Biomedical subjects

G S Kopf

Publications and source records attributed to G S Kopf.

At least 91 records · Page 5Linked to original sources

PubMed1993

Molecular events mediating sperm activation.

Fertilization is a unique event in the life of an organism and results from requisite and reciprocal cell-induced sperm and egg activation events mediated by unique cellular and environmental cues associated with either the gametes or the reproductive tract/environment. Sperm interaction with the female reproductive tract/environment and sperm-egg interaction both at a distance and in close proximity represent a series of integrated processes designed to deliver sperm with optimal fertilizing potential to the site of fertilization. The idea that sperm and eggs "communicate" prior to fertilization, and that these communicative processes are requisite for successful fertilization, has been proposed for nearly a century. Recent studies have revealed that many aspects of gamete activation prior and subsequent to fertilization have similarities to intercellular and intracellular signaling systems utilized by somatic cells. The nature of these signaling systems is now being elucidated at the molecular level in both vertebrate and nonvertebrate systems and has revealed some unique aspects of communication and transmembrane signaling between gametes. This review will consider the mechanisms by which eggs communicate with sperm to initiate sperm activation events that are essential for fertilization. It will focus particularly on some of the most recent work surrounding these issues, and new directions and questions that are relevant to this most highly specialized interaction between gametes will be discussed.

Acrosome↗

PubMed1993

Complete mouse egg activation in the absence of sperm by stimulation of an exogenous G protein-coupled receptor.

Sperm-induced egg activation has several features in common with signal transduction pathways mediated by ligand-receptor-effector systems. G proteins are postulated to act as signal transducers regulating the plethora of cellular responses constituting egg activation that occur in response to sperm. We report that acetylcholine treatment of mouse eggs expressing the human m1 muscarinic receptor, a G protein-coupled receptor, results in the full complement of events of egg activation, including ZP2 to ZP2f conversion, recruitment of maternal mRNAs, pronuclear formation with subsequent DNA synthesis, cleavage to the two-cell stage, and activation of the embryonic genome. The induction of a full complement of sperm-induced egg activation events by ACh in the absence of sperm strongly suggests that sperm-induced egg activation is mediated by egg G proteins.

Acetylcholine↗

PubMed1993

Linear left ventricular aneurysmectomy: modern imaging studies reveal improved morphology and function.

It remains uncertain whether left ventricular aneurysmectomy (LVA) improves ventricular function and whether LVA improves or distorts left ventricular contour. We applied the powerful imaging techniques of multiple-gated acquisition scanning, intraoperative transesophageal echocardiography, and magnetic resonance imaging to assess functional and morphologic changes after LVA in 75 consecutive patients undergoing LVA by conventional resection and linear closure. Fifty-two patients (69%) underwent concomitant coronary artery bypass grafting, 25 (33%) had directed endocardial resection, and 4 (5%) had valve replacement. The hospital mortality rate was 6.7% (5/75). Actuarial survival rates were 86%, 80%, and 64% at 1 year, 2 years, and 5 years, respectively. Mean anginal class improved from 3.49 to 1.24 (p < 0.0001). Mean congestive heart failure class improved from 3.04 to 1.70 (p < 0.0001). By multiple-gated acquisition scan (48 patients), mean ejection fraction improved from 0.25 preoperatively to 0.33 postoperatively (p < 0.0001). Intraoperative transesophageal echocardiography (28 patients) revealed no cases of distortion and demonstrated normalization of left ventricular contour in 69% of patients. Mean wall motion score improved from 16.4 to 18.8 (p < 0.001). Mean cross-sectional area of the left ventricle decreased from 18.7 cm2 to 12.8 cm2 (p < 0.006). Magnetic resonance imaging confirmed normalization of left ventricular contour without distortion. We conclude that linear LVA is clinically effective and objectively improves left ventricular morphology and function. On this basis, we have extended application of LVA to include patients with at least moderate-sized aneurysms undergoing coronary artery bypass grafting, despite the absence of traditional indications of arrhythmia, embolism, and frank congestive heart failure.

