Genome-wide association study of NMDA receptor coagonists in human cerebrospinal fluid and plasma.

The N-methyl-D-aspartate receptor (NMDAR) coagonists glycine, D-serine and L-proline play crucial roles in NMDAR-dependent neurotransmission and are associated with a range of neuropsychiatric disorders. We conducted the first genome-wide association study of concentrations of these coagonists and their enantiomers in plasma and cerebrospinal fluid (CSF) of human subjects from the general population (N=414). Genetic variants at chromosome 22q11.2, located in and near PRODH (proline dehydrogenase), were associated with L-proline in plasma (ß=0.29; P=6.38 × 10(-10)). The missense variant rs17279437 in the proline transporter SLC6A20 was associated with L-proline in CSF (ß=0.28; P=9.68 × 10(-9)). Suggestive evidence of association was found for the D-serine plasma-CSF ratio at the D-amino-acid oxidase (DAO) gene (ß=-0.28; P=9.08 × 10(-8)), whereas a variant in SRR (that encodes serine racemase and is associated with schizophrenia) constituted the most strongly associated locus for the L-serine to D-serine ratio in CSF. All these genes are highly expressed in rodent meninges and choroid plexus, anatomical regions relevant to CSF physiology. The enzymes and transporters they encode may be targeted to further construe the nature of NMDAR coagonist involvement in NMDAR gating. Furthermore, the highlighted genetic variants may be followed up in clinical populations, for example, schizophrenia and 22q11 deletion syndrome. Overall, this targeted metabolomics approach furthers the understanding of NMDAR coagonist concentration variability and sets the stage for non-targeted CSF metabolomics projects.

Genome-wide association study of monoamine metabolite levels in human cerebrospinal fluid.

Studying genetic determinants of intermediate phenotypes is a powerful tool to increase our understanding of genotype-phenotype correlations. Metabolic traits pertinent to the central nervous system (CNS) constitute a potentially informative target for genetic studies of intermediate phenotypes as their genetic underpinnings may elucidate etiological mechanisms. We therefore conducted a genome-wide association study (GWAS) of monoamine metabolite (MM) levels in cerebrospinal fluid (CSF) of 414 human subjects from the general population. In a linear model correcting for covariates, we identified one locus associated with MMs at a genome-wide significant level (standardized ß=0.32, P=4.92 × 10(-8)), located 20 kb from SSTR1, a gene involved with brain signal transduction and glutamate receptor signaling. By subsequent whole-genome expression quantitative trait locus (eQTL) analysis, we provide evidence that this variant controls expression of PDE9A (ß=0.21; P unadjusted=5.6 × 10(-7); P corrected=0.014), a gene previously implicated in monoaminergic transmission, major depressive disorder and antidepressant response. A post hoc analysis of loci significantly associated with psychiatric disorders suggested that genetic variation at CSMD1, a schizophrenia susceptibility locus, plays a role in the ratio between dopamine and serotonin metabolites in CSF. The presented DNA and mRNA analyses yielded genome-wide and suggestive associations in biologically plausible genes, two of which encode proteins involved with glutamate receptor functionality. These findings will hopefully contribute to an exploration of the functional impact of the highlighted genes on monoaminergic transmission and neuropsychiatric phenotypes.

Quality of intraoperatively salvaged unwashed blood in hip arthroplasty.

BACKGROUND AND OBJECTIVES: Transfusion of autologous whole blood is one available method to reduce the need for allogenic blood transfusion. The aim of the present study was to investigate the quality of shed blood collected in a new intraoperative autotransfusion system. MATERIALS AND METHODS: In this prospective study fifteen consecutive patients who were scheduled for elective total hip arthroplasty were included. Shed blood was collected with a novel intraoperative autologous blood transfusion system (Sangvia®, AstraTech) from the surgical wound. Blood samples were taken from the transfusion bag. RESULTS: Mean blood loss during operation was 364ml (190-750ml) and mean transfused blood volume was 200ml (30-700ml). Mean haemoglobin concentration was 62g/l (17-91g/l) and mean plasma free haemoglobin concentration was 6.7g/l (1.9-12.7g/l) in transfusion blood. CONCLUSION: The basic laboratory characteristics of intraoperatively salvaged blood with the Sangvia® system are generally in the same range as reported in the studies on the postoperative transfusion of unwashed blood. From a blood quality point of view, our study indicates that transfusion of intraoperatively salvaged unwashed blood with the Sangvia® system in patients undergoing total hip arthroplasty is expected to be safe.

