAVs is established by molecular profiling after systemic AAV delivery in mouse. Over 30% of enhancer-AAVs produce specific expression in the targeted subclass, including both excitatory and inhibitory subclasses. We present a collection of Parvalbumin (PVALB) enhancer-AAVs that show highly enriched expression not only in cortical PVALB cells but also in some subcortical PVALB populations. Five vectors maintain PVALB-enriched expression in primate neocortex. These results demonstrate how genome-wide open chromatin data mining and cross-species AAV validation can be used to create the next generation of non-species-restricted viral genetic tools.

Stereotactic Brain Biopsy Hemorrhage Risk Factors and Implications for Postoperative Care at a Single Institution: An Argument For Postoperative Imaging.

OBJECTIVE: To determine preoperative factors contributing to postoperative hemorrhage after stereotactic brain biopsy (STB), clinical implications of postoperative hemorrhage, and the role of postoperative imaging in clinical management. METHODS: Retrospective review of STB (2005-2018) across 2 institutions including patients aged >18 years undergoing first STB. Patients with prior craniotomy, open biopsy, or prior STB were excluded. Preoperative variables included age, sex, neurosurgeon seniority, STB method. Postoperative variables included pathology, postoperative hemorrhage on computed tomography, immediate and 30-day postoperative seizure, infection, postoperative hospital stay duration, and 30-day return to operating room (OR). Analysis used the Fisher exact tests for categorical variables. RESULTS: Overall, 410 patients were included. Average age was 56.5 (±16.5) years; 60% (n = 248) were men. The majority of biopsies were performed by senior neurosurgeons (66%, n = 270); frontal lobe (42%, n = 182) and glioblastoma (45%, n = 186) were the most common location and pathology. Postoperative hemorrhage occurred in 28% (114) of patients with 20% <0.05 cm3 and 8% >0.05 cm3. Postoperative hemorrhage of any size was associated with increased rate of postoperative deficit within both 24 hours and 30 days, postoperative seizure, and length of hospital stay when controlling for pathology. Hemorrhages >0.05 cm3 had a 16% higher rate of return to the OR for evacuation, due to clinical deterioration as opposed to radiographic progression. CONCLUSIONS: Postbiopsy hemorrhage was associated with higher risk of immediate and delayed postoperative deficit and seizure. Postoperative computed tomography should be used to determine whether STB patients can be discharged same day or admitted for observation; clinical evaluation should determine return to OR for evacuation.

Evaluating angioarchitectural characteristics of glial and metastatic brain tumors with conventional magnetic resonance imaging.

Primary and metastatic brain tumors can overlap in traditional imaging features detected on preoperative conventional magnetic resonance imaging (MRI). The research objective was to determine whether morphological vascular characteristics present in routine preoperative imaging using traditional MRI sequences are predictive of primary versus metastatic brain tumors; secondarily to determine association of conventional and vascular-related imaging parameters with intraoperative blood loss, pathological invasion, and World Health Organization (WHO) tumor grade. A retrospective review analyzed 100 consecutive intracranial tumor surgeries, 50 WHO grade II-IV gliomas and 50 intracranial metastases. Two blinded expert readers independently evaluated preoperative MRIs, obtained via standard morphological imaging sequences, for adjacent or intra-tumoral arterial aneurysm, peritumoral venous ectasia, prominence, or engorgement ("aberrant peritumoral vessels"), and prominent intra-tumoral flow voids. Multivariate analysis was performed to develop models predictive of glioma and glioblastoma (GBM). Aberrant peritumoral vessels and prominent intra-tumoral flow voids were statistically significant predictors of glioma in univariate analyses (p = 0.048, p = 0.001, respectively) and when combined in multivariate analysis (OR = 5.23, p = 0.001), particularly for GBM (OR = 9.08, p < 0.001). Multivariate modeling identified prominent intra-tumoral flow voids and FLAIR invasion as the strongest combined predictors of gliomas and GBM. Aberrant peritumoral vessels and larger tumor volume predicted higher intraoperative blood loss in all analyses. No vascular-related parameters predicted pathological invasion on multivariate analysis. Aberrant peritumoral vessels and prominent intra-tumoral flow voids were predictive of gliomas, specifically GBM. These vascular characteristics, evaluated on routine clinical preoperative MRI imaging, may aid in distinguishinggliomafrom brainmetastases andmay predict intraoperative blood loss.

