Although genomic anomalies in glioblastoma (GBM) have been well studied for over a decade, its 5-year survival rate remains lower than 5%. We seek to expand the molecular landscape of high-grade glioma, composed of IDH-wildtype GBM and IDH-mutant grade 4 astrocytoma, by integrating proteomic, metabolomic, lipidomic, and post-translational modifications (PTMs) with genomic and transcriptomic measurements to uncover multi-scale regulatory interactions governing tumor development and evolution. Applying 14 proteogenomic and metabolomic platforms to 228 tumors (212 GBM and 16 grade 4 IDH-mutant astrocytoma), including 28 at recurrence, plus 18 normal brain samples and 14 brain metastases as comparators, reveals heterogeneous upstream alterations converging on common downstream events at the proteomic and metabolomic levels and changes in protein-protein interactions and glycosylation site occupancy at recurrence. Recurrent genetic alterations and phosphorylation events on PTPN11 map to important regulatory domains in three dimensions, suggesting a central role for PTPN11 signaling across high-grade gliomas.

Glioblastoma (GBM) is the most aggressive nervous system cancer. Understanding its molecular pathogenesis is crucial to improving diagnosis and treatment. Integrated analysis of genomic, proteomic, post-translational modification and metabolomic data on 99 treatment-naive GBMs provides insights to GBM biology. We identify key phosphorylation events (e.g., phosphorylated PTPN11 and PLCG1) as potential switches mediating oncogenic pathway activation, as well as potential targets for EGFR-, TP53-, and RB1-altered tumors. Immune subtypes with distinct immune cell types are discovered using bulk omics methodologies, validated by snRNA-seq, and correlated with specific expression and histone acetylation patterns. Histone H2B acetylation in classical-like and immune-low GBM is driven largely by BRDs, CREBBP, and EP300. Integrated metabolomic and proteomic data identify specific lipid distributions across subtypes and distinct global metabolic changes in IDH-mutated tumors. This work highlights biological relationships that could contribute to stratification of GBM patients for more effective treatment.

Vascular dementia (VaD) is cognitive decline linked to reduced cerebral blood perfusion, yet there are few therapeutic options to protect cognitive function following cerebrovascular accidents. The purpose of this study was to profile gene expression changes unique to VaD to identify and characterize disease relevant changes that could offer clues for future therapeutic direction. Microarray-based profiling and validation studies of postmortem frontal cortex samples from VaD, Alzheimer disease, and age-matched control subjects revealed that the oxytocin receptor (OXTR) was strongly and differentially upregulated in VaD. Further characterization in fixed tissue from the same cases showed that OXTR upregulation occurs de novo around and within microinfarcts in peri-infarct reactive astrocytes as well as within vascular profiles, likely on microvascular endothelial cells. These results indicate that increased OXTR expression in peri-infarct regions may be a specific response to microvascular insults. Given the established OXTR signaling cascades that elicit antioxidant, anti-inflammatory, and pro-angiogenic responses, the present findings suggest that de novo OXTR expression in the peri-infarct space is a tissue-protective response by astroglial and vascular cells in the wake of ischemic damage that could be exploited as a therapeutic option for the preservation of cognition following cerebrovascular insults.

Tuberous sclerosis complex (TSC) is a rare genetic disease causing multisystem growth of benign tumours and other hamartomatous lesions, which leads to diverse and debilitating clinical symptoms. Patients are born with TSC1 or TSC2 mutations, and somatic inactivation of wild-type alleles drives MTOR activation; however, second hits to TSC1/TSC2 are not always observed. Here, we present the genomic landscape of TSC hamartomas. We determine that TSC lesions contain a low somatic mutational burden relative to carcinomas, a subset feature large-scale chromosomal aberrations, and highly conserved molecular signatures for each type exist. Analysis of the molecular signatures coupled with computational approaches reveals unique aspects of cellular heterogeneity and cell origin. Using immune data sets, we identify significant neuroinflammation in TSC-associated brain tumours. Taken together, this molecular catalogue of TSC serves as a resource into the origin of these hamartomas and provides a framework that unifies genomic and transcriptomic dimensions for complex tumours.

