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PubMed1985

A new algorithm for voltage clamp by iteration: a learning control of a nonlinear neuronal system.

Voltage-clamp of excitable membrane allows the measurement of membrane currents associated with electrical potential changes across the membrane. However, it has been impossible in practice to apply the conventional analog feedback voltage-clamp circuits to single electrode voltage clamping in central neurons. The reason for this is that the feedback system becomes unstable because of the positive feedback required for compensation of capacitative loss through the wall of the microelectrode. Park et al. (1981) proposed a new iterative technique to solve this problem. It requires that the potential to be clamped repeats itself with little or no change. The amount of current needed to clamp the membrane potential is not determined at once, but in a step-wise, trial and error fashion in the course of a set of repetitions. Since the feedback loop is open in real time, the system has great stability, and this advantage can be exploited in single electrode preparations. The computation algorithm which calculates the current waveform based on the voltage deviation during the last trial is the central part of the iterative voltage-clamp system. In this paper, we propose a new algorithm, which has several theoretical and practical advantages over the original one proposed by Park et al. First, two parameters used in the new algorithm are predetermined by a current-clamp experiment. Second, the speed of convergence of the new algorithm is faster than that of the Park's original algorithm. This was shown by computer simulation of iterative voltage clamp of artificial membrane following Hodgkin-Huxley equations for squid axon membrane and Rall's compartment model for a neuron with dendrites. Finally, we offer proof that the new algorithm is certain to converge for the general cases of voltage-clamp experiments with active membrane properties, synaptic membranes, etc. Consequently, the new algorithm for iterative voltage clamp is very suitable for single electrode voltage clamp in the central neurons. The new algorithm has been successfully applied to voltage-clamp experiments on rubrospinal neurons of cats (Tsukahara, Murakami, Kawato, Oda, and Etoh, in preparation).

Animals↗
PubMed1983

Electrocardiographic responses of rat fetuses with clamped or intact umbilical cords to acute maternal uterine ischemia.

Uterine ischemia results in severe cardiac disturbances in the fetus. It has been postulated that these effects are due to interaction between the fetus and the ischemic uterus or placenta, and not to hypoxia or buildup of metabolites in the fetus. Rat fetal cardiac responses to uterine clamping and umbilical cord clamping were compared by electrocardiography. On day 21 of gestation, fetuses in a total of 14 pregnant rats were exposed to umbilical clamping, uterine clamping, or no clamping. Electrocardiograms were recorded for each fetus immediately after clamping and at 10, 20, and 30 minutes after clamping. Immediately after clamping, the uterine clamping group alone showed severe sinus bradycardia. At all other observation times, fetuses exposed to uterine or umbilical clamping showed bradycardia and other electrocardiographic changes typically associated with hypoxia. Therefore, umbilical clamping protected the fetuses from the adverse effects of uterine clamping for only a very short time.

Animals↗
PubMed1999

Myocardial protection in operations requiring more than 2 h of aortic cross-clamping.

OBJECTIVE: Long periods of aortic cross-clamping time during cardiac surgery are associated with high rates of morbidity and mortality because of damage to the myocardium. Recently, we have used a method of myocardial protection based on the principles of hyperkalemic cardioplegic arrest. We use antegrade administration of warm, undiluted blood followed by continuous retrograde infusion of tepid, undiluted blood supplemented with potassium and magnesium. In this study, we have retrospectively reviewed our experience with this method of cardioprotection in operations requiring more than 2 h of cross-clamp time. METHODS: We retrospectively reviewed the medical records of 1280 patients who underwent myocardial revascularization, valve repair or replacement, or a combination of both operations between January 1, 1994 and December 31, 1997. Patients were divided into two groups: the short cross-clamp group (SXC) (n = 1144) had cross-clamp times < 120 min (mean, 78 +/- 20 min; range, 35-119 min) and the long cross-clamp group (LXC) (n = 136) had cross-clamp times > 120 min (mean, 154 +/- 31 min; range, 120-277 min). We compared preoperative, operative, and postoperative variables between the two groups. RESULTS: Significantly more patients in the long cross-clamp group (43.4%) underwent the combined operation than in the short cross-clamp group (2.3%), and the rate of reoperation was significantly higher in the long cross-clamp group (12%) than in the short cross-clamp group (5%). Despite these differences in operative complexity, we found no difference in hospital mortality rates between the two groups. The only significant postoperative differences were that the long cross-clamp group had a greater need for inotropic agents (43 vs. 29%), higher serum levels of creatine kinase (880 +/- 583 vs. 613 +/- 418) and CK-MB (10.9 +/- 6.4 vs. 5.9 +/- 5.2), and a longer hospital stay (9.6 vs. 6.1 days). CONCLUSION: Long, complex operations requiring more than 2 h of cross-clamping can be performed safely with our method of cardioprotection based on continuous retrograde infusion of tepid, hyperkalemic, undiluted blood.

