Genomic imputation and evaluation using high-density Holstein genotypes.

Genomic evaluations for 161,341 Holsteins were computed by using 311,725 of 777,962 markers on the Illumina BovineHD Genotyping BeadChip (HD). Initial edits with 1,741 HD genotypes from 5 breeds revealed that 636,967 markers were usable but that half were redundant. Holstein genotypes were from 1,510 animals with HD markers, 82,358 animals with 45,187 (50K) markers, 1,797 animals with 8,031 (8K) markers, 20,177 animals with 6,836 (6K) markers, 52,270 animals with 2,683 (3K) markers, and 3,229 nongenotyped dams (0K) with >90% of haplotypes imputable because they had 4 or more genotyped progeny. The Holstein HD genotypes were from 1,142 US, Canadian, British, and Italian sires, 196 other sires, 138 cows in a US Department of Agriculture research herd (Beltsville, MD), and 34 other females. Percentages of correctly imputed genotypes were tested by applying the programs findhap and FImpute to a simulated chromosome for an earlier population that had only 1,112 animals with HD genotypes and none with 8K genotypes. For each chip, 1% of the genotypes were missing and 0.02% were incorrect initially. After imputation of missing markers with findhap, percentages of genotypes correct were 99.9% from HD, 99.0% from 50K, 94.6% from 6K, 90.5% from 3K, and 93.5% from 0K. With FImpute, 99.96% were correct from HD, 99.3% from 50K, 94.7% from 6K, 91.1% from 3K, and 95.1% from 0K genotypes. Accuracy for the 3K and 6K genotypes further improved by approximately 2 percentage points if imputed first to 50K and then to HD instead of imputing all genotypes directly to HD. Evaluations were tested by using imputed actual genotypes and August 2008 phenotypes to predict deregressed evaluations of US bulls proven after August 2008. For 28 traits tested, the estimated genomic reliability averaged 61.1% when using 311,725 markers vs. 60.7% when using 45,187 markers vs. 29.6% from the traditional parent average. Squared correlations with future data were slightly greater for 16 traits and slightly less for 12 with HD than with 50K evaluations. The observed 0.4 percentage point average increase in reliability was less favorable than the 0.9 expected from simulation but was similar to actual gains from other HD studies. The largest HD and 50K marker effects were often located at very similar positions. The single-breed evaluation tested here and previous single-breed or multibreed evaluations have not produced large gains. Increasing the number of HD genotypes used for imputation above 1,074 did not improve the reliability of Holstein genomic evaluations.

Factors influencing MAC reduction after cardiopulmonary bypass in dogs.

BACKGROUND: Anaesthetic requirements may be reduced following surgery employing cardiopulmonary bypass (CPB). This study, in dogs, determined the role of a) volatile agents (enflurane [E] vs isoflurane [I]), b) oxygenator (bubble [B] vs membrane [M]), and c) presence [FL] vs absence [NoFL] of an in-line arterial filter in the bypass circuit in altering anaesthetic requirements following CPB. METHODS: Male mongrel dogs were anaesthetized with either enflurane (n = 24) or isoflurane (n = 24). They were randomly assigned to one of eight groups (n = 6 per group); Group 1 (E/B/FL), Group 2 (E/M/FL), Group 3 (E/M/NoFL), Group 4 (E/B/NoFL), Group 5 (I/M/FL), Group 6 (I/B/FL), Group 7 (I/M/NoFL) or Group 8 (I/B/NoFL). MAC was determined using the tail-clamp method at hourly intervals, twice before and three times after a one hour normothermic perfusion using aortoatrial cannulation and CPB. RESULTS: Prior to CPB, MAC was reproducible (enflurane: MAC1 2.17 +/- 0.29 vs MAC2 2.14 +/- 0.28%; isoflurane: MAC1 1.42 +/- 0.31 vs MAC2 1.41 +/- 0.33%) and differed among groups only for the volatile agent employed. Following CPB, MAC was reduced in all groups (P < 0.05 vs pre-CPB measurements) except Group 1 (E/B/FL). The degree of MAC reduction in other groups ranged from 39-64% and was not different based on type of agent employed, use of a membrane or bubble oxygenator, or presence or absence of an in-line arterial filter. CONCLUSION: In dogs, MAC reduction following CPB was variable, not related to type of volatile agent employed, use of a membrane or bubble oxygenator, or presence or absence of an in-line arterial filter. The explanation for reductions in anaesthetic requirements following CPB in this model remains speculative.

Formalin-induced nociceptive behavior and edema: involvement of multiple peripheral 5-hydroxytryptamine receptor subtypes.