Next-generation sequencing (NGS) capabilities can affect therapeutic decisions in patients with complex, advanced, or refractory cancer. We report the feasibility of a tumor sequencing advisory board at a regional cancer center. Specimens were analyzed for approximately 2800 mutations in 50 genes. Outcomes of interest included tumor sequencing advisory board function and processes, timely discussion of results, and proportion of reports having potentially actionable mutations. NGS results were successfully generated for 15 patients, with median time from tissue processing to reporting of 11.6 days (range, 5 to 21 days), and presented at a biweekly multidisciplinary tumor sequencing advisory board. Attendance averaged 19 participants (range, 12 to 24) at 20 days after patient enrollment (range, 10 to 30 days). Twenty-seven (range, 1 to 4 per patient) potentially actionable mutations were detected in 11 of 15 patients: TP53 (n = 6), KRAS (n = 4), MET (n = 3), APC (n = 3), CDKN2A (n = 2), PTEN (n = 2), PIK3CA, FLT3, NRAS, VHL, BRAF, SMAD4, and ATM. The Hotspot Panel is now offered as a clinically available test at our institution. NGS results can be obtained by in-house high-throughput sequencing and reviewed in a multidisciplinary tumor sequencing advisory board in a clinically relevant manner. The essential components of a center for personalized cancer care can support clinical decisions outside the university.

The effects of electrical stimulation of the lateral vestibular nucleus (LVN) and medullary reticular formation (RF) on electromyographic activity in axial muscles medial longissimus (ML) and lateral longissimus (LL) in the rat were studied. Long trains (150–500 ms) at 200–330 Hz and 20–100 μA were sufficient to activate ML and LL at latencies of 20–100 ms from the beginning of the train. Results of stimulation at 200–330 Hz to RF or LVN showed that muscle units were activated at a fixed latency from any effective pulse in the stimulus train. Using high frequency (1 kHz) trains of 3–6 pulses to LVN, EMG activity was detected at minimum latencies of 3.5–6 ms. When conduction times from the medulla to the spinal cord, and the spinal cord to the muscle are subtracted, this latency range is consistent with monosynaptic activation. In many cases, muscle units were recruited in order of size, with both RF and LVN stimulation. Combined stimulation of LVN and RF sites in n. gigantocellularis led to EMG activity in ML and LL at currents which were insufficient to evoke activity when presented singly. When stimulation of one site (300–400 ms train) was just sufficient to evoke a response, a shorter, overlapping train (100–150 ms) to the other site led to a higher rate of muscle activity that continued through the end of the long train, even after the short train had ended. In all cases, the effect of RF facilitating LVN was similar to the effect of LVN facilitating RF. The evidence for convergence between these two systems in the medulla and the spinal cord is discussed.

The neuropeptide galanin coexists in 80-90% of the norepinephrine-containing neurons in the locus coeruleus. In situ hybridization histochemistry was used to examine the effects of reserpine treatment or swim stress on tyrosine hydroxylase and galanin mRNA concentrations in locus coeruleus neurons. Reserpine administration significantly increased tyrosine hydroxylase and galanin mRNA levels in the locus coeruleus. The reserpine-induced increase in tyrosine hydroxylase mRNA was significantly correlated with the reserpine-induced increase in galanin mRNA. Three consecutive days of swim stress did not significantly alter either tyrosine hydroxylase or galanin mRNA concentrations in the locus coeruleus. These data suggest that both tyrosine hydroxylase and galanin gene expression in locus coeruleus neurons may be regulated by a reserpine-sensitive mechanism.

