scholarly journals A Kinetic Analysis of Calcium-Triggered Exocytosis

2001 ◽  
Vol 118 (2) ◽  
pp. 145-156 ◽  
Author(s):  
Paul S. Blank ◽  
Steven S. Vogel ◽  
James D. Malley ◽  
Joshua Zimmerberg
Keyword(s):  
Function Analysis ◽  
Secretory Vesicle ◽  
Free Calcium ◽  
Vesicle Docking ◽  
Calcium Dependence ◽  
Calcium Dependent ◽  
Sea Urchin Egg ◽  
Vesicle Exocytosis ◽  
Free Calcium Concentration

Although the relationship between exocytosis and calcium is fundamental both to synaptic and nonneuronal secretory function, analysis is problematic because of the temporal and spatial properties of calcium, and the fact that vesicle transport, priming, retrieval, and recycling are coupled. By analyzing the kinetics of sea urchin egg secretory vesicle exocytosis in vitro, the final steps of exocytosis are resolved. These steps are modeled as a three-state system: activated, committed, and fused, where interstate transitions are given by the probabilities that an active fusion complex commits (α) and that a committed fusion complex results in fusion, p. The number of committed complexes per vesicle docking site is Poisson distributed with mean n. Experimentally, p and n increase with increasing calcium, whereas α and the pn ratio remain constant, reducing the kinetic description to only one calcium-dependent, controlling variable, n. On average, the calcium dependence of the maximum rate (Rmax) and the time to reach Rmax (Tpeak) are described by the calcium dependence of n. Thus, the nonlinear relationship between the free calcium concentration and the rate of exocytosis can be explained solely by the calcium dependence of the distribution of fusion complexes at vesicle docking sites.

scholarly journals Submaximal Responses in Calcium-triggered Exocytosis Are Explained by Differences in the Calcium Sensitivity of Individual Secretory Vesicles

1998 ◽  
Vol 112 (5) ◽  
pp. 559-567 ◽  
Author(s):  
Paul S. Blank ◽  
Myoung-Soon Cho ◽  
Steven S. Vogel ◽  
Doron Kaplan ◽  
Albert Kang ◽  
...  
Keyword(s):  
Calcium Sensitivity ◽  
Secretory Vesicle ◽  
Secretory Vesicles ◽  
Calcium Dependent ◽  
Sea Urchin Egg ◽  
Dependent Inactivation ◽  
Graded Response ◽  
Vesicle Exocytosis

A graded response to calcium is the defining feature of calcium-regulated exocytosis. That is, there exist calcium concentrations that elicit submaximal exocytotic responses in which only a fraction of the available population of secretory vesicles fuse. The role of calcium-dependent inactivation in defining the calcium sensitivity of sea urchin egg secretory vesicle exocytosis in vitro was examined. The cessation of fusion in the continued presence of calcium was not due to calcium-dependent inactivation. Rather, the calcium sensitivity of individual vesicles within a population of exocytotic vesicles is heterogeneous. Any specific calcium concentration above threshold triggered subpopulations of vesicles to fuse and the size of the subpopulations was dependent upon the magnitude of the calcium stimulus. The existence of multiple, stable subpopulations of vesicles is consistent with a fusion process that requires the action of an even greater number of calcium ions than the numbers suggested by models based on the assumption of a homogeneous vesicle population.

scholarly journals Rhynchophylline Protects Cultured Rat Neurons against Methamphetamine Cytotoxicity

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Dan Dan Xu ◽  
Robin Hoeven ◽  
Rong Rong ◽  
William Chi-Shing Cho
Keyword(s):  
Mtt Assay ◽  
Neuronal Death ◽  
Time Course ◽  
Inhibitory Effect ◽  
Neuroprotective Activity ◽  
Free Calcium ◽  
Neuronal Cultures ◽  
Death Time ◽  
Free Calcium Concentration