Adult↗

PubMed1993

Regulation of the polyspermy block in the mouse egg: maturation-dependent differences in cortical granule exocytosis and zona pellucida modifications induced by inositol 1,4,5-trisphosphate and an activator of protein kinase C.

Germinal vesicle (GV)-intact fully grown mouse oocytes do not undergo cortical granule (CG) exocytosis in response to A23187 treatment, whereas metaphase II (MII)-arrested eggs do. This differential response may reflect the development of the ability of the egg to undergo CG exocytosis, which is responsible for the biochemical modification of the glycoprotein ZP2 in the zona pellucida. Accordingly, we compared in these two stages the ability of 12-O-tetradecanoyl phorbol 13-acetate (TPA) or inositol 1,4,5-trisphosphate (IP3) to promote CG exocytosis and/or the ZP2 to ZP2f conversion; these agents are known to stimulate early events of mouse egg activation. TPA (10 ng/ml) treatment for 60 and 120 min resulted in a 25% and 52% CG loss in GV-intact oocytes and a 38% and 76% loss in MII eggs, respectively; fertilization resulted in a CG loss of approximately 70-80%. Although a similar extent of ZP2 to ZP2f conversion was observed in oocytes and eggs after a 120-min TPA treatment (approximately 70-80%), a greater extent of conversion was observed in oocytes after a 60-min treatment (80% for oocytes, 50% for eggs). Microinjection of IP3 (final concentration 1 microM) into MII eggs resulted in an extent of ZP2 conversion similar to that observed following fertilization, whereas little conversion occurred in GV-intact oocytes similarly injected. These results indicate that a protein kinase C sensitivity develops prior to meiotic maturation, whereas responsiveness to IP3 develops after maturation has resumed. We propose that the regulatory mechanism involving an IP3-mediated calcium release is deficient in GV-stage oocytes.

Animals↗

PubMed1993

Modifications of the mouse zona pellucida during oocyte maturation: inhibitory effects of follicular fluid, fetuin, and alpha 2HS-glycoprotein.

Mouse oocyte maturation in vivo or in vitro is accompanied by a precocious release of cortical granules. When oocytes are matured in vitro in the absence of serum, cortical granule exocytosis results in modifications of the zona pellucida (ZP) that are known to constitute the zona pellucida block to polyspermy. Fetuin, a glycoprotein found in newborn calf serum, can inhibit these maturation-associated changes in the zona pellucida. In this report we demonstrate that the human fetuin homologue, alpha 2-HS-glycoprotein, is present in human follicular fluid. Human follicular fluid, or purified human alpha 2-HS-glycoprotein, inhibits the maturation-associated conversion of mouse ZP2 to ZP2f in a concentration-dependent manner. Although the concentration of alpha 2HS-glycoprotein in human follicular fluid is high enough to account for the inhibitory effect of human follicular fluid on this maturation-associated conversion, human follicular fluid immunodepleted of alpha 2HS-glycoprotein retains its full inhibitory activity. Thus, other inhibitory molecules are likely to be present in follicular fluid and contribute to the inhibition of the maturation-associated conversion of ZP2.

Animals↗

PubMed1992

Activation of a Gi protein in mouse sperm membranes by solubilized proteins of the zona pellucida, the egg's extracellular matrix.