Aged↗
PubMed2004

Structural analysis of the inactive state of the Escherichia coli DNA polymerase clamp-loader complex.

Clamp-loader complexes are heteropentameric AAA+ ATPases that load sliding clamps onto DNA. The structure of the nucleotide-free Escherichia coli clamp loader had been determined previously and led to the proposal that the clamp-loader cycles between an inactive state, in which the ATPase domains form a closed ring, and an active state that opens up to form a "C" shape. The crystal structure was interpreted as being closer to the active state than the inactive state. The crystal structure of a nucleotide-bound eukaryotic clamp loader [replication factor C (RFC)] revealed a different and more tightly packed spiral organization of the ATPase domains, raising questions about the significance of the conformation seen earlier for the bacterial clamp loader. We describe crystal structures of the E. coli clamp-loader complex bound to the ATP analog ATPgammaS (at a resolution of 3.5 A) and ADP (at a resolution of 4.1 A). These structures are similar to that of the nucleotide-free clamp-loader complex. Only two of the three functional ATP-binding sites are occupied by ATPgammaS or ADP in these structures, and the bound nucleotides make no interfacial contacts in the complex. These results, along with data from isothermal titration calorimetry, molecular dynamics simulations, and comparison with the RFC structure, suggest that the more open form of the E. coli clamp loader described earlier and in the present work corresponds to a stable inactive state of the clamp loader in which the ATPase domains are prevented from engaging the clamp in the highly cooperative manner seen in the fully ATP-loaded RFC-clamp structure.

Adenosine Diphosphate↗
PubMed2007

A function for the psi subunit in loading the Escherichia coli DNA polymerase sliding clamp.

Crystal structures of an Escherichia coli clamp loader have provided insight into the mechanism by which this molecular machine assembles ring-shaped sliding clamps onto DNA. The contributions made to the clamp loading reaction by two subunits, chi and psi, which are not present in the crystal structures, were determined by measuring the activities of three forms of the clamp loader, gamma(3)deltadelta', gamma(3)deltadelta'psi, and gamma(3)deltadelta'psichi. The psi subunit is important for stabilizing an ATP-induced conformational state with high affinity for DNA, whereas the chi subunit does not contribute directly to clamp loading in our assays lacking single-stranded DNA-binding protein. The psi subunit also increases the affinity of the clamp loader for the clamp in assays in which ATPgammaS is substituted for ATP. Interestingly, the affinity of the gamma(3)deltadelta' complex for beta is no greater in the presence than in the absence of ATPgammaS. A role for psi in stabilizing or promoting ATP- and ATPgammaS-induced conformational changes may explain why large conformational differences were not seen in gamma(3)deltadelta' structures with and without bound ATPgammaS. The beta clamp partially compensates for the activity of psi when this subunit is not present and possibly serves as a scaffold on which the clamp loader adopts the appropriate conformation for DNA binding and clamp loading. Results from our work and others suggest that the psi subunit may introduce a temporal order to the clamp loading reaction in which clamp binding precedes DNA binding.

Adenosine Triphosphatases↗
PubMed1997

Acute cortisol responses of lambs to castration using a rubber ring and/or a castration clamp with or without local anaesthetic.

