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.

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.

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.

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.

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.

In vivo visualization of hemodialysis-induced alterations in leukocyte-endothelial interactions.

BACKGROUND: The aim of this study was to develop a model for hemodialysis (HD) in small animals using conventional dialysis equipment that would allow the intravital microscopic observation of leukocyte-endothelial interactions in vivo. METHODS: Cuprophan dialyzers were adapted to obtain a similar ratio of membrane area to blood volume as in clinical HD. A silicone ring was inserted into the dialyzer's inlet to limit the number of blood-perfused capillaries. Rabbits were dialyzed for one hour without a dialysate flow. RESULTS: Extracorporeal circulation with the cuprophan dialyzer resulted in a transient leukopenia and complement activation. At the nadir of leukopenia, leukocytes that rolled along the venular wall were scarcely observed, whereas rolling was abundant (54 +/- 9 per min) prior to extracorporeal circulation. The adhesion of leukocytes to the vascular endothelium was not induced. After 60 minutes, rolling of leukocytes was still reduced by 73 +/- 5.5%, despite the full recovery of circulating leukocyte counts. Extracorporeal circulation without a dialyzer also tended to reduce leukocyte rolling, although systemic leukocyte counts were not affected. CONCLUSIONS: The use of adapted conventional cuprophan hemodialyzers in rabbits yielded a transient leukopenia similar to that in clinical HD. Using intravital microscopy, we demonstrated impairment of leukocyte-endothelial interactions. In addition, our data indicate that tissues, in which leukocytes can roll and adhere, are not automatically sites of leukocyte sequestration during HD-induced leukopenia.

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.

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.

Minoxidil-induced cardiac hypertrophy in guinea pigs.

To investigate whether during cardiac hypertrophy changes occur in contractile protein composition and in mechanical and energetic properties of the myocardium, contractile protein composition, isometric force and adenosine triphosphate (ATP) consumption were studied in control and hypertrophied guinea-pig hearts. Cardiac hypertrophy was induced by adding minoxidil (120 or 200 mg/l) to the drinking water. Protein analysis was performed by one-dimensional gel electrophoresis. The myosin heavy-chain (MHC) composition was determined in an enzyme-linked immunosorbent assay (ELISA). ATP consumption and force development were simultaneously measured during isometric contraction in chemically skinned trabeculae. Histochemical analysis of cross-sectional area of cardiomyocytes and interstitial space was performed on the left ventricular tissue of 200 mg/l minoxidil-treated and control guinea pigs. Minoxidil treatment (120 and 200 mg/l) significantly increased left ventricular dry weight normalized for body weight by 19 +/- 4 and 24 +/- 4%, respectively. No significant differences were found in the cellular cross-sectional area, while interstitial space was slightly decreased in minoxidil-treated hearts. In left ventricular trabeculae of 200 mg/l minoxidil-treated guinea pigs, ATPase activity was slightly less than in those of control guinea pigs, whereas force did not differ significantly. Calcium sensitivity of force and ATPase activity were not affected by minoxidil treatment. Gel electrophoresis revealed no difference in contractile protein composition, but a tendency towards a lower amount of alpha-MHC in the minoxidil-treated hearts was found in ELISA.

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.

Vasodilation by shear-induced platelet aggregation in extracorporeal circuits.

Extracorporeal circulation may have adverse effects on vascular reactivity. To reduce such effects, we recently coated a tube connecting the carotid and the distal femoral artery of rats with albumin. When we partially occluded this perfusion line, the reduction of flow was followed by a marked increase, which seemed not to be caused by autoregulation but by release of a vasodilator at the site of occlusion. In the present study, we investigated whether this vasodilator could originate from platelets aggregating under the influence of increased shear stress at the site of occlusion. Blood distal to the site of occlusion indeed contained numerous platelet aggregates that were not present before occlusion. Continuous recording with a photometric device showed that aggregation in the tube started before flow increased and ended before flow decreased again. Blockade of serotonin S1- and S2-receptors with methiothepin prevented the flow response. Estimated shear stress (231 +/- 17 dyn/cm2) and shear rate (6,370 +/- 478 s-1) at the site of occlusion were of the magnitude known to elicit platelet aggregation. Others have recently demonstrated that shear-induced platelet aggregation is mediated by binding of von Willebrand factor to platelet glycoprotein Ib, which is inhibited by aurintricarboxylic acid. This drug (35 mg/kg iv) completely abolished both platelet aggregation and flow increase in our experiments. These results suggest that the vasodilation during partial tube occlusion is mediated by serotonin released from platelets that aggregate as a result of high shear stress.

Extracorporeal circuits and autoregulation: effect of albumin coating.

