scholarly journals Purinergic signaling triggers endfoot high-amplitude Ca2+ signals and causes inversion of neurovascular coupling after subarachnoid hemorrhage

2016 ◽  
Vol 36 (11) ◽  
pp. 1901-1912 ◽  
Author(s):  
Anthony C Pappas ◽  
Masayo Koide ◽  
George C Wellman
Keyword(s):  
Subarachnoid Hemorrhage ◽  
Purinergic Signaling ◽  
Brain Slices ◽  
Neurovascular Coupling ◽  
P2y Receptors ◽  
High Amplitude ◽  
Differential Interference Contrast Microscopy ◽  
Purine Nucleotides ◽  
Interference Contrast Microscopy ◽  
And Inversion

Neurovascular coupling supports brain metabolism by matching focal increases in neuronal activity with local arteriolar dilation. Previously, we demonstrated that an emergence of spontaneous endfoot high-amplitude Ca2+ signals (eHACSs) caused a pathologic shift in neurovascular coupling from vasodilation to vasoconstriction in brain slices obtained from subarachnoid hemorrhage model animals. Extracellular purine nucleotides (e.g., ATP) can trigger astrocyte Ca2+ oscillations and may be elevated following subarachnoid hemorrhage. Here, the role of purinergic signaling in subarachnoid hemorrhage-induced eHACSs and inversion of neurovascular coupling was examined by imaging parenchymal arteriolar diameter and astrocyte Ca2+ signals in rat brain slices using two-photon fluorescent and infrared-differential interference contrast microscopy. We report that broad-spectrum inhibition of purinergic (P2) receptors using suramin blocked eHACSs and restored vasodilatory neurovascular coupling after subarachnoid hemorrhage. Importantly, eHACSs were also abolished using a cocktail of inhibitors targeting Gq-coupled P2Y receptors. Further, activation of P2Y receptors in brain slices from un-operated animals triggered high-amplitude Ca2+ events resembling eHACSs and disrupted neurovascular coupling. Neither tetrodotoxin nor bafilomycin A1 affected eHACSs suggesting that purine nucleotides are not released by ongoing neurotransmission and/or vesicular release after subarachnoid hemorrhage. These results indicate that purinergic signaling via P2Y receptors contributes to subarachnoid hemorrhage-induced eHACSs and inversion of neurovascular coupling.

scholarly journals Acute changes in neurovascular reactivity after subarachnoid hemorrhage in vivo

2016 ◽  
Vol 37 (1) ◽  
pp. 178-187 ◽  
Author(s):  
Matilde Balbi ◽  
Masayo Koide ◽  
Susanne M Schwarzmaier ◽  
George C Wellman ◽  
Nikolaus Plesnila
Keyword(s):  
Subarachnoid Hemorrhage ◽  
Neuronal Damage ◽  
Brain Slices ◽  
Neurovascular Coupling ◽  
Differential Interference Contrast Microscopy ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Normal Increase ◽  
Acute Changes

Subarachnoid hemorrhage causes acute and long-lasting constrictions of pial arterioles. Whether these vessels dilate normally to neuronal activity is of great interest since a mismatch between delivery and consumption of glucose and oxygen may cause additional neuronal damage. Therefore, we investigated neurovascular reactivity of pial and parenchymal arterioles after experimental subarachnoid hemorrhage. C57BL/6 mice were subjected to subarachnoid hemorrhage by filament perforation or sham surgery. Neurovascular reactivity was assessed 3 h later by forepaw stimulation or inhalation of 10% CO2. Diameters of cerebral arterioles were assessed using two-photon microscopy. Neurovascular coupling and astrocytic endfoot Ca2+ were measured in brain slices using two-photon and infrared-differential interference contrast microscopy. Vessels of sham-operated mice dilated normally to CO2 and forepaw stimulation. Three hours after subarachnoid hemorrhage, CO2 reactivity was completely lost in both pial and parenchymal arterioles, while neurovascular coupling was not affected. Brain slices studies also showed normal neurovascular coupling and a normal increase in astrocytic endfoot Ca2+ acutely after subarachnoid hemorrhage. These findings suggest that communication between neurons, astrocytes, and parenchymal arterioles is not affected in the first few hours after subarachnoid hemorrhage, while CO2 reactivity, which is dependent on NO signaling, is completely lost.

