Hypothermic storage in salt-free preservative solution alter motility duration in sterlet sperm

2021 ◽  
pp. 64-79
Author(s):  
Dmitry Alexandrovich Isaev ◽  
Alexander Pavlovich Glebov ◽  
Marina Yurievna Martynova ◽  
Elena Ivanovna Shishanova
Keyword(s):  
Cell Death ◽  
Sperm Motility ◽  
Physiological State ◽  
Genetic Research ◽  
Progressive Decrease ◽  
Initial Value ◽  
Preservative Solution ◽  
Hypothermic Storage ◽  
Gametic Selection ◽  
Atp Supply

Together with concentration, motility is one of the most important characteristics of sturgeon sperm, determining its quality and suitability for insemination. After activation in water, the duration of progressive sperm motility is also important, and this time should not be less than that required for fertilization. Motility of spermatozoa depends on their physiological state, maturity, age and intracellular reserves of macroergic substances. During hypothermic storage, the percentage of spermatozoa that can be activated decreases progressively due to depletion of ATP supply or cell death. To improve the hypothermic storage of sterlet sperm, we have developed salt-free preservative solution ISGT-80 based on glucose and trehalose. During storage of sterlet sperm specimens from 20 males in ISGT-80 for 18 days, we observed, along with a progressive decrease in the percentage of motile spermatozoa, an alteration in the duration of their motility. On the 3rd to 6th day of storage, the time of half-loss of motility (τ50) increased significantly by approximately 1 min on average compared with fresh samples, then gradually decreased, however, not descending to the initial value. The reasons for this prolongation of motility are not clear, but we do not exclude the predominant death of spermatozoa with a short motility duration in the first days of storage and selection in favor of long-lived spermatozoa. Such gametic selection can lead to a shift in allele frequencies at heterozygous loci in the offspring. Thus, hypothermic storage of sperm could become an attractive subject for genetic research with the aim of developing new selection tools in sturgeon breeding.

scholarly journals Prolonged high intracellular free calcium concentrations induced by ATP are not immediately cytotoxic in isolated rat hepatocytes. Changes in biochemical parameters implicated in cell toxicity

1989 ◽  
Vol 263 (2) ◽  
pp. 347-353 ◽  
Author(s):  
J F Nagelkerke ◽  
P Dogterom ◽  
H J G M De Bont ◽  
G J Mulder
Keyword(s):  
Cell Death ◽  
Rat Hepatocytes ◽  
Free Calcium ◽  
Cell Toxicity ◽  
Protein Thiol ◽  
Initial Value ◽  
Isolated Rat Hepatocytes ◽  
Control Value ◽  
Mitochondrial Functions ◽  
Isolated Rat

Isolated rat hepatocytes were incubated with ATP to induce high intracellular free Ca2+ concentrations as determined with the Quin-2 method. Immediately after addition of ATP, the intracellular concentration of Ca2+ rose from 200 nM to more than 2.5 microM. It stayed at this value during the first 1/2 h; the rise was absolutely dependent on extracellular Ca2+. After the first 1/2 h the Ca2+ concentration decreased to 1-2 microM (5-10 times the control value). These high intracellular free Ca2+ concentrations did not lead to an immediate loss of cell viability. Only after 2 h of incubation a substantial number of cells lost viability. This was preceded by a decrease in cellular NADH (greater than 40%) and accompanied by a sharp increase in the intracellular Ca2+ concentration. Under these conditions the NADPH concentration was not affected. Cellular GSH was decreased to 30% of the initial value, but no lipid peroxidation or protein-thiol depletion was observed. Intracellular ATP, ADP and AMP were increased in the presence of extracellular ATP. Ca2+-dependent proteases seemed not to be involved in cell death. These observations are consistent with a collapse of mitochondrial functions as a final trigger of cell death.

scholarly journals The inhibition of pyruvate transport across the plasma membrane of the bloodstream form of Trypanosoma brucei and its metabolic implications

1995 ◽  
Vol 312 (2) ◽  
pp. 479-484 ◽  
Author(s):  
E A C Wiemer ◽  
P A M Michels ◽  
F R Opperdoes
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Trypanosoma Brucei ◽  
Physiological State ◽  
Bloodstream Form ◽  
Eukaryotic Cells ◽  
Monocarboxylate Transporters ◽  
Facilitated Diffusion ◽  
Selective Inhibitors

