Biology subjects

Nelson, M. T.

Publications and source records attributed to Nelson, M. T..

15 recordsLinked to original sources

bioRxiv2026

Neurons entrain and shape an intrinsic arteriole rhythm that distributes blood across the cortical mantle

The cerebral arteriole network distributes blood across cortex through branches whose diameters oscillate near a frequency fvaso ~ 0.1 Hz. We asked whether the spatiotemporal pattern of these vaso-oscillations reflects neuronal entrainment of an intrinsic vascular oscillation. Three findings support this: First, suppressing endothelial hyperpolarization left resting-state oscillations intact but weakened the vascular response to neural activity, showing that vasomotion originates within the arteriole network rather than following neuronal drive. Second, arterioles amplify sensory drive selectively rather than faithfully: neural activity near 0.1 Hz is resonantly enhanced to form a strong peak in the vascular response. Third, neuronal frequency sets spatial reach, and areas compete: when two neighboring cortical areas are driven simultaneously, the one nearer fvaso captures more vascular territory. Neurons therefore entrain an intrinsic vascular rhythm and stimulation frequency governs how vasomotion distributes blood, shapes functional brain images, and may influence interstitial fluid transport.

neuroscience↗

bioRxiv2026

Spatial uncoupling of endothelial calcium signaling from phospha-tidylserine externalization in vascular injury

Ca2+ is both the signal that spreads injury information through the endothelium and the trigger for phospholipid scrambling that makes a membrane procoagulant. How a blood vessel can disseminate the first without broadcasting the second is unknown. Here, we combine single-cell laser injury in endothelial monolayers with imaging of intact mesenteric vessels in vitro and intravital Ca2+ imaging in vivo. Injury generated a coherent Ca2+ wavefront that spread rapidly before progressively slowing and terminating at a reproducible boundary 197 {+/-} 7 m from the wound, engaging 50 {+/-} 8 cells. The first ring of neighboring cells exhibited a distinct response that was prevented by carbenoxolone, whereas propagation beyond this ring required extracellular ATP, purinergic receptor activation, and IP3-dependent Ca2+ release. Wavefront kinematics were consistent with ATP diffusion. In vivo, injury elicited vasoconstriction and bidirectional Ca2+ waves propagating 205 {+/-} 28 m before terminating. Remarkably, despite widespread Ca2+ elevations, phosphatidylserine (PS) externalization remained confined to the wounded cell in monolayers and intact vessels in vitro. In contrast, ionomycin-induced Ca2+ overload exposed phosphatidylserine in nearly all endothelial cells. These findings reveal an unexpected spatial uncoupling between endothelial Ca2+ signaling and phospholipid scrambling, demonstrating that Ca2+ dependent scramblase activation is subject to additional spatial regulation. The endothelium can therefore propagate injury signals across a broad field of cells while maintaining a highly localized procoagulant membrane response.

physiology↗

bioRxiv2026

G-protein coupled receptor activity mediates detrusor smooth muscle phasic contractility through regulation of membrane potential

The urinary bladder functions to store and release urine, yet how the sensation of bladder fullness is conveyed and perceived to the central nervous system is not understood. During bladder filling, the detrusor smooth muscle (DSM) generates phasic contractions, resulting in pressure fluctuations within the bladder. These transient pressure events drive bursts of afferent nerve activity, yet the underlying mechanism leading to rhythmic contractions remains unclear. Here, we examined the role of Gq protein-coupled receptor (GqPCR) activity on DSM excitability and contractility. Using ex vivo pressurized urinary bladder preparations and sharp microelectrode experiments on bladder strips from mice, we evaluated whole bladder transient pressure events, whole bladder DSM Ca2+ activity, and membrane potential in bladder strips. We found that global inhibition of urinary bladder GqPCR activity with YM-254890 abates phasic contractility and transient pressure events through a reduction in DSM Ca2+ activity and propagation of Ca2+ waves. Further, we found inhibition of GqPCR significantly hyperpolarizes DSM, reducing action potentials and decreasing excitability, and activation of protein kinase C restores membrane potential to baseline levels. These findings highlight that GqPCR activity mediates DSM excitability and contractility in such a way as to result in phasic detrusor contractions and transient pressure events.

