Intra- and postoperative low-dose ketamine for adolescent idiopathic scoliosis surgery: a randomized controlled trial.

BACKGROUND: In this randomized controlled trial, we examined whether intra- and postoperative infusion of low-dose ketamine decreased postoperative morphine requirement and morphine-related adverse effects as nausea and vomiting after scoliosis surgery. METHODS: After IRB approval and informed consent, 36 patients, aged 10-19 years, undergoing posterior correction surgery for adolescent idiopathic scoliosis, were randomly allocated into two groups: intra- and postoperative ketamine infusion at a rate of 2 µg/kg/min until 48 h after surgery (ketamine group, n = 17) or infusion of an equal volume of saline (placebo group, n = 19). All patients were administered total intravenous anesthesia with propofol and remifentanil during surgery and intravenous morphine using a patient-controlled analgesia device after surgery. The primary outcome was cumulative morphine consumption in the initial 48 h after surgery. Pain scores (Numerical Rating Scale, NRS, 0-10), sedation scales, incidence of postoperative nausea and vomiting (PONV), and antiemetic consumption were recorded by nurses blinded to the study protocol for 48 h after surgery. RESULTS: Patient characteristics did not differ between the two groups. Cumulative morphine consumption for 48 h after surgery was significantly lower in the ketamine group compared to the placebo group (0.89 ± 0.08 mg/kg vs. 1.16 ± 0.07 mg/kg, 95% confidence interval for difference between the means, 0.03-0.48 mg/kg, P = 0.019). NRS pain, sedation scales, and incidence of PONV did not differ between the two groups. Antiemetic consumption was significantly smaller in ketamine group. CONCLUSIONS: Intra- and postoperative infusion of low-dose ketamine reduced cumulative morphine consumption and antiemetic requirement for 48 h after surgery.

Mapping of a quantitative trait locus for beef marbling on bovine chromosome 9 in purebred Japanese black cattle.

The goal of this study is to detect quantitative trait loci (QTL) for carcass traits applicable for a DNA-based breeding system in a Japanese Black cattle population. A purebred paternal half-sib family from a commercial line composed of 65 steers was initially analyzed using 188 informative microsatellites giving a 16-cM average interval covering 29 autosomes. A significant QTL for marbling was detected in the centromeric portion of bovine chromosome (BTA) 9. After additional marker genotyping across a larger sample size composed of 169 individuals, this locus was refined to a 20-cM confidence interval between microsatellites BM1227 (24 cM) and DIK2741 (50 cM) at a 1% chromosome-wise threshold. The allele substitution effect between Q and q for a beef marbling standard score (1 to 12 range) on BTA9 was 1.0 (5.7% of total phenotypic variance in QTL contribution in this family). This result provides a primary platform for a marker-assisted selection system of the beef marbling trait within the Japanese Black (Wagyu) cattle population.

Psychological and endocrine factors and pain after mastectomy.

BACKGROUND: This prospective study was designed to examine the associations of demographic, clinical, psychological and neuroendocrine factors with acute and chronic post-operative pain following partial mastectomy. METHODS: Sixty-four female patients scheduled for partial mastectomy were enrolled. Pre-operative anxiety/depression was assessed, using the Hospital Anxiety and Depression Scale (HADS). Pre-operative 24-h urinary cortisol levels were measured 2 days before surgery. Post-operative pain was examined using a visual analog scale (VAS) for acute pain on 0-2 post-operative day (POD), and a short-form McGill Pain Questionnaire for chronic pain at 6 months after surgery. In the last 29 subjects, post-operative 24-h urinary cortisol levels were also measured on 0 POD and were subjected to correlation analysis. RESULTS: Multivariate logistic regression analysis revealed that lower pre-operative cortisol secretion and greater pre-operative anxiety were significantly associated with an increased risk of moderate to severe acute post-operative pain [Odds Ratio (95% Confidence Interval); 0.96 (0.92-0.98), and 1.24 (1.04-1.54)], and that patients with greater pre-operative anxiety and moderate to severe acute pain were more likely to develop chronic post-operative pain [OR (95% CI); 1.63 (1.23-2.40), and 5.07 (1.30-24.6)]. Correlational analysis demonstrated that the post-operative cortisol level was inversely correlated with pre-operative anxiety and the intensity of acute post-operative pain (r = -0.40, p < 0.05, and r = -0.50, p < 0.01), but not with the intensity of chronic pain. CONCLUSIONS: This study confirms that pre-operative anxiety is associated with both acute and chronic post-operative pain after partial mastectomy. It also suggests that lower perioperative cortisol secretion might be associated with greater acute post-operative pain. SIGNIFICANCE: Although the associations between psychological stress/stress hormone levels and chronic post-operative pain remain to be determined, pre-operative psychological stress and perioperative cortisol levels are correlated with acute post-operative pain.