Zona pellucida (ZP)-induced acrosomal exocytosis in mammalian spermatozoa is thought to be mediated by signal transduction cascades similar to those found in hormonally responsive cells. In order to characterize this process further, we have examined the role of GTP-binding regulatory proteins (G proteins) in coupling sperm-ZP interaction to intracellular second messenger systems in mouse sperm. An in vitro signal transduction assay was developed to assess ZP-G protein dynamics in sperm membrane preparations. Guanosine 5'-3-O-(thio)triphosphate (GTP gamma S), a poorly hydrolyzable analogue of GTP, bound to these membranes in a specific and concentration-dependent fashion which reached saturation at 100 nM. Incubation of the membrane preparations with heat-solubilized ZP resulted in a significant increase in specific GTP gamma S binding in a concentration-dependent fashion with a half-maximal response at 1.25-2 ZP/microliters. Solubilized ZP also caused a significant increase in high affinity GTPase activity in the membranes over basal levels. Mastoparan increased specific GTP gamma S binding to the sperm membranes and stimulated high-affinity membrane GTPase activity to levels consistently greater than that seen with the solubilized ZP. Mastoparan, together with solubilized ZP, gave the same level of stimulation of GTP gamma S binding as mastoparan alone. Pertussis toxin completely inhibited the ZP-stimulated GTP gamma S binding, but only decreased mastoparan-stimulated GTP gamma S binding by 70-80%. Purified ZP3, the ZP component which possesses quantitatively all of the acrosomal exocytosis-inducing activity of the intact ZP, stimulated GTP gamma S binding to the same level as solubilized ZP; ZP1 and ZP2 did not stimulate GTP gamma S binding. ZP from fertilized eggs (ZPf), which does not possess acrosome reaction-inducing activity, also failed to stimulate GTP gamma S binding to sperm membranes. These data demonstrate the direct activation of a Gi protein in sperm membrane preparations in response to the ZP glycoprotein, ZP3, that induces the acrosome reaction. These data imply that Gi protein activation is an early event in the signal sequence leading to sperm acrosomal exocytosis.

Acrosome↗

PubMed1992

Mechanical circulatory support decreases neurologic complications in the treatment of traumatic injuries of the thoracic aorta.

The surgical treatment of traumatic injuries of the thoracic aorta is controversial because a number of technical approaches have been recommended. Despite the technique employed, spinal cord ischemia continues to be a persistent problem. Nineteen patients with confirmed aortic injuries secondary to blunt trauma were treated at the Yale-New Haven (Conn) Medical Center from 1984 to 1991. The patients were analyzed in two groups: group 1 (n = 10) underwent repair using mechanical circulatory support and group 2 (n = 9) underwent repair without mechanical circulatory support. Sixteen patients survived. Three patients died of complications of multiple trauma. The groups were comparable with respect to aortic cross-clamp time, preoperative systolic blood pressure, and Injury Severity Score. Three patients in the nonmechanical support group developed neurologic complications (P less than .05). No patient in the mechanical support group had a neurologic complication. We believe that mechanical circulatory support reduces the incidence of neurologic complications following traumatic injuries of the thoracic aorta and should be used whenever clinically feasible.

Academic Medical Centers↗

PubMed1992

Is cardiopulmonary bypass effective for treatment of hypothermic arrest due to drowning or exposure?

Various techniques have been advocated for resuscitation from hypothermic arrest caused by ice-cold freshwater drowning or exposure. We have resuscitated five such patients after emergency hospital admission using cardiopulmonary bypass initiated via median sternotomy. All patients presented to our facility with core temperatures less than 26 degrees C. Three patients had been in full cardiopulmonary arrest for more than 30 minutes prior to arrival. The fourth patient presented in ventricular fibrillation; the fifth was admitted to the hospital in sinus bradycardia that quickly deteriorated to asystole. All had cardiopulmonary bypass emergently initiated via median sternotomy. All were rewarmed on bypass to 37 degrees C and all survived at least 24 hours. Three of the five patients are currently alive and well with normal neurologic function. Cardiopulmonary bypass is an effective technique for resuscitation after hypothermic arrest due to near drowning and/or exposure.

Adult↗

PubMed1992

A guanine nucleotide-binding regulatory protein in human sperm mediates acrosomal exocytosis induced by the human zona pellucida.