Autoregulation of muscle blood flow often is difficult to demonstrate when extracorporeal perfusion is used. This could be caused by contact of blood and foreign material. Accordingly, we tested whether autoregulation is preserved when the system is coated with albumin. Polyurethane tubing between the carotid and distal femoral artery of rats was partially occluded for 1-2 min. With the system uncoated (n = 6 rats) autoregulation was absent. With coated systems (n = 6 rats) the closed-loop gain (Gc) of the apparent autoregulation (0 < Gc < 1) for the pressure range from control (137 +/- 14) to 80 mmHg was 0.40 +/- 0.24 (mean +/- SD). In most cases autoregulation was preceded by a period of "superregulation": after a variable delay flow rose above control. When the distance between occluder and tip of femoral cannula was varied in another group of rats (n = 6), the delay (20-75 s) was linearly related to transit time (10-35 s) of blood. Flow increase thus seemed to be caused by a blood-borne vasodilator originating at the occlusion site and not by a myogenic or metabolic response to decreased pressure and flow. The vasodilator did not originate from the albumin coating. Partial occlusion of an extracorporeal shunt thus can increase flow to the perfused tissue when the system is coated with albumin. The reaction readily disappears when the system is not coated.

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)

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.

Short-term systemic autoregulation.

We studied total systemic autoregulation in closed-chest, chloralose-anesthetized dogs. Cardiac out-put (previously implanted electromagnetic flow probe on ascending aorta) and aortic pressure were varied by reducing venous return using a balloon catheter in the vena cava. Compensatory action of the baroreflex was prevented by bilateral vagotomy and isolation of both carotid sinuses. To avoid high vessel tone carotid sinus pressure was set at the original baseline value using a pressurized blood reservoir. With each balloon inflation aortic flow and aortic pressure decreased and stabilized in about 1 min. Pressure and flow were allowed to return to base-line values after each balloon inflation in an attempt to minimize the activation of slower regulatory mechanisms. The steady-state pressure-flow relations could be fitted with a sigmoidal curve. The mean quality (0 less than Q less than 1) of autoregulation in eight dogs was 0.41 +/- 0.08 (SD). Autoregulation was found in the pressure range from 42 to 140 mmHg. The early appearance of total systemic autoregulation suggests that, in the intact animal, it may counteract baroreflex control.

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.

Pump perfusion abolishes autoregulation possibly via prostaglandin release.

The influence of pump perfusion on autoregulation was studied in the hindleg of the halothane- and chloralose-anesthetized cat. Flow was measured with an electromagnetic flow probe in a tube between aorta and the vascularly isolated, denervated leg and varied with a calibrated occluder. Perfusion pressure was measured via a T-piece distal to the occluder. The steady-state pressure-flow relations could be fit with a sigmoidal curve. The mean closed-loop gain (0 less than Gc less than 1) for autoregulation in six cats was 0.46 +/- 0.11 (SD). When in these cats a roller pump was used, an almost linear pressure-flow relation was found (Gc = 0.01 +/- 0.09), while the resistance at control flow was decreased by 15 +/- 4%. Administration of indomethacin (5 mg/kg iv), a cyclooxygenase inhibitor, partly restored autoregulation during pumping (Gc = 0.34 +/- 0.09) and slowly increased the resistance to above its original value (20 +/- 13%). In six other cats, pump perfusion had no influence on autoregulation when started after indomethacin administration but resistance increased. This increase could not be prevented with ketanserin, a specific serotonin 2 receptor blocker. We conclude that pump perfusion abolishes autoregulation and decreases resistance via a process that involves prostaglandins. Blockade of the prostaglandin synthesis unmasks a slow vasoconstrictor influence in the bed.

Cardiac alpha-1 adrenoceptors are not involved in heart rate control of the anaesthetized dog.

To study the possible role of cardiac postsynaptic alpha-1 adrenoceptors in heart rate control of the anaesthetized open-chest dog we injected a specific alpha-1 agonist (amidephrine) into the right coronary artery or stimulated electrically the right stellate ganglion. Reflex influences were minimized by bilateral cervical vagotomy and de-afferentiation of both stellate ganglia. Activation of alpha-2, beta- and muscarinic receptors was prevented by intravenous administration of yohimbine, propranolol and atropine, respectively. Since alpha-1 receptor stimulation could affect heart rate indirectly via coronary constriction, a continuous intracoronary infusion of adenosine (0.25 mg/kg/h) was given. Amidephrine did not affect heart rate at the lower dose (1-10 microgram). After the highest dose (100 micrograms) the maximum variation in heart rate was an increase of 2.2 +/- 1.1 bpm at 3 min after injection (mean +/- SEM; P less than 0.05). This slight cardioacceleration was simultaneous with an aortic pressure rise of 13.8 +/- 3.4 mm Hg and it was abolished by alpha-1 blockade with prazosin (1 mg/kg i.v.). After propranolol (1 mg/kg +0.5 mg/kg/h) the residual positive chronotropic effect of sympathetic stimulation (12.2 +/- 4.0 bpm) was not significantly altered (13.8 +/- 5.7 bpm) by prazosin administration. Similar results were recorded without adenosine infusion. We conclude that in the anaesthetized dog chronotropic effects directly mediated by alpha-1 adrenoceptors either do not exist or lack physiological significance.