scholarly journals Transmembrane H+ fluxes and the regulation of neural induction in Xenopus laevis

Zygote ◽  
2021 ◽  
pp. 1-12
Author(s):  
Ho Chi Leung ◽  
Catherine Leclerc ◽  
Marc Moreau ◽  
Alan M. Shipley ◽  
Andrew L. Miller ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Ex Vivo ◽  
Neural Induction ◽  
Dorsal Side ◽  
The Other ◽  
Bafilomycin A1 ◽  
Selective Electrode ◽  
Solvent Control ◽  
Dorsal Ectoderm

Summary It has previously been reported that in ex vivo planar explants prepared from Xenopus laevis embryos, the intracellular pH (pHi) increases in cells of the dorsal ectoderm from stage 10.5 to 11.5 (i.e. 11–12.5 hpf). It was proposed that such increases (potentially due to H+ being extruded, sequestered, or buffered in some manner), play a role in regulating neural induction. Here, we used an extracellular ion-selective electrode to non-invasively measure H+ fluxes at eight locations around the equatorial circumference of intact X. laevis embryos between stages 9–12 (˜7–13.25 hpf). We showed that at stages 9–11, there was a small H+ efflux recorded from all the measuring positions. At stage 12 there was a small, but significant, increase in the efflux of H+ from most locations, but the efflux from the dorsal side of the embryo was significantly greater than from the other positions. Embryos were also treated from stages 9–12 with bafilomycin A1, to block the activity of the ATP-driven H+ pump. By stage 22 (24 hpf), these embryos displayed retarded development, arresting before the end of gastrulation and therefore did not display the usual anterior and neural structures, which were observed in the solvent-control embryos. In addition, expression of the early neural gene, Zic3, was absent in treated embryos compared with the solvent controls. Together, our new in vivo data corroborated and extended the earlier explant-derived report describing changes in pHi that were suggested to play a role during neural induction in X. laevis embryos.

Antimalarial activity of the bisquinoline trans-N1,N2-bis (7-chloroquinolin-4-yl)cyclohexane-1,2-diamine: comparison of two stereoisomers and detailed evaluation of the S,S enantiomer, Ro 47-7737.

1997 ◽  
Vol 41 (3) ◽  
pp. 677-686 ◽  
Author(s):  
R G Ridley ◽  
H Matile ◽  
C Jaquet ◽  
A Dorn ◽  
W Hofheinz ◽  
...  
Keyword(s):  
Antimalarial Activity ◽  
Ex Vivo ◽  
The Other ◽  
Parasite Resistance ◽  
Curative Effect ◽  
Specific Process ◽  
Terminal Half Life ◽  
Detailed Evaluation ◽  
Oral Doses

The S,S enantiomer of the bisquinoline trans-N1,N2-bis(7-chloroquinolin-4-yl)cyclohexane-1,2-diamine, Ro 47-7737, is significantly more potent against chloroquine-resistant Plasmodium falciparum than the R,R enantiomer and the previously described racemate. Both the enantiomers and the racemate are more potent inhibitors of heme polymerization than chloroquine, and their activities are probably mediated by inhibition of this parasite-specific process. The S,S enantiomer, Ro 47-7737, was studied in more detail and proved to be a potent antimalarial in the treatment of P. vivax ex vivo and P. berghei in vivo. Its suppression of P. berghei growth in a mouse model (50% effective dose, 2.3 mg/kg of body weight) was equal to that of chloroquine and mefloquine, and Ro 47-7737 was found to be more potent than these two drugs in the Rane test, in which the curative effect of a single dose is monitored. The dose at which 50% of animals were permanently cured (34 mg/kg) was markedly superior to those of chloroquine (285 mg/kg) and mefloquine (> 250 mg/kg). When administered orally at 50 mg/kg, Ro 47-7737 also showed a faster clearance of parasites than either chloroquine or mefloquine, and unlike the other two compounds, Ro 47-7737 showed no recrudescence. In a study to compare prophylactic efficacies of oral doses of 50 mg/kg, Ro 47-7737 provided protection for 14 days compared to 3 days for mefloquine and 1 day for chloroquine. The good curative and prophylactic properties of the compound can be explained in part by its long terminal half-life. The ability to generate parasite resistance to Ro 47-7737 was also assessed. With a rodent model, resistance could be generated over eight passages. This rate of resistance generation is comparable to that of mefloquine, which has proved to be an effective antimalarial for many years. Toxicity liabilities, however, ruled out this compound as a candidate for drug development.