The pyruvate produced by glycolysis in the bloodstream form of the trypanosome is excreted into the host bloodstream by a facilitated diffusion carrier. The sensitivity of pyruvate transport for alpha-cyano-4-hydroxycinnamate and the compound UK5099 [alpha-cyano-beta-(1-phenylindol-3-yl)acrylate], which are known to be selective inhibitors of pyruvate (monocarboxylate) transporters present in mitochondria and the plasma membrane of eukaryotic cells, was examined. The trypanosomal pyruvate carrier was found to be rather insensitive to inhibition by alpha-cyano-4-hydroxycinnamate (Ki = 17 mM) but could be completely blocked by UK5099 (Ki = 49 microM). Inhibition of pyruvate transport resulted in the retention, and concomitant accumulation, of pyruvate within the trypanosomes, causing acidification of the cytosol and osmotic destabilization of the cells. Our results indicate that this physiological state has serious metabolic consequences and ultimately leads to cell death; thereby identifying the pyruvate carrier as a possible target for chemotherapeutic intervention.

Factors influencing the Interaction of insecticidal Mists on flying Insects

1946 ◽  
Vol 37 (2) ◽  
pp. 177-190 ◽  
Author(s):  
W. A. L. David ◽  
P. Bracey
Keyword(s):  
Blood Meal ◽  
Maximum Dose ◽  
Physiological State ◽  
Progressive Decrease ◽  
Relative Resistance ◽  
Male And Female ◽  
Practical Applications ◽  
Flying Insects ◽  
Factors Influencing ◽  
Spray Droplets

Experiments are described which illustrate some of the ways in which the results of exposing insects to a spray mist are dependent on either their behaviour or physiological state. Only fine mist dispersals with particles below 10 microns in diameter are considered and in such mists the flight movements of the insects are very largely responsible for the collisions between the insect and the droplets. An increase in flight activity leads to the accumulation of a greater number of spray droplets and consequently a higher kill. Under these conditions insecticides such as pyrethrins which exert a strong irritating action on Aëdes in minimal amounts are superior to DDT which does not have this effect. For practical applications under circumstances where the sprays depend on mist action to produce their effect it is advantageous to mix pyrethrum with the DDT sprays so that the maximum dose of the latter may be accumulated.Further experiments are concerned with the relative resistance of male and female Aëdes aegypti to insecticides and the change of resistance which takes place on ageing or as a result of a blood meal. It is shown that the male is innately more susceptible to both pyrethrins and DDT and that the resistance of the females is higher after a blood meal. There is also a progressive decrease in resistance with age.

Capacitation mechanisms, and the role of capacitation as seen in eutherian mammals

1996 ◽  
Vol 8 (4) ◽  
pp. 581 ◽  
Author(s):  
RA Harrison
Keyword(s):  
Cell Death ◽  
Reproductive Tract ◽  
Physiological State ◽  
Hormonal Control ◽  
Female Reproductive Tract ◽  
Functional Heterogeneity ◽  
Vitro Fertilization ◽  
Range Of Functions

Capacitation, the process whereby spermatozoa are rendered capable of interacting with and fertilizing the egg, was discovered more than 40 years ago. However, our understanding of it is still far from satisfactory. Several factors conspire to obfuscate studies of capacitation mechanisms: the inherent functional heterogeneity of sperm populations, the range of functions used as parameters of capacitation (whence the endpoint of the process has become conceptually uncertain), and the several profound differences between model in vitro fertilization (IVF) systems and the situation in vivo in the female reproductive tract. Recent investigations in the author's laboratory have shown that bicarbonate/CO2, an essential component for successful IVF, causes rapid changes in lipid architecture of the sperm plasma membrane and slower changes in surface coating. These changes are accompanied by membrane destabilization and cell death. Evidence suggests that bicarbonate's actions are mediated through cyclic nucleotide signalling. Of particular note is the heterogeneity in rate of response to bicarbonate shown by individual cells in the sperm populations. Taken together with other observations, the findings suggest that capacitation is a series of positive destabilizing events that eventually lead to cell death. The 'capacitated' state would then be a window of destabilization within which spermatozoa can undergo a zona-induced acrosome reaction and display hyperactivated motility. Further along the destabilization pathway, spontaneous acrosome reactions would occur before total membrane degeneration. In vivo, capacitation would be a conflict between destabilization and sperm survival. Concentrations of bicarbonate are maintained low in the cauda epididymidis, where sperm survive for long periods, and one may speculate that hormonal control of local bicarbonate/CO2 in oviducal 'storage' sites in the female tract could allow 'safe' sequestering of live spermatozoa until around the time of ovulation; the environment may then change to produce a 'capacitating' effect, whence, due to the inherent functional heterogeneity of the sequestered population, small numbers of capacitated spermatozoa are released sequentially. In this way, a succession of spermatozoa in the correct physiological state may be provided for the freshly ovulated egg.