Detection of the MYD88 mutation by the combination of the allele-specific PCR and quenching probe methods.

INTRODUCTION: The MYD88 missense mutation c.794T>C, p.Leu265Pro, is found in patients with Waldenstörm's macroglobulinemia and lymphoma. Direct sequencing, allele-specific PCR (AS-PCR), PCR-restriction fragment length polymorphism (PCR-RFLP), and high-resolution melting analysis (HRM) are currently used to detect the mutation; however, they are either time-consuming or have low detection sensitivity. Here, we developed a novel highly sensitive and rapid detection method based on the quenching probe (QP) technique and AS-PCR. METHOD: A lymphoma cell line heterozygous for the MYD88 mutation, two wild-type cell lines, and two samples from Waldenstörm's macroglobulinemia patients were analyzed by AS-PCR, PCR-RFLP, HRM, and QP, and their detection sensitivity was examined using the mixtures of the mutant and wild-type DNA. RESULTS: For mutation-carrying heterozygous samples, the QP method produced W-shaped melting profiles presenting curves derived from the wild-type and mutant alleles. The QP analysis was performed in 2 h and demonstrated the detection limit of 5%, which was similar to that of the other methods. However, the combination of AS-PCR and QP (AS-QP) improved the sensitivity to 0.62% of the mutant allele. CONCLUSION: The AS-QP analysis is rapid and minimally improves detection sensitivity compared to the AS-PCR.

Photoperiodic time measurement in tau mutant hamsters.

Photoperiodic regulation of testicular function was investigated in homozygous tau mutant hamsters; these animals have an innate circadian period of about 20 h. In 20-h light:dark (LD) cycles, the minimum photoperiod required to prevent testicular regression was between 10.0 and 11.5 h per 20-h cycle (equivalent to 12.0-13.8 circadian hours). This was proportionally similar to the minimum photoperiod necessary to prevent regression in wild-type hamsters maintained in 24-h LD cycles. To examine the shape of the photoperiodic photosensitivity curve in homozygous tau mutant hamsters, the authors measured the effects of different T cycles on testicular maintenance. Entrainment to LD 1:18.0 and LD 1:20.5 partially or completely prevented gonadal regression in homozygous tau mutant hamsters, but LD 1:19.4 did not prevent regression. When considered in terms of circadian time, the photoperiodic photosensitivity curve for homozygous tau mutant hamsters was similar to that described previously for wild-type hamsters. The results indicate that, as in wild-type hamsters, photoperiodic regulation of reproduction is regulated by circadian photosensitivity in homozygous tau mutant hamsters. Because tau mutant hamsters measure day length against a time base of 20 h, the circadian pacemaker that measures day length might be the same as that which generates circadian rhythmicity in locomotor activity. The authors' data leave open the question of whether the tau mutation has had effects on the control of reproduction that are not directly attributable to its effects on the period of the circadian oscillator.

Circadian behavior and plasticity of light-induced c-fos expression in SCN of tau mutant hamsters.