Histological features of neural induction in Xenopus laevis

Development ◽  
1971 ◽  
Vol 26 (3) ◽  
pp. 543-570
Author(s):  
D. Tarin
Keyword(s):  
Xenopus Laevis ◽  
Neural Induction ◽  
Neural Plate ◽  
Paraxial Mesoderm ◽  
Intestinal Tube ◽  
Spinal Cord Regions ◽  
The Central Nervous System ◽  
Dorsal Ectoderm ◽  
Two Stages ◽  
The Brain

It was first established by grafting experiments that neural induction occurs in Xenopus laevis and that it is the mesoderm in the dorsal lip of the blastopore which normally exercises this function. The subsequent histological work provided the following information: At stage 10½ mesodermal invagination was already well under way, in advance of the formation of the archenteric cavity. This confirms the earlier observations of Nieuwkoop & Florschutz(1950). The first evidence of neural induction, thickening of the mid-dorsal ectoderm combined with the development of an inner tier of columnar cells, occurred at stage 11½. By stage 12 there was generalized thickening of the dorsal ectoderm and between stage 12½ and 13 the brain and spinal cord regions of the neural plate became distinguishable. The dorsal mesoderm segregated into notochord rudiment and two lateral masses at stage 13 and the latter further subdivided into paraxial mesoderm and lateral plates by stage 14. The margins of the neural plate were clearly distinguished from presumptive epidermis by stage 15 and the median neural groove was also well marked. In the next two stages the folding of the neural plate in the line of this groove proceeded rapidly. The dorsoventral enlargement of the somites and the relative shrinkage of the notochord were considered to contribute to the mechanism of neurulation. Regionalization of the brain into prosencephalon, mesencephalon and rhombencephalon was in progress at stages 18 and 19. These results indicate that induction consists of an initial activation of dorsal ectoderm (generalized thickening) followed by gradual transformation of the neural plate to form the different parts of the central nervous system (regionalization). Intercellular metachromatic material was noted in various parts of the embryo. This was most plentiful between stage 10½ and stage 13 and thereafter gradually decreased. It was the only feature which persisted long enough to represent a possible inductive agent. At all stages the archenteron was lined with a continuous layer of endoderm. This indicates that the mode of formation of the gastro-intestinal tube in Xenopus is different to that in urodeles. It further implies that the mesoderm is not present on the blastular surface prior to gastrulation but lies in deeper layers.

The Effect of Dietary Cholesterol on Platelet Survival in the Rabbit – A Study Using 14C-Serotonin and 51Chromium Double-Labelled Platelets

1984 ◽  
Vol 52 (01) ◽  
pp. 085-089 ◽  
Author(s):  
Ian R Wanless
Keyword(s):  
Ex Vivo ◽  
Dietary Cholesterol ◽  
The Other ◽  
Cholesterol Feeding ◽  
Control Groups ◽  
Platelet Survival ◽  
Double Label ◽  
And Control

SummaryHypercholesterolemic men and monkeys have shortened platelet survival but attempts to demonstrate this in rabbits have been unsuccessful. The present study examines platelet survival in cholesterol-fed rabbits. Platelets were double-labelled in vitro with 51Cr and 14C-serotonin or in vivo by intravenous injection of 14C-serotonin.In the double-label experiments 51Cr survival was always shorter than 14C survival but changes in survival of one label were accompanied by similar changes in the other label. Survival was shortened after 4 and 7 weeks of cholesterol feeding. This was demonstrated when the donor platelets were from normal rabbits for both cholesterol and control recipients but no shortening was demonstrated when cholesterol-rich platelets were injected into cholesterol-fed recipients and normal platelets were injected into the control rabbits. When 14C-serotonin was injected intravenously 14C survival was the same as when platelets were labelled ex vivo prior to injection and 14C survival was shortened in rabbits fed cholesterol for 1,2, and 6 weeks.These results indicate that cholesterol-feeding shortens platelet survival. This effect may be masked if the test platelets are not identical in both test and control groups.