scholarly journals Propentdyopents as Heme Degradation Intermediates Constrict Mouse Cerebral Arterioles and Are Present in the Cerebrospinal Fluid of Patients With Subarachnoid Hemorrhage

2019 ◽  
Vol 124 (12) ◽  
Author(s):  
Alexander Joerk ◽  
Marcel Ritter ◽  
Niklas Langguth ◽  
Raphael Andreas Seidel ◽  
Diana Freitag ◽  
...  
Keyword(s):  
Cerebrospinal Fluid ◽  
Subarachnoid Hemorrhage ◽  
Potassium Channels ◽  
Intrathecal Injection ◽  
Brain Slices ◽  
Differential Interference Contrast Microscopy ◽  
Cerebral Arterioles ◽  
Arteriolar Diameter

Rationale: Delayed ischemic neurological deficit is the most common cause of neurological impairment and unfavorable prognosis in patients with subarachnoid hemorrhage (SAH). Despite the existence of neuroimaging modalities that depict the onset of the accompanying cerebral vasospasm, preventive and therapeutic options are limited and fail to improve outcome owing to an insufficient pathomechanistic understanding of the delayed perfusion deficit. Previous studies have suggested that BOXes (bilirubin oxidation end products), originating from released heme surrounding ruptured blood vessels, are involved in arterial vasoconstriction. Recently, isolated intermediates of oxidative bilirubin degradation, known as PDPs (propentdyopents), have been considered as potential additional effectors in the development of arterial vasoconstriction. Objective: To investigate whether PDPs and BOXes are present in hemorrhagic cerebrospinal fluid and involved in the vasoconstriction of cerebral arterioles. Methods and Results: Via liquid chromatography/mass spectrometry, we measured increased PDP and BOX concentrations in cerebrospinal fluid of SAH patients compared with control subjects. Using differential interference contrast microscopy, we analyzed the vasoactivity of PDP isomers in vitro by monitoring the arteriolar diameter in mouse acute brain slices. We found an arteriolar constriction on application of PDPs in the concentration range that occurs in the cerebrospinal fluid of patients with SAH. By imaging arteriolar diameter changes using 2-photon microscopy in vivo, we demonstrated a short-onset vasoconstriction after intrathecal injection of either PDPs or BOXes. Using magnetic resonance imaging, we observed a long-term PDP-induced delay in cerebral perfusion. For all conditions, the arteriolar narrowing was dependent on functional big conductance potassium channels and was absent in big conductance potassium channels knockout mice. Conclusions: For the first time, we have quantified significantly higher concentrations of PDP and BOX isomers in the cerebrospinal fluid of patients with SAH compared to controls. The vasoconstrictive effect caused by PDPs in vitro and in vivo suggests a hitherto unrecognized pathway contributing to the pathogenesis of delayed ischemic deficit in patients with SAH.

scholarly journals Inversion of neurovascular coupling after subarachnoid hemorrhage in vivo

2017 ◽  
Vol 37 (11) ◽  
pp. 3625-3634 ◽  
Author(s):  
Matilde Balbi ◽  
Masayo Koide ◽  
George C Wellman ◽  
Nikolaus Plesnila
Keyword(s):  
Blood Flow ◽  
Cerebral Blood Flow ◽  
Subarachnoid Hemorrhage ◽  
Neuronal Activity ◽  
Ex Vivo ◽  
Brain Slices ◽  
Neurovascular Coupling ◽  
Vessel Diameter ◽  
Cerebral Arterioles