In hamsters homozygous for the circadian clock mutation tau, the photic history dramatically affects the magnitude of light-induced circadian phase shifts. The maximum amplitude of phase shifts produced by 1-h light pulses presented at CT 14 was less than 2 h in animals that had been in DD for 2 days, whereas animals that had been kept in DD for 49 days could be shifted by more than 8 h. In this study, the authors compared the effect of previous light history on the amplitude of circadian phase shifts and on c-fos expression in the SCN of tau mutant hamsters. Although the maximum amplitude of behavioral phase shifts was drastically different between animals that had been held for either 2 or 49 days in DD, maximal fos induction was not significantly different in these two groups. However, photic thresholds for light-induced behavioral phase shifts, c-fos mRNA, and Fos immunoreactivity were closely correlated within both groups, and these thresholds were lower (more sensitive to light) after 49 than after 2 days in DD. The correlation between phase shifting and Fos induction thresholds, under conditions where both responses are dramatically altered by the previous light history, demonstrates an association between changes in circadian behavioral phase-shifting responses of tau mutant hamsters and plasticity of light-induced c-fos expression in SCN. However, because the maximum amplitudes of Fos induction and phase shifting were not correlated in animals that had been in DD for 2 days, we speculate that the level of c-fos expression does not directly determine phase shift amplitude.

Construction and characterization of a rad51rad52 double mutant as a host for YAC libraries.

RAD52 or RAD51 recombination-deficient yeast strains stabilize otherwise unstable YACs containing ribosomal DNA or the human color vision locus (Kohno et al., 1994). Thus the RAD52RAD51 pathways(s) are apparently involved in the instability of YACs containing tandem repeat loci, presumably by promoting recombination-based deletion formation. Some other genomic loci are still unstable or unrecoverable in those strains, but we now find that greater stability is observed in a rad51rad52 double mutant strain that we have newly constructed. YACs containing a highly unstable region around DXS49 or centromeric regions throw off a variety of products in single mutants, but are much more stable in the rad51rad52 strain, which could therefore provide a better host for library construction and maintenance.

Developmental changes of gene expression in heme metabolic enzymes in rat placenta.

Transcription levels of the non-specific delta-aminolevulinate synthase (ALAS-N) and heme oxygenase-1 (HO-1) in the placenta at the terminal stage of pregnancy were comparable to those in the female adult liver and in the spleen, respectively. Immunohistochemical studies demonstrated that both enzymes were exclusively expressed in the trophoblast. During gestation, transcript of ALAS-N slightly increased, while HO-1 mRNA significantly decreased. Induced acute fetal hypoxia resulted in an increase in ALAS-N mRNA and in a decrease in HO-1 mRNA. These findings indicate that placental heme metabolism is influenced by the oxygen supply.

Flow cytometric study of injuries in cultured endothelial cells by neutrophils of the inherited cataract rats.

We examined the mechanism of endothelial injuries in the inherited cataract rats (ICR), which have a number of age-associated spontaneous injuries in the aortic endothelium. Cell cycle traverse rate of endothelial cells of ICR was shorter than that of Wistar rats. The rate was estimated from bromodeoxyuridine (BrdU) incorporation into cell nuclei measured periodically after BrdU pulse labeling. Next we established the method for measurement of cultured endothelial cell injury by neutrophils with flow cytometry by assessing the regeneration of injured endothelial cells. By the use of the gate analysis method, contaminated neutrophils were excluded from the analysis. Endothelial cell injury by neutrophils of Wistar rats was detectable at 1 x 10(5) neutrophils (1 neutrophil to 1 endothelial cell) when stimulated with 10 ng/ml phorbol myristate acetate (PMA). Extent of injury increased with an increasing number of neutrophils and the concentration of a stimulator, PMA. We detected endothelial cell injury by ICR neutrophils not only when they were stimulated but also in a resting condition, and ICR neutrophils yielded more injury than Wistar rat neutrophils. Number of adhered neutrophils to endothelial cells and effects of plasma or lymphocytes were the same between two strain rats. Scavengers of hydrogen peroxide and singlet oxygen inhibited the ICR neutrophil-induced endothelial cell injury. These findings indicate that an increase of generation of excited oxygen species from neutrophils, particularly of singlet oxygen, may cause injury of endothelial cells in this specific strain of rats.