Changes in plasma cortisol concentrations during the first 4 hours after castration of 6-week-old lambs by one of four methods were measured. The methods were application of a rubber ring above the testes, application of a ring to shorten the scrotum below the testes, use of a castration clamp plus a ring, and use of a castration clamp alone. The efficacy of local anaesthetic in reducing or abolishing the cortisol responses to castration by all these methods except short scrotum was assessed. Local anaesthetic was injected into the neck of the scrotum, both spermatic cords, the scrotal neck plus spermatic cords, or into both testes. The combined clamp plus ring method was used to test whether or not clamp damage to afferent nerves from the testes would reduce the cortisol response to ring castration. The short scrotum method elicited a significantly lower cortisol response than that caused by ring castration. Injection of local anaesthetic into both spermatic cords marginally reduced the cortisol response to ring castration, but the response was virtually abolished by prior scrotal neck, or scrotal neck plus spermatic cords, or intra-testicular local anaesthetic injections. The clamp plus ring method, where each spermatic cord and the associated scrotal tissue were crushed separately for 1, 5 or 10 seconds with no overlap between the two crush lines, did not elicit lower cortisol responses than the ring-only method. Local anaesthetic injected into the scrotal neck virtually abolished, and spermatic cord injections markedly reduced, the cortisol responses to combined clamp plus ring castration. The IO-second clamp application caused a more protracted cortisol response than did ring-only castration, whereas the cortisol responses to 1-second clamp application and to ring-only castration were similar. Local anaesthetic injected into the scrotal neck or spermatic cords did not affect significantly the cortisol response to the IO-second clamp application. The following conclusions were drawn from this work: the testes as well as the scrotum were sources of noxious sensory input after ring application; the clamp plus ring method used here was ineffective in reducing the pain-induced distress (as indicated by cortisol concentrations) caused by ring-only castration; injection of local anaesthetic into the scrotal neck or into the testes prior to ring application blocked most noxious sensory input from both the scrotum and the testes; and on the basis of the present cortisol responses the 10-second clamp applications method could not be recommended in reference to the ring-only method, but the l-second clamp application method might be equally acceptable for castrating 6-week-old-lambs. Of the methods examined, injections of local anaesthetic into the scrotal neck or both testes were the most effective in reducing the pain-induced distress caused by ring-only castration.

Journal Article↗
PubMed1998

The effect of prolonged clamping and vascular stasis on the patency of arterial and venous microanastomoses.

During free tissue transfer, an artery and one or two veins must be sutured. Either the artery or the vein can be repaired first, but the optimal order of vascular repair has not been established. Maintaining a clamp on a vessel is associated with vessel injury, and releasing the clamp may cause bleeding and vascular stasis. The purpose of this study was to determine if the order of vessel repair and the length of vascular clamping affects surgical outcome. Free flaps were created on Sprague-Dawley rats (400-450 g) as follows: Bilateral abdominal skin flaps (3 x 3 cm) based on the superficial inferior epigastric vessels were elevated. The femoral artery and vein were tied and divided distal to the branching of the inferior epigastric vessels. Proximal to the branching, the artery and vein were divided. The animals were then randomized into four groups as follows: In group I (N=16), the artery was repaired and then the clamps were released to revascularize the flap. Venous stasis occurred as the vein was being repaired because of the venous clamp. In group II (N=15), the artery was repaired, but the clamp was maintained to prevent blood from coming in contact with the fresh arterial anastomosis and to prevent venous stasis. The vein was then repaired. In group III (N=15), the vein was repaired first but the venous clamp was not released until the artery was repaired. In group IV (N=15), the vein was repaired first and the clamps were released, allowing venous blood to contact the fresh anastomosis while the artery was being repaired. After final clamp removal, all anastomoses were assessed immediately for evidence of thrombosis. Five days postoperatively the skin flap was evaluated for evidence of necrosis and the anastomosis was inspected for evidence of late thrombosis. The anastomoses were resected for histopathological evaluation. Flap success was compared between groups using chi-squared analysis. Eleven of 16 flaps failed (69%) in group I, and 3 of 15 flaps failed (20%) in each of groups II, III, and IV. Statistical significance was reached in comparing group I with the other three groups (p < 0.01). All failures in group I were caused by immediate venous thrombosis. The other failures were secondary to arterial thrombosis. Histopathological analysis failed to demonstrate any differences between the groups. According to these results, arterial repair followed by clamp release prior to the completion of the venous repair results in a low success rate, probably secondary to venous stasis within the draining vein while the venous anastomosis is being completed. If the arterial repair is performed first, then it is recommended that the arterial clamp be maintained until the venous repair is completed. If the vein is repaired first, then it can be clamped or unclamped with similar results.

