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Comparative Biochemistry and... Jun 2022For amphibian species that display external fertilization in an aquatic environment, hypoosmotic shock to sperm cells can quickly result in damage to cellular structure...
For amphibian species that display external fertilization in an aquatic environment, hypoosmotic shock to sperm cells can quickly result in damage to cellular structure and function. This study sought to determine how fertilization media osmolality, temperature, and time impact the stability of the mitochondrial vesicle's association with the sperm head and thus motility and quality of forward progression. The presence of the mitochondrial vesicle and its relationship with sperm motility and quality of forward progression were analyzed in sperm samples from the Fowler's toad (Anaxyrus fowleri) (n = 10) when held for six hours under two temperatures and four osmolalities. Results indicated that the presence of the mitochondrial vesicle is needed for sperm motility over time (p < 0.001), where higher osmolalities (p < 0.001) and lower temperatures (p < 0.001) correlated with maintaining the vesicle attachment to the spermatozoa. The higher osmolality of spermic urine was the most important factor for maintaining higher quality of forward progressive motility (p < 0.01) of spermatozoa. Sperm samples held at 4 °C and 40 mOsm/kg had the longest half-life for motility (540 min) and quality of forward progression (276 min), whereas sperm held at 22 °C and 2.5 mOsm/kg had the shortest half-life for motility (7 min) and quality of forward progression (18 min). Special attention should be given to the osmolality and temperature of fertilization solutions, or breeding tank water, when developing cold storage protocols for toad sperm or reproducing animals to ensure the retention of the mitochondrial vesicle for maximum fertilization capability.
Topics: Animals; Bufonidae; Cryopreservation; Male; Osmolar Concentration; Sperm Motility; Spermatozoa
PubMed: 35321851
DOI: 10.1016/j.cbpa.2022.111191 -
Communicative & Integrative Biology Sep 2010Neurotransmitter release relies on the fusion of synaptic vesicles with the plasma membrane of synaptic boutons, which is followed by the recycling of vesicle components...
Neurotransmitter release relies on the fusion of synaptic vesicles with the plasma membrane of synaptic boutons, which is followed by the recycling of vesicle components and formation of new vesicles. It is not yet clear whether upon fusion the vesicles persist as multimolecular patches in the plasma membrane, or whether they segregate into individual components. Evidence supporting each of these two models has been suggested in recent years. Using diffraction-unlimited imaging (stimulated emission depletion, or STED) of native synaptic vesicle proteins, we have proposed that vesicle proteins remain in clusters on the neuronal surface. These clusters do not appear to intermix. We discuss here these findings in the context of previous studies on synaptic vesicle fusion, and we propose a recycling model which accounts for most of the recent findings on the post-fusion fate of synaptic vesicle components.
PubMed: 21057631
DOI: 10.4161/cib.3.5.12132 -
Langmuir : the ACS Journal of Surfaces... Jul 2015Fusion between suspended lipid vesicles is difficult to achieve without membrane proteins or ions because the vesicles have extremely low equilibrium membrane tension...
Fusion between suspended lipid vesicles is difficult to achieve without membrane proteins or ions because the vesicles have extremely low equilibrium membrane tension and high poration energy. Nonetheless, vesicle fusion in the absence of mediators can also be achieved by mechanical forcing that is strong enough to induce membrane poration. Here, we employ a strong fluid shear stress to achieve vesicle fusion. By utilizing a unique vortex formation phenomenon in branched channels as a platform for capturing, stressing, and fusing the lipid vesicles, we directly visualize using high-speed imaging the vesicle fusion events, induced solely by shear, on the time scale of submilliseconds. We show that a large vesicle with a size of up to ∼10 μm can be achieved by the fusion of nanoscale vesicles. This technique has the potential to be utilized as a fast and simple way to produce giant unilamellar vesicles and to serve as a platform for visualizing vesicle interactions and fusions in the presence of shear.
