Systemic steroid pretreatment improves cerebral protection after circulatory arrest.

BACKGROUND: This study evaluates whether systemic steroid pretreatment enhances neuroprotection during deep hypothermic circulatory arrest (DHCA) compared with steroid in cardiopulmonary bypass (CPB) prime. METHODS: Four-week-old piglets randomly placed into two groups (n = 5 per group) were given methylprednisolone (30 mg/kg) into the pump prime (group PP), or pretreated intravenously 4 hours before CPB (group PT). All animals underwent 100 minutes of DHCA (15 degrees C), were weaned off CPB, and were sacrificed 6 hours later. Postoperative changes in body weight, bioimpedance, and colloid oncotic pressure (COP) were measured. Cerebral trypan blue content, immunohistochemical evaluation of transforming growth factor-beta1 (TGF-beta1) expression, and caspase-3 activity were performed. RESULTS: Percentage weight gain (group PP 25.0% +/- 10.4% versus group PT 12.5% +/- 4.0%; p = 0.036), and percentage decrease in bioimpedance (PP 37.2% +/- 14.5% versus PT 15.6% +/- 7.9%; p = 0.019) were significantly lower, whereas postoperative COP was significantly higher in group PT versus group PP (PT 15.3 +/- 1.8 mm Hg versus PP 11.6 +/- 0.8 mm Hg; p = 0.003). Cerebral trypan blue (ng/g dry tissue) was significantly lower in group PT (PT 5.6 x 10(-3) +/- 1.1 x 10(-3) versus PP 9.1 x 10(-3) +/- 5.7 x 10(-4); p = 0.001). Increased TGF-beta1 expression and decreased caspase-3 activity were shown in group PT. CONCLUSIONS: Systemic steroid pretreatment significantly reduced total body edema and cerebral vascular leak and was associated with better immunohistochemical indices of neuroprotection after DHCA.

Endothelin-1 myocardial clearance, production, and effect on capillary permeability in vivo.

Myocardial metabolism of endothelin-1 (ET-1) and its effect on coronary microcirculatory exchanges were obtained in anesthetized dogs by combining the indicator-dilution technique with immunoreactive ET-1 measurements. The myocardium extracted 17.7 +/- 4.6% of tracer ET-1 (n = 12). Simultaneously measured ET-1 levels in the aorta (0.97 +/- 0.46 pg/ml) and coronary sinus (0.96 +/- 0.53 pg/ml) were not different, supporting a production of ET-1 by the heart that balances the amount extracted. Intracoronary infusion of ET-1 (5 ng.kg-1.min-1) increased coronary sinus ET-1 levels approximately 50-fold, decreased coronary blood flow per unit of interstitial space by approximately 30% (P = 0.006), and increased myocardial microcirculatory transit times (n = 6). Permeability to albumin was unaffected by ET-1, whereas the permeability-surface area product for sucrose decreased following derecruitment of myocardial capillaries. We conclude that there is a normal myocardial metabolic balance of ET-1 and that the heart marginally contributes to circulating ET-1. Pharmacological doses of ET-1 may adversely affect myocardial metabolism by reducing blood flow and the permeability-surface area product for small circulating substances.

Tracer oxygen distribution is barrier-limited in the cerebral microcirculation.

The kinetics of tracer oxygen distribution in the brain microcirculation of the awake dog were investigated with the multiple indicator dilution technique. A bolus containing 51Cr-labeled red blood cells, previously totally desaturated and then resaturated with [18O]2 (oxygen), 125I-albumin, 22Na, and [3H]water, was injected into the carotid artery, and serial anaerobic blood samples were collected from the sagittal sinus over the next 30 seconds. The outflow recovery curves were analyzed with a distributed-in-space two-barrier model for water and a one-barrier model for oxygen. The analysis provided an estimate of flow per gram brain weight as well as estimates for the tracer water and oxygen rate constants for blood-to-brain exchange and tracer oxygen parenchymal sequestration. Flow to tissue was found to vary between different animals, in concert with parallel changes in oxygen consumption. The 18O2 outflow curves showed an early peak, coincident with and more than half the magnitude of its vascular reference curve (labeled red blood cells), whereas the [3H]water curve increased abruptly to a low-in-magnitude curve at low flow values and to a small early peak at high flow values. Analysis indicates that the transfers of both 18O2 and [3H]water indicators from blood to brain are barrier-limited, with the former highly so because of the large red blood cell capacity for oxygen, and that the proportion of the tracer oxygen returning to the circulation from tissue is a small fraction of the total tracer emerging at the outflow.

Effects of flow, perfusion pressure, and oxygen consumption on cardiac capillary exchange.