physiology↗

bioRxiv2026

Electromechanical Dynamics and Myogenic Responses in Cerebral Smooth Muscle Cells and Capillary Pericytes

Cerebral blood flow (CBF) control is essential for normal brain function and is disrupted in pathological conditions. Arterial diameters are tightly regulated to provide on demand increases in blood flow in regions of neuronal activity. Pericytes (PCs) exhibit robust myogenic tone and may also respond to neuronal activity to fine-tune local resistance and blood flow. Thus, mural control of microcirculatory resistance may extend beyond arteries and arterioles. Yet, PCs electrophysiology and contractility have not been thoroughly characterized, and this prohibits an integrated view of brain blood flow control. In this study, we develop a detailed mathematical model of mural cell electrophysiology, Ca2+ dynamics and biomechanics. The model is informed by electrophysiological data in smooth muscle cells (SMCs) or PCs and predictions are compared against pressure-induced responses in isolated arterioles and capillaries, respectively. Simulations recapitulate myogenic constrictions and examine differences in contractile dynamics as we move from arterioles to proximal and distal capillaries. In arteriole-to-capillary transitional (ACT) zone PCs, increased mechanosensitivity, more Ca2+ influx through non-selective cation (NSC) channels and/or a higher sensitivity of the contractile apparatus to Ca2+ can compensate for reduced L-type voltage-operated (VOCC) Ca2+ influx and allow for robust constrictions at the lower operating pressures of capillaries relative to the arterioles. A significant Ca2+ influx through NSC relative to VOCC, however, can decouple the PCs contractile apparatus from electrical signaling. Vasoactivity to chemomechanical stimuli along the arteriole to capillary axis is progressively driven by VOCC-independent Ca2+ influx and Ca2+ sensitization with slow kinetics. The proposed cell model can form the basis for detailed multiscale and multicellular models that will examine physiological function at a single vessel or vascular network levels and investigate CBF control in health and in disease. Key pointsO_LIA mural cell model of electrophysiology, calcium (Ca2+) dynamics and biomechanics is informed by data and adapted for modeling cerebral arteriole smooth muscle cells and capillary pericytes. C_LIO_LIIon channel activities are characterized by patch-clamp electrophysiology in isolated cerebral smooth muscle cell and pericytes, and capillary and arteriole electromechanical responses to transmural pressure changes are assessed using novel ex vivo preparations. C_LIO_LIMyogenic constrictions in arterioles can be reproduced by pressure-induced non-selective cation channel (NSC) activation that depolarizes the cell, opens L-type Ca2+ channels (VOCCs) and increases Ca2+ influx. C_LIO_LIRobust myogenic constrictions in arteriole-to-capillary transition (ACT) zone pericytes may reflect significant Ca2+ influx through NSC, increased mechanosensitivity, or higher sensitivity of the contractile apparatus to Ca2+, potentially compensating for reduced VOCC density relative to arteriolar smooth muscle. C_LIO_LIA significant contribution of NSC relative to VOCC in Ca2+ influx, can decouple the contractile apparatus from electrical signaling. C_LIO_LIThe model shows how gradients in ionic activities, mechanosensitivity and/or Ca2+ sensitivity can alter contractile phenotype and electromechanical coupling along the arteriole to capillary continuum. C_LIO_LIThe proposed model can form the basis for detailed multiscale and multicellular models that will investigate cerebral blood flow control in health and in disease. C_LI