Synchronous identification of a dysembryoplastic neuroepithelial tumor (DNET) and an oligodendroglioma in a patient: A case report.

Synchronous gliomas of different histopathology are quite rare in non-syndromic, non-irradiated patients. Although "mixed" gliomas are not infrequent, and malignant gliomas often contain areas of disparate differentiation (e.g., glioblastoma with ependymal differentiation), it is unusual to find gliomas of different lineage presenting concurrently. We present a case of synchronous gliomas, one dysembryoplastic neuroepithelial tumor (DNET) and the other oligodendroglioma.

Patterns of Failure After Stereotactic Radiosurgery for Recurrent High-Grade Glioma: A Single Institution Experience of 10 Years.

BACKGROUND: Stereotactic radiosurgery (SRS) is a treatment modality that is frequently used as salvage therapy for small nodular recurrent high-grade gliomas (HGG). Due to the infiltrative nature of HGG, it is unclear if this highly focused technique provides a durable local control benefit. OBJECTIVE: To determine how demographic or clinical factors influence the pattern of failure following SRS for recurrent high-grade gliomas. METHODS: We retrospectively reviewed clinical, radiographic, and follow-up information for 47 consecutive patients receiving SRS for recurrent HGG at our institution between June 2006 and July 2016. All patients initially presented with an HGG (WHO grade III and IV). Following SRS for recurrence, all patients experienced treatment failure, and we evaluated patterns of local, regional, and distant failure in relation to the SRS 50% isodose line. RESULTS: Most patients with recurrent HGG developed "in-field" treatment failure following SRS (n = 40; 85%). Higher SRS doses were associated with longer time to failure (hazards ratio = 0.80 per 1 Gy increase; 95% confidence interval 0.67-0.96; P = .016). There was a statistically significant increase in distant versus in-field failure among older patients (P = .035). This effect was independent of bevacizumab use (odds ratio = 0.54, P = 1.0). CONCLUSION: Based on our experience, the majority of treatment failures after SRS for recurrent HGG were "in-field." Older patients, however, presented with more distant failures. Our results indicate that higher SRS doses delivered to a larger area as fractioned or unfractioned regimen may prolong time to failure, especially in the older population.

h-Channels Contribute to Divergent Intrinsic Membrane Properties of Supragranular Pyramidal Neurons in Human versus Mouse Cerebral Cortex.

Gene expression studies suggest that differential ion channel expression contributes to differences in rodent versus human neuronal physiology. We tested whether h-channels more prominently contribute to the physiological properties of human compared to mouse supragranular pyramidal neurons. Single-cell/nucleus RNA sequencing revealed ubiquitous HCN1-subunit expression in excitatory neurons in human, but not mouse, supragranular layers. Using patch-clamp recordings, we found stronger h-channel-related membrane properties in supragranular pyramidal neurons in human temporal cortex, compared to mouse supragranular pyramidal neurons in temporal association area. The magnitude of these differences depended upon cortical depth and was largest in pyramidal neurons in deep L3. Additionally, pharmacologically blocking h-channels produced a larger change in membrane properties in human compared to mouse neurons. Finally, using biophysical modeling, we provide evidence that h-channels promote the transfer of theta frequencies from dendrite-to-soma in human L3 pyramidal neurons. Thus, h-channels contribute to between-species differences in a fundamental neuronal property.

Sparse recurrent excitatory connectivity in the microcircuit of the adult mouse and human cortex.