Rhynchophylline (Rhy) is an active component isolated from species of the genusUncariawhich has been used for the treatment of ailments to the central nervous system in traditional Chinese medicine. Besides acting as a calcium channel blocker, Rhy was also reported to be able to protect against glutamate-induced neuronal death. We thus hypothesize that Rhy may have neuroprotective activity against methamphetamine (MA). The primary neurons were cultured directly from the cerebral cortex of neonatal rats, acting asin vitromodel in the present study. The neurotoxicity of MA and the protective effect of Rhy were evaluated by MTT assay. The effects of MA, Rhy or their combination on intracellular free calcium concentration ([Ca2+]i) were determined in individual neocortical neurons by the Fluo-3/AM tracing method. The MTT assay demonstrated that MA has a dose-dependent neurotoxicity in neuronal cultures. The addition of Rhy prior to the exposure to MA prevented neuronal death. Time course studies with the Fluo-3/AM probe showed that Rhy significantly decreased neuronal [Ca2+]iwhich was elevated by the exposure to MA. Our results suggested that Rhy can protect the neuronal cultures against MA exposure and promptly attenuate intracellular calcium overload triggered by MA challenge. This is the first report demonstrating an inhibitory effect of Rhy against MA impairment in cultured neuronsin vitro.

Toxicity of Industrially Relevant Chlorinated Organic Solvents In Vitro

2013 ◽  
Vol 32 (2) ◽  
pp. 136-145 ◽  
Author(s):  
Catherine McDermott ◽  
James J.A. Heffron
Keyword(s):  
Cell Proliferation ◽  
Organic Solvents ◽  
Caspase 3 ◽  
Free Calcium ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Chlorinated Organic ◽  
Free Calcium Concentration ◽  
Ros Formation

The cytotoxic effects of 4 industrially important chlorinated organic solvents, dichloromethane (DCM), 1,2-dichloroethane (DCE), trichloroethylene (TCE), and tetrachloroethylene (PERC) in vitro, were investigated. Jurkat T cells were exposed to the solvents individually for 72 hours and changes in reactive oxygen species (ROS) formation, cell proliferation, intracellular free calcium concentration ([Ca2+]), and caspase-3 activity were measured. There was a concentration-dependent increase in the ROS formation and intracellular free [Ca2+] following exposure to each of the solvents. This was accompanied by a decrease in the cell proliferation. Solvent potency decreased in the following order: PERC > TCE > DCM > DCE. Caspase-3 activity was increased in a concentration-dependent manner by TCE and PERC but was not significantly altered by DCM or DCE. n-Acetyl-l-cysteine pretreatment showed that changes in the intracellular free [Ca2+] and caspase-3 activity were independent of ROS formation. However, increased ROS formation did play a causal role in the decreased cell proliferation observed.

scholarly journals Bryophyllum pinnatum Compounds Inhibit Oxytocin-Induced Signaling Pathways in Human Myometrial Cells

2021 ◽  
Vol 12 ◽  
Author(s):  
Stefanie Santos ◽  
Leonie Zurfluh ◽  
Mónica Mennet ◽  
Olivier Potterat ◽  
Ursula von Mandach ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Calcium Concentration ◽  
Inhibitory Effect ◽  
Intracellular Free Calcium ◽  
Free Calcium ◽  
Myometrial Cells ◽  
Intracellular Free Calcium Concentration ◽  
Free Calcium Concentration ◽  
Bryophyllum Pinnatum

Bryophyllum pinnatum has been used in the treatment of premature labor, first in anthroposophic hospitals and, recently, in conventional settings as an add-on medication. In vitro work with hTERT human myometrial cells showed that B. pinnatum leaf press juice inhibits the increase of intracellular free calcium concentration induced by oxytocin, a hormone known to play a role in labor. Our aim was to identify fractions/compounds in B. pinnatum press juice that contribute to this inhibitory effect, and to investigate their effect on oxytocin-driven activation of the MAPK cascade. Several fractions/compounds from B. pinnatum press juice led to a concentration-dependent decrease of oxytocin-induced increase of intracellular free calcium concentration, but none of them was as strong as B. pinnatum press juice. However, the combination of a bufadienolide and a flavonoid-enriched fraction was as effective as B. pinnatum press juice, and their combination had a synergistic effect. B. pinnatum press juice inhibited oxytocin-driven activation of MAPKs SAPK/JNK and ERK1/2, an effect also exerted by the bufadienolide-enriched fraction. The effect of B. pinnatum press juice on oxytocin-induced signaling pathways was comparable to that of the oxytocin-receptor antagonist and tocolytic agent atosiban. Our findings further substantiate the use of B. pinnatum press juice preparations in the treatment of preterm labor.