Anastomosis, Surgical↗
PubMed1984

Acute regulation of human lymphocyte insulin receptors. Analysis by the glucose clamp.

The T lymphocyte insulin receptor model has been used to explore the regulation of insulin receptor appearance in that lymphocytes do not bear the insulin receptor in the circulation and thus are not amenable to regulation by virtue of ligand binding. Such cells synthesize insulin receptors when stimulated by antigen in vivo or in vitro. In these studies, the glucose clamp technique was employed to isolate perturbations in plasma glucose and plasma insulin as potential mediators of the regulation of the mitogen-induced T lymphocyte insulin receptor. Nondiabetic, normal weight individuals volunteered for 10 hyperglycemic clamp studies and nine euglycemic clamp studies with five individuals studied by both protocols. Hyperglycemia and hyperinsulinemia were created by the hyperglycemic clamp (basal plasma glucose was increased from 89 +/- 2 mg/dl to 230 +/- 2 mg/dl and an insulin of 99 +/- 8 microU/ml was reached). Blood was removed for isolation of T lymphocytes at 0, 1, 3, and 4 h of the clamped condition. After 1 h of hyperglycemia accompanied by an elevated plasma insulin, T cell insulin binding fell from 9.9 +/- 0.9 pg/10(6) lymphocytes to 8.5 +/- 0.9 pg/10(6), and reached a nadir of 19 +/- 4% at the conclusion of the clamp. Scatchard analysis of binding data from two of the subjects who underwent the hyperglycemic clamp demonstrated a reduction of the number of binding sites per cell without a change in the affinity of ligand for receptor. To separate the effects of glucose and insulin and the manner in which insulin is provided, the 4-h euglycemic clamp was performed in which fasting plasma glucose was maintained (95 +/- 2 mg/dl) while constant hyperinsulinemia was created (80 +/- 3 microU/ml). Insulin binding to activated, cultured T lymphocytes demonstrated a similar fall in insulin binding. Scatchard analysis of three additional studies again revealed a reduction in receptor number to approximately 40% of base line. These studies reveal that T cell insulin receptor regulation is achieved by hyperinsulinemia independent of the glucose level achieved. The reduction in insulin binding and receptor number could not be accounted for by variations in the strength of lectin stimulation, the time course of lectin response, or by the stress of the clamp itself. The effect of the clamp was specific for the lymphocyte insulin receptor in that the clamp had little effect on the interleukin II receptor activation marker. Acute changes in plasma insulin by the glucose clamp technique are perceived by the T lymphocyte and displayed in tissue culture by an alteration in lectin-induced insulin receptors. One can conclude that rapid changes in ambient in vivo insulin concentrations can regulate the synthesis of T lymphocyte insulin receptors generated in vitro.

Adult↗
PubMed2006

Crystal structure of a DNA polymerase sliding clamp from a Gram-positive bacterium.

BACKGROUND: Sliding DNA clamps are processivity factors that are required for efficient DNA replication. DNA polymerases maintain proximity to nucleic acid templates by interacting with sliding clamps that encircle DNA and thereby link the polymerase enzyme to the DNA substrate. Although the structures of sliding clamps from Gram-negative bacteria (E. coli), eukaryotes, archaea, and T4-like bacteriophages are well-known, the structure of a sliding clamp from Gram-positive bacteria has not been reported previously. RESULTS: We have determined the crystal structure of the dimeric beta subunit of the DNA polymerase III holoenzyme of Streptococcus pyogenes. The sliding clamp from this Gram-positive organism forms a ring-shaped dimeric assembly that is similar in overall structure to that of the sliding clamps from Gram-negative bacteria, bacteriophage T4, eukaryotes and archaea. The dimer has overall dimensions of approximately 90 A x approximately 70 A x approximately 25 A with a central chamber that is large enough to accommodate duplex DNA. In comparison to the circular shape of other assemblies, the S. pyogenes clamp adopts a more elliptical structure. CONCLUSION: The sequences of sliding clamps from S. pyogenes and E. coli are only 23% identical, making the generation of structural models for the S. pyogenes clamp difficult in the absence of direct experimental information. Our structure of the S. pyogenes beta subunit completes the catalog of clamp structures from all the major sequence grouping of sliding clamps. The more elliptical rather than circular structure of the S. pyogenes clamp implies that the topological nature of encircling DNA, rather than a precise geometric shape, is the most conserved aspect for this family of proteins.

