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The adult brain exhibits a local increase in cortical blood flow in response to external stimulus. However, broadly varying hemodynamic responses in the brains of newborn and young infants have been reported. Particular controversy exists over whether the \"true\" neonatal response to stimulation consists of a decrease or an increase in local deoxyhemoglobin, corresponding to a positive (adult-like) or negative blood oxygen level-dependent (BOLD) signal in functional magnetic resonance imaging (fMRI), respectively. A major difficulty with previous studies has been the variability in human subjects and measurement paradigms. Here, we present a systematic study in neonatal rats that charts the evolution of the cortical blood flow response during postnatal development using exposed-cortex multispectral optical imaging. We demonstrate that postnatal-day-12-13 rats (equivalent to human newborns) exhibit an \"inverted\" hemodynamic response (increasing deoxyhemoglobin, negative BOLD) with early signs of oxygen consumption followed by delayed, active constriction of pial arteries. We observed that the hemodynamic response then matures via development of an initial hyperemic (positive BOLD) phase that eventually masks oxygen consumption and balances vasoconstriction toward adulthood. We also observed that neonatal responses are particularly susceptible to stimulus-evoked systemic blood pressure increases, leading to cortical hyperemia that resembles adult positive BOLD responses. We propose that this confound may account for much of the variability in prior studies of neonatal cortical hemodynamics. Our results suggest that functional magnetic resonance imaging studies of infant and child development may be profoundly influenced by the maturing neurovascular and autoregulatory systems of the neonatal brain.

Astrocytes are increasingly believed to play an important role in neurovascular coupling. Recent in vivo studies have shown that intracellular calcium levels in astrocytes correlate with reactivity in adjacent diving arterioles. However, the hemodynamic response to stimulation involves a complex orchestration of vessel dilations and constrictions that spread rapidly over wide distances. In this work, we study the three-dimensional cytoarchitecture of astrocytes and their interrelations with blood vessels down through layer IV of the mouse somatosensory cortex using in vivo two-photon microscopy. Vessels and astrocytes were visualized through intravenous dextran-conjugated fluorescein and cortically applied sulforhodamine 101 (SR101), respectively. In addition to exploring astrocyte density, vascular proximity, and microvascular density, we found that sheathing of subpial vessels by astrocyte processes was continuous along all capillaries, arterioles, and veins, comprising a highly interconnected pathway through which signals could feasibly be relayed over long distances via gap junctions. An inner SR101-positive sheath noted along pial and diving arterioles was determined to be nonastrocytic, and appears to represent selective SR101 staining of arterial endothelial cells. Our findings underscore the intimate relationship between astrocytes and all cortical blood vessels, and suggest that astrocytes could influence neurovascular regulation at a range of sites, including the capillary bed and pial arterioles.

While a range of cellular mechanisms have been proposed to underlie control of neurovascular coupling, a comprehensive, reconciliatory model has yet to be determined. To fit with such a model, it is essential that candidate mechanisms exhibit reaction times, spatial ranges, and speeds of propagation that are consistent with the vascular manifestations of the ‘hemodynamic response’. Understanding these vascular dynamics is therefore a critical step towards developing a robust model of neurovascular coupling. In this study, we utilize high-speed optical imaging of exposed rodent somatosensory cortex to explore and characterize the spatiotemporal dynamics of surface vessels during functional hyperemia. Our high-speed, high-resolution optical imaging approach allows us to study the hemodynamic response independently in individual vessels, and in discrete regions of the parenchyma with enough resolution to precisely characterize subtle spatial and temporal features of the response. Specifically, we explore when and where the first hemodynamic changes occur in response to stimuli, the direction and speed at which these changes propagate in arterioles and regions of the parenchyma, and the relative timing at which each of these compartments returns to its original baseline state. From these results, we are able to conclude that the hemodynamic response appears to initiate in the parenchyma and then spreads rapidly to surface arterioles. Following the initial onset we find evidence that the response spreads spatially outwards via the dilation of targeted arterioles. This propagation of vasodilation is independent of the direction of blood flow within each arteriole. We also find evidence of a decay phase that acts with a more uniform spatial dependence, rather than along targeted vessels, causing the periphery of the responding region to return to baseline first. We hypothesize that different underlying cellular mechanisms/signaling pathways are responsible for the response initiation and the response decay. Our results advance the fundamental understanding of the hemodynamic response, as well as our ability to evaluate potential cellular mechanisms for their involvement in neurovascular coupling. ►High frame rate optical imaging resolves rapid dynamics of cortical vasculature. ►Functional response begins in the parenchyma, before dilation of pial arterioles. ►Response proceeds with retrograde propagation of vasodilation in pial arterioles. ►Direction of propagation in arterioles is independent of direction of blood flow. ►Return to baseline may have different underlying mechanism compared to onset.