scholarly journals Activation of metabolic and stress responses during subtoxic expression of the type I toxin hok in Erwinia amylovora

2021 ◽  
Author(s):  
Jingyu Peng ◽  
Lindsay R. Triplett ◽  
George Sundin
Keyword(s):  
Cell Death ◽  
Stress Responses ◽  
Erwinia Amylovora ◽  
Physiological State ◽  
Fruit Trees ◽  
Toxin Gene ◽  
Type I ◽  
Pome Fruit ◽  
Membrane Rupture ◽  
High Level

Abstract Background: Toxin-antitoxin (TA) systems, abundant in prokaryotes, are composed of a toxin gene and its cognate antitoxin. Several toxins are implied to affect the physiological state and stress tolerance of bacteria in a population. We previously identified a chromosomally encoded hok-sok type I TA system in Erwinia amylovora, the causative agent of fire blight disease on pome fruit trees. A high-level induction of the hok gene was lethal to E. amylovora cells through unknown mechanisms. The molecular targets or regulatory roles of Hok were unknown.Results: Here, we examined the physiological and transcriptomic changes of Erwinia amylovora cells expressing hok at subtoxic levels that were confirmed to confer no cell death, and at toxic levels that resulted in killing of cells. In both conditions, hok caused membrane rupture and collapse of the proton motive force in a subpopulation of E. amylovora cells. We demonstrated that induction of hok resulted in upregulation of ATP biosynthesis genes, and caused leakage of ATP from cells only at toxic levels. We showed that overexpression of the phage shock protein gene pspA largely reversed the cell death phenotype caused by high levels of hok induction. We also showed that induction of hok at a subtoxic level rendered a greater proportion of stationary phase E. amylovora cells tolerant to the antibiotic streptomycin. Conclusions: We characterized the molecular mechanism of toxicity by high-level of hok induction and demonstrated that low-level expression of hok primes the stress responses of E. amylovora against further membrane and antibiotic stressors.

scholarly journals Effects of Decalcifying Agents of Variable Duration on PD-L1 Immunohistochemistry

2019 ◽  
Vol 153 (2) ◽  
pp. 258-265 ◽  
Author(s):  
Amanda L Strickland ◽  
Sara Blacketer ◽  
Kyle Molberg ◽  
John Markantonis ◽  
Elena Lucas
Keyword(s):  
Cell Death ◽  
Programmed Cell Death ◽  
Formic Acid ◽  
Staining Intensity ◽  
Progressive Decrease ◽  
Progressive Decline ◽  
Variable Duration ◽  
Formalin Fixed ◽  
All Treatment

Abstract Objectives To evaluate the effects of decalcifying agents on programmed cell death ligand 1 (PD-L1) immunohistochemistry (IHC). Methods Fragments of 10 placentas (high PD-L1 expressor) and 10 lungs (lower PD-L1 expressor) were formalin-fixed and subjected to four decalcifying solutions (EDTA, formic acid/MasterCal IM Plus [FA/MC], 12% HCl, and Decal STAT/23% HCl) for 1, 2, 6, or 24 hours. H&E staining and PD-L1 using IHC 22C3 pharmDx were performed, and PD-L1 staining was assessed. Results Minimal to no change in staining intensity or proportion of stained cells was seen with EDTA or FA/MC at all decalcifying durations. Both HCl-based decalcifiers demonstrated a progressive decrease in percentage of positive cells and staining intensity with longer decalcifying duration, particularly with Decal STAT. Conclusions EDTA and FA/MC have little effect on PD-L1 expression. 12% HCl causes a progressive decline in staining. Decal STAT dramatically reduced staining with all treatment durations, especially at 24 hours.