Guanine nucleotide-binding regulatory proteins play key intermediary roles in regulating zona pellucida-mediated acrosomal exocytosis in mouse and bull sperm. Since human sperm possess a Gi-like protein and undergo the acrosome reaction in response to the human zona pellucida, we investigated whether this G protein plays a regulatory role in this exocytotic process. Zonae pellucidae isolated from eggs that had been inseminated but had shown no signs of fertilization after retrieval for in vitro fertilization and embryo transfer were pooled into groups of greater than or equal to 50 in order to reduce variability in biological responses due to the possible presence of ZP that had undergone modifications associated with the polyspermy block. Acid-solubilized zonae pellucidae were incubated with capacitated sperm, and the sperm then assessed for the acrosome reaction using both the P. sativum agglutinin and chlortetracycline fluorescence assays; both assays gave similar results. Sperm incubated with solubilized zonae pellucidae at a final concentration of 2, 4, or 6 ZP/microliter underwent acrosomal exocytosis to a similar extent as compared with A-23187. Sperm were incubated with 1 microgram/ml pertussis toxin during capacitation to functionally inactivate the Gi-like protein. Pertussis toxin treatment of sperm did not affect sperm motility and the ability of the cells to bind to structurally intact zonae pellucidae. Pertussis toxin, however, completely inhibited the percentage acrosome reactions induced by solubilized zonae pellucidae. By contrast, the A-23187-induced acrosome reaction was insensitive to PT treatment. Pertussis toxin inhibition of the zona pellucida-induced acrosome reaction occurred in a concentration-dependent manner with maximal effects observed at 100 ng/ml PT.(ABSTRACT TRUNCATED AT 250 WORDS)

Acrosome↗

PubMed1992

Activation of a G protein in mouse sperm by the zona pellucida, an egg-associated extracellular matrix.

Mammalian sperm possess a guanine nucleotide-binding regulatory protein (G protein), with properties similar to Gi, that appears to be involved in the signal transduction pathway required for zona pellucida (ZP)-mediated acrosomal exocytosis. Mouse sperm treated with pertussis toxin (PT), a toxin that functionally inactivates Gi proteins, bind to the ZP of mouse eggs but are inhibited from undergoing acrosomal exocytosis. We have measured high-affinity GTPase activity and GTP gamma [35S] binding in mouse sperm homogenates incubated in the absence and presence of ZP glycoproteins isolated from either ovulated eggs or from ovarian homogenates to determine whether this extracellular matrix can activate the sperm-associated Gi protein. An increase in GTP hydrolysis (approximately 50% over basal activity) and GTP gamma [35S] binding (approximately 25-60% over basal activity) is observed when sperm homogenates are incubated in the presence of solubilized ZP glycoproteins, and the increase in GTPase activity is dependent on the concentration of ZP added to the homogenates. Accompanying this increase is a reduction in the ability of PT to catalyze in vitro [32P]ADP-ribosylation of a Mr = 41,000 sperm Gi protein, suggesting that the increase in GTPase activity and GTP gamma [35S] binding is associated with the activation of a PT-sensitive sperm G protein(s). The ability of the ZP to stimulate high-affinity GTPase activity in these homogenates appears to be dependent on the capacitation state of the sperm from which the homogenates are prepared. These data suggest that a component(s) of the ZP may function in a manner similar to that of other ligands by binding to a sperm surface-associated receptor and subsequently activating a G protein coupled to an intracellular signal transduction cascade(s) required for induction of acrosomal exocytosis.

Animals↗

PubMed1992

Long-term experience with descending aortic dissection: the complication-specific approach.