Comparative Analysis of Infused “Static”, Ex Vivo-Generated Platelets Vs. Infused Megakaryocytes-Generated Platelets: A Cautionary Tale

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2881-2881
Author(s):  
Yuhuan Wang ◽  
Vincent Hayes ◽  
Danuta Jarocha ◽  
Mortimer Poncz
Keyword(s):  
Stem Cells ◽  
Size Distribution ◽  
Half Life ◽  
Ex Vivo ◽  
The Other ◽  
Human Donor ◽  
Nsg Mice ◽  
Other Hand

Abstract Ex vivo-generated (EV) platelets beginning with embryonic stem cells or induced pluripotent stem cells (iPSCs) or hematopoietic progenitors cells (HPCs) may have clinical utility over donor-derived platelets, and efforts to produce such EV-platelets have been pursued in several laboratories under static megakaryocyte (Meg) culture conditions. Success in generating these has been reported, even demonstrating EV-platelet incorporation into growing thrombi in murine models. We have pursued an alternative strategy for thrombopoiesis using EV-Megs, grown from either human adult HPCs or from iPSCs or fetal livers, and directly infusing them into NOD-SCID gamma-interferon-deficient (NSG) mice. These studies were based on our prior observation that infused murine EV-Megs into wildtype mice are entrapped in the pulmonary bed and over the subsequent 1-4 hours release a wave of functional platelets at a significant level. We now show that infusion of human EV-Megs do the same in NSG mice, but resulting in two different pools of derived platelets: (1) A pool of young (as determined by thiazole orange staining) platelets having the same bell-shaped size distribution was seen as after infusion of human donor-derived platelets in these mice. These platelets take several hours to appear, but then have the same half-life as donor-derived platelets. These platelets are derived from the infused EV-Megs and were termed in vivo-generated (IV)-platelets. (2) A second pool of mostly older platelets was present that originated during the static growth of the EV-Megs, and these EV-platelets varied widely in size and age. Initially, these platelets accounted for a third of all the human platelets seen. Unlike IV-platelets, EV-platelets are immediately present and circulate with a markedly short half-life of 2-3 hours unless the recipient NSG mice were pre-treated with clodronate-ladened liposomes to delete their macrophage pools. Rapid removal of EV-platelets by macrophages is due to their being preactivated as determined by surface P-selectin expression in whole mice blood. These EV-platelets also had very limited further responsiveness to convulxin activation. On the other hand, human IV-platelets were quiescent prior to agonist stimulation in whole mice blood and responded strongly to agonist, similar to human donor-derived platelets infused into NSG mice. The IV-platelets were also selectively incorporated into cremaster arteriole laser injury thrombi over EV-platelets. Finally, directly harvested “platelets” from EV static-grown Megs were isolated and analyzed both in vitro and in vivo. Only a third of these particles are CD41+/CD42+ platelets and approximately half are actually CD41-/CD42-. Both pools showed the same wide size distribution in vitro and in vivo after infusion into mice. The CD41+/CD42+ fraction behaved just as the EV-platelets, but the CD41-/CD42- fraction half-life was unaffected by pre-treatment with clodronate-ladened liposomes. In summary, infused human Megs grown under static growth conditions released platelets in a recipient mouse’s lung with features characteristic of donor-derived platelets. On the other hand, “platelets” harvested from the same Megs were predominantly not even platelets as measured using CD41/CD42 markers. The portion that were CD41+/CD42+ platelets were preactivated, poorly responsive to agonists, and cleared rapidly. These findings set a standard on how to judge the potential clinical value of platelets derived from EV-Megs and also raise concerns whether direct visual imaging of “platelet release” in static culture is biologically meaningful given that most particles released were not CD41+/CD42+ platelets, and the ones that were CD41+/CD42+ were mis-sized and functionally limited. Disclosures No relevant conflicts of interest to declare.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