The roles of blood flow, local oxygen consumption, and perfusion pressure on cardiac transcapillary exchange were characterized in closed-chest anesthetized dogs by use of the multiple-indicator dilution technique. Occlusion of the carotid arteries or injection of dipyridamole increased coronary flow to significantly higher values compared with a group of animals in a basal state obtained in a previous study. Carotid occlusion resulted in a significant increase in perfusion pressure and myocardial oxygen consumption, whereas these two variables were significantly reduced after dipyridamole. For the whole group of animals, the capillary permeability-surface area product for sucrose increased with coronary flow, which appeared to be the important controller for this microcirculatory exchange parameter. Perfusion pressure and myocardial oxygen consumption also regulated permeability-surface area product values, although to a lesser extent than flow. The heterogeneity of transit times in the capillaries was reduced at high coronary flow values, despite large differences in the cardiac utilization of oxygen. The data suggest that cardiac capillary exchange responds mostly to hemodynamic changes originating at the precapillary level.

Microsphere and dilution measurements of flow and interstitial space in dog heart.

The effects of coronary flow on cardiac capillary permeability-surface area products and interstitial spaces were examined at rest and after hemodilution in the canine heart. Multiple-indicator-dilution experiments and left atrial injections of microspheres were carried out in closed-chest anesthetized animals at rest and after plasma expansion with dextran. Plasma expansion was utilized to produce a large increase in coronary perfusion compared with control conditions. Values for plasma flow per unit interstitial space, derived from analysis of the indicator-dilution data, were found to correlate closely with average vascular plasma flow per gram, calculated from the cardiac microsphere data; the one reflects the other. With an increase in flow, cardiac capillary permeability-surface area product values were found to increase substantially, whereas the average sucrose extravascular or cardiac interstitial spaces remained stable. Consequently the dilution parameter, flow per unit interstitial space, which is independent of tracer loss, provided a good reflection of flow per weight of tissue in the heart, without the additional requirement for a flow probe.

The capillary transport system for free fatty acids in the heart.

The nature of the process by which free fatty acids, which are tightly bound to albumin, traverse the endothelium of cardiac capillaries to reach the cardiac muscle cells, so that they are extracted to a net extent of approximately 40%, needs clarification. Previous studies have indicated that a membrane fatty acid-binding protein provides for carrier-mediated uptake of free fatty acids by isolated hepatocytes, cardiomyocytes, and jejunal mucosal cells. A monoclonal monospecific antibody was prepared against purified membrane fatty acid-binding protein from rat liver. Multiple-indicator dilution experiments were carried out in the isolated rat heart with labeled albumin, sucrose, and palmitate in the presence of control perfusate or perfusate containing either specific antibody or comparable nonspecific myeloma cell supernatant (each of the latter containing additional albumin, in identical concentrations). Analysis of the labeled-sucrose curves provided a permeability-surface area product for sucrose to which that for palmitate could be compared. In comparison with control supernatants, myeloma supernatant produced a minor inhibition of palmitate uptake, as a result of the increase in albumin concentration. The specific antibody, which contained identical albumin concentrations, produced a major inhibition of palmitate uptake, significantly greater than with the myeloma supernatant. The data indicate that the membrane fatty acid-binding protein mediates the transfer of free fatty acid across the endothelial cells of cardiac capillaries for presentation to heart muscle. Passive intramembrane lateral diffusion of palmitate could not provide an explanation for the findings.

Constitutive nonexocytotic norepinephrine release in sympathetic curves of in situ canine heart.

Nonexocytotic norepinephrine bulk overflow from sympathetic nerves has been previously demonstrated in the perfused isolated heart only after inhibition of sympathetic nerve metabolism by hypoxia, ischemia, or metabolic inhibitors. The measurement, however, ignores simultaneous uptake of norepinephrine by sympathetic nerves. We quantitated simultaneous norepinephrine uptake and release in pentobarbital-anesthetized dogs by the use of a transient tracer approach, the multiple indicator dilution technique, combined with measurement of endogenous arterial and venous plasma norepinephrine levels. Sympathetic vesicles were previously depleted by reserpine, and desipramine was used to inhibit the neuronal membrane pump. Labeled albumin, sucrose, and norepinephrine were injected into the coronary artery, and sequential samples were collected from the coronary sinus. Reserpine pretreatment significantly depleted tissue and decreased plasma norepinephrine levels; tracer norepinephrine uptake increased slightly, and there was a substantial decrease in the rate of release of unlabeled norepinephrine. Desipramine then decreased tracer uptake and virtually eliminated norepinephrine release. We conclude that desipramine-suppressible, constitutive nonexocytotic norepinephrine release is present in cardiac sympathetic nerves.

A method of dynamic analysis of iodine-123-metaiodobenzylguanidine scintigrams in cardiac mechanical overload hypertrophy and failure.

Cardiac sympathetic neuronal degeneration accompanies mechanical overload heart failure. We hypothesized that sympathetic nerve and myocyte failure share a common etiology and that 123I-metaiodobenzylguanidine (MIBG) might provide a precise method of detecting failure in chronic mechanical overload. Our aim was to develop a method for the dynamic analysis of 123I-MIBG scintigrams which could yield a quantitative index of myocardial sympathetic neuronal function in this condition. We performed serial 123I-MIBG scintigraphy in 33 volunteers, 10 orthotopic cardiac transplant recipients and 26 patients with chronic mechanical overload of the left ventricle. We constructed a compartmental model in which total heart activity represents the sum of cardiac sympathetic vesicular and cytosolic pools. Patients with antecedent mechanical overload heart failure or myocardial dysfunction had accelerated myocardial egress of tracer that we ascribed to a specific impairment in vesicular storage rather than to a more rapid turnover of an intact vesicular pool.