physiology↗

bioRxiv2026

Electro-Calcium uncoupling precedes neurodegeneration in Alzheimers disease

Alzheimers disease (AD) is categorized as a neurodegenerative disease, but there is a growing recognition of the vascular components in AD pathophysiology. Reduction in cerebral perfusion is routinely observed in AD patients and preclinical models prior to overt clinical symptoms. However, there is a limited mechanistic understanding of the early neurovascular deficits in AD, and how these may ultimately contribute to pathology. Here, we investigated the mechanisms of early neurovascular dysfunction in AD by using 3-month-old 5xFAD mice, a familial mouse model of AD. Functional hyperemia--the increase in cerebral blood flow (CBF) in response to neuronal activity--is driven by inward rectifier K+ (Kir2.1)-mediated hyperpolarizing (electrical) signals and Ca2+-dependent nitric oxide production within the capillary endothelial cells (cECs). Electrical and Ca2+ signals are tightly coupled through cECs membrane potential, referred to as Electro-Calcium (E-Ca) coupling. We hypothesize that E-Ca uncoupling contributes to impaired functional hyperemia in 5xFAD mice and that these neurovascular deficits precede the neurodegeneration and cognitive decline. At three months of age, 5xFAD mice did not exhibit any impairment in spatial learning and memory, or neuronal density. However, whisker stimulation-induced functional hyperemia was significantly reduced in 5xFAD mice compared to controls. Functional hyperemia exhibited a bimodal response in controls--consisting of fast and slow phases--with the slow phase being significantly reduced in 5xFAD mice. To identify mechanisms underlying these deficits, we measured cortical neuronal and endothelial Ca2+ activity using in-vivo imaging. Neuronal Ca2+ activity was comparable between controls and 5xFAD mice, while cECs Ca2+ activity was significantly reduced in 5xFAD mice. Moreover, Kir2.1 channel blocker, barium (100 M) significantly suppressed cECs Ca2+ activity in controls, but not in 5xFAD mice, consistent with crippled E-Ca coupling. Despite these vascular functional impairments, capillary density was preserved in 5xFAD mice. TRPV4 channels are one of the major Ca2+ entry pathways in cECs and potentiate E-Ca coupling. cECs TRPV4 current density was significantly reduced in 5xFAD mice while Kir2.1 current density was unchanged, indicating that impaired TRPV4 function underlies the E-Ca uncoupling. In summary, early E-Ca uncoupling leads to impaired functional hyperemia in 5xFAD mice and may contribute to later neuronal and cognitive decline.

neuroscience↗

bioRxiv2025

Endothelial Arf6 sustains capillary electrical signaling and cerebral blood flow through PIP2 regeneration and activation of Kir2.1 channels.

Brain capillaries sense neural activity and direct blood flow to active regions--a process termed neurovascular coupling that underlies activity-dependent increases in local perfusion (functional hyperemia). A key contributor to functional hyperemic responses is the capillary endothelial cell (cEC) inward rectifier K+ (Kir2.1) channel, which is activated by neuronal activity-derived extracellular K+ and initiates vasodilatory electrical signals that propagate through the vascular network. Kir2.1 channel function requires continual production of its lipid cofactor, phosphatidylinositol-4,5-bisphosphate (PIP2), and is compromised in mouse models of cerebral small vessel (cSVD). Although decreased PIP2 availability is a common feature of cSVD, mechanisms underlying PIP2 synthesis remain poorly understood. We hypothesized that Arf6, a small GTPase expressed in cECs that stimulates PIP2 production, is critical for this process. Using patch-clamp electrophysiology, we demonstrate that inhibiting Arf6 activity progressively decreased cEC Kir2.1 channel activity. This deficit corresponded to loss of capillary-to-arteriole electrical signaling in isolated vessels and diminished functional hyperemia in vivo. Exogenously provided PIP2 restored Kir2.1 currents and functional hyperemia after Arf6 inhibition or genetic knockdown. Collectively, our data indicate that cEC Arf6 sustains Kir2.1 activity by maintaining PIP2 levels and demonstrate that diminished PIP2 synthesis is sufficient to impair functional hyperemia. Furthermore, we identify Arf6 as a mechanistic link between PIP2 production and endothelial electrical signaling, highlighting Arf6 as a potential therapeutic target for restoring functional hyperemia. Significance StatementActive brain regions send electrical signals through capillaries to dilate upstream arterioles and increase blood flow. The resulting activity-dependent increase in local blood flow (functional hyperemia) is mediated through inward-rectifier potassium (Kir2.1) channels. These channels-- and hence functional hyperemia--require continuous regeneration of the lipid cofactor PIP2 (phosphatidylinositol-4,5-bisphosphate). If PIP2 is deficient, electrical signaling fails--a defect characteristic of models of cerebral small vessel disease and Alzheimers disease. We identify the small GTPase Arf6 as a key to maintaining PIP2 and thus preserving capillary Kir2.1 activity and functional hyperemia. Our findings reveal an important pathway for PIP2 homeostasis and position Arf6 as a cornerstone upholding functional hyperemic responses, highlighting Arf6 as a target for restoring cerebral blood flow in disease.