The role of 5-hydroxytryptamine and its receptor subtypes in the development of acute inflammation was investigated using the rat paw formalin test as a model for pain (measured by flinching behavior) and edema formation (measured by plethysmometry). The role of endogenously released 5-hydroxytryptamine was assessed using 5-hydroxytryptamine receptor subtype-selective antagonists co-injected with 2.5% formalin, while the receptor subtypes involved in the inflammatory process were further defined by co-injection of 5-hydroxytryptamine or 5-hydroxytryptamine receptor subtype-selective agonists with 0.5% formalin in anticipation of an augmented response. When co-administered with 2.5% formalin, propranolol, tropisetron or GR113808A, but not ketanserin, effectively blocked nociceptive behavior. In the presence of 0.5% formalin, 5-carboxamidotryptamine, 1-(m-chlorophenyl) biguanide or 5-methoxytryptamine, but not (+/-)-1-4-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane, augmented the flinching response. These data suggest involvement of 5-hydroxytryptamine1, 5-hydroxytryptamine3 and 5-hydroxytryptamine4 receptors in peripheral nociception. There may be some dissociation of nociception and edema formation, since no single 5-hydroxytryptamine receptor antagonist inhibited edema formation with 2.5% formalin; however, with 0.5% formalin, edema formation was enhanced by co-administration of 5-hydroxytryptamine, 5-carboxamidotryptamine, (+/-)-1-4-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane or 5-methoxytryptamine, but not 1-(m-chlorophenyl) biguanide. These data suggest involvement of 5-hydroxytryptamine1, 5-hydroxytryptamine2 and possibly 5-hydroxytryptamine4 receptors in edema formation. These results confirm the involvement of 5-hydroxytryptamine1 and 5-hydroxytryptamine3 receptor subtypes in peripheral nociception associated with acute inflammation and further suggest an involvement of the more recently characterized 5-hydroxytryptamine4 receptor in this process. There appears to be a dissociation in 5-hydroxytryptamine receptors involved in peripheral nociception and edema formation.

Adenosine A3 receptor activation produces nociceptive behaviour and edema by release of histamine and 5-hydroxytryptamine.

This study evaluated the pain enhancing properties of the adenosine A3 receptor agonist N6-benzyl-5'-N-ethylcarboxamidoadenosine (N6-benzyl-NECA) by assessing behavioural effects following s.c. administration alone to the dorsal hindpaw of the rat, or in combination with a low concentration of formalin (0.5%). Edema formation was monitored by determining paw volume with plethysmometry. N6-benzyl-NECA (0.005-10 nmol) produced a dose-related increase in intrinsic flinching behaviours, as well as an increase in phase 2A flinch responses in the presence of formalin. Intrinsic effects were blocked by the histamine H1 receptor antagonist mepyramine and the 5-hydroxytryptamine2 (5-HT2) receptor antagonist ketanserin, but not by other 5-HT receptor antagonists or adenosine A1 or A2 receptor antagonists. N6-benzyl-NECA also produced an increase in paw volume, both alone and in the presence of formalin, with higher doses being required to produce this effect than for the flinch response. The increase in paw volume was also blocked by mepyramine and ketanserin but not by other antagonists. These results indicate both a nociceptive response and a proinflammatory response resulting in edema formation following activation of adenosine A3 receptors which is mediated by both 5-HT and histamine released most likely from mast cells.

Complex role of peripheral adenosine in the genesis of the response to subcutaneous formalin in the rat.

When applied peripherally, adenosine has been shown to be pronociceptive in a number of animal and human models. Recent evidence has implicated adenosine as a significant mediator in the inflammatory process. In this study using rats, we have examined the effect of adenosine and of selective adenosine A1 and A2 receptor agonists and antagonists on the response to a subcutaneous injection of formalin into the rat hindpaw. Adenosine co-injected with formalin 0.5% significantly increased flinching in both phases in a dose-dependent manner. The highest dose of adenosine had no behavioral effect on its own. The adenosine A2 receptor agonist 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamido adenosine hydrochloride (CGS-21680), at a dose of 1.5 nmol, increased flinching associated with 0.5% formalin injection but at higher doses produced depressant effects due to systemic absorption. The adenosine A1 receptor agonist N6-cyclohexyladenosine produced only systemic behavioral effects as determined by contralateral application. The flinching response to 2.5% formalin was significantly decreased by the adenosine A2 receptor antagonist 3,7-dimethyl-1-propargylxanthine (DMPX). In contrast, 8-cyclopentyl-1,3-dimethylxanthine (CPT), the selective adenosine A1 receptor antagonist augmented the response to 2.5% formalin. The non-selective adenosine receptor antagonist caffeine had no significant effect over a wide range of doses. In summary, exogenous adenosine enhances nociception in the formalin test, probably via a peripheral A2 receptor-mediated action. Endogenous adenosine, acting at both A1 and A2 receptors, appears to be involved in the formalin-induced inflammatory response.(ABSTRACT TRUNCATED AT 250 WORDS)

Does hemoglobin concentration affect perioperative myocardial lactate flux in patients undergoing coronary artery bypass surgery?

