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L A Jaffe

Publications and source records attributed to L A Jaffe.

53 records · Page 3Linked to original sources
PubMed1985

Fertilization potential and polyspermy prevention in the egg of the nemertean, Cerebratulus lacteus.

We investigated the electrical properties of the egg of the nemertean worm Cerebratulus, and found evidence that an electrically-mediated polyspermy block operates for a period of about 1 hr after fertilization. At fertilization, in natural or artificial sea water, the membrane potential shifts from its resting level of about -66 mV to a peak of about +43 mV, and in most cases remains greater than 0 mV for more than 1 hr. The average potential during the first 30 min is +22 +/- 8 mV (SD, n = 12). When the external Na+ concentration is reduced from 486 to 51 mM (choline substituted) the fertilization potential amplitude is reduced; the average potential during the first 30 min is -27 +/- 21 mV (SD, n = 5). Eggs inseminated in 51 mM Na+ sea water become polyspermic, indicating that polyspermy prevention depends on an electrically-mediated mechanism. The electrical block is required for about 60 min, since transfer to 51 mM Na+ sea water during this period results in polyspermy. During the first hour following fertilization, the egg is also developing a permanent, nonelectrical block; the degree of polyspermy which results upon transfer to low Na+ sea water decreases progressively with time. The permanent block appears to be at the level of the egg plasma membrane or glycocalyx, since the egg envelope is not a barrier to sperm penetration, nor does its removal induce polyspermy. Electron micrographs show no obvious changes in the morphology of the extracellular layers, plasma membrane or cortex of the egg after fertilization.

Animals↗
PubMed1985

Fertilization-induced ionic conductances in eggs of the frog, Rana pipiens.

Fertilization of the frog egg (Rana pipiens) elicits a positive-going shift in membrane potential (fertilization potential) that lasts 10-20 min and functions as a fast block to polyspermy. We examined the ion conductances underlying the fertilization potential, using the voltage-clamp technique. We measured the membrane capacitance during the fertilization potential by applying an alternating current. We also determined the intracellular K and Cl concentrations in the egg, using ion-selective micro-electrodes. The conductance is largest in the first 2 min after fertilization. Regardless of whether the stimulus is provided by one or by more than one sperm or by artificial activation, the size of the conductance increase is the same, reaching a maximum of about 40 microseconds. Two separate conductances are involved at fertilization: Cl and K. [K]i = 121 mM and [Cl]i = 44 mM. The natural external medium is pond water (approximated in our experiments by 10% Ringer solution); therefore, an increase in K and Cl conductances leads to an efflux of both ions. The equilibrium potential of the fertilization current is between the Cl and K equilibrium potentials (ECl and EK), closer to ECl. 10 mM-external tetraethylammonium (TEA) brings the equilibrium potential close to ECl and reduces the maximum conductance by about half. The Cl conductance is not blocked by 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS). The time courses of the K and Cl conductances are similar. The TEA-resistant conductance (primarily Cl conductance) activated at fertilization increases as the membrane potential becomes more positive. A voltage-sensitive Na conductance present in the unfertilized egg disappears after fertilization. During fertilization this conductance is too small to contribute significantly to the fertilization potential. The membrane capacitance increases by an average of 1.9 times in the first 2 min following the rise of the fertilization potential, during the period of cortical vesicle exocytosis. Capacitance then gradually decreases; at 1 h after fertilization, capacitance is 82% of the value in the unfertilized egg. The conductance increase precedes the capacitance increase by several seconds. Therefore the initial appearance of Cl and K channels cannot be accounted for by addition of membrane by cortical vesicle exocytosis. The conductance subsequently decreases, suggesting that the disappearance of the Cl and K channels is not caused by membrane removal.

Action Potentials↗
PubMed1985

Propagating potassium and chloride conductances during activation and fertilization of the egg of the frog, Rana pipiens.

Fertilization or artificial activation of the frog egg (Rana pipiens) elicits a positive-going shift in membrane potential which results from an opening of Cl- and K+ channels in the egg membrane. We examined the spatial localization of the currents produced by the opening of these channels, using large patch electrodes (tip diameters 3-10 microns). We also used small patch electrodes (tip diameters about 1 micron) to study the single K+ channel currents. After activation, with the patch electrode at any position on the egg surface, we observed a transient current, with a main peak lasting several seconds. This activation current occurred after a variable delay of 0-3 min following the rise of the activation potential. With 10% Ringer solution in the bath and pipette, the current was usually outward, although it sometimes had an inward component. With one patch electrode on the animal surface of the egg and another patch electrode on the vegetal surface, we observed that the activation current propagated over the egg surface. In experiments where the egg was activated by applying a hyperpolarizing pulse, the response in the animal half preceded that in the vegetal half by an average of about 1 min. The amplitude of the peak outward current was similar for animal and vegetal recordings (1-2 mA/cm2). Tetraethylammonium (11 mM) in the patch pipette blocked most of the outward component of the activation current and revealed an underlying inward component. The inward component of the activation current was carried by Cl-, since it could be reversed by raising the Cl- concentration in the pipette. The Cl- component of the activation current propagated over the egg surface, with timing similar to that of the total current. The average amplitude of the peak Cl- current was six or more times larger at the animal than the vegetal surface. Fertilization caused a current to propagate from the animal to the vegetal surface, like the current observed during activation. With a small patch electrode, single channel currents of the K+ component of the activation current could be seen. The probability that the channels were open increased at more positive potentials. The single channel conductance was estimated to be 25 pS, and the reversal potential to be -150 mV. Single Cl- channel currents have not yet been seen. Activation or fertilization of the frog egg resulted in a wave-like opening of Cl- and K+ channels, which spread from the animal to the vegetal half of the egg.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1984

Electrical properties of vertebrate oocyte membranes.