neuroscience↗

bioRxiv2025

Endothelial Trauma Depends on Surface Charge and Extracellular Calcium Levels

We tested the hypothesis that the ubiquitous store-operated Ca2+ entry (SOCE) pathway contributes to histone-induced endothelial Ca2+ events. We also considered an alternate hypothesis: cationic electrostatic interactions between histones and negatively charged phospholipids deform endothelial membranes and thereby allow extracellular Ca2+ entry. A role for SOCE in histone responses was ruled out by genetic ablation of the ORAI1/2/3 channel trio; yet, histone effects were blocked by application of the multivalent cation gadolinium Gd3+. Using live cell video microscopy of endothelial cells labeled with membrane dye FM1-43, we recorded plasma membrane movements including vesiculation, blebbing, and ruffling of lamellipodia over 60 minutes following histone exposure. These cell membrane theatrics were markedly different from the uniform pattern of exocytosis and subsequent blebbing produced by calcium overload with ionomycin. The membrane permeabilization produced by histones, and not ionomycin, was transient and a subset of cells recovered membrane integrity within 1 hour. Removal of extracellular Ca2+ prevented histone-induced intracellular Ca2+ overload while surprisingly exacerbating plasma membrane deformation. Conversely, decreasing the density of the negative charge surface by adding calcium or or increasing extracellular Ca2+ levels effectively screened common membrane phospholipids from interactions with labeled histones and prevented endothelial damage in cells exposed to histones. Collectively these results indicate that low extracellular Ca2+ levels enhance interactions between histones and endothelial cell membrane phospholipids to increase cytotoxicity. Importantly, this supports the concept of aggressive Ca2+ repletion during resuscitation to prevent hypocalcemia, stabilize endothelial cell membranes and improve cardiovascular recovery from shock. SignificanceIn acute critical illness, the rapid collapse of vascular endothelial functions drives aberrant blood clotting and organ failure through mechanisms that are not understood. Emerging evidence that early administration of donor plasma improves survival of trauma patients has transformed the massive transfusion protocols used in surgical settings, but the sodium citrate included in transfused blood products to prevent coagulation often produces significant and severe hypocalcemia. Here, we demonstrate that cytotoxic trauma factors that are elevated in the blood during resuscitation interact electrostatically with endothelial cell phospholipids, and that low Ca2+ exacerbates toxicity by increasing this interaction. Using high speed video imaging, we demonstrate fast endothelial cell membrane movements in response to injury, including protrusion and ruffling of lamellipodia, release and reuptake of extracellular vesicles, and blebbing. These findings provide important insights into the nature of shock-induced endotheliopathy and highlight the potential cardiovascular risk associated with chelation-induced hypocalcemia during resuscitation.

physiology↗

bioRxiv2025

Adenosine and acute low oxygen conditions suppress urinary bladder contractility through the activation of adenosine 2B receptors and large-conductance calcium-activated potassium channels