Increasing concern over complications related to blood transfusions has prompted a reevaluation of what constitutes an "adequate" perioperative hemoglobin concentration, particularly in patients undergoing coronary artery bypass graft (CABG) surgery. Data from 224 patients with preserved ventricular function (ejection fraction > 50%), undergoing CABG surgery, previously studied under a variety of anesthetic protocols, were reexamined to determine the effect of hemoglobin (HGB) concentration on myocardial lactate flux (MLF) (as an index of ischemia). The interaction of MLF and HGB concentration, anesthetic technique (ANES), and hemodynamic variables (including systemic and pulmonary arterial pressures (SAP and PAP), cardiac output (CO), and myocardial oxygen consumption (MVO2) was determined from a pool of 1598 data sets obtained from 224 patients. Data were collected from just prior to induction of anesthesia until 24 h postoperatively. Univariate analysis revealed a statistically significant relationship between MLF and HGB concentration (P < 0.001) but the correlation coefficient was only 0.09. Multiple regression analysis did not determine HGB concentration to be a significant independent term affecting MLF in either the overall group or in a subgroup of 22 patients having an adverse outcome (myocardial infarction, stroke, or death). For patients undergoing CABG surgery, HGB concentrations within the range of 58-172 g/L were not a significant variable in production of global myocardial ischemia as evidenced by MLF. This suggests that HGB concentrations as low as 60-70 g/L in the perioperative period are well tolerated and are not associated with an increased incidence of myocardial ischemia.

Myocardial metabolic and hemodynamic changes during propofol anesthesia for cardiac surgery in patients with reduced ventricular function.

This study examined the hypothesis that the use of propofol for induction and maintenance of anesthesia in patients with reduced ejection fraction (< 0.5) undergoing coronary artery revascularization would not be associated with a greater degree or incidence of myocardial ischemia as compared to patients receiving a moderate dose sufentanil-enflurane anesthetic technique. Two groups of patients were assigned randomly to receive one of two propofol anesthetic regimes. Group A (n = 21) received propofol 1-2 mg/kg as the induction drug and sufentanil 0.03 micrograms.kg-1.min-1 (fixed rate) plus propofol 50-200 micrograms.kg-1.min-1 (variable rate) infusions for maintenance of anesthesia. Group B (n = 21) received sufentanil 5 micrograms/kg for induction and propofol 50-200 micrograms.kg-1.min-1 (variable rate) infusion for maintenance of anesthesia. For comparison, a third group (Group C, n = 18) was studied subsequently. This group received sufentanil 5 micrograms/kg for induction of anesthesia which was maintained with enflurane. Adverse hemodynamic changes (hypertension, tachycardia) were managed by additional propofol (Groups A and B), sufentanil (Group C), or vasopressors (hypotension). Hemodynamic and myocardial metabolic profiles were measured when awake and sedated and at postinduction, postintubation, postincision, poststernotomy, and precardiopulmonary bypass times. Ischemia was assessed by measuring myocardial lactate production. The incidence of myocardial lactate production was reduced in Group B as compared to Group C (Group A, 45/126; Group B, 23/126; Group C, 58/107; P < 0.05). Myocardial lactate flux declined in all groups as surgery progressed; but apart from the reduction in flux (indicative of increased ischemia) noted in Group C versus Group B postinduction, no between-group differences were detected.(ABSTRACT TRUNCATED AT 250 WORDS)

Oral clonidine premedication attenuates the haemodynamic effects associated with ketamine anaesthetic induction in humans.

Induction of anaesthesia is often associated with undesirable variations in blood pressure and heart rate. Clonidine has been demonstrated to attenuate many of these undesirable effects when used as a premedicant. Other alpha 2 adrenergic agonists have been used to ameliorate the cardiostimulatory effects of ketamine in animals but there are few data on the use of this combination in humans. The effect of oral clonidine premedication, 5 micrograms.kg-1 on the haemodynamic changes induced by i.v. ketamine was studied in 42 patients volunteers. Ninety minutes before surgery, patients randomly received clonidine (C), diazepam (D), or a placebo (P) in a double-blinded fashion. Anaesthesia was induced with a ketamine infusion of 1 mg.kg-1 x min-1 until loss of consciousness. Heart rate and phasic blood pressure were measured noninvasively prior to induction, before and up to seven minutes after tracheal intubation. There were no differences in demographics or baseline vital signs among the three groups. With ketamine administration, increases in heart rate and blood pressure were less in those patients given C preoperatively than in those who received either D or P. The peak increase in mean blood pressure was 39% (C) versus 70% (D) and 55% (P) (P < 0.01). Heart rate increased by a maximum of 20% (C) versus 41% (D) and 46% (P) (P < 0.01). We conclude that oral clonidine attenuates the hyperdynamic effects of anaesthetic induction with i.v. ketamine.