Topics: Hydrodynamics; Mechanical Phenomena; Stress, Mechanical; Unilamellar Liposomes
PubMed: 26098933
DOI: 10.1021/acs.langmuir.5b01752 -
Acta Urologica Belgica Jun 1997Ejaculatory duct(s) obstruction(s) (EDO) may be responsible for as much as one third of azoospermia- or severe oligospermia-related infertility; it's clinical... (Review)
Review
Ejaculatory duct(s) obstruction(s) (EDO) may be responsible for as much as one third of azoospermia- or severe oligospermia-related infertility; it's clinical presentation also includes some low urinary tract irritative symptoms, such as repeated epididymitis, pelvi-perineal pain, hematospermia and other ejaculatory disturbances. The diagnosis of EDO is based on patient's history, semen analysis (hypospermia, azoospermia, low fructose level), and transrectal ultrasound (TRUS), which can demonstrate seminal vesicle(s), vas ampulla(s) and/or ejaculatory duct(s) dilatation, Müllerian or utricular cyst, and ejaculatory duct(s) or seminal calcification(s). Confirmation of the suspected diagnosis, if needed, requires classical vasography or TRUS-guided seminal tract puncture and vesiculography. Treatment is usually successfully achieved with transurethral endoscopic procedures: retrograde ejaculatory duct(s) catheterisation, dilatation, incision or resection; seminal tract endoscopy is seldom performed. Very few complications occur; evaluation of long term results is lacking. Indications of such endoscopic procedures remain to be defined, especially in cases of partial EDO.
Topics: Blood; Calcinosis; Catheterization; Cysts; Dilatation; Dilatation, Pathologic; Ejaculatory Ducts; Endoscopy; Epididymitis; Genital Diseases, Male; Humans; Infertility, Male; Male; Medical History Taking; Oligospermia; Pain; Prostatic Diseases; Radiography; Semen; Seminal Vesicles; Testicular Diseases; Ultrasonography; Vas Deferens
PubMed: 9324904
DOI: No ID Found -
Materials (Basel, Switzerland) Dec 2021The spontaneous formation and fusion of raspberry vesicles was studied using the dissipative particle dynamics (DPD) method. The vesicles were formed through the...
The spontaneous formation and fusion of raspberry vesicles was studied using the dissipative particle dynamics (DPD) method. The vesicles were formed through the self-assembly of amphiphilic EOF star terpolymers in selective solvent. E and F blocks are solvophobic and the O block is solvophilic. The shortest F block plays a major role in the formation of raspberry vesicles. Distinct vesicle formation mechanisms were observed at different polymer concentrations. At higher concentrations, vesicles form via the bending and closure of an oblate F-bump-E bilayer. At lower concentrations, the formation pathway contains: the initial formation of a vesicle with a core, the combination of such vesicles into cylindrical micelles, and the bending of the cylindrical micelles to form a hollow vesicle. In addition, raspberry vesicle fusion is regulated by F bumps through the continuous coalescence of them from apposed vesicle membranes. The contact area bends, followed by the formation of a fusion pore and a tilted inner layer. As the pore sealed, the hemifusion structure appears, which further restructures to form a vesicle. Our results provide guidance on understanding the dynamic processes of complex vesicles and biological membrane fusion.
PubMed: 34947284
DOI: 10.3390/ma14247690 -
PloS One 2015Chromaffin cells release catecholamines by exocytosis, a process that includes vesicle docking, priming and fusion. Although all these steps have been intensively...
Chromaffin cells release catecholamines by exocytosis, a process that includes vesicle docking, priming and fusion. Although all these steps have been intensively studied, some aspects of their mechanisms, particularly those regarding vesicle transport to the active sites situated at the membrane, are still unclear. In this work, we show that it is possible to extract information on vesicle motion in Chromaffin cells from the combination of Langevin simulations and amperometric measurements. We developed a numerical model based on Langevin simulations of vesicle motion towards the cell membrane and on the statistical analysis of vesicle arrival times. We also performed amperometric experiments in bovine-adrenal Chromaffin cells under Ba2+ stimulation to capture neurotransmitter releases during sustained exocytosis. In the sustained phase, each amperometric peak can be related to a single release from a new vesicle arriving at the active site. The amperometric signal can then be mapped into a spike-series of release events. We normalized the spike-series resulting from the current peaks using a time-rescaling transformation, thus making signals coming from different cells comparable. We discuss why the obtained spike-series may contain information about the motion of all vesicles leading to release of catecholamines. We show that the release statistics in our experiments considerably deviate from Poisson processes. Moreover, the interspike-time probability is reasonably well described by two-parameter gamma distributions. In order to interpret this result we computed the vesicles' arrival statistics from our Langevin simulations. As expected, assuming purely diffusive vesicle motion we obtain Poisson statistics. However, if we assume that all vesicles are guided toward the membrane by an attractive harmonic potential, simulations also lead to gamma distributions of the interspike-time probability, in remarkably good agreement with experiment. We also show that including the fusion-time statistics in our model does not produce any significant changes on the results. These findings indicate that the motion of the whole ensemble of vesicles towards the membrane is directed and reflected in the amperometric signals. Our results confirm the conclusions of previous imaging studies performed on single vesicles that vesicles' motion underneath plasma membranes is not purely random, but biased towards the membrane.