Astrocytes are increasingly believed to play an important role in neurovascular coupling. Recent in vivo studies have shown that intracellular calcium levels in astrocytes correlate with reactivity in adjacent diving arterioles. However, the hemodynamic response to stimulation involves a complex orchestration of vessel dilations and constrictions that spread rapidly over wide distances. In this work, we study the three-dimensional cytoarchitecture of astrocytes and their interrelations with blood vessels down through layer IV of the mouse somatosensory cortex using in vivo two-photon microscopy. Vessels and astrocytes were visualized through intravenous dextran-conjugated fluorescein and cortically applied sulforhodamine 101 (SR101), respectively. In addition to exploring astrocyte density, vascular proximity, and microvascular density, we found that sheathing of subpial vessels by astrocyte processes was continuous along all capillaries, arterioles, and veins, comprising a highly interconnected pathway through which signals could feasibly be relayed over long distances via gap junctions. An inner SR101-positive sheath noted along pial and diving arterioles was determined to be nonastrocytic, and appears to represent selective SR101 staining of arterial endothelial cells. Our findings underscore the intimate relationship between astrocytes and all cortical blood vessels, and suggest that astrocytes could influence neurovascular regulation at a range of sites, including the capillary bed and pial arterioles.

Neurovascular coupling is the relationship between neural activity and blood flow that allows the brain to exhibit increases in blood flow to areas of elevated neural activity during sensory stimulation. It is these localized changes in blood flow, collectively known as the hemodynamic response, that are detected by modern neuroimaging techniques such as functional magnetic resonance imaging (fMRI). Intact neurovascular coupling is imperative to neural health as de-coupling of neural activity from blood flow modulations has been implicated in many neurodegenerative diseases such as Alzheimer's disease, dementia, traumatic brain injury, and ischemic stroke. Despite the importance of neurovascular coupling for both fMRI interpretation and neurological disease, the mechanisms underlying the control of blood flow in the brain remain poorly understood. While previous studies have proposed a range of different cellular mechanisms capable of mediating vascular changes in the brain, it remains difficult to reconcile these mechanisms with a unified theory that is also consistent with the complex spatiotemporal features of the hemodynamic response. The goal of this dissertation is to study the vascular components of the hemodynamic response and the cellular mechanisms that orchestrate them. Using novel high-speed multi-spectral optical imaging of the exposed rodent somatosensory cortex, a detailed characterization of the cortical hemodynamic response is conducted. These observations guide cellular level two-photon microscopy of neural and glial cell activity. The precise spatiotemporal characteristics of the neurovascular response elucidated in these in vivo studies are then used to construct and constrain a conceptual framework for the signaling and actuation pathways that orchestrate the hemodynamic response. To test this framework, targeted light-dye treatment and optogenetic stimulation are used to selectively activate or deactivate targeted signaling pathways. The findings of this research strongly suggest that at least two different mechanisms control the sensory-evoked blood flow response, the first of which critically depends on the vascular endothelium.

The adult brain exhibits a local increase in cortical blood flow in response to external stimulus. However, broadly varying hemodynamic responses in the brains of newborn and young infants have been reported. Particular controversy exists over whether the “true” neonatal response to stimulation consists of a decrease or an increase in local deoxyhemoglobin, corresponding to a positive (adult-like) or negative blood oxygen level-dependent (BOLD) signal in functional magnetic resonance imaging (fMRI), respectively. A major difficulty with previous studies has been the variability in human subjects and measurement paradigms. Here, we present a systematic study in neonatal rats that charts the evolution of the cortical blood flow response during postnatal development using exposed-cortex multispectral optical imaging. We demonstrate that postnatal-day-12–13 rats (equivalent to human newborns) exhibit an “inverted” hemodynamic response (increasing deoxyhemoglobin, negative BOLD) with early signs of oxygen consumption followed by delayed, active constriction of pial arteries. We observed that the hemodynamic response then matures via development of an initial hyperemic (positive BOLD) phase that eventually masks oxygen consumption and balances vasoconstriction toward adulthood. We also observed that neonatal responses are particularly susceptible to stimulus-evoked systemic blood pressure increases, leading to cortical hyperemia that resembles adult positive BOLD responses. We propose that this confound may account for much of the variability in prior studies of neonatal cortical hemodynamics. Our results suggest that functional magnetic resonance imaging studies of infant and child development may be profoundly influenced by the maturing neurovascular and autoregulatory systems of the neonatal brain.