scholarly journals Activation of metabolic and stress responses during subtoxic expression of the type I toxin hok in Erwinia amylovora

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Jingyu Peng ◽  
Lindsay R. Triplett ◽  
George W. Sundin
Keyword(s):  
Cell Death ◽  
Stress Responses ◽  
Erwinia Amylovora ◽  
Physiological State ◽  
Fruit Trees ◽  
Toxin Gene ◽  
Type I ◽  
Pome Fruit ◽  
Membrane Rupture ◽  
High Level

Abstract Background Toxin-antitoxin (TA) systems, abundant in prokaryotes, are composed of a toxin gene and its cognate antitoxin. Several toxins are implied to affect the physiological state and stress tolerance of bacteria in a population. We previously identified a chromosomally encoded hok-sok type I TA system in Erwinia amylovora, the causative agent of fire blight disease on pome fruit trees. A high-level induction of the hok gene was lethal to E. amylovora cells through unknown mechanisms. The molecular targets or regulatory roles of Hok were unknown. Results Here, we examined the physiological and transcriptomic changes of Erwinia amylovora cells expressing hok at subtoxic levels that were confirmed to confer no cell death, and at toxic levels that resulted in killing of cells. In both conditions, hok caused membrane rupture and collapse of the proton motive force in a subpopulation of E. amylovora cells. We demonstrated that induction of hok resulted in upregulation of ATP biosynthesis genes, and caused leakage of ATP from cells only at toxic levels. We showed that overexpression of the phage shock protein gene pspA largely reversed the cell death phenotype caused by high levels of hok induction. We also showed that induction of hok at a subtoxic level rendered a greater proportion of stationary phase E. amylovora cells tolerant to the antibiotic streptomycin. Conclusions We characterized the molecular mechanism of toxicity by high-level of hok induction and demonstrated that low-level expression of hok primes the stress responses of E. amylovora against further membrane and antibiotic stressors.

scholarly journals Vaginal lubricants in the couple trying-to-conceive: assessing healthcare professional recommendations and effect on in vitro sperm function

2018 ◽  
Author(s):  
Scott C Mackenzie ◽  
Steven A Gellatly
Keyword(s):  
Cell Death ◽  
Healthcare Professional ◽  
Sperm Motility ◽  
Online Survey ◽  
Healthcare Professionals ◽  
Sperm Function ◽  
Clinical Guidance ◽  
Progressive Motility ◽  
Vaginal Lubricants

Vaginal lubricants are commonly used by couples trying-to-conceive. However, most vaginal lubricants are sperm toxic and therefore should not be used by couples trying-to-conceive. Despite this, lubricant sperm toxicity is insufficiently reported and guidance for healthcare professionals (HCPs) are absent. In this study, lubricant-related practices of fertility-based HCPs in Scotland were sampled via an online survey. Lubricants identified as being utilised in the fertility setting were subsequently incubated with prepared sperm samples to establish effects on sperm motility. HCP recommendations (n=32) on lubricant use were varied although knowledge related to sperm toxicity was generally poor. HCPs infrequently asked about lubricant use and were unaware of guidance in this area. Aquagel, the only prescribed lubricant identified in this study, reduced sperm progressive motility to 49% of control after 10 minutes, even at concentrations as low as 5%. Vitality testing suggested the deterioration in progressive motility with Aquagel was not as a result of cell death. Conversely, Pré Vaginal Lubricant, a ‘sperm-safe’ lubricant, did not significantly affect any markers of sperm function assessed. Development of clinical guidance in this area is recommended to ensure HCPs deliver informed advice as lubricant use in couples trying-to-conceive may inadvertently contribute to delay in conception.