Peripheral blood gene expression profiles linked to monoamine metabolite levels in cerebrospinal fluid.

The blood-brain barrier separates circulating blood from the central nervous system (CNS). The scope of this barrier is not fully understood which limits our ability to relate biological measurements from peripheral to central phenotypes. For example, it is unknown to what extent gene expression levels in peripheral blood are reflective of CNS metabolism. In this study, we examine links between central monoamine metabolite levels and whole-blood gene expression to better understand the connection between peripheral systems and the CNS. To that end, we correlated the prime monoamine metabolites in cerebrospinal fluid (CSF) with whole-genome gene expression microarray data from blood (N=240 human subjects). We additionally applied gene-enrichment analysis and weighted gene co-expression network analyses (WGCNA) to identify modules of co-expressed genes in blood that may be involved with monoamine metabolite levels in CSF. Transcript levels of two genes were significantly associated with CSF serotonin metabolite levels after Bonferroni correction for multiple testing: THAP7 (P=2.8 × 10-8, ß=0.08) and DDX6 (P=2.9 × 10-7, ß=0.07). Differentially expressed genes were significantly enriched for genes expressed in the brain tissue (P=6.0 × 10-52). WGCNA revealed significant correlations between serotonin metabolism and hub genes with known functions in serotonin metabolism, for example, HTR2A and COMT. We conclude that gene expression levels in whole blood are associated with monoamine metabolite levels in the human CSF. Our results, including the strong enrichment of brain-expressed genes, illustrate that gene expression profiles in peripheral blood can be relevant for quantitative metabolic phenotypes in the CNS.

Closed-loop baroreflex control of total peripheral resistance in the cat: identification of gains by aid of a model.

The baroreflex regulation of total peripheral resistance was quantified in closed-loop conditions. To vary arterial pressure cardiac output was reduced by graded inferior caval vein occlusion or by arterial bleeding. In eight lightly anaesthetised cats the static relation between mean arteriovenous pressure gradient and mean flow could be described by a curve that was convex to the pressure axis and had zero intercept. The ratio of the change in resistance to a given change in arterial pressure was taken as resistance gain (GR). The value of this gain was estimated with the aid of a model which predicts pressure from flow when the right parameter value for gain is filled in. It consists of a non-linear negative-feedback control system with control pressure as reference point and with a constant gain. The estimation was carried out with the aid of an automatic identification procedure. GR varied from 0.002 to 0.010 min . ml-1 in different animals under light anaesthesia. With deeper anaesthesia gain decreased by 35 to 50% and became zero with very deep anaesthesia or barodenervation. Assuming that the reflex is a linear system about control pressure and flow we linearised our model and computed the static overall open-loop gain (GO). Within this narrow range GO varied from 0.64 to 2.30 for different cats under light anaesthesia and decreased by the same percentage as GR with deeper anaesthesia.

Endogenous adenosine enhances vagal negative chronotropic effect during hypoxia in the anaesthetised rabbit.