Antithrombotic Effects and Bleeding Time Prolongation with Synthetic Platelet GPIIb/llla Inhibitors in Animal Models of Platelet-Mediated Thrombosis

1994 ◽  
Vol 71 (01) ◽  
pp. 095-102 ◽  
Author(s):  
Désiré Collen ◽  
Hua Rong Lu ◽  
Jean-Marie Stassen ◽  
Ingrid Vreys ◽  
Tsunehiro Yasuda ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Bleeding Time ◽  
Bolus Injection ◽  
Cell Injury ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Femoral Vein ◽  
Thrombotic Occlusion ◽  
Antithrombotic Effects

SummaryCyclic Arg-Gly-Asp (RGD) containing synthetic peptides such as L-cysteine, N-(mercaptoacetyl)-D-tyrosyl-L-arginylglycyl-L-a-aspartyl-cyclic (1→5)-sulfide, 5-oxide (G4120) and acetyl-L-cysteinyl-L-asparaginyl-L-prolyl-L-arginyl-glycyl-L-α-aspartyl-[0-methyltyrosyl]-L-arginyl-L-cysteinamide, cyclic 1→9-sulfide (TP9201) bind with high affinity to the platelet GPIIb/IIIa receptor.The relationship between antithrombotic effect, ex vivo platelet aggregation and bleeding time prolongation with both agents was studied in hamsters with a standardized femoral vein endothelial cell injury predisposing to platelet-rich mural thrombosis, and in dogs with a carotid arterial eversion graft inserted in the femoral artery. Intravenous administration of G4120 in hamsters inhibited in vivo thrombus formation with a 50% inhibitory bolus dose (ID50) of approximately 20 μg/kg, ex vivo ADP-induccd platelet aggregation with ID50 of 10 μg/kg, and bolus injection of 1 mg/kg prolonged the bleeding time from 38 ± 9 to 1,100 ± 330 s. Administration of TP9201 in hamsters inhibited in vivo thrombus formation with ID50 of 30 μg/kg, ex vivo platelet aggregation with an ID50 of 50 μg/kg and bolus injection of 1 mg/kg did not prolong the template bleeding time. In the dog eversion graft model, infusion of 100 μg/kg of G4120 over 60 min did not fully inhibit platelet-mediated thrombotic occlusion but was associated with inhibition of ADP-induccd ex vivo platelet aggregation and with prolongation of the template bleeding time from 1.3 ± 0.4 to 12 ± 2 min. Infusion of 300 μg/kg of TP9201 over 60 min completely prevented thrombotic occlusion, inhibited ex vivo platelet aggregation, but was not associated with prolongation of the template bleeding time.TP9201, unlike G4120, inhibits in vivo platelet-mediated thrombus formation without associated prolongation of the template bleeding time.

Significance of Platelet β-Adrenoceptors for Platelet Responses In Vivo and In Vitro

1992 ◽  
Vol 68 (06) ◽  
pp. 687-693 ◽  
Author(s):  
P T Larsson ◽  
N H Wallén ◽  
A Martinsson ◽  
N Egberg ◽  
P Hjemdahl
Keyword(s):  
Ex Vivo ◽  
High Dose ◽  
Platelet Aggregability ◽  
Adrenoceptor Agonist ◽  
Dose Infusion ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Factor Antigen

SummaryThe significance of platelet β-adrenoceptors for platelet responses to adrenergic stimuli in vivo and in vitro was studied in healthy volunteers. Low dose infusion of the β-adrenoceptor agonist isoprenaline decreased platelet aggregability in vivo as measured by ex vivo filtragometry. Infusion of adrenaline, a mixed α- and β-adrenoceptor agonist, increased platelet aggregability in vivo markedly, as measured by ex vivo filtragometry and plasma β-thromboglobulin levels. Adrenaline levels were 3–4 nM in venous plasma during infusion. Both adrenaline and high dose isoprenaline elevated plasma von Willebrand factor antigen levels β-Blockade by propranolol did not alter our measures of platelet aggregability at rest or during adrenaline infusions, but inhibited adrenaline-induced increases in vWf:ag. In a model using filtragometry to assess platelet aggregability in whole blood in vitro, propranolol enhanced the proaggregatory actions of 5 nM, but not of 10 nM adrenaline. The present data suggest that β-adrenoceptor stimulation can inhibit platelet function in vivo but that effects of adrenaline at high physiological concentrations are dominated by an α-adrenoceptor mediated proaggregatory action.