Topics: Algorithms; Animals; Biological Transport; Catecholamines; Cattle; Chromaffin Cells; Computer Simulation; Electrophysiological Phenomena; Exocytosis; Models, Biological; Models, Statistical; Transport Vesicles
PubMed: 26675312
DOI: 10.1371/journal.pone.0144045 -
ACS Measurement Science Au Dec 2021In this work, we introduce a novel method for visualization and quantitative measurement of the vesicle opening process by correlation of vesicle impact electrochemical...
In this work, we introduce a novel method for visualization and quantitative measurement of the vesicle opening process by correlation of vesicle impact electrochemical cytometry (VIEC) with confocal microscopy. We have used a fluorophore conjugated to lipids to label the vesicle membrane and manipulate the membrane properties, which appears to make the membrane more susceptible to electroporation. The neurotransmitters inside the vesicles were visualized by use of a fluorescence false neurotransmitter 511 (FFN 511) through accumulation inside the vesicle via the neuronal vesicular monoamine transporter 2 (VMAT 2). Optical and electrochemical measurements of single vesicle electroporation were carried out using an in-house, disk-shaped, gold-modified ITO (Au/ITO) microelectrode device (5 nm thick, 33 μm diameter), which simultaneously acted as an electrode surface for VIEC and an optically transparent surface for confocal microscopy. As a result, the processes of adsorption, electroporation, and opening of single vesicles followed by neurotransmitter release on the Au/ITO surface have been simultaneously visualized and measured. Three opening patterns of single isolated vesicles were frequently observed. Comparing the vesicle opening patterns with their corresponding VIEC spikes, we propose that the behavior of the vesicular membrane on the electrode surface, including the adsorption time, residence time before vesicle opening, and the retention time after vesicle opening, are closely related to the vesicle content and size. Large vesicles with high content tend to adsorb to the electrode faster with higher frequency, followed by a shorter residence time before releasing their content, and their membrane remains on the electrode surface longer compared to the small vesicles with low content. With this approach, we start to unravel the vesicle opening process and to examine the fundamentals of exocytosis, supporting the proposed mechanism of partial or subquantal release in exocytosis.
PubMed: 34939075
DOI: 10.1021/acsmeasuresciau.1c00021 -
Microbiology Spectrum Mar 2023Outer membrane vesicles (OMVs) are a primary means of communication for Gram-negative bacteria. The specific role of vesicle components in cellular communication and how...
Outer membrane vesicles (OMVs) are a primary means of communication for Gram-negative bacteria. The specific role of vesicle components in cellular communication and how components are packaged are still under investigation, but a correlation exists between OMV biogenesis and content. The two primary mechanisms of OMV biogenesis are membrane blebbing and explosive cell lysis, and vesicle content is based on the biogenesis mechanism. Hypervesiculation, which can be induced by stress conditions, also influences OMV content. Norovirus interaction with Enterobacter cloacae induces stress responses leading to increased OMV production and changes in DNA content, protein content, and vesicle size. The presence of genomic DNA and cytoplasmic proteins in these OMVs suggests some of the vesicles are formed by explosive cell lysis, so reduction or loss of these components indicates a shift away from this mechanism of biogenesis. Based on this, further investigation into bacterial stability and OMV content was conducted. Results showed that norovirus induced a dramatic shift in OMV lipid content. Specifically, the increased accumulation of phospholipids is associated with increased blebbing, thereby supporting previous observations that noroviruses shift the mechanism of OMV biogenesis. Slight differences in OMV metabolite content were also observed. While norovirus induced changes in OMV content, it did not change the lipid content of the bacterial outer membrane or the metabolite content of the bacterial cell. Overall, these results indicate that norovirus induces significant changes to OMV lipid architecture and cargo, which may be linked to a change in the mechanism of vesicle biogenesis. Extracellular vesicles from commensal bacteria are recognized for their importance in modulating host immune responses, and vesicle content is related to their impact on the host. Therefore, understanding how vesicles are formed and how their content shifts in response to stress conditions is necessary for elucidating their downstream functions. Our recent work has demonstrated that interactions between noroviruses and Enterobacter cloacae induce bacterial stress responses leading to hypervesiculation. In this article, we characterize and compare the lipid and metabolomic cargo of E. cloacae vesicles generated in the presence and absence of norovirus and show that viral interactions induce significant changes in vesicle content. Furthermore, we probe how these changes and changes to the bacterial cell may be indicative of a shift in the mechanism of vesicle biogenesis. Importantly, we find that noroviruses induce significant changes in vesicle lipid architecture and cargo that may be responsible for the immunogenic activity of these vesicles.