STUDY OBJECTIVE: Hypoxia potentiates the negative chronotropic effect of efferent vagal stimulation. A similar potentiation is evoked by exogenous adenosine. The aim of this study was to verify whether vagal potentiation during hypoxia is caused by endogenous adenosine. DESIGN: In anaesthetised rabbits the peripheral end of the right vagus was stimulated once every 20 s for 1 s, during normoxia and during systemic hypoxia, before and after adenosine receptor blockade. Hypoxia was induced by lowering oxygen content of the inspired air for 6 min. EXPERIMENTAL MATERIAL: 12 rabbits were anaesthetised with chloralose (50 mg.kg-1, intravenously) and halothane (0.3 vol%) and artificially ventilated. Reflex influences on heart rate were minimised by bilateral cervical vagotomy and administration of atenolol (1 mg.kg-1, followed by 0.25 mg.kg-1.h-1). Hypoxia was repeated before and after 8-phenyltheophylline administration (19.5 mumol.kg-1, intravenously) in seven rabbits, or before and after vehicle injection in five rabbits (time control). MEASUREMENTS AND RESULTS: The PaO2 attained at the end of the hypoxic period was 19(SEM 1) mm Hg [2.5(0.1) kPa]. Before adenosine receptor blockade, arterial pressure increased during hypoxia [14(6)mm Hg after 1 min], then decreased [7.3(8.8) mm Hg below control after 4 min]. Heart rate fell by 38.3(12.1) beats.min-1 in the last 3 min of hypoxia. Vagal negative chronotropic effect increased from -30.3(1.8) beats.min-1 during control to -58.7(4.6) beats.min-1 during the last 5 min of hypoxia, ie, a potentiation of 93.2(9)%. Administration of 8-phenyltheophylline reduced the effects of hypoxia on spontaneous heart rate and vagal bradycardia: heart rate decreased by 14.2(7.8) beats.min-1 and vagal negative chronotropic effect increased from -32.2(2.1) to -39.3(3.7) beats.min-1, ie, a potentiation of 21.5(10)%. Blood pressure showed a stronger increase [19.1(4.4) mm Hg after 2 min], but no decrease. These differences were not seen in the five control rabbits, in which hypoxia was repeated without adenosine receptor blockade. CONCLUSIONS: These results show that adenosine does play a role in hypoxia induced bradycardia and vagal potentiation.

Extracorporeal circulation can induce hypotension by both blood-material contact and pump-induced platelet aggregation.

OBJECTIVE: Use of extracorporeal systems in cardiopulmonary bypass and dialysis induces vascular reactions, which can lead to hypotension and lung edema. METHODS: To study the contribution of blood-material contact and use of a roller pump, as well as prevention of their adverse effects, we perfused a rat hind leg with a tube connecting a carotid and a femoral artery. RESULTS: Autoperfusion of an uncoated tube caused a fall of aortic pressure and femoral resistance to 66% +/- 16% and 76% +/- 15%, respectively, of their initial values within 2 hours, whereas in control animals without a shunt, these variables hardly changed (to 94% +/- 2.8% and 99% +/- 2.8%, respectively). Lung water content became significantly higher than that found in control animals (79.4% +/- 1.50% versus 77. 0% +/- 1.67%). If we coated the tube with albumin, these changes were largely prevented. When the coated tube was placed in a roller pump, aortic pressure and femoral resistance immediately fell to 79% +/- 17.2% and 63% +/- 13.5%, respectively, whereas lung water content did not increase. The vasodilation was caused by platelet aggregation and could be prevented with aurintricarboxylic acid, which inhibits shear-induced platelet aggregation by blocking the binding of von Willebrand factor to platelet glycoprotein Ib receptors. CONCLUSIONS: Extracorporeal circulation may induce hypotension and lung edema by means of blood-material contact. Hypotension can be prevented by coating the system with albumin but can still result from pump-induced platelet aggregation.

Pump-induced platelet aggregation in albumin-coated extracorporeal systems.