The electrical properties of vertebrate oocyte and egg cell membranes are reviewed. Ion channels of these oocytes generate transcellular currents and action potentials as well as responses to neurotransmitters. Electrical properties change during meiotic maturation and fertilization. Available information about the electrical properties of sperm is also discussed.

Action Potentials↗
PubMed1983

Absence of an electrical polyspermy block in the mouse.

To examine the possibility of an electrical polyspermy block in the mouse, we recorded the electrophysiological properties of zona-free mouse eggs during fertilization. Starting from an unfertilized value of -41 +/- 4 mV (SD), the membrane potential undergoes an oscillation (seen in 8 of 11 records) of 4 +/- 1 mV in amplitude, starting 7 +/- 5 min after insemination, and lasting about 1 min. However, except for this small oscillation, the membrane potential is constant during the 60 min following insemination; the average range (11 +/- 4 mV) is not significantly different from that which is observed in 60-min recordings from unfertilized eggs. These results indicate that the polyspermy block which is established during this period (D. P. Wolf, 1978, Dev. Biol. 64, 1-10) is not electrically mediated. Consistent with this finding, reduction of the sodium or calcium concentration in the external medium does not induce polyspermy. As a consequence of fertilization, the resistance of the egg membrane decreases from 96 +/- 34 to 44 +/- 15 M omega; this change accompanies the voltage oscillation.

Animals↗
PubMed1983

Studies of the voltage-dependent polyspermy block using cross-species fertilization of amphibians.

Fertilization of frog eggs by frog sperm is inhibited if the egg's membrane potential is positive (N.L. Cross and R.P. Elinson, 1980, Dev. Biol. 75, 187-198); however, fertilization of salamander eggs by salamander sperm does not depend on membrane potential (M. Charbonneau, M. Moreau, B. Picheral, J.P. Vilain, and P. Guerrier, 1983, Dev. Biol. 98, 304-318). Since salamander sperm can fertilize frog eggs, we have investigated whether this cross-fertilization is voltage dependent. If, during insemination with Notophthalmus sperm, Xenopus eggs were voltage clamped between +7 and +20 mV, fertilization proceeded in 7/10 (70%) of the clamped eggs, compared to 38/48 (79%) of the neighboring eggs. In control experiments in which voltage-clamped Xenopus eggs were inseminated with Xenopus sperm, fertilization proceeded in only 1/10 (10%) of the clamped eggs, compared to 59/60 (98%) of the neighbors. Similar results were obtained with cross-fertilization experiments between Notophthalmus sperm and Rana eggs. These experiments indicate that the voltage dependence of fertilization depends on the species of sperm.

Amphibians↗
PubMed1982

Studies of the mechanism of the electrical polyspermy block using voltage clamp during cross-species fertilization.

Prevention of polyspermic fertilization in sea urchins (Jaffe, 1976, Nature (Lond.). 261:68-71) and the worm Urechis (Gould-Somero, Jaffe, and Holland, 1979, J. Cell Biol. 82:426-440) involves an electrically mediated fast block. The fertilizing sperm causes a positive shift in the egg's membrane potential; this fertilization potential prevents additional sperm entries. Since in Urechis the egg membrane potential required to prevent fertilization is more positive than in the sea urchin, we tested whether in a cross-species fertilization the blocking voltage is determined by the species of the egg or by the species of the sperm. With some sea urchin (Strongylocentrotus purpuratus) females, greater than or equal to 90% of the eggs were fertilized by Urechis sperm; a fertilization potential occurred, the fertilization envelope elevated, and sometimes decondensing Urechis sperm nuclei were found in the egg cytoplasm. After insemination of sea urchin eggs with Urechis sperm during voltage clamp at +50 mV, fertilization (fertilization envelope elevation) occurred in only nine of twenty trials, whereas, at +20 mV, fertilization occurred in ten of ten trials. With the same concentration of sea urchin sperm, fertilization of sea urchin eggs occurred, in only two of ten trials at +20 mV. These results indicate that the blocking voltage for fertilization in these crosses is determined by the sperm species, consistent with the hypothesis that the fertilization potential may block the translocation within the egg membrane of a positively charged component of the sperm.

Animals↗
PubMed1980

Actin, microvilli, and the fertilization cone of sea urchin eggs.