Medullary reticular stimulation can activate deep back muscle EMG in urethane-anesthetized female rats. Midbrain central gray stimulation can facilitate brainstem reticular control over deep back muscles. Since these deep back muscles lateral longissimus (LL) and medial longissimus (ML) execute the vertebral dorsiflexion of lordosis behavior, and since the motor control hierarchy sketched above parallels lordosis behavior circuitry, we tested the hypothesis that medial hypothalamic lesions (which, in behavioral experiments, decrease lordosis) can also reduce medullary reticular activation of deep back muscle EMG. Urethane-anesthetized rats were tested systematically for amplitude of lateral longissimus (LL) and medial longissimus (ML) EMG responses to electrical stimulus trains applied to the nucleus gigantocellularis (NGC) of the medullary reticular formation, before and after electrolytic lesions of the ventromedial hypothalamus (n = 18) or control sites (n = 30). Bilateral ventromedial hypothalamic lesions were able to greatly reduce EMG responses in LL and ML, often with a time course similar to previous lordosis behavioral results. Surprisingly, lesions at the anterior ventromedial nucleus pole were particularly effective, and may reflect importance of intraventromedial local neurons. Although, on the average, various control lesions were less effective, the ventromedial hypothalamic effect was not unique. For example, it was possible to see an EMG decrease following lesions of the dorsomedial thalamus. Nevertheless, EMG loss was not well correlated with changes in the cortical EEG, and thus does not appear to be a simple consequence of changes in “arousal.” In conclusion, it appears that ventromedial hypothalamic neurons can affect medullary reticular control of back muscle EMG, but must share this role with other forebrain elements.

The studies described herein were designed to test the hypothesis that a neuroleptic, haloperidol, may alter the level of expression of the tyrosine hydroxylase and cholecystokinin genes in discrete brain regions.

A human cDNA clone containing the 5' coding region of the GABAA/benzodiazepine receptor alpha subunit was used to quantify and visualize receptor mRNA in various regions of the rat brain. Using a [32P]CTP-labelled antisense RNA probe (860 bases) prepared from the alpha subunit cDNA, multiple mRNA species were detected in Northern blots using total and poly A rat brain RNA. In all brain regions, mRNAs of 4.4 and 4.8 kb were observed, and an additional mRNA of 3.0 kb was detected in the cerebellum and hippocampus. The level of GABAA/benzodiazepine receptor mRNA was highest in the cerebellum followed by the thalamus = frontal cortex = hippocampus = parietal cortex = hypothalamus much greater than pons = striatum = medulla. In situ hybridization revealed high levels of alpha subunit mRNA in cerebellar gray matter, olfactory bulb, thalamus, hippocampus/dentate gyrus, and the arcuate nucleus of the hypothalamus. These data suggest the presence of multiple GABAA/benzodiazepine receptor alpha subunit mRNAs in rat brain and demonstrate the feasibility of studying the expression of genes encoding the GABAA/benzodiazepine receptor after pharmacological and/or environmental manipulation.

EMG responses were recorded from axial muscles transversospinalis, medial longissimus, and lateral longissimus in urethane-anesthetized rats during combined electrical stimulation of the reticular formation and midbrain central gray. Central gray stimulation facilitated reticular formation-evoked EMG activity in the back muscles of the rat. Electrical stimulation of the central gray lowered the threshold for reticulospinal activation of axial muscles and could maintain firing in these muscles after the end of a reticular formation train. Units were recruited in order of size from small to large. In only one case, central gray stimulation activated axial muscles directly without reticular formation stimulation. The central gray may be important in relaying hypothalamic influences to the reticular formation, which has direct access to the axial muscles responsible for lordosis behavior.

Electrical stimulation in the midbrain central gray in urethane-anesthetized female rats increased responses of the deep back muscles lateral longissimus and medial longissimus to stimulation of the lateral vestibular nucleus (LVN). During central gray stimulation, LVN stimuli led to larger muscle responses, recruitment of new motor units, and decreased latency of muscle response. Effective central gray sites are hypothesized to act through axons descending to medullary reticular formation. Results are consistent with participation of these neuronal groups in the activation of lordosis behavior, a vertebral dorsiflexion that requires deep back muscle contraction, but these electromyographic results could also be relevant for other behaviors that require vertebral postural adjustments.

Neurons in the central nervous system concentrate estrogen, progesterone, testosterone or dihydrotestosterone, and corticosterone. The location of these neurons in the brain has been determined by using autoradiographic (Pfaff 1968; Pfaff and Keiner 1973; Stumpf et al. 1975; Morrell and Pfaff 1981) and steroid receptorbinding techniques (McEwen et al. 1979). Steroid sex hormone-binding cells are found in the medial preoptic area, medial hypothalamus, and limbic forebrain areas in the vertebrate brain; in general, these neurons are found in brain regions that participate in behavioral and pituitary reproductive functions (McEwen et al. 1979). In contrast corticosterone-concentrating neurons are found primarily in the hippocampus.