Enhancement of ADP-induced Platelet Aggregation by Adrenaline in Vivo and Its Prevention

1973 ◽  
Vol 29 (02) ◽  
pp. 490-498 ◽  
Author(s):  
Hiroh Yamazaki ◽  
Itsuro Kobayashi ◽  
Tadahiro Sano ◽  
Takio Shimamoto
Keyword(s):  
Platelet Count ◽  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
The Other ◽  
Endothelial Surface ◽  
Important Interaction ◽  
Blood Coagulability ◽  
The Difference

SummaryThe authors previously reported a transient decrease in adhesive platelet count and an enhancement of blood coagulability after administration of a small amount of adrenaline (0.1-1 µg per Kg, i. v.) in man and rabbit. In such circumstances, the sensitivity of platelets to aggregation induced by ADP was studied by an optical density method. Five minutes after i. v. injection of 1 µg per Kg of adrenaline in 10 rabbits, intensity of platelet aggregation increased to 115.1 ± 4.9% (mean ± S. E.) by 10∼5 molar, 121.8 ± 7.8% by 3 × 10-6 molar and 129.4 ± 12.8% of the value before the injection by 10”6 molar ADP. The difference was statistically significant (P<0.01-0.05). The above change was not observed in each group of rabbits injected with saline, 1 µg per Kg of 1-noradrenaline or 0.1 and 10 µg per Kg of adrenaline. Also, it was prevented by oral administration of 10 mg per Kg of phenoxybenzamine or propranolol or aspirin or pyridinolcarbamate 3 hours before the challenge. On the other hand, the enhancement of ADP-induced platelet aggregation was not observed in vitro, when 10-5 or 3 × 10-6 molar and 129.4 ± 12.8% of the value before 10∼6 molar ADP was added to citrated platelet rich plasma (CPRP) of rabbit after incubation at 37°C for 30 second with 0.01, 0.1, 1, 10 or 100 µg per ml of adrenaline or noradrenaline. These results suggest an important interaction between endothelial surface and platelets in connection with the enhancement of ADP-induced platelet aggregation by adrenaline in vivo.

In Vivo Effect of Suloctidil as an Antiplatelet Agent

1979 ◽  
Vol 41 (03) ◽  
pp. 465-474 ◽  
Author(s):  
Marcia R Stelzer ◽  
Thomas S Burns ◽  
Robert N Saunders
Keyword(s):  
Ex Vivo ◽  
Antiplatelet Agent ◽  
Ratio Method ◽  
Platelet Aggregate ◽  
Permanent Effect ◽  
Platelet Aggregates ◽  
5 Ht Uptake ◽  
High Doses

SummaryThe relationship between the effects of suloctidil in vivo as an antiplatelet agent and in vitro as a modifier of platelet serotonin (5-HT) parameters was investigated. Suloctidil was found to be effective in reducing platelet aggregates formation in the retired breeder rat as determined using the platelet aggregate ratio method (PAR) with an ED50 of 16.1 mg/kg 24 hours post administration. In contrast to the hypothesis that 5-HT depletion is involved in the anti-aggregatory mechanism of suloctidil, no correlation was found between platelet 5- HT content and this antiplatelet activity. Reduction of platelet 5-HT content required multiple injections of high doses (100 mg/kg/day) of suloctidil. Suloctidil administration for 8 days at 100 mg/kg/day, which lowered platelet 5-HT content by 50%, resulted in no permanent effect on ex vivo platelet 5-HT uptake or thrombin-induced release, nor alteration in the plasma 5-HT level. However, these platelets exhibited a short-lived, significant increase in percent leakage of 5-HT after 30 minutes of incubation. Therefore, suloctidil treatment at high doses may with time result in platelet 5-HT depletion, however this effect is probably not related to the primary anti-aggregatory activity of the drug.

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