Spontaneous injuries in the aortic endothelium of the inherited cataract rats and their prevention by tocopherol. A study by scanning electron microscopy.

The aortic endothelium of inherited cataract rats (ICR), which spontaneously develop cataracts and neutrophilia, was examined by scanning electron microscopy using silver nitrate staining and pressure fixation. In ICR aged 4 weeks, the luminal surface of the aorta was similar to that in Wistar rats from which they had been derived. However, 8 weeks after birth, endothelial cells were upraised and partially detached from an underlying tissue. At 16 weeks, morphological changes exhibited by such detaching cells were more evident than at 8 weeks and fibrin was seen to be adhering to the surface of these cells; no platelet involvement was noted, however. Oral administration of DL-alpha-tocopheryl acetate for 2 weeks resulted in a reduction in the extent of endothelial injury and the luminal surface of the aorta became similar to that seen in 4- or 8-week-old animals. We found that the number of age-associated spontaneous injuries occurring in the aortic endothelium of ICR could be reduced by tocopherol administration.

Effects of a circadian mutation on seasonality in Syrian hamsters (Mesocricetus auratus).

In Syrian hamsters, exposure to short photoperiods or constant darkness induces a decrease in gonadotrophin secretion and gonadal regression. After 10-12 weeks, animals undergo spontaneous gonadal reactivation, gonadotrophin concentrations rise, and in males, testes size increases and spermatogenesis resumes. The tau mutation shortens the period of circadian wheel-running activity by 4 h in the homozygote. Here, we examine the impact of this mutation on the reproductive response to photoperiod change. Seventeen adult tau mutant and nine adult wild-type males were housed in complete darkness for 25 weeks and testes size determined at weekly intervals. Gonadal regression and subsequent recrudescence occurred in both groups of animals. Regression occurred more rapidly in tau mutants, with a nadir significantly earlier than wild-types but after a similar number of circadian cycles. Rates of testicular recrudescence were similar in both groups. Our data suggest that an acceleration of the circadian period increases the rate of reproductive inhibition in animals exposed to inhibitory photoperiods. Once initiated, the rate of spontaneous reactivation may be independent of the circadian axis.

Hypothalamic digitalis-like substance is released with sodium-loading in rats.

Role of the hypothalamic digitalis-like substance (EDLS) on the hypertension associated with an excess intake of sodium and the releasing mechanism were investigated. The blood pressure in rats fed with a sodium diet increased significantly after 4 weeks of the treatment compared to the control rats fed with a regular diet, which was accompanied by increased urinary output of the EDLS. Electrical lesions of the AV3V area in the hypothalamus significantly decreased both the urinary EDLS level and the blood pressure elevated by the sodium-loading. With the immunohistochemistry using digoxin antibody, the immunoreactives were localized in the neurons of paraventricular and supraoptic nuclei and some other hypothalamic areas, and were also seen in the nerve fibers distributed in the basal hypothalamus, infundibulum, and pituitary posterior lobe. Assuming that the CSF sodium is responsible for the release of EDLS, hypertonic NaCl (2.5 M) was infused into the lateral ventricle for 30 minutes. Blood pressure increased gradually, attaining peak rises about 30 minutes later. The plasma content of the EDLS was significantly greater in the hypertonic NaCl group than the control group treated with either the artificial CSF or 2.5 M of urea solution. On the other hand, the hypothalamic content decreased with the infusion of the hypertonic saline. Furthermore, the continuous intracerebroventricular infusions of the hypertonic NaCl with osmotic minipumps in conscious rats significantly increased the arterial pressure after 6 days. Thereby, the plasma level of the EDLS was significantly greater than the control rats that received only the artificial CSF.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunohistochemical investigations of the influence of reserpine on the serotonin neuron system in the rat brain.