Under healthy conditions the urinary bladder undergoes relatively long periods of filling with well-spaced voiding events to ensure proper storage and removal of urine respectively. During the filling phase, distinct contractile events in detrusor urinary smooth muscle (UBSM) elicit transient non-voiding pressure events and associated bursts in afferent nerve activity to relay the sensation of bladder fullness. The mechanisms that regulate UBSM excitability and associated non-voiding pressure events under physiological and pathological conditions are poorly understood. Here we investigated the role of adenosine signaling in regulating urinary bladder contractility. Using an ex vivo pressurized bladder preparation from mice and patch-clamp electrophysiology in isolated UBSM we evaluated whole bladder transient pressure events, whole bladder detrusor Ca2+ activity, and single UBSM ion channel activity. We found that adenosine suppresses bladder activity through activation of A2B adenosine receptors and downstream activation of large-conductance calcium-activated potassium (BKCa) channels. We further demonstrated that acute exposure to low oxygen conditions using a chemical oxygen scavenger potently suppresses bladder contractility through the A2B receptor pathway. These results highlight the prominent role adenosine receptors and downstream potassium channels play in regulating urinary bladder contractility in physiological and pathological contexts.

physiology↗

bioRxiv2023

Epidermal growth factor receptors in vascular endothelial cells contribute to functional hyperemia in the brain

Functional hyperemia - activity-dependent increases in local blood perfusion - underlies the on-demand delivery of blood to regions of enhanced neuronal activity, a process that is crucial for brain health. Importantly, functional hyperemia deficits have been linked to multiple dementia risk factors, including aging, chronic hypertension, and cerebral small vessel disease (cSVD). We previously reported crippled functional hyperemia in a mouse model of genetic cSVD that was likely caused by depletion of phosphatidylinositol 4,5-bisphosphate (PIP2) in capillary endothelial cells (EC) downstream of impaired epidermal growth factor receptor (EGFR) signaling. Here, using EC-specific EGFR-knockout (KO) mice, we directly examined the role of endothelial EGFR signaling in functional hyperemia, assessed by measuring increases in cerebral blood flow in response to contralateral whisker stimulation using laser Doppler flowmetry. Molecular characterizations showed that EGFR expression was dramatically decreased in freshly isolated capillaries from EC-EGFR-KO mice, as expected. Notably, whisker stimulation-induced functional hyperemia was significantly impaired in these mice, an effect that was rescued by exogenous administration of PIP2, but not by the EGFR ligand, HB-EGF. These data suggest that the deletion of the EGFR specifically in ECs depletes PIP2 and attenuates functional hyperemia, underscoring the central role of the endothelial EGFR signaling in cerebral blood flow regulation.

physiology↗

bioRxiv2023

Pathogenic soluble tau peptide disrupts endothelial calcium signaling and vasodilation in the brain microvasculature

The accumulation of the microtubule-associated tau protein in and around blood vessels contributes to brain microvascular dysfunction through mechanisms that are incompletely understood. Delivery of nutrients to active neurons in the brain relies on capillary inositol 1,4,5-triphosphate receptor (IP3R)-mediated calcium (Ca2+) signals to direct blood flow. The initiation and amplification of endothelial cell IP3R-mediated Ca2+ signals requires an intact microtubule cytoskeleton. Since tau accumulation in endothelial cells disrupts native microtubule stability, we reasoned that tau-induced microtubule destabilization would impair endothelial IP3-evoked Ca2+ signaling. We tested the hypothesis that tau disrupts the regulation of local cerebral blood flow by reducing endothelial cell Ca2+ signals and endothelial-dependent vasodilation. We used a pathogenic soluble tau peptide (T-peptide) model of tau aggregation and mice with genetically encoded endothelial Ca2+ sensors to measure cerebrovascular endothelial responses to tau exposure. T-peptide significantly attenuated endothelial Ca2+ activity and cortical capillary blood flow in vivo within 120 seconds. Further, T-peptide application constricted pressurized cerebral arteries and inhibited endothelium-dependent vasodilation. This study demonstrates that pathogenic tau alters cerebrovascular function through direct attenuation of endothelial Ca2+ signaling and endothelium-dependent vasodilation.