Subarachnoid hemorrhage (SAH) induces acute changes in the cerebral microcirculation. Recent findings ex vivo suggest neurovascular coupling (NVC), the process that increases cerebral blood flow upon neuronal activity, is also impaired after SAH. The aim of the current study was to investigate whether this occurs also in vivo. C57BL/6 mice were subjected to either sham surgery or SAH by filament perforation. Twenty-four hours later NVC was tested by forepaw stimulation and CO2 reactivity by inhalation of 10% CO2. Vessel diameter was assessed in vivo by two-photon microscopy. NVC was also investigated ex vivo using brain slices. Cerebral arterioles of sham-operated mice dilated to 130% of baseline upon CO2 inhalation or forepaw stimulation and cerebral blood flow (CBF) increased. Following SAH, however, CO2 reactivity was completely lost and the majority of cerebral arterioles showed paradoxical constriction in vivo and ex vivo resulting in a reduced CBF response. As previous results showed intact NVC 3 h after SAH, the current findings indicate that impairment of NVC after cerebral hemorrhage occurs secondarily and is progressive. Since neuronal activity-induced vasoconstriction (inverse NVC) is likely to further aggravate SAH-induced cerebral ischemia and subsequent brain damage, inverse NVC may represent a novel therapeutic target after SAH.

The Role of Purinergic Signaling in Trichomonas vaginalis Infection

2020 ◽  
Vol 20 ◽  
Author(s):  
Micheli Ferla ◽  
Tiana Tasca
Keyword(s):  
Drug Targets ◽  
Trichomonas Vaginalis ◽  
Hiv Transmission ◽  
Cell Damage ◽  
Purinergic Signaling ◽  
Host Cells ◽  
Purine Nucleotides ◽  
Cellular Effects ◽  
Adverse Pregnancy ◽  
Purine Salvage Pathways

: Trichomoniasis, one of the most common non-viral sexually transmitted infections worldwide, is caused by the parasite Trichomonas vaginalis. The pathogen colonizes the human urogenital tract and the infection is associated with complications such as adverse pregnancy outcomes, cervical cancer, and an increase in HIV transmission. The mecha-nisms of pathogenicity are multifactorial, and controlling immune responses is essential for infection maintenance. Extra-cellular purine nucleotides are released by cells in physiological and pathological conditions, and they are hydrolyzed by enzymes called ecto-nucleotidases. The cellular effects of nucleotides and nucleosides occur via binding to purinoceptors, or throughthe uptake by nucleoside transporters. Altogether, enzymes, receptors and transporters constitute the purinergic signaling, a cellular network that regulates several effects in practically all systems including mammals, helminths, proto-zoa, bacteria, and fungi. In this context, this review updates the data on purinergic signaling involved in T. vaginalis biol-ogy and interaction with host cells, focusing on the characterization of ecto-nucleotidases and on purine salvage pathways. The implications of the final products, the nucleosides adenosine and guanosine, for human neutrophil response and vagi-nal epithelial cell damage reveal the purinergic signaling as a potential new mechanism for alternative drug targets.

scholarly journals Minus-end capture of preformed kinetochore fibers contributes to spindle morphogenesis

2003 ◽  
Vol 160 (5) ◽  
pp. 671-683 ◽  
Author(s):  
Alexey Khodjakov ◽  
Lily Copenagle ◽  
Michael B. Gordon ◽  
Duane A. Compton ◽  
Tarun M. Kapoor
Keyword(s):  
Three Dimensional ◽  
Differential Interference Contrast ◽  
Differential Interference Contrast Microscopy ◽  
Spindle Formation ◽  
Capture Mechanism ◽  
Contrast Microscopy ◽  
Interference Contrast Microscopy ◽  
Kinesin Eg5 ◽  
Mitotic Kinesin Eg5 ◽  
Dependent Mechanism

Near-simultaneous three-dimensional fluorescence/differential interference contrast microscopy was used to follow the behavior of microtubules and chromosomes in living α-tubulin/GFP-expressing cells after inhibition of the mitotic kinesin Eg5 with monastrol. Kinetochore fibers (K-fibers) were frequently observed forming in association with chromosomes both during monastrol treatment and after monastrol removal. Surprisingly, these K-fibers were oriented away from, and not directly connected to, centrosomes and incorporated into the spindle by the sliding of their distal ends toward centrosomes via a NuMA-dependent mechanism. Similar preformed K-fibers were also observed during spindle formation in untreated cells. In addition, upon monastrol removal, centrosomes established a transient chromosome-free bipolar array whose orientation specified the axis along which chromosomes segregated. We propose that the capture and incorporation of preformed K-fibers complements the microtubule plus-end capture mechanism and contributes to spindle formation in vertebrates.

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