A peroxidase-antiperoxidase immunohistochemical method with serotonin antiserum was employed to investigate the influence of reserpine on serotonin neurons of rats which were sacrificed at various times after injection (10 mg/kg i.p.). The disappearance of serotonin immunoreactivity induced by reserpine was detected only in the perikarya after 15 min, and then rapidly proceeded to the terminals. Between 2 and 4 h, immunoreactivity completely disappeared throughout the brain. The immunoreactivity reappeared in the perikarya after 6 h, and progressed toward the terminals gradually. However, there was an obvious difference in the rate of recovery of immunoreactivity between areas. After 7 days, the immunoreactivity returned to control levels.

Machado-Joseph disease gene products carrying different carboxyl termini.

Three cDNA clones for the Machado-Joseph disease gene (MJD1) were isolated, two of which have a new exon sequence and a distinct 3' terminal nucleotide sequence resulting in a new carboxyl terminal domain in the translated product. The nucleotide sequence of the other one is similar to the previously published one except for five polymorphisms, one of which is a single nucleotide substitution resulting in a change from the stop codon (TAA; allele A) to a tyrosine residue (TAC; allele C). Genetic analysis results suggest that Japanese MJD mutations are associated with allele A.

Reinnervation of serotonin fibers in the denervated rat subcommissural organ by fetal raphe transplants. An immunohistochemical study.

The ability of axonal outgrowth of serotonin neurons in the implanted brain tissue of subcommissural organ (SCO) was immunohistochemically studied. The serotonin neuron system of the experimental rats was completely destroyed by the intraventricular injection of 5,6-dihydroxytryptamine. The raphe region of normal fetal rats was implanted into the caudal part of the third ventricle of the neurotoxic drug pretreated rats. The host brain was examined 3 months after transplantation. The numerous serotonin fibers were distributed in the SCO and periventricular region of the third ventricle of the host brain. The outgrowing serotonin fibers from the raphe transplant seemed to innervate the SCO with the target specificity.

Transplantation of fetal mesencephalic and medullary raphe tissues into the cerebellum of denervated adult rats--an immunohistochemical study.

Pieces of mesencephalic and medullary raphe tissues were transplanted into the cerebella of 5,6-dihydroxytryptamine-treated adult rats. The extent of axonal outgrowth of serotonergic and dopaminergic neurons in the grafts was immunohistochemically studied. At 3 months after transplantation, numerous dopaminergic neurons with many processes extending within the graft were detected in the mesencephalic raphe graft, but not in the medullary raphe graft. In contrast, both the mesencephalic and medullary raphe grafts contained numerous serotonergic neurons and a dense plexus of their fibers. The outgrowing serotonergic fibers from the mesencephalic raphe graft showed a hyperinnervation pattern in the cerebellar cortex adjacent to the graft. Furthermore, a glomerulus-like accumulation of serotonergic fibers was observed in the granular layer. In the cases of medullary raphe grafts, the relatively abundant outgrowing serotonergic fibers showed a laminar organization in the cerebellar cortex near the graft, which was similar to the normal distributional pattern. These results indicate that serotonergic and dopaminergic neurons located within the mesencephalic raphe graft clearly differed from each other in their ability to extend their processes into the host cerebellum, which provides further evidence for the existence of specific interactions between outgrowing serotonergic fibers and their terminal fields (targets).

Distribution of the endogenous digitalis-like substance (EDLS)-containing neurons labeled by digoxin antibody in hypothalamus and three circumventricular organs of dog and macaque.

Endogenous digitalis-like substance (EDLS) is a newly discovered humoral agent which causes sodium-diuresis. EDLS is well known to have inhibitory activity to Na+,K(+)-ATPase and cross-immunoreactivity to digoxin antibody; however, its precise chemical structure has not yet been determined. We had previously developed a histochemical technique for EDLS, i.e., digoxin-immunohistochemistry, and demonstrated that EDLS was produced in the hypothalamic neurons. In the present study, the distribution of EDLS-containing neurons in the hypothalamus of dog and macaque was investigated using this technique, because anti-EDLS antibody cannot be obtained yet. In both species, EDLS neuronal somata were mainly localized in the paraventricular nucleus and the supraoptic nucleus and its accessory nuclei. A number of somata were also scattered in the other hypothalamic areas. The processes of these neurons ran from the area where the somata were located, through the lateral and basal area of the hypothalamus, to the infundibulum. These nerve fibers with varicosities were associated with the primary capillaries of hypophysial portal veins. A few immunopositive nerve fibers were also seen in the pituitary posterior lobe of both species. Intensive immunoreactivities were observed in the subfornical organ and organum vasculosum laminae terminalis. There were no differences between dog and macaque.