neuroscience↗

bioRxiv2021

The polyanionic drug suramin neutralizes histones and prevents endotheliopathy

Drugs are needed to protect against the neutrophil-derived histones responsible for endothelial injury in acute inflammatory conditions such as trauma and sepsis. Heparin and other polyanions can neutralize histones but may cause secondary, deleterious effects such as excessive bleeding. Here, we demonstrate that suramin--a widely available polyanionic drug--completely neutralizes the toxic effects of histones. The sulfate groups on suramin form stable electrostatic interactions with hydrogen bonds in the histone octamer with a dissociation constant of 250 nM. In cultured endothelial cells (Ea.Hy926), histone-induced thrombin generation was significantly decreased by suramin. In isolated murine blood vessels, suramin abolished aberrant endothelial cell calcium signals and rescued impaired endothelial-dependent vasodilation caused by histones. Suramin significantly decreased pulmonary endothelial cell ICAM-1 expression and neutrophil recruitment caused by infusion of sub-lethal doses of histones in vivo. Suramin also prevented lung edema, intra-alveolar hemorrhage and mortality in mice receiving a lethal dose of histones. Protection of vascular endothelial function from histone-induced damage is a novel mechanism of action for suramin with therapeutic implications for conditions characterized by elevated histone levels. Significance StatementPathologic levels of circulating histones cause acute endotheliopathy, characterized by widespread disruption of critical endothelial functions and thromboinflammation. We discovered that suramin binds histones and prevents histone-induced endothelial dysfunction, thrombin generation, lung injury, and death. Histone binding is a novel mechanism of action for suramin, considered among the safest and most effective drugs by the World Health Organization. These results support the use of suramin for protection of blood vessels in conditions exacerbated by circulating histones including trauma and sepsis.

immunology↗

bioRxiv2021

Electro-Metabolic Sensing Through Capillary ATP-Sensitive K+ Channels and Adenosine to Control Cerebral Blood Flow

The dense network of capillaries composed of capillary endothelial cells (cECs) and pericytes lies in close proximity to all neurons, ideally positioning it to sense neuro/glial-derived compounds that regulate regional and global cerebral perfusion. The membrane potential (VM) of vascular cells serves as the essential output in this scenario, linking brain activity to vascular function. The ATP-sensitive K+ channel (KATP) is a key regulator of vascular VM in other beds, but whether brain capillaries possess functional KATP channels remains unknown. Here, we demonstrate that brain capillary ECs and pericytes express KATP channels that robustly control VM. We further show that the endogenous mediator adenosine acts through A2A receptors and the Gs/cAMP/PKA pathway to activate capillary KATP channels. Moreover, KATP channel stimulation in vivo causes vasodilation and increases cerebral blood flow (CBF). These findings establish the presence of KATP channels in cECs and pericytes and suggest their significant influence on CBF. HIGHLIGHTSO_LICapillary network cellular components--endothelial cells and pericytes--possess functional KATP channels. C_LIO_LIActivation of KATP channels causes profound hyperpolarization of capillary cell membranes. C_LIO_LICapillary KATP channels are activated by exogenous adenosine via A2A receptors and cAMP-dependent protein kinase. C_LIO_LIKATP channel activation by adenosine or synthetic openers increases cerebral blood flow. C_LI

neuroscience↗

bioRxiv2021

Traumatic Brain Injury Impairs Systemic Vascular Function Through Altered Lipid Metabolism and Disruption of Inward-Rectifier Potassium (Kir2.1) Channels