Generating a comprehensive description of cortical networks requires a large-scale, systematic approach. To that end, we have begun a pipeline project using multipatch electrophysiology, supplemented with two-photon optogenetics, to characterize connectivity and synaptic signaling between classes of neurons in adult mouse primary visual cortex (V1) and human cortex. We focus on producing results detailed enough for the generation of computational models and enabling comparison with future studies. Here, we report our examination of intralaminar connectivity within each of several classes of excitatory neurons. We find that connections are sparse but present among all excitatory cell classes and layers we sampled, and that most mouse synapses exhibited short-term depression with similar dynamics. Synaptic signaling between a subset of layer 2/3 neurons, however, exhibited facilitation. These results contribute to a body of evidence describing recurrent excitatory connectivity as a conserved feature of cortical microcircuits.

Defining Glioblastoma Resectability Through the Wisdom of the Crowd: A Proof-of-Principle Study.

BACKGROUND: Extent of resection (EOR) correlates with glioblastoma outcomes. Resectability and EOR depend on anatomical, clinical, and surgeon factors. Resectability likely influences outcome in and of itself, but an accurate measurement of resectability remains elusive. An understanding of resectability and the factors that influence it may provide a means to control a confounder in clinical trials and provide reference for decision making. OBJECTIVE: To provide proof of concept of the use of the collective wisdom of experienced brain tumor surgeons in assessing glioblastoma resectability. METHODS: We surveyed 13 academic tumor neurosurgeons nationwide to assess the resectability of newly diagnosed glioblastoma. Participants reviewed 20 cases, including digital imaging and communications in medicine-formatted pre- and postoperative magnetic resonance images and clinical vignettes. The selected cases involved a variety of anatomical locations and a range of EOR. Participants were asked about surgical goal, eg, gross total resection, subtotal resection (STR), or biopsy, and rationale for their decision. We calculated a "resectability index" for each lesion by pooling responses from all 13 surgeons. RESULTS: Neurosurgeons' individual surgical goals varied significantly ( P = .015), but the resectability index calculated from the surgeons' pooled responses was strongly correlated with the percentage of contrast-enhancing residual tumor ( R = 0.817, P < .001). The collective STR goal predicted intraoperative decision of intentional STR documented on operative notes ( P < .01) and nonresectable residual ( P < .01), but not resectable residual. CONCLUSION: In this pilot study, we demonstrate the feasibility of measuring the resectability of glioblastoma through crowdsourcing. This tool could be used to quantify resectability, a potential confounder in neuro-oncology clinical trials.

Safety and efficacy of carmustine (BCNU) wafers for metastatic brain tumors.

BACKGROUND: Carmustine (BCNU) wafers (Gliadel) prolongs local disease control and progression-free survival (PFS) in patients with malignant gliomas. However, in metastatic brain tumors, there is a paucity of evidence in support of its safety and efficacy. The goal of this study was to assess the safety and efficacy of Gliadel wafers in patients with metastatic brain tumors. METHODS: We retrospectively reviewed the University of Washington experience with Gliadel wafers for metastatic brain tumors between 2000 and 2015. RESULTS: Gliadel wafers were used in 14 patients with metastatic brain tumors during the period reviewed. There were no postoperative seizures, strokes, or hemorrhages. There was one postoperative wound infection necessitating return to the operating room. The mean time to tumor progression (n = 7) and death (n = 5) after Gliadel wafer implantation was 2.5 and 2.9 years, respectively. Age was the only variable affecting PFS in patients receiving Gliadel wafers. Patients <53 years old (n = 7) had a PFS of 0.52 years, whereas patients >53 years old (n = 7) had a PFS of 4.29 years (P = 0.02). There was no significant difference in PFS in relation to presenting Karnofsky Performance Status (P = 0.26), number of brain metastasis (P = 0.82), tumor volume (P = 0.54), prior surgery (P = 0.57), or prior radiation (P = 0.41). There were no significant differences in the mean survival in relationship to any variable including age. CONCLUSIONS: BCNU wafers are a safe and a potentially efficacious adjunct to surgery and radiation for improving local disease control in metastatic brain tumors. Larger studies, however, are needed to examine overall efficacy and tumor specific efficacy.

Flow-Related Aneurysm within Glioblastoma: A Case Report and Review of Literature.