OBJECTIVE: Coating of extracorporeal systems with heparin does not prevent platelet activation and subsequent bleeding disorders. We investigated whether this could be due to elevated shear stress caused by a roller pump. METHODS: Human or rat blood was made to flow through an uncoated or an albumin-coated medical polyvinyl chloride tube with or without a roller pump. Aggregation of platelets in the tubing was recorded continuously with a photometric device. RESULTS: Although in vitro gravitational flow in uncoated tubes caused immediate platelet aggregation and platelet loss, this remained absent in coated tubes. When the pump was started in experiments with a coated tube strong platelet aggregation was observed and platelet count fell within 5 minutes to 78% +/- 2% and 71% +/- 3% of control values in human and rat blood, respectively. In vivo, no aggregation was observed during spontaneous flow in rats with an albumin-coated tube running from the carotid artery to the femoral artery, but aggregation started as soon as the blood was pumped. Pump-induced platelet aggregation, both in vitro and in vivo, could be prevented with aurintricarboxylic acid, which specifically inhibits shear-induced platelet aggregation as has recently been shown. Pump perfusion of blood in an uncoated tube did not elicit platelet aggregation. CONCLUSIONS: Pump perfusion of blood in coated systems elicits shear-induced platelet aggregation, which may be prevented by administration of substances that block the binding of von Willebrand factor to glycoprotein Ib receptors on the platelets. The effects of pumping on platelets are masked in uncoated circuits because of the dominant influence of blood-material contact.

Inhibition of efferent sympathetic nerve activity by centrally administered paraoxon in the cat.

We recently found that central administration of the cholinesterase inhibitor paraoxon lowered blood pressure substantially. It was postulated that the decrease in pressure was mediated by a reduction of sympathetic outflow. In the present study, efferent splanchnic nerve activity in anaesthetized and paralysed cats was recorded, and quantified by measuring the variance of signal amplitude. After administration of 8 micrograms paraoxon into the vertebral arteries, blood pressure and splanchnic nerve activity decreased simultaneously. A mean fall of 46 +/- 6% and 45 +/- 13% (mean +/- S.E.M.) respectively was reached within 12 min and was maintained during the period studied (30 min). When the effect of paraoxon was antagonized by dexetimide, both blood pressure and splanchnic nerve activity returned to control values. Since previous work has shown that the depressor action could not be prevented by efferent vagal blockade it seems likely that the fall in blood pressure after paraoxon was mainly caused by a decreased sympathetic outflow. In addition, we varied the amplifier band width in recording splanchnic nerve activity. The measurement of frequencies between 10 and 225 Hz appeared to be sufficient for studying the change in activity after paraoxon.

Systemic autoregulation counteracts the carotid baroreflex.

The interaction between autoregulation and baroregulation and its effect on the gains of the short-term pressure regulatory system was studied by performing both open- and closed-loop experiments in the same five anesthetized, vagotomized dogs, and by analyzing the data making use of a new model. With carotid pressure constant (no baroregulation) the pressure-flow data were convex to the flow axis, thus indicating the presence of autoregulation. When baroregulation was present the data were convex to the pressure axis. Our model was able to fit the data as measured in both cases. From the fitting procedure the zero-flow pressure intercept Pzf, the autoregulation resistance gain Gra, and the baroregulation resistance gain Grb were estimated. Pzf was about 20 mmHg in three dogs and about zero in the other two. Average values of Gra and Grb were 13.0 +/- 3.5 mmHg min2/L2 and 0.83 +/- 0.25 min/L, respectively. The two curves which fitted the data points collected in the presence and in the absence of baroreflex intersected at a point (Qo, Po) generally different from the control point. We determined the open-loop gain, Goc = GrbQo, about the point (Qo, Po). The averaged value was 2.23 +/- 0.84. When autoregulation was neglected, the resistance gain Grb and the open-loop gain Goc obtained from the same closed-loop method were underestimated (0.32 +/- 0.15 min/L and 0.88 +/- 0.48, respectively). In the open-loop preparation the carotid sinuses were isolated and the aortic (P) versus carotid (Pca) pressure data were collected. A third-order polynomial was fitted to these data.(ABSTRACT TRUNCATED AT 250 WORDS)

Pump perfusion causes vasodilation by activation of platelets.