Sea urchin eggs and oocytes at the germinal vesicle stage were fixed at various times after insemination, and thin sections were examined. Actin filaments can first be found in the cortical cytoplasm 1 min after insemination, and by 2 min enormous numbers of filaments are present. At these early stages, the filaments are only occasionally organized into bundles, but one end of many filaments contacts the plasma membrane. By 3 min, and even more dramatically by 5 min after insemination, the filaments become progressively more often found in bundles that lie parallel to the long axis of the microvilli and the fertilization cones. By 7 min, the bundles of filaments in the cone are maximally pronounced, with virtually all the filaments lying parallel to one another. Decoration of the filaments with subfragment 1 of myosin shows that, in both the microvilli and the cones, the filaments are unidirectionally polarized with the arrowheads pointing towards the cell center. The efflux of H+ from the eggs was measured as a function of time after insemination. The rapid phase of H+ efflux occurs at the same time as actin polymerization. From these results it appears that the formation of bundles of actin filaments in microvilli and in cones is a two-step process, involving actin polymerization to form filaments, randomly oriented but in most cases having one end in contact with the plasma membrane, followed by the zippering together of the filaments by macromolecular bridges.

Actins↗
PubMed1979

Electrically mediated fast polyspermy block in eggs of the marine worm, Urechis caupo.

Previous work has established that the polyspermy block in Urechis acts at the level of sperm-egg membrane fusion. (J. Exp. Zool. 196:105). Present results indicate that during the first 5--10 min after insemination the block is mediated by a positive shift in membrane potential (the fertilization potential) elicited by the penetrating sperm, since holding the membrane potential of the unfertilized egg positive by passing current reduces the probability of sperm entry, while progressively reducing the amplitude of the fertilization potential by decreasing external Na+ progressively enhances multiple sperm penetrations. Also, a normal fertilization potential is correlated with a polyspermy block even under conditions (pH 7) in which eggs do not develop. We have investigated the mechanism of the electrical polyspermy block by quantifying the relationship between sperm incorporation, membrane potential and ion fluxes. Results indicate that the polyspermy block is mediated by the electrial change per se and not by the associated fluxes of Na+, Ca++, and H+.

Animals↗
PubMed1979

Ionic mechanism of the fertilization potential of the marine worm, Urechis caupo (Echiura).

Microelectrode and tracer flux studies of the Urechis egg during fertilization have shown: (a) insemination causes a fertilization potential; the membrane potential rises from an initial level of -33 +/- 6 mV to a peak at +51 +/- 6 mV (n = 16), falls to a plateau of about +30 mV, then returns to the original resting potential 9 +/- 1 min (n - 10) later; (b) the fertilization potential results from an increase in Na+ permeability, which is amplified during the first 15 s by a Ca++ action potential; (c) the maximum amplitude of the fertilization potential, excluding the first 15 s, changes by 51 mV for a 10-fold change in external [Na+]; (d) in the 10 min period after insemination, both Na+ and Ca++ influxes increase relative to unfertilized egg values by factors of 17 +/- 7 (n = 6) and 34 +/- 14 (n = 4), respectively; the absolute magnitude of the Na+ influx is 16 +/- 6 times larger than that of Ca++; (e) in the absence of sperm these same electrical and ionic events are elicited by trypsin; thus, the ion channels responsible must preexist in the unfertilized egg membrane; (f) increased Na+ influx under conditions of experimentally induced polyspermy indicates that during normal monospermic fertilization, only a fraction of available Na+ channels are opened; we conclude that these channels are sperm-gated; (g) Ca++ influx at fertilization is primarily via the membrane potential-gated channel, because kinetics are appropriate, and influx depends on potential in solutions of varying [Na+], but is independent of number of sperm incorporations in normal sea water.

Action Potentials↗
PubMed1975

Calcium accumulations within the growing tips of pollen tubes.

Pollen of L. longiflorum was grown in 45Ca-labeled medium and washed with nonradioactive medium. Whole, labeled pollen was then frozen and autoradiographed at -78 degrees C. The autoradiographs show striking accumulations of 45Ca in the growing tips of the pollen tubes. This result is obtained when the pollen is labeled for times as short as 1 min, or as long as 5 h. In most cases, the tip concentration is about two to four times greater than that in the bulk of the pollen tube, and extends for a length of about 20 mum. In autoradiographs of tubes longer than 1 mm, a small fraction of cells show a distinctly larger 45Ca accumulation, the tip containing more than 100 times that in the rest of the cell. The 1- to 5-h labeling experiments show that calcium is relatively concentrated within the cytoplasm of the growing tip. The 1- to 3-min labeling experiments suggest that calcium may enter the tip faster than it enters other regions. These patterns of calcium accumulation and flux may be related to the localized secretion of vesicles at the grow;ng tip.

Calcium↗
PubMedDate not supplied

Fertilization increases the polyphosphoinositide content of sea urchin eggs.

Fertilization of the sea urchin egg stimulates a wave of exocytosis of cortical vesicles, but the mechanism by which fertilization regulates this secretion is not fully understood. We describe here experiments which suggest that polyphosphoinositide metabolism could be a factor in this regulation. We find that the cortical vesicle exocytosis in the egg of Strongylocentrotus purpuratus is preceded by a 40% increase in its content of triphosphoinositide (TPI) and a 22% increase of diphosphoinositide (DPI).

Animals↗