227 PHOSPHOINOSITIDE-3-KINASE SIGNALING AND THE CONTROL OF SPERM MOTILITY IN THE PORCINE OVIDUCT

2008 ◽  
Vol 20 (1) ◽  
pp. 193 ◽  
Author(s):  
N. Satake ◽  
P. F. Watson ◽  
W. V. Holt
Keyword(s):  
Signaling Pathway ◽  
Sperm Motility ◽  
Intracellular Signaling ◽  
Physiological State ◽  
Apical Plasma Membrane ◽  
Mitochondrial Activity ◽  
Intracellular Signaling Pathway ◽  
Pi3k Inhibitors ◽  
Phosphoinositide 3 Kinase ◽  
Boar Spermatozoa

Prior to ovulation, interactions of spermatozoa with the oviductal epithelium lead to the formation of a sperm reservoir; here we investigate the hypothesis that the physiological state of the stored spermatozoa is modulated by the epithelial cells. In the pig, where modulation of sperm motility is sensitive to environmental components, the high bicarbonate content of the oviductal fluid would be expected to stimulate motility to its maximal extent; however, as this is incompatible with reservoir formation, we postulate that a signaling pathway counteracts bicarbonate stimulation and depresses motility (Satake et al. 2006 J. Exp. Biol. 209, 1560–1572). We chose to investigate a signaling mechanism which may suppress sperm motility through phosphoinositide-3-kinase (PI3K). Boar spermatozoa were exposed to solubilized proteins derived from the oviduct epithelial apical plasma membrane (sAPM). We exploited the motility activation effects of bicarbonate on Percoll-washed boar spermatozoa to see whether it was countered by sAPM and reversed by PI3K inhibitors. Washed spermatozoa (n = 9 boars) were incubated (10 min at 38�C) in Tyrode's medium in the presence of sAPM (25 and 50 µg mL–1) or PI3K inhibitors (LY294002 or wortmannin) prior to activation with 15 mm bicarbonate/CO2, and then incubated for a further 35 min. Sperm motility was monitored at 5-min intervals using a Hobson Sperm Tracker (Hobson Tracker, Ltd., Sheffield, UK). Samples were also analyzed with a flow cytometer for mitochondrial activity using JC-1. PATN analysis (Belbin 1993 PATN Pattern Analysis Package, Div. Wildlife & Ecol., CSIRO, Canberra, Australia) was used to distinguish four motility-based sperm subgroups within the data. The proportions of fast and progressive spermatozoa showed significant and rapid (<5 min) bicarbonate-induced increases when there was no sAPM treatment, or when PI3K inhibitors were present. However, there was no significant bicarbonate-induced increase in the presence of sAPM alone, or when spermatozoa were exposed to a mixture of wortmannin and sAPM. PI3K inhibition by LY294002 reversed the sAPM-induced motility suppression (P < 0.001). Mitochondrial membrane potential was also suppressed by sAPM and restored by the presence of LY294002 (P < 0.01). These results support the hypothesis that the sperm–oviduct interaction is modulated by an intracellular signaling pathway involving PI3K and appropriate receptors.

scholarly journals The cytosolic concentration of phosphate determines the maximal rate of glycogenolysis in perfused rat liver

1990 ◽  
Vol 266 (1) ◽  
pp. 207-212 ◽  
Author(s):  
F Vanstapel ◽  
M Waebens ◽  
P Van Hecke ◽  
C Decanniere ◽  
W Stalmans
Keyword(s):  
Cyclic Amp ◽  
Rat Liver ◽  
Glycogen Phosphorylase ◽  
Perfused Rat Liver ◽  
Maximal Rate ◽  
Free Medium ◽  
Progressive Decrease ◽  
Dibutyryl Cyclic Amp ◽  
Initial Value ◽  
Contrast Exposure

Glycogenolysis was studied in glycogen-rich perfused livers in which glycogen phosphorylase was fully converted into the a form by exposure of the livers to dibutyryl cyclic AMP. We monitored intracellular Pi by 31P n.m.r. Perfusion with Pi-free medium during 30 min caused a progressive decrease of the Pi signal to 50% of its initial value. In contrast, exposure of the livers to KCN and/or 2,4-dinitrophenol resulted in a rapid doubling of the Pi signal. Alterations in the intracellular Pi coincided with proportional changes in the rate of hepatic glycogenolysis (measured as the output of glucose plus lactate). The results indicate that the rate of glycogenolysis catalysed by phosphorylase a depends linearly on the hepatic Pi concentration. Hence the Km of phosphorylase a for its substrate Pi must be considerably higher than the concentrations that occur in the cytosol, even during hypoxia.

Sign in / Sign up

Export Citation Format

Share Document