Changes of immunoreactive neuropeptide Y, somatostatin and corticotropin-releasing factor (CRF) in the brain of a novel epileptic mutant rat, Ihara's genetically epileptic rat (IGER).

Ihara's genetically epileptic rat (IGER) is a rat mutant with genetically scheduled spontaneous convulsions mimicking human limbic seizures. In the present study, the possible changes of three neuropeptides, neuropeptide Y (NPY), somatostatin (SRIF) and corticotropin-releasing factor (CRF), in the brains of IGER were investigated. Increased contents of immunoreactive (IR) NPY were found only in the hippocampus of 2-month IGERs before developing convulsive seizures, while similar increases of IR-NPY were discovered in the striatum and pyriform and entorhinal cortex as well as hippocampus in 8-month IGERs with repetitive seizures. There were no significant differences in the brain contents of IR-SRIF and IR-CRF between IGERs and the controls at both ages. These findings indicate an enhanced rate of NPY synthesis in this experimental model of epilepsy which may play a critical role in the development of epileptogenesis.

Altered hippocampal expression of neuropeptide Y, somatostatin, and glutamate decarboxylase in Ihara's epileptic rats and spontaneously epileptic rats.

By in situ hybridization and immunocytochemistry, expression of neuropeptide Y (NPY), somatostatin and glutamate decarboxylase 65 (GAD65) was studied in the hippocampus of two different epileptic mutant rats, Ihara's epileptic rat (IER) and the spontaneously epileptic rat (SER). GAD65 mRNA expression was enhanced in interneurons of the hippocampus in young IER, that had not yet developed generalized seizures. In older IER and older SER that both showed spontaneous seizures, marked increases of NPY mRNA in hippocampal granule cells and interneurons were found, as well as elevated GAD65 mRNA levels in interneurons. NPY immunoreactivity was enhanced in hilar interneurons and the dentate gyrus of older IER. In addition, some older IER stained heavily for NPY in mossy fibers. These findings suggest that up-regulation of NPY and GAD65 synthesis may be important in epileptogenesis.

Cell killing and mutation induction by heavy ion beams.

Carbon beam radiotherapy for cancer patients was initiated in Japan in June 1994. This study attempts to clarify the radiobiological effects of heavy ion beams. In this study, human cancer cell lines (RMG-1, MDA-MB231) and V79 cells were used. The cell killing was determined by colony forming assay, and mutation induction was determined by counting the number of 6-thioguanine resistant colonies (hprt locus mutation assay). The cell lines were irradiated with carbon (20 or 80 keV/microm) or neon beams (80 keV/microm). Carbon ions with a higher LET value (80 keV/microm) had an enhanced cytotoxic effect compared to those with a lower LET value (20 keV/microm). Carbon beams produced a slightly stronger cytotoxic effect than neon beams when irradiated at the same LET level (80 keV/microm), but the difference was not remarkable. The mutant fraction was significantly higher in all cell lines when they were irradiated with heavy ion beams, compared to the results for X-ray irradiation. The mutant fraction increased when the LET of the carbon beams increased. At equivalent LET values, the mutant fraction was lower for neon beams than for carbon beams. Fractionation of carbon beam irradiation had no effect on survival, but reduced the mutant fraction. Neon beams might be more appropriate for heavy ion therapy, especially when higher doses are being used. In addition, the fractionation of heavy ion beam administration might be appropriate for reducing the mutant fraction.