α1-Adrenoceptor subtypes in rat renal resistance vessels: in vivo and in vitro studies

2000 ◽  
Vol 278 (1) ◽  
pp. F138-F147 ◽  
Author(s):  
Max Salomonsson ◽  
Kristina Brännström ◽  
William J. Arendshorst
Keyword(s):  
Induced Response ◽  
Free Calcium ◽  
Sprague Dawley ◽  
In Vivo Experiments ◽  
Resistance Vessels ◽  
Free Calcium Concentration ◽  
Afferent Arterioles ◽  
High Concentrations

This study provides new information about the relative importance of different α1-adrenoceptors during norepinephrine (NE) activation in rat renal resistance vessels. In Sprague-Dawley rats, we measured renal blood flow (RBF) using electromagnetic flowmetry in vivo and the intracellular free calcium concentration ([Ca2+]i) utilizing ratiometric photometry of fura 2 fluorescence in isolated afferent arterioles. Renal arterial bolus injection of NE produced a transient 46% decrease in RBF. In microdissected afferent arterioles, NE (1 μM) elicited an immediate square-shaped increase in [Ca2+]i, from 90 to 175 nM ( P < 0.001). Chloroethylclonidine (CEC) (50 μM) had no chronic irreversible alkylating effect in vitro but exerted acute reversible blockade on norepinephrine (NE) responses both on [Ca2+]i in vitro and on RBF in vivo. The RBF response was attenuated by ∼50% by the putative α1A-adrenoceptor and α1D-adrenoceptor antagonists 5-methylurapidil (5-MU), and 8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4.5]decane-7,9-dione dihydrochloride (BMY-7378) (12.5 and 62.5 μg/h), respectively. The in vitro [Ca2+]i response to NE was blocked ∼25% and 50% by 5-MU (100 nM and 1 μM). BMY-7378 (100 nM and 1 μM) attenuated the NE-induced response by ∼40% and 100%. The degree of inhibition in vitro was similar to the in vivo experiments. In conclusion, 5-MU and BMY-7378 attenuated the NE-induced responses, although relatively high concentrations were required, suggesting involvement of both the α1A-adrenoceptor and α1D-adrenoceptor. Participation of the α1B-adrenoceptor is less likely, as we found no evidence for CEC-induced alkylation.

scholarly journals Exocytosis of sea urchin egg cortical vesicles in vitro is retarded by hyperosmotic sucrose: kinetics of fusion monitored by quantitative light-scattering microscopy.

1985 ◽  
Vol 101 (6) ◽  
pp. 2398-2410 ◽  
Author(s):  
J Zimmerberg ◽  
C Sardet ◽  
D Epel
Keyword(s):  
Light Scattering ◽  
Sea Urchin ◽  
Freeze Fracture ◽  
Calcium Dependent ◽  
Sea Urchin Egg ◽  
Attachment Sites ◽  
En Face ◽  
Cortical Vesicles ◽  
Kinetics Of

We have used the isolated planar cortex of sea urchin eggs to examine the role of osmotic forces in exocytosis by morphological and physiological methods. Electron micrographs of rotary-shadowed replicas show an en face view of exocytosis and demonstrate fusion of cortical vesicles to the underlying oolemma upon addition of calcium. Freeze-fracture replicas of rapidly frozen cortices reveal specialized attachment sites between cortical vesicles and the oolemma, and between the cortical vesicles themselves. We describe a novel light scattering assay for the kinetics of fusion which allows rapid changes of solutions and monitors exocytosis in real time. The rate and extent of fusion are found to be calcium dependent. The removal of calcium halts exocytosis. The validation of exocytosis in this system and development of tools for kinetic analysis allowed us to test predictions of the osmotic hypothesis of exocytosis: hyperosmotic media should inhibit exocytosis; calcium should cause vesicular swelling. Cortical vesicles were found to be permeant to sucrose, glucose, and urea. In media made hyperosmotic with 1.7 M sucrose, cortical vesicles were seen to shrink. Addition of calcium in hyperosmotic media led to a 10-fold decrease in the rate of exocytosis compared with the isotonic rate. The rate, while triggered by calcium, was no longer calcium-dependent. This slowing of exocytosis allowed us to photograph the swelling of cortical vesicles caused by calcium. Removal of calcium had no effect on subsequent exocytosis. Return of cortices to isotonic medium without calcium led to immediate exocytosis. These results are consistent with the idea that swelling of cortical vesicles is required for fusion of biological membranes.

Sign in / Sign up

Export Citation Format

Share Document