Amino Acid Sequence↗
PubMed1992

Simultaneous assessment of insulin secretion and insulin sensitivity using a hyperglycemia clamp.

A hyperglycemic clamp is an established method to assess insulin secretion and is generally used only for this purpose. To determine whether it could also be used to assess insulin sensitivity, we compared insulin sensitivity indices (ISI) obtained during euglycemic and hyperglycemic clamp experiments in 22 nonobese volunteers (body mass index, 23.9 +/- 0.6 kg/m2) and in 20 obese individuals (body mass index, 30.8 +/- 1.3 kg/m2) matched for age and gender. The ISI values (micromoles per kg.min/pmol) of the obese group assessed during hyperglycemic (0.088 +/- 0.011) and euglycemic (0.050 +/- 0.005) clamp experiments were both significantly lower than the ISI of the nonobese group assessed in hyperglycemic and euglycemic clamp experiments (0.179 +/- 0.024 and 0.096 +/- 0.009, respectively; both P less than 0.01). Although the ISI values obtained with hyperglycemic clamps were consistently greater than those obtained with euglycemic clamp (0.137 +/- 0.016 vs. 0.075 +/- 0.007; P less than 0.001), they were highly correlated (r = 0.84; P less than 0.0001). Moreover, when these indices were converted to clearance rates, thereby correcting for the mass action effects of glucose on glucose disposal, the values obtained with the hyperglycemic clamp (0.0137 +/- 0.0016 mL/kg.min/pmol) were statistically identical to those obtained with the euglycemic clamp (0.0142 +/- 0.0013 mL/kg.min/pmol), as indicated by a regression equation having an intercept of 0 and a slope (1.03) not different from 1. We, therefore, conclude that the hyperglycemic clamp and the euglycemic clamp yield comparable estimates of insulin sensitivity and that, under appropriate conditions, the hyperglycemic clamp technique may be used to assess both insulin sensitivity and insulin secretion in the same individual in a single experiment.

Blood Glucose↗
PubMed1994

A comparison between the minimal model and the glucose clamp in the assessment of insulin sensitivity across the spectrum of glucose tolerance. Insulin Resistance Atherosclerosis Study.

An insulin-modified frequently sampled intravenous glucose tolerance test (FSIGTT) with minimal model analysis was compared with the glucose clamp in 11 subjects with normal glucose tolerance (NGT), 20 with impaired glucose tolerance (IGT), and 24 with non-insulin-dependent diabetes mellitus (NIDDM). The insulin sensitivity index (SI) was calculated from FSIGTT using 22- and 12-sample protocols (SI(22) and SI(12), respectively). Insulin sensitivity from the clamp was expressed as SI(clamp) and SIP(clamp). Minimal model parameters were similar when calculated with SI(22) and SI(12). SI could not be distinguished from 0 in approximately 50% of diabetic patients with either protocol. SI(22) correlated significantly with SI(clamp) in the whole group (r = 0.62), and in the NGT (r = 0.53), IGT (r = 0.48), and NIDDM (r = 0.41) groups (P < 0.05 for each). SI(12) correlated significantly with SI(clamp) in the whole group (r = 0.55, P < 0.001) and in the NGT (r = 0.53, P = 0.046) and IGT (r = 0.58, P = 0.008) but not NIDDM (r = 0.30, P = 0.085) groups. When SI(22), SI(clamp), and SIP(clamp) were expressed in the same units, SI(22) was 66 +/- 5% (mean +/- SE) and 50 +/- 8% lower than SI(clamp) and SIP(clamp), respectively. Thus, minimal model analysis of the insulin-modified FSIGTT provides estimates of insulin sensitivity that correlate significantly with those from the glucose clamp. The correlation was weaker, however, in NIDDM. The insulin-modified FSIGTT can be used as a simple test for assessment of insulin sensitivity in population studies involving nondiabetic subjects. Additional studies are needed before using this test routinely in patients with NIDDM.