Use of a pump in extracorporeal circuits depresses autoregulation and vascular tone. To study whether platelets are involved, we perfused rat hindlegs by means of an extracorporeal shunt between carotid and femoral artery. Autoperfusion could instantaneously be replaced by pump perfusion. To avoid interference by effects caused by blood-material contact, the circuit was coated with albumin. Spontaneous flow did not elicit platelet aggregation as recorded continuously with a photometric device inserted into the tubing, nor did it affect femoral vascular resistance. However, pump perfusion immediately evoked strong platelet aggregation that stabilized at a lower level after 2-3 minutes. Femoral resistance rose slightly during the first 2 minutes, but thereafter fell to 63% of control and stayed at approximately 70% for the next 2 hours. Pump induced platelet aggregation and fall in vascular resistance could be prevented with aurintricarboxylic acid, which specifically inhibits shear induced platelet aggregation. We conclude that pump perfusion with blood in coated systems elicits shear-induced platelet aggregation that, in turn, leads to vasodilation in the perfused vascular bed. These effects can be prevented by blocking the binding of von Willebrand factor to the platelet glycoprotein Ib receptors.

Small changes in heart rate following alpha-1 adrenoceptor stimulation in the anaesthetized dog.

In a previous work (1) we observed a weak alpha-1 adrenoceptor mediated chronotropic effect in anaesthetized dogs: the intracoronary injection of 100 micrograms of amidephrine, an alpha-1 agonist, increased heart rate by 2.5 +/- 0.8 bpm (mean +/- SEM). Since these experiments had been performed in the presence of alpha-2 blockade with yohimbine, one could argue that alpha-1 adrenoceptors had been partially blocked as well. To test for this possibility 5 additional experiments were performed with the same protocol, just omitting yohimbine administration. The chronotropic effect of amidephrine was larger (6.2 +/- 1.9 bpm after i.c. injection of 100 micrograms), but the difference was not significant. This confirms our earlier finding that alpha-1 adrenoceptors are not involved in heart rate control of the anaesthetized dog.

Seasonal variation of serotonin turnover in human cerebrospinal fluid, depressive symptoms and the role of the 5-HTTLPR.

Studying monoaminergic seasonality is likely to improve our understanding of neurobiological mechanisms underlying season-associated physiological and pathophysiological behavior. Studies of monoaminergic seasonality and the influence of the serotonin-transporter-linked polymorphic region (5-HTTLPR) on serotonin seasonality have yielded conflicting results, possibly due to lack of power and absence of multi-year analyses. We aimed to assess the extent of seasonal monoamine turnover and examined the possible involvement of the 5-HTTLPR. To determine the influence of seasonality on monoamine turnover, 5-hydroxyindoleacetic acid (5-HIAA) and homovanillic acid (HVA) were measured in the cerebrospinal fluid of 479 human subjects collected during a 3-year period. Cosine and non-parametric seasonal modeling were applied to both metabolites. We computed serotonin (5-HT) seasonality values and performed an association analysis with the s/l alleles of the 5-HTTLPR. Depressive symptomatology was assessed using the Beck Depression Inventory-II. Circannual variation in 5-HIAA fitted a spring-peak cosine model that was significantly associated with sampling month (P=0.0074). Season of sampling explained 5.4% (P=1.57 × 10(-7)) of the variance in 5-HIAA concentrations. The 5-HTTLPR s-allele was associated with increased 5-HIAA seasonality (standardized regression coefficient=0.12, P=0.020, N=393). 5-HIAA seasonality correlated with depressive symptoms (Spearman's rho=0.13, P=0.018, N=345). In conclusion, we highlight a dose-dependent association of the 5-HTTLPR with 5-HIAA seasonality and a positive correlation between 5-HIAA seasonality and depressive symptomatology. The presented data set the stage for follow-up in clinical populations with a role for seasonality, such as affective disorders.