BACKGROUND AND PURPOSETrauma can lead to widespread vascular endothelial dysfunction, but the underlying mechanisms remain largely unknown. Strong inward-rectifier potassium channels (Kir2.1) play a critical role in the dynamic regulation of regional perfusion and blood flow. Kir2.1 channel activity is modulated by phosphatidylinositol 4,5-bisphosphate (PIP2), a minor membrane phospholipid that is degraded by phospholipase A2 (PLA2) in conditions of oxidative stress or severe inflammation. We hypothesized that PLA2-induced depletion of PIP2 impairs Kir2.1 channel function. METHODSA fluid percussion injury model of traumatic brain injury (TBI) in rats was used to study mesenteric resistance arteries 24 hours after injury. Patch-clamp electrophysiology in freshly isolated endothelial and smooth muscle cells was performed to monitor Kir2.1 conductance, and the functional responses of intact arteries were assessed using pressure myography. We analyzed circulating PLA2, hydrogen peroxide (H2O2), and metabolites to identify alterations in signaling pathways associated with PIP2 in TBI. RESULTSElectrophysiology analysis of endothelial and smooth muscle cells revealed a significant reduction of Ba2+-sensitive Kir2.1 currents after TBI. Additionally, dilations to elevated extracellular potassium and BaCl2- or ML 133-induced constrictions in pressurized arteries were significantly decreased following TBI, consistent with an impairment of Kir2.1 channel function. The addition of a PIP2 analog to the patch pipette successfully rescued endothelial Kir2.1 currents after TBI. Both H2O2 and PLA2 activity were increased after injury. Metabolomics analysis demonstrated altered lipid metabolism signaling pathways, including increased arachidonic acid, and fatty acid mobilization after TBI. CONCLUSIONSOur findings support a model in which increased H2O2-induced PLA2 activity after trauma hydrolyzes endothelial PIP2, resulting in impaired Kir2.1 channel function.

physiology↗

bioRxiv2020

Vascular control of the CO2/H+ dependent drive to breathe

Respiratory chemoreceptors regulate breathing in response to changes in tissue CO2/H+. Blood flow is a fundamental determinant of tissue CO2/H+, yet little is known regarding how regulation of vascular tone in chemoreceptor regions contributes to respiratory behavior. Previously, we showed in rat that CO2/H+-vasoconstriction in the retrotrapezoid nucleus (RTN) supports chemoreception by a purinergic-dependent mechanism (Hawkins et al. 2017). Here, we show in mice that CO2/H+ dilates arterioles in other chemoreceptor regions, thus demonstrating CO2/H+ vascular reactivity in the RTN is unique. We also identify P2Y2 receptors in RTN smooth muscle cells as the substrate responsible for this response. Specifically, pharmacological blockade or genetic deletion of P2Y2 from smooth muscle cells blunted the ventilatory response to CO2, and re-expression of P2Y2 receptors only in RTN smooth muscle cells fully rescued the CO2/H+ chemoreflex. These results identify P2Y2 receptors in RTN smooth muscle cells as requisite determinants of respiratory chemoreception. Significance StatementDisruption of vascular control as occurs in cardiovascular disease leads to compromised chemoreceptor function and unstable breathing. Despite this, virtually nothing is known regarding how regulation of vascular tone in chemoreceptor regions contributes to respiratory behavior. Here, we identify P2Y2 receptors in RTN vascular smooth muscle cells as a novel vascular element of respiratory chemoreception. Identification of this mechanism may facilitate development of treatments for breathing problems including those associated with cardiovascular disease.

neuroscience↗

bioRxiv2020

Interplay of the Trypanosome Lytic Factor and innate immune cells in the resolution of cutaneous Leishmania infection