Analysis of Variance↗
PubMed1983

Catecholamine release during and after cross clamping of descending thoracic aorta.

During graft replacement of descending thoracic and thoracoabdominal aneurysms, aortic cross clamping without the use of bypass or shunts is accompanied by underperfusion of distal vascular beds. This study, comprising 10 patients, was based on the hypothesis that ischemia of the lower spinal cord and adrenal glands might precipitate release of catecholamines. Arterial blood samples, obtained before (control), during, and after arterial cross clamping, were analyzed for epinephrine (E) and norepinephrine (NE). The E/NE ratio was used as an index of ischemic sympathoadrenal activation. During cross clamping, mean E and E/NE increased by a factor of 4.5 (P = 0.03) and 2.4 (P = 0.001), respectively. The greatest increases were observed in the 5 min postclamp sample: relative to control, mean E increased 22-fold (P = 0.011), NE 3-fold (P = 0.009), and E/NE 8.4-fold (P = 0.013). During the immediate postclamp period, mean E/NE fell exponentially with an average "half-life" of 12.05 +/- 5.83 min (SD). A second-order polynomial related (P = 0.004) log E/NE in first postclamp sample to the ratio between clamp time and mean proximal arterial pressure during clamping (T/AP). Left-ventricular-minute-work-function correlations: (1) positive with log E during clamping (P = 0.043); and (2) none with log E (P = 0.563) or log (E + NE) (P = 0.641) 5 min post-clamp; (3) positive with log (E + NE) 30 min post-clamp (P = 0.016). It is concluded that (1) distal ischemia caused by cross clamping the descending thoracic aorta without bypass or shunts, results in distal regional sympathoadrenal activation, independent of known central reflex mechanisms; (2) this activation leads to marked increases of E/NE and is, in part, dependent upon T/AP during clamping; (3) the activation is probably transitory; (4) by indirect evidence, cardiodepressant factor(s) may transiently be present following declamping; (5) by its magnitude, sympathoadrenal activation is likely to provide compensation for deleterious factors that may affect cardiovascular functions during cross clamping and after declamping; alternatively, it might contribute to postoperative hypertension and cardiac complications such as myocardial ischemia and arrhythmias.

Adrenal Glands↗
PubMed1990

Sodium nitroprusside decreases spinal cord perfusion pressure during descending thoracic aortic cross-clamping in the dog.

Paraplegia is a devastating complication of surgery on the descending thoracic aorta. During surgical repair, the aorta is cross-clamped, and nitroprusside is often used to treat arterial hypertension that can occur above the cross-clamp. Twenty-one dogs were studied to determine the effects of nitroprusside on intraspinal pressures, mean aortic pressures below the cross-clamp, and spinal cord perfusion pressure. Perfusion pressure in spinal radicular arteries originating below the aortic cross-clamp was estimated as the distal aortic pressure minus intraspinal pressure. Nitroprusside was used to return the mean arterial pressure above the cross-clamp to values similar to the pre-cross-clamp levels in 7 dogs. Fourteen animals did not receive sodium nitroprusside. Aortic cross-clamping resulted in small but significant increases in intraspinal pressure (4.3 +/- 0.8 to 7.5 +/- 0.9 mm Hg in non-nitroprusside-treated dogs, and 3.4 +/- 1.0 to 5.6 +/- 1.5 mm Hg in the nitroprusside group before nitroprusside). Nitroprusside caused a further increase in intraspinal pressure (5.6 +/- 1.5 to 8.3 +/- 2.2 mm Hg) and a decrease in aortic pressure below the cross-clamp (26 +/- 5 to 18 +/- 4 mm Hg). The increase in intraspinal pressure and the decrease in aortic pressure below the cross-clamp after nitroprusside resulted in a decrease in spinal cord perfusion pressure from 19 +/- 5 mm Hg to 11 +/- 4 mm Hg. Because nitroprusside decreases spinal cord perfusion pressure and may increase the risk of spinal cord ischemia, the avoidance of large doses of nitroprusside to arbitrarily return mean arterial pressure above the cross-clamp to pre-cross-clamp levels is recommended.