We analyzed long-term results in 71 patients (45 men and 26 women) treated over 17 years for documented descending aortic dissection. Forty-nine patients were treated medically and 22, surgically. Actuarial survival was 65% at 1 year, 57% at 3 years, 50% at 5 years, and 28% at 10 years for the whole group. For the group treated medically, survival was 73%, 63%, 58%, and 25% at 1 year, 3 years, 5 years, and 10 years, respectively, and for the group treated surgically, 47%, 40%, and 28% at 1 year, 3 years, and 5 years, respectively. Ten (20.4%) of the 49 medically treated patients died early (5 of rupture), and 14 (28.6%) died late (8 of dissection). Five medically treated patients crossed over to surgical management for complications of dissection. Among the surgically treated patients, 6 underwent standard graft replacement of the proximal descending aorta, 8 underwent the fenestration procedure (with a standardized retroperitoneal abdominal approach), and 4 underwent the thromboexclusion operation. Specific analysis of fenestration in 14 patients (including some with persistent descending aortic dissection after replacement of the ascending aorta for dissection) found it to be safe and effective. Actuarial survival after fenestration was 77%, 77%, and 53% at 1 year, 3 years, and 5 years, respectively. Thromboexclusion was found effective, and postoperative studies confirmed thrombosis of the descending aorta with preservation of the lowest intercostal arteries. Fifteen of the 21 surviving medically treated patients agreed to return for follow-up imaging. Nine had thrombosis of the false lumen. An interesting radiographic finding was that 4 of the 15 restudied patients had a saccular aneurysm in the aorta at the level of the left subclavian artery. We recommend a complication-specific approach to the management of descending aortic dissection. Uncomplicated dissection is treated medically, whereas complicated dissection is treated surgically, with realized rupture treated by standard graft replacement, limb ischemia treated by fenestration, and enlargement or impending rupture treated by thromboexclusion.

Adult↗

PubMed1992

Developmental expression of G protein alpha subunits in mouse spermatogenic cells: evidence that G alpha i is associated with the developing acrosome.

Guanine nucleotide-binding proteins (G proteins) are important signal transducing molecules found in all cells. G proteins are associated with the plasma membrane/outer acrosomal membrane region of acrosome-intact sperm and at least one G protein is involved in the zona pellucida-induced acrosome reaction. With the goal of elucidating the functions of these proteins during spermatogenesis, we investigated the types of G proteins present in spermatogenic cells and when they first become associated with the developing acrosome. Using bacterial toxin-catalyzed [32P]ADP-ribosylation in conjunction with immunoprecipitation and immunofluorescence utilizing antibodies directed against specific regions of various G protein isotypes, the alpha subunits of Gi1, Gi2, Gi3, and G(o) were detected in mouse spermatocytes and spermatids. An antiserum recognizing a conserved sequence of G alpha i subtypes localized to the proacrosomal granules of spermatocytes and the developing acrosome of spermatids. Levels of G alpha o diminished as spermatocytes developed into spermatids such that G alpha o was not detected in cauda epididymal sperm. Immunoreactivity using G alpha o-specific antisera did not display a distinct regionalization within any of the spermatogenic cell types. G alpha s was not detected in the developing spermatogenic cells or sperm. The association of G alpha i with the developing acrosome suggests a role for G proteins may have a role in acrosome biogenesis as well as being part of a complex required later for signal transduction leading to acrosomal exocytosis.

Acrosome↗

PubMed1992

Temporal pattern of synthesis of the mouse cortical granule protein, p75, during oocyte growth and maturation.

We previously demonstrated that a protein of M(r) 75,000 (p75) is localized to cortical granules (CGs) in mouse oocytes and eggs and is released upon activation or fertilization of eggs (K.E. Pierce, M. C. Siebert, G. S. Kopf, R. M. Schultz, and P. G. Calarco, 1990, Dev. Biol. 141, 381-392). To examine the temporal pattern of synthesis of p75 during the early stages of CG formation, growing oocytes, which were isolated from juvenile mice, were incubated for 4 hr in medium containing [35S]methionine, and radiolabeled proteins were immunoprecipitated using an antiserum that detects p75. Synthesis of p75 is detected at low levels in the smallest oocytes examined (less than 20 microns). Synthesis of p75 relative to total protein synthesis increases about 12-fold during oocyte growth from the 20-40 microns size and then remains constant throughout the remaining period of oocyte growth (40-70 microns). In the fully grown, germinal vesicle (GV)-intact oocyte (70-80 microns), immunoprecipitated p75 comprises approximately 1.5% of the total amount of radiolabeled protein. Three hours after the transfer of these oocytes to a medium that supports resumption of meiosis and GV breakdown in vitro, oocytes subjected to a 1-hr labeling pulse display a 35% decrease in the relative level of p75 synthesis. By 15 hr of maturation, p75 synthesis was reduced to 14% of that in the fully grown, GV-intact oocyte and this is similar to the level of p75 synthesis in ovulated eggs. The level of p75 synthesis following in vitro translation of total egg RNA is only 38% lower than that obtained from total oocyte RNA. In addition, synthesis of p75 is observed following in vitro translation of oocyte, but not egg, poly(A)+ RNA. These results are consistent with p75 synthesis during oocyte maturation being under translational control.