Flow-limited tracer oxygen distribution in the isolated perfused rat liver: effects of temperature and hematocrit.

We used the multiple-indicator dilution technique to examine the kinetics of tracer oxygen distribution and uptake in the rat liver perfused in a nonrecirculating fashion with blood. 51Cr-labeled 18O2-saturated erythrocytes, labeled albumin, sucrose and water (the tracers for oxygen and vascular, interstitial and cellular references) were injected simultaneously into the portal vein. Timed anaerobic samples were collected from the hepatic vein and analyzed by mass spectrometry for relative 18O2 enrichment and radioactivity. In a set of experiments performed at 32 degrees C, oxygen uptake was substantially diminished; tracer oxygen profiles approached those expected for a completely recovered, flow-limited substance. At 37 degrees C, much larger tracer oxygen sequestration occurred. Experiments were carried out at each temperature at higher and lower hematocrit, and oxygen consumption at each temperature was found to be independent of hematocrit. The tissue space of distribution for tracer oxygen relative to the total sinusoidal vascular content was influenced by the hematocrit: it was smaller at higher hematocrit and larger at lower hematocrit, as expected. The derived partition coefficient of oxygen for liver cells relative to plasma (expressed in terms of the liver and plasma water spaces) was, on average, 2.62 ml/ml; it was independent of the hematocrit. Analysis of the indicator dilution experiments indicates that the tracer oxygen is distributed into tissue in a flow-limited rather than a barrier-limited fashion, and that with this, an ongoing concomitant intracellular sequestration of tracer can be seen.

Cardiac microcirculatory effects of beta-adrenergic blockade during sympathetic stimulation.

The effects of beta-adrenergic blockade on cardiac transcapillary exchange were examined at rest and during sympathetic stimulation. Multiple indicator dilution experiments were carried out in closed-chest anesthetized dogs at rest and during carotid occlusion, either under basal conditions or after beta-adrenergic blockade with alprenolol. beta-Adrenergic blockade at rest had no effect on coronary flow or transcapillary exchange in comparison with the control situation, but it abolished the increase in coronary flow and in the permeability/surface area product for labeled sucrose produced by carotid occlusion. High coronary resistance values in beta-blocked animals with carotid occlusion were associated with a high degree of heterogeneity in capillary transit times, but the overall relation between coronary flow and the capillary permeability/surface area product was unchanged. The findings indicate that beta-blockade increases coronary resistance during sympathetic stimulation and, simultaneously, decreases the coronary blood flow and capillary permeability/surface area product while increasing the heterogeneity of capillary transit times.

Use of norepinephrine uptake to measure lung capillary recruitment with exercise.

We used the multiple indicator-dilution technique with norepinephrine, a vascular endothelium surface marker, to study the pulmonary vascular changes in awake exercising dogs. The vascular space tracers, labeled erythrocytes and albumin, and a water space tracer, 1,8-octanediol, were injected with the norepinephrine, and right atrium-aortic root dilution curves were obtained in nine dogs, at rest and at two increasing levels of exercise. Extravascular lung water multiple tracer dilutional estimates increased with flow and rapidly approached a maximal asymptotic value representing 75% of the postmortem lung weight. The ratio of the extravascular lung water measured in this way to that measured gravimetrically also increased, to reach an asymptotic proportion of close to 100%. The transit time-defined central vascular space increased linearly with flow; the ratio of lung tissue space to lung vascular space, therefore, decreased with increasing flow. The mean tracer upslope norepinephrine extractions at rest and at the two levels of exercise were 17 +/- 1.2, 14 +/- 0.8, and 15 +/- 0.8% (SE). With the use of the Crone approximation, we computed permeability-surface area products for norepinephrine; these increased linearly with flow. If permeability does not change, the increase in the permeability-surface area product with flow can be attributed to capillary recruitment. We conclude that when all lung tissue has become accessible to 1,8-octanediol delivered via the perfused vascular space, there is nevertheless further recruitment, with increase in flow, of vascular surface that can extract norepinephrine.

Handling of tracer bicarbonate by the liver. The relative impermeability of hepatocyte cell membranes to the ionic species.

The multiple indicator dilution technique was used to study transfer of labeled HCO3- across the hepatocyte membrane in the anesthetized mongrel dog. A bolus of H[14C]O3-, 51Cr-labeled erythrocytes, [36Cl-] and/or [3H]sucrose, and [3H]OH was injected through a catheter in the portal vein, and timed anaerobic blood samples were obtained from a catheter in the hepatic vein. Experiments were carried out in untreated controls and after intravenous infusion of acetazolamide (100