BACKGROUND: Glioblastoma multiforme (GBM) is the most common primary brain tumor in adults, with a median survival of 13 months despite surgery and chemoradiation. GBMs are often hypervascular tumors caused by abnormal oversecretion of growth factors such as vascular endothelial growth factor. These angiogenic factors are hypothesized to promote increased blood flow and possibly secondary changes to arterial walls, thus facilitating the formation of flow-related aneurysms. CASE DESCRIPTION: A 59-year-old woman presented with headaches, confusion, nausea and emesis. Computed tomography and magnetic resonance imaging revealed a hypervascular lesion, likely high-grade glioma, in the right frontal lobe, with a dilated vessel within the tumor. Cerebral angiography demonstrated a flow-related aneurysm on the right frontopolar artery supplying the tumor. The aneurysm was embolized with coils and the patient later underwent craniotomy for near total resection of the lesion without complications. Final pathology returned GBM with dilated vessels noted. CONCLUSIONS: Hypervascular lesions, such as GBMs, may be associated with flow-related aneurysms on feeding arteries, but aneurysms within the gross tumor are unusual. Although rare, this finding needs to be recognized on preoperative imaging before tumor resection to prevent potentially catastrophic intraoperative complications.

Recurrent Supplementary Motor Area Syndrome Following Repeat Brain Tumor Resection Involving Supplementary Motor Cortex.

BACKGROUND: Supplementary motor area (SMA) syndrome occurs after surgery involving the SMA and is characterized by contralateral hemiparesis with or without speech impairment (dependent on involvement of the dominant SMA), which is transient and characteristically resolves over the course of weeks to months. Recurrent SMA syndrome after repeat craniotomy has not been previously described. OBJECTIVE: To describe the presentation and clinical course of patients who developed recurrent SMA syndrome after redo resection of tumors involving the SMA. METHODS: We performed a retrospective review of 15 patients who underwent repeated resection of low-grade glioma from the superior and middle frontal gyrus. Of these patients, we identified 6 cases of recurrent SMA syndrome. RESULTS: Six patients had a documented SMA syndrome occurring after initial and subsequent resection of tumor in proximity to the SMA. Intraoperative localization of eloquent motor and language cortex was achieved in each patient by using a combination of somatosensory evoked potentials and electrocortical stimulation mapping. Location of tumor and extent of resection was examined with magnetic resonance imaging. CONCLUSION: This series demonstrates that recurrent SMA syndrome occurs in patients undergoing repeat resection of tumors involving the SMA. The presence of recurrent SMA syndrome provides support for reorganization of SMA function to adjacent ipsilateral cortex after resection. Patients with recurrent neoplasms of the SMA should be counseled on the possibility of recurrent SMA syndrome.

Outcomes in Reoperated Low-Grade Gliomas.

BACKGROUND: Low-grade gliomas (LGGs) comprise a diverse set of intrinsic brain tumors that correlate strongly with survival. Data on the effect of reoperation are sparse. OBJECTIVE: To evaluate the effect of reoperation on patients with LGG. METHODS: Fifty-two consecutive patients with reoperated LGGs treated at the University of Washington between 1986 and 2004 were identified and evaluated in a retrospective analysis. RESULTS: The average overall survival (OS) for this cohort was 12.95 ± 0.96 years. The overall 10-year survival rate was 57%. The absence of any residual tumor at either the first or second operation was associated with significantly increased OS. Negative prognostic variables for OS included the use of upfront radiation and pathology at recurrence. The average overall progression-free survival to the first recurrence (PFS1) was 6.23 ± 0.51 years. Positive prognostic factors for improved PFS1 included the use of upfront radiation therapy. Variables not associated with differences in PFS1 included the use of upfront chemotherapy, enhancement, pathology, extent of resection, the presence of residual tumor, and Karnofsky Performance Scale score <80. The average overall progression-free survival to the second recurrence was 2.73 ± 0.39 years. Pathology at recurrence was associated with significant differences in progression-free survival to the second recurrence, as was extent of resection at time of first recurrence, and Karnofsky Performance Scale score <80. CONCLUSION: This is among the largest studies to assess variables associated with outcome in patients with reoperated LGG. Reresection appears to provide significant benefit, and extent of resection remains the strongest predictor of OS.

Treatment of Gliomas: How did we get here?