Animals↗

PubMed1992

Role of G proteins in mouse egg activation: stimulatory effects of acetylcholine on the ZP2 to ZP2f conversion and pronuclear formation in eggs expressing a functional m1 muscarinic receptor.

Sperm-mediated egg activation may be analogous to ligand-mediated signal transduction through G protein-coupled receptors. We investigated this possibility in the mouse egg by microinjecting mouse oocytes with an m1 muscarinic receptor mRNA. Following oocyte maturation in vitro, the metaphase II-arrested eggs were treated with acetylcholine and its effect was examined on zona pellucida modifications and pronuclear formation, which are end points of early and late egg activation, respectively. Treatment of these eggs with acetylcholine reveals that both the ZP2 to ZP2f conversion and pronuclear formation occur. Atropine and microinjected GDP beta S block the acetylcholine-induced ZP2 conversion, suggesting that the acetylcholine effects are mediated via a functional G protein-coupled m1 receptor. The acetylcholine-induced ZP2 conversion, however, is not inhibited by pertussis toxin under conditions in which greater than 90% of the endogenous Gi is inactivated by ADP ribosylation. The presence of a pertussis toxin-insensitive G protein, Gq, is detected by immunoblotting; this G protein could be a candidate to mediate the pertussis toxin-insensitive effects of acetylcholine. Results of these experiments are consistent with the hypothesis that receptor-mediated G protein activation may play a role in egg activation.

Acetylcholine↗

PubMed1992

Hemodynamic evaluation before and after closure of fenestrated Fontan. An acute study of changes in oxygen delivery.

BACKGROUND: Acute changes in hemodynamics and oxygen delivery accompanying temporary occlusion of atrial defects in 14 patients after a fenestrated Fontan procedure were evaluated at a median interval of 32 days after surgery to identify candidates for permanent transcatheter closure of their defects. METHODS AND RESULTS: Patients ranged in age from 9 months to 33 years and in weight from 7.9 to 69 kg. Right atrial (RA), left atrial (LA), and aortic pressures, mixed venous (SmvO2) and aortic (SaO2) oxygen saturation, and whole-body oxygen consumption (VO2) were measured, and systemic blood flow (Qs), systemic oxygen transport (SOT), and oxygen extraction were calculated before and after occlusion. SmvO2, VO2, and RA pressures did not change, but SaO2 increased from 84 +/- 6% to 95 +/- 3% (p less than 0.05), and LA pressures fell from 5.1 +/- 3.6 to 3.7 +/- 2.2 mm Hg (p less than 0.05). Qs fell from 2.4 +/- 0.7 to 1.8 +/- 0.41.min-1.m-2 (p less than 0.05), SOT fell from 425 +/- 154 to 366 +/- 112 ml.O2.min-1.m-2 (p less than 0.05), and oxygen extraction increased from 0.40 +/- 0.12 to 0.46 +/- 0.13 (p less than 0.05). Only one patient did not undergo definitive closure of his defect because of a marked decrease in Qs and SOT with a significant rise in RA pressure. CONCLUSIONS: Although delayed closure of an atrial defect in these patients improved morbidity and mortality, the complete separation of the venous and systemic circulations was accomplished at the expense of decreased Qs and oxygen delivery despite the improved level of arterial oxygenation. The resting levels of oxygen extraction are elevated and will be associated with decreased exercise capability. The long-term benefits of closure of these fenestrations are yet to be shown.

Adolescent↗

PubMed1992

Neonatal correction of transposition of the great arteries: the Connecticut experience.