Animals↗
PubMed2005

Ergonomic redesign and evaluation of a clamping tool handle.

The handle of a commercial bar clamp was redesigned using ergonomic principles and then compared with an original clamp. Ten male and ten female students participated in simulated clamping tasks under various conditions, including different clamping heights, clamping methods, and handle-gripping methods, with respect to the dependent variables of clamping and handle-squeezing forces. The results showed that the redesigned clamp produced larger clamping force with lower handle-squeezing forces than the original clamp. As expected, males exerted more force than females in both clamping and squeezing forces. A pistol grip method was superior to an upside-down handle-gripping method. Two-handed operation was recommended for this type of clamp by simultaneously pulling the trigger and sliding the bar in order to initially tighten around objects. This study shows that the application of ergonomic guidelines increases the efficiency and usability of manual handtools.

Adult↗
PubMed2005

Myocardial protection with intermittent cross-clamp fibrillation: does preconditioning play a role?

OBJECTIVE: Previously, we showed intermittent cross-clamp fibrillation afforded equivalent protection to cardioplegia. This study examined whether protection induced by intermittent cross-clamp fibrillation involves an ischemic preconditioning mechanism. METHODS: Isolated Langendorff-perfused rat hearts were subjected to three different studies to determine: Study 1, whether a single intermittent cross-clamp fibrillation episode (10 min) and reperfusion (10 min) before prolonged ischemia acts as a preconditioning trigger for protection; Study 2, whether cardioprotection induced by intermittent cross-clamp fibrillation alone (no prolonged ischemia) involves a preconditioning mechanism; Study 3, whether intermittent cross-clamp fibrillation cardioprotection can be prevented by targeting putative components of the preconditioning mechanism (protein kinase C or the mitochondrial ATP-sensitive potassium (K(ATP)) channel). Hearts were reperfused (60 min) and recovery of function (left ventricular developed pressure measured using an intraventricular balloon) and myocardial injury (creatine kinase leakage) were measured. RESULTS: In Study 1, recovery of function in the single intermittent cross-clamp fibrillation hearts was 61+/-3% (mean+/-SEM) (p<0.05) compared to 41+/-2% in control group; glibenclamide (a non-specific ATP-sensitive potassium (K(ATP))-channel blocker) prevented this preconditioning protection (37+/-4%). In Study 2, recovery of function in intermittent cross-clamp fibrillation hearts (62+/-3%) was significantly (p<0.05) higher than intermittent cross-clamp fibrillation hearts treated with glibenclamide (33+/-2%) and ischemia hearts (30+/-5%). In Study 3, protection by intermittent cross-clamp fibrillation (60+/-3%; p<0.05) was attenuated by protein kinase C inhibition (chelerythrine, 34+/-3%) and mitochondrial K(ATP)-channel blockade (5-hydroxydecanoate, 27+/-4%) to levels not significantly different from that of ischemia hearts (25+/-4%). CONCLUSIONS: The cardioprotective efficacy of intermittent cross-clamp fibrillation was attenuated by protein kinase C inhibition or K(ATP)-channel blockade. Involvement of these putative preconditioning cascade components in association with cardioprotection induced by intermittent cross-clamp fibrillation, suggests a role for the ischemic preconditioning mechanism.

Animals↗
PubMed2003

Yeast Rad17/Mec3/Ddc1: a sliding clamp for the DNA damage checkpoint.

The Saccharomyces cerevisiae Rad24 and Rad17 checkpoint proteins are part of an early response to DNA damage in a signal transduction pathway leading to cell cycle arrest. Rad24 interacts with the four small subunits of replication factor C (RFC) to form the RFC-Rad24 complex. Rad17 forms a complex with Mec3 and Ddc1 (Rad1731) and shows structural similarities with the replication clamp PCNA. This parallelism with a clamp-clamp loader system that functions in DNA replication has led to the hypothesis that a similar clamp-clamp loader relationship exists for the DNA damage response system. We have purified the putative checkpoint clamp loader RFC-Rad24 and the putative clamp Rad1731 from a yeast overexpression system. Here, we provide experimental evidence that, indeed, the RFC-Rad24 clamp loader loads the Rad1731 clamp around partial duplex DNA in an ATP-dependent process. Furthermore, upon ATP hydrolysis, the Rad1731 clamp is released from the clamp loader and can slide across more than 1 kb of duplex DNA, a process which may be well suited for a search for damage. Rad1731 showed no detectable exonuclease activity.