PubMed: 36943087
DOI: 10.1128/spectrum.04691-22 -
Human Molecular Genetics Apr 2014Mutations in the dysferlin gene resulting in dysferlin-deficiency lead to limb-girdle muscular dystrophy 2B and Myoshi myopathy in humans. Dysferlin has been proposed as...
Mutations in the dysferlin gene resulting in dysferlin-deficiency lead to limb-girdle muscular dystrophy 2B and Myoshi myopathy in humans. Dysferlin has been proposed as a critical regulator of vesicle-mediated membrane resealing in muscle fibers, and localizes to muscle fiber wounds following sarcolemma damage. Studies in fibroblasts and urchin eggs suggest that trafficking and fusion of intracellular vesicles with the plasma membrane during resealing requires the intracellular cytoskeleton. However, the contribution of dysferlin-containing vesicles to resealing in muscle and the role of the cytoskeleton in regulating dysferlin-containing vesicle biology is unclear. Here, we use live-cell imaging to examine the behavior of dysferlin-containing vesicles following cellular wounding in muscle cells and examine the role of microtubules and kinesin in dysferlin-containing vesicle behavior following wounding. Our data indicate that dysferlin-containing vesicles move along microtubules via the kinesin motor KIF5B in muscle cells. Membrane wounding induces dysferlin-containing vesicle-vesicle fusion and the formation of extremely large cytoplasmic vesicles, and this response depends on both microtubules and functional KIF5B. In non-muscle cell types, lysosomes are critical mediators of membrane resealing, and our data indicate that dysferlin-containing vesicles are capable of fusing with lysosomes following wounding which may contribute to formation of large wound sealing vesicles in muscle cells. Overall, our data provide mechanistic evidence that microtubule-based transport of dysferlin-containing vesicles may be critical for resealing, and highlight a critical role for dysferlin-containing vesicle-vesicle and vesicle-organelle fusion in response to wounding in muscle cells.
Topics: Animals; Cell Line; Cell Membrane; Cytoplasmic Vesicles; Green Fluorescent Proteins; Heterocyclic Compounds, 4 or More Rings; Kinesins; Lysosomes; Membrane Fusion; Membrane Proteins; Microtubules; Muscle Cells; Muscle Fibers, Skeletal; Muscle Proteins; Muscle, Skeletal; Muscular Dystrophies, Limb-Girdle; Myosin Type II; Nocodazole; Rats; Tubulin Modulators
PubMed: 24203699
DOI: 10.1093/hmg/ddt557 -
Physical Review. E, Statistical,... Jun 2012Dynamics of a vesicle under simple shear flow is studied in the limit of small capillary number. A perturbative approach is used to derive the equation of vesicle...
Dynamics of a vesicle under simple shear flow is studied in the limit of small capillary number. A perturbative approach is used to derive the equation of vesicle dynamics. The expansions are shown to converge for significantly deflated vesicles (with excess area from the sphere as high as 2). In particular, we provide an explicit analytical expression for the tank-treading to tumbling bifurcation point. This expression is valid for excess areas up to 2.5. The results are compared with full 3D numerical simulations. The proposed method can be used for analytical or numerical solution of vesicle dynamics under weak flow of general form.
Topics: Computer Simulation; Microfluidics; Models, Chemical; Models, Molecular; Stress, Mechanical; Transport Vesicles
PubMed: 23005142
DOI: 10.1103/PhysRevE.85.061922