The treatment of transposition of the great arteries, a common congenital cardiac defect, has undergone significant development. Prior to 1989 a surgical approach which repaired the transposition at the atrial level (Senning's operation), but did not restore normal anatomy, was the procedure of choice. Since 1989 a surgical approach that restores normal anatomy (Jatene's arterial switch) has been followed. Forty-four patients have been corrected since 1979 (N = 26 Senning's operation, N = 18 arterial switch). The arterial switch patients are corrected at an earlier age, have a longer, more complex operation without a significant increase in operative mortality, intensive care, or duration of hospitalization. The prevalence or frequency of normal ventricular function and normal sinus rhythm is significantly increased over the repair at the atrial level. The frequency of pulmonary stenosis is increased. The duration of follow-up for these patients is significantly shorter than for those with atrial level repair.

Connecticut↗

PubMed1992

Fenestrated Fontan operation with delayed transcatheter closure of atrial septal defect. Improved results in high-risk patients.

Ten patients, each with two or more risk factors for morbidity and death, underwent a fenestrated Fontan procedure in which a 4 to 6 mm circular fenestration was left between the systemic and pulmonary venous chambers. None died; a similar group of high-risk patients without fenestration had a mortality rate of 2 of 8. Patients with fenestration had significantly less drainage from the chest tube, less need for inotropic support, and shorter intensive care and hospital stays than did patients without fenestration. Comparison with a group of low-risk patients undergoing the Fontan operation showed no statistical difference in these postoperative parameters. Fenestrations were closed in all 10 patients at from 9 days to 6 months after operation by means of the transcatheter clamshell occluder device. Two patients had left pulmonary artery balloon angioplasty and three patients had other atrial communications closed with additional clamshell devices. During short-term follow-up periods averaging 18 months, all patients were clinically well; however, one patient with mitral atresia required reoperation for obstruction between the left atrium and the tricuspid valve, not related to the clamshell device. These data indicate that fenestration may be one method of achieving lower morbidity and mortality rates among high-risk patients undergoing the Fontan procedure.

Adolescent↗

PubMed1991

Evidence for the involvement of a pertussis toxin-insensitive G-protein in egg activation of the frog, Xenopus laevis.

Activation responses of the frog egg at fertilization include the release of calcium from intracellular stores and the opening of calcium-dependent chloride channels, which produce the fertilization potential. To investigate the presence of guanine nucleotide-binding proteins (G-proteins), and their role in initiation of these events in the egg of the frog Xenopus laevis, we assayed for pertussis and cholera toxin substrates, and applied activators and inhibitors of G-proteins. Pertussis toxin catalyzed the [32P]ADP ribosylation of a Mr 40,000 component, but no cholera toxin substrates were demonstrated. Injection of greater than or equal to 25 pmole of guanosine-5'-O-(3-thiotriphosphate) GTP-gamma-S), an activator of G-proteins, produced a change in membrane potential that mimicked the fertilization potential and also caused cortical granule exocytosis and cortical contraction. Injections of up to 600 pmole of guanosine 3':5'-cyclic monophosphate or 9 nmole of guanosine-5'-(beta-gamma-imido)triphosphate did not active eggs. The membrane potential response to GTP-gamma-S injection showed the same peak and chloride dependence as the fertilization potential, although the duration of the GTP-gamma-S response was somewhat greater. GTP-gamma-S did not activate eggs if the calcium rise was prevented by prior injection of the calcium chelator BAPTA. Injection of up to 200 ng of cholera toxin did not activate eggs. However, eggs were activated by applying 1 nM serotonin to eggs that had been injected with a specific mRNA for the serotonin 1c receptor, a member of the class of receptors that act by way of G-proteins. Egg activation in response to either sperm or serotonin was not inhibited by pertussis toxin, under experimental conditions where approximately 80-90% of the toxin substrate was ADP-ribosylated. These results support the hypothesis that sperm activate Xenopus eggs at fertilization by way of a pertussis and cholera toxin-insensitive G-protein.

Adenosine Diphosphate Ribose↗