Does hypothermia or hyperventilation affect enflurane MAC reduction following partial cardiopulmonary bypass in dogs?

This study in dogs determined the effect of systemic cooling and arterial hypocarbia during cardiopulmonary bypass (CPB) on the requirements for enflurane anaesthesia (MAC) before and after CPB. Twelve mongrel dogs were each anaesthetized with enflurane in oxygen on two separate occasions. End-tidal enflurane concentration was measured with a Puritan-Bennett Anaesthesia Agent Monitor. Using the tail-clamp method, MAC was determined twice with a one-hour interval between measurements (MAC 1 and MAC 2). Partial CPB was then initiated using femoral arterio-venous cannulation and maintained for one hour. Following separation from CPB, MAC was again determined twice with a one hour interval between measurements (MAC 3 and MAC 4). Dogs were randomly assigned according to PaCO2 management during CPB (low, 17.6 +/- 8.6 mmHg vs high, 38.9 +/- 11.5 mmHg), and then subjected to two experimental conditions. The first experiment on each dog was undertaken using normothermia during CPB (warm, 35-37 degrees C) while the second experiment (at least two weeks later) was conducted using hypothermia during CPB (cold, 30 degrees C). Analysis of the data, using ANOVA for repeated measures, revealed MAC 3 (1.95 +/- 0.33%, post-CPB) to be reduced when compared with MAC 1 (2.18 +/- 0.28%, P < 0.01) or MAC 2 (2.10 +/- 0.22%, P < 0.01), determined before CPB. Multivariate repeated measures analysis revealed no independent effects of hypothermia or arterial hypocarbia during CPB, on MAC reduction. By the time of the second experiment in each dog (two weeks later), MAC had returned to baseline levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Stimulation of bull sperm hyaluronidase by polycations.

The activity of bull sperm hyaluronidase (hyaluronate 3-glycanohydrolase, EC 3.2.1.36) is increased by the inclusion of polycations in the assay mixture. At pH 3.8, bovine serum albumin and histone give the greatest stimulation, while protamine sulfate, spermine, spermidine and hyamine 2389 stimulate to a lesser extent. Enzyme activity increases with serum albumin concentration to a nearly constant, high level at serum albumin concentrations greater than 1 mg/ml. Other stimulatory compounds show a similar concentration dependence except that inhibition of enzyme activity occurs at high concentrations of stimulator. The degree of stimulation depends on the pH, sample concentration and substrate concentration. Enzyme preparations with a low protein content give the greatest stimulation, while preparations with a high protein content show little stimulation. The concentration of serum albumin required for maximum stimulation increases with increased hyaluronic acid concentration. The results suggest that the stimulation of sperm hyaluronidase is nonspecific and results from an interaction of the polycation with hyaluronic acid. Since protein in the enzyme preparation substitutes for exogenous stimulator to a varying degree, serum albumin should be included in the assay mixture for sperm and testicular hyaluronidase to assure measurement of maximum enzyme activity.

Isolation of the outer acrosomal membrane from bull sperm.

A procedure is described for subcellular fractionation of bull sperm which allows the isolation of outer acrosomal membrane without the use of detergent. After washing to remove seminal plasma contaminants, the acrosomal membrane is removed by homogenization and separated on a two-step sucrose gradient. The isolated membranes have been characterized by light and electron microscopy and enzyme analysis. While the acrosomal enzymes hyaluronidase and acrosin are bound to the isolated membranes, they represent only a small percentage of the total activity and therefore do not provide reliable marker enzymes for this fraction. Subcellular fractionation of sperm also yields information on the solubility of acrosomal enzymes. Two types of acrosomal enzymes have been identified on the basis of their distribution in gradient fractions. Both alpha-fucosidase and beta-N-acetyl glucosaminidase are concentrated in the soluble fraction of the gradient. In contrast, over 70% of the acrosin and hyaluronidase activity remains associated with the sperm pellet. These differences in solubility of these enzymes may reflect differences in their function in fertilization.