Trypanosome Lytic Factor (TLF) is a primate-specific high-density lipoprotein complex that contains APOL1, the lytic component. Human TLF confers sterile immunity to many animal-infective extracellular Trypanosoma Ssp, which have been extensively investigated. Here, we have dissected the underappreciated role of TLF and neutrophils against intracellular Leishmania in intradermal infection. Our data show that mice producing human or baboon TLF have reduced parasite burdens when infected intradermally with metacyclic promastigotes of L. major. This TLF-mediated reduction in parasite burden was lost in neutrophil-depleted TLF mice, suggesting that early recruitment of neutrophils is required for TLF-mediated killing of L. major. Neutrophils and macrophages are the predominant phagocytes recruited to the site of infection. Our data show that acidification of the macrophage phagosome is essential for TLF-mediated lysis of metacyclic promastigotes. In vitro we find that only metacyclic promastigotes co-incubated with TLF in an acidic milieu were lysed. However, amastigotes were not killed by TLF at any pH. These findings correlated with binding experiments, revealing that labeled TLF binds specifically to the surface of metacyclic promastigotes, but not to amastigotes. During differentiation to the amastigote stage, the parasites shed their surface glycoconjugates. Metacyclic promastigotes of L. major deficient in the synthesis of surface glycoconjugates (lpg1- and lpg5A-/lpg5B-) were partially resistant to TLF lysis. We propose that TLF binds to the outer surface glycoconjugates of metacyclic promastigotes, whereupon APOL1 forms a pH-gated ion channel in the plasma membrane, resulting in osmotic lysis. We hypothesize that resistance to TLF requires shedding of the surface glycoconjugates, which occurs upon phagocytosis by immune cells. Author SummaryLeishmaniasis is a common term used for disease caused by parasites of the genus Leishmania. Depending on the parasite species and the clinical outcome of the disease, leishmaniasis can be divided into cutaneous, muco-cutaneous and visceral. Of the three, cutaneous leishmaniasis is the most common form, which is usually characterized by a localized lesion due to the infection of immune cells, primarily macrophages of the dermis and local lymph nodes. Sometimes, infected individuals can remain asymptomatic and do not show visible lesions. Moreover, the time between the infection and appearance of lesions are also variable and range from a few weeks to months and a few years in some cases. This subclinical stage of leishmaniasis depends on a variety of factors: parasite virulence, infectious dose, and host immune response. Therefore, it is important to understand the host-parasite interaction and its role in the clinical outcome of the disease. Here, we analyze the interaction between a cutaneous strain of Leishmania and a host innate immune factor called Trypanosome Lytic Factor (TLF). TLF is a type of High-Density Lipoprotein (HDL) complex that circulates in our plasma. TLF kills extracellular African Trypanosomes by lysing the parasites. The lytic ability of TLF is due to the primate specific protein APOL1 that forms pH gated ion channels. APOL1 inserts into biological membranes at acidic pH and forms a closed ion-channel that opens when the membrane associated APOL1 is exposed to neutral pH. Using transgenic mice producing primate TLF, we show both human and baboon TLFs ameliorate cutaneous Leishmania major infection. The reduction in parasite burden correlated with: 1. infectious dose of metacyclic promastigotes and 2. the concentration of circulating TLF in mouse plasma. The early recruitment of neutrophils at the site of infection was required for the reduction of parasite burden by TLF. Macrophages, another major cell that phagocytoses metacyclic promastigotes at the site of infection require an acidified phagosome for TLF mediated killing of L. major. The acidification step is also essential for TLF mediated lysis of axenic metacyclic promastigotes of Leishmania in vitro. The susceptibility of metacyclic promastigotes to TLF mediated lysis is governed by the surface glycoconjugates of Leishmania. We find that surface glycoconjugate deficient Leishmania are resistant to TLF mediated killing. Based on these data, we conclude that the shedding of surface glycoconjugates while transitioning from metacyclic promastigotes to amastigotes results in parasite resistance to TLF mediated lysis. Whether TLF is effective at killing metacyclic promastigotes of other experimentally tractable Leishmania sp. such as L. infantum, and L. donovani, which have slightly different surface glycoconjugate structures is yet to be tested. Our data raise the possibility that TLF can have lytic activity against a broad range of pathogens such as bacteria, viruses, fungi and parasites with surface glycoconjugates that transit through intracellular acidic compartments.

microbiology↗