Peripheral resistance after cardiac output reduction in the barodenervated cat.

Studies on the nervous or humoral control of total peripheral resistance are often complicated by concomitant changes in cardiac output. We studied the influence of cardiac output on peripheral resistance in the absence of modulating reflexes. In barodenervated and vagotomized cats, cardiac output was varied by graded inferior caval vein occlusion or by arterial bleeding. Total peripheral resistance was obtained with an analogue device which continuously divided the pressure difference between aorta and caval vein by cardiac output (electromagnetic flowmeter). Cardiac output reduction caused a decrease of peripheral resistance, followed within 2 minutes by a slow increase. Resistance stabilized at preocclusion levels within 5.8 (range 4-9) minutes. The relative changes in resistance and cardiac output were linearly related, when cardiac output was reduced by less than 40%. With larger reductions, the relation became nonlinear, and with a drop of more than 65%, no further change was noticed. These changes in resistance could not be explained by variations in blood viscosity as measured by Hct. They were nonnervous in nature: when all reflexes were abolished by ganglionic blockade, a similar pattern was found. Humoral mechanisms like the vasopressin or the renin-angiotensin system, known to be activated by hypotension, probably played no role, since arterial osmolality remained stable and captopril did not influence the resistance response. The involvement of metabolic autoregulation could not be excluded, but was unlikely because O2 consumption and serum lactate did not change.

Respiratory fluctuations in pupil size.

Regular fluctuations in pupil size of the cat were measured and the properties, nervous pathways, and origin of these oscillations were investigated. The rhythm of pupil movements under control conditions appeared to be either locked to the central respiratory cycle or to the artificial ventilatory cycle. These movements were only seen in lightly anesthetized or tranquilized cats, but not in alert or deeply anesthetized cats (ether, halothane or pentobarbital). The fluctuations proved to be independent of sympathetic innervation but related to variations in parasympathetic outflow. At least two sources for pupil oscillations appeared to be involved: central respiratory activity and respiratory blood pressure fluctuations that modulated pupil width via sinoaortic baroreceptors. Lung movements per se, as a third possible factor, did not modulate pupil width, whereas electrical stimulation of the afferent lung vagi did; therefore the role of this mechanical factor is not clear. A review of the pertinent literature shows that in the organism there are many phenomena exhibiting respiratory oscillations. It seems likely that these oscillations have the same origin as the respiratory pupil fluctuations.

Relationship between strength of short-term systemic autoregulation and initial resistance.

The relationship between strength of short-term whole body autoregulation and peripheral resistance in the reference state (initial resistance) was investigated in 9 anesthetized closed-chest dogs and 18 anesthetized open-chest cats. Baroreflex regulation was abolished in one of three ways: barodenervation, ganglionic blockade, or setting pressure constant in the isolated carotid sinuses after vagotomy. Ascending aortic pressure and flow and venous pressure were measured in the reference state and 1-3 min after partial occlusions of the inferior vena cava. Cardiac output and peripheral resistance (ratio between arteriovenous pressure difference and cardiac output) were normalized for body weight. Strength of autoregulation was quantified by a resistance gain (Gra), defined as the ratio between change in normalized peripheral resistance and corresponding change in normalized cardiac output. A broad range of values for peripheral resistance in the reference state (Ro) was obtained as a result of the different interventions used to abolish baroreflex regulation. Arteriovenous pressure difference and normalized cardiac output during multiple vena cava occlusions in the 9 dogs and in 8 of the cats were fitted with a parabola convex to the flow axis. From the best fit, Gra was estimated. In the remaining 10 cats Gra was estimated from a single occlusion of vena cava. When data of all dogs and cats were taken together, we found a linear relationship between Gra and Ro: Gra = K1.Ro + K2. The constants K1 and K2 were 17.9 x 10(-3) min.kg.ml-1 and -14.5 x 10(-3) mmHg.min2.kg2.ml-2, respectively. The correlation coefficient was 0.9.