Over the past 30 years, the treatment of gliomas has become more multi-modality with clinical trials demonstrating that adjuvant chemo-radiation following surgery improves survival of patients. Unfortunately, this advance in therapeutic intervention has had a modest impact on patient survival, with only a 3-6 month improvement in survival during this time period. In this review, we discuss the progress made in each key aspect of glioma treatment; chemotherapy, surgery and radiation therapy. We present key clinical trials that were used as basis for current management guidelines for patients with gliomas. Ultimately, it is clear that future treatments of patients with gliomas will entail specific chronologic combinations of these three modalities in personalized regimens designed for individual patient tumor sub-type.

O6-methylguanine-DNA methyltransferase activity is associated with response to alkylating agent therapy and with MGMT promoter methylation in glioblastoma and anaplastic glioma.

BACKGROUND: CpG methylation in the O6-methylguanine-DNA methyltransferase (MGMT) promoter is associated with better outcome following alkylating agent chemotherapy in glioblastoma (GBM) and anaplastic glioma (AG). To what extent improved response reflects low or absent MGMT activity in glioma tissue has not been unequivocally assessed. This information is central to developing anti-resistance therapies. METHODS: We examined the relationship of MGMT activity in 91 GBMs and 84 AGs with progression-free survival (PFS) following alkylator therapy and with promoter methylation status determined by methylation-specific PCR (MSP). RESULTS: Cox regression analysis revealed that GBMs with high activity had a significantly greater risk for progression in dichotomous (P ≤ 0.001) and continuous (P ≤ 0.003) models, an association observed for different alkylator regimens, including concurrent chemo-radiation with temozolomide. Analysis of MGMT promoter methylation status in 47 of the GBMs revealed that methylated tumors had significantly lower activity (P ≤ 0.005) and longer PFS (P ≤ 0.036) compared to unmethylated tumors, despite overlapping activities. PFS was also significantly greater in methylated vs. unmethylated GBMs with comparable activity (P ≤ 0.005), and among unmethylated tumors with less than median activity (P ≤ 0.026), suggesting that mechanisms in addition to MGMT promote alkylator resistance. Similar associations of MGMT activity with PFS and promoter methylation status were observed for AGs. CONCLUSIONS: Our results provide strong support for the hypotheses that MGMT activity promotes alkylator resistance and reflects promoter methylation status in malignant gliomas. GENERAL SIGNIFICANCE: MGMT activity is an attractive target for anti-resistance therapy regardless of methylation status.

Patient-specific metrics of invasiveness reveal significant prognostic benefit of resection in a predictable subset of gliomas.

OBJECT: Malignant gliomas are incurable, primary brain neoplasms noted for their potential to extensively invade brain parenchyma. Current methods of clinical imaging do not elucidate the full extent of brain invasion, making it difficult to predict which, if any, patients are likely to benefit from gross total resection. Our goal was to apply a mathematical modeling approach to estimate the overall tumor invasiveness on a patient-by-patient basis and determine whether gross total resection would improve survival in patients with relatively less invasive gliomas. METHODS: In 243 patients presenting with contrast-enhancing gliomas, estimates of the relative invasiveness of each patient's tumor, in terms of the ratio of net proliferation rate of the glioma cells to their net dispersal rate, were derived by applying a patient-specific mathematical model to routine pretreatment MR imaging. The effect of varying degrees of extent of resection on overall survival was assessed for cohorts of patients grouped by tumor invasiveness. RESULTS: We demonstrate that patients with more diffuse tumors showed no survival benefit (P = 0.532) from gross total resection over subtotal/biopsy, while those with nodular (less diffuse) tumors showed a significant benefit (P = 0.00142) with a striking median survival benefit of over eight months compared to sub-totally resected tumors in the same cohort (an 80% improvement in survival time for GTR only seen for nodular tumors). CONCLUSIONS: These results suggest that our patient-specific, model-based estimates of tumor invasiveness have clinical utility in surgical decision making. Quantification of relative invasiveness assessed from routinely obtained pre-operative imaging provides a practical predictor of the benefit of gross total resection.