Adenosine Triphosphatases↗
PubMed2006

Influence of the descending thoracic aortic cross clamping on bispectral index value and plasma propofol concentration in humans.

BACKGROUND: In this study, the authors investigated changes in Bispectral Index (BIS) values and plasma propofol concentrations (Cp) after aortic cross clamping in the descending thoracic aortic aneurysm repair surgery during propofol anesthesia. METHODS: Prospectively, in 10 patients undergoing thoracic aortic surgery during total intravenous anesthesia with propofol, BIS values were recorded during cross clamping of the descending thoracic aorta. In this study, the rate of propofol infusion was controlled to keep the BIS value between 30 and 60 throughout surgery. Simultaneously, Cp values in the blood samples taken from the right radial artery (area proximal to cross clamping) and the left femoral artery (area distal to cross clamping) were measured. RESULTS: Approximately 15 min after initiating aortic cross clamping, BIS values in all cases started to decrease abruptly. Cp values of samples taken from the radial artery after cross clamping of the aorta were significantly (P < 0.05) increased compared with pre-cross clamp values (1.8 +/- 0.4 microg/ml), and the mean Cp after aortic cross clamping varied between 3.0 and 5.3 microg/ml. In addition, there were significant differences in the Cp values between radial arterial and femoral arterial blood samples throughout aortic cross clamping. Cp values in samples from the radial artery were approximately two to seven times higher than those from the femoral artery. CONCLUSIONS: This study showed that Cp values increased and BIS values decreased rapidly after aortic cross clamping in thoracic aortic aneurysm repair surgery during propofol anesthesia. These findings suggested that all anesthesiologists should control the infusion rate carefully, taking the abrupt changes in its pharmacokinetics into consideration, especially during cross clamping of the descending thoracic aorta.

Adult↗
PubMed2003

The optimal sequence of microvascular repair during prolonged clamping in free flap transfer.

During free flap transfer, the surgeon may decide to begin with repair of the artery or the vein(s) and to unclamp the first vessel as soon as repair is completed or maintain the clamping of both vessels until completion of all repairs. Complications can lead to prolonged clamping times, potentially increasing the risk of tissue ischemia, vascular damage, and thrombosis. The goals of the present study were to determine whether the sequence of vessel repair and the duration of clamping affect the success of free flap transfer in cases requiring prolonged clamping. Sixty abdominal fasciocutaneous free flaps based on the superficial inferior epigastric vessels were created in Sprague-Dawley rats. To model clinical situations in which prolonged clamping is necessary, the study used a 1-hour delay before the repair of the second vessel. Flaps were randomized into four groups. In group I (n = 15), the artery was repaired first, and the arterial clamp was removed immediately to allow arterial inflow. In group II (n = 15), the arterial repair was first, and the arterial clamp was maintained until completion of venous repair. In group III (n = 15), venous repair was first, with venous clamping maintained until completion of the arterial repair. In group IV (n = 15), initial venous repair was followed by immediate unclamping, before arterial repair. On release of all clamps, the patency of arteries and veins was confirmed immediately and after 1 hour using a "milking" test. On the fifth postoperative day, each flap was assessed for necrosis and for patency of the anastomoses. Of 15 flaps in each group, five (33 percent) failed in group I, four (27 percent) failed in groups II and III, and six (40 percent) failed in group IV. Differences between groups were not statistically significant (p = 0.8). These results demonstrate that in cases requiring prolonged occlusive clamping (2 to 3 hours), factors such as venous congestion, possible clamp injury, and presence of static blood in contact with the new anastomosis have relatively equivalent contributions to the risk of failure. Accordingly, no advantage seems to be gained by beginning with the artery or the vein or by using early or delayed unclamping of the first vessel repaired.

Abdominal Wall↗