Fred Wright, Patrick Sullivan and colleagues present the results of a large expression QTL study of peripheral blood using a classic twin design with follow-up replication in independent samples. Their results enable a more precise estimate of the heritability of gene expression and provide a useful resource for exploring the genetic control of transcription. We assessed gene expression profiles in 2,752 twins, using a classic twin design to quantify expression heritability and quantitative trait loci (eQTLs) in peripheral blood. The most highly heritable genes (∼777) were grouped into distinct expression clusters, enriched in gene-poor regions, associated with specific gene function or ontology classes, and strongly associated with disease designation. The design enabled a comparison of twin-based heritability to estimates based on dizygotic identity-by-descent sharing and distant genetic relatedness. Consideration of sampling variation suggests that previous heritability estimates have been upwardly biased. Genotyping of 2,494 twins enabled powerful identification of eQTLs, which we further examined in a replication set of 1,895 unrelated subjects. A large number of non-redundant local eQTLs (6,756) met replication criteria, whereas a relatively small number of distant eQTLs (165) met quality control and replication standards. Our results provide a new resource toward understanding the genetic control of transcription.

The first James Webb Space Telescope (JWST) Near InfraRed Camera imaging in the field of the galaxy cluster PLCK G165.7+67.0 (z = 0.35) uncovered a Type Ia supernova (SN Ia) at z = 1.78, called “SN H0pe.” Three different images of this one SN were detected as a result of strong gravitational lensing, each one traversing a different path in spacetime, thereby inducing a relative delay in the arrival of each image. Follow-up JWST observations of all three SN images enabled photometric and rare spectroscopic measurements of the two relative time delays. Following strict blinding protocols which oversaw a live unblinding and regulated postunblinding changes, these two measured time delays were compared to the predictions of seven independently constructed cluster lens models to measure a value for the Hubble constant, H0 = 71.8 + 9.2 − 8.1 km s−1 Mpc−1. The range of admissible H0 values predicted across the lens models limits further precision, reflecting the well-known degeneracies between lens model constraints and time delays. It has long been theorized that a way forward is to leverage a standard candle, but this has not been realized until now. For the first time, the lens models are evaluated by their agreement with the SN absolute magnifications, breaking degeneracies and producing our best estimate, H0 = 75.7−5.5+8.1 km s−1 Mpc−1. This is the first precise measurement of H0 from a multiply imaged SN Ia and only the second from any multiply imaged SN.

Two patients, one with transfusion-dependent β-thalassemia and the other with sickle cell disease, received autologous CD34+ cells edited with CRISPR-Cas9 targeting of BCL11A . Their clinical course over the following 16 to 18 months supports further experimental testing of CRISPR-Cas9 gene editing to treat these diseases.

High-throughput screening and structure-guided design identified small-molecule inhibitors that prevent the interaction between N-terminally acetylated E2 conjugating enzyme UBE2M and DCN1, an E3 ligase for the ubiquitin-like protein Nedd8. N-terminal acetylation is an abundant modification influencing protein functions. Because ∼80% of mammalian cytosolic proteins are N-terminally acetylated, this modification is potentially an untapped target for chemical control of their functions. Structural studies have revealed that, like lysine acetylation, N-terminal acetylation converts a positively charged amine into a hydrophobic handle that mediates protein interactions; hence, this modification may be a druggable target. We report the development of chemical probes targeting the N-terminal acetylation–dependent interaction between an E2 conjugating enzyme (UBE2M or UBC12) and DCN1 (DCUN1D1), a subunit of a multiprotein E3 ligase for the ubiquitin-like protein NEDD8. The inhibitors are highly selective with respect to other protein acetyl-amide–binding sites, inhibit NEDD8 ligation in vitro and in cells, and suppress anchorage-independent growth of a cell line with DCN1 amplification. Overall, our data demonstrate that N-terminal acetyl-dependent protein interactions are druggable targets and provide insights into targeting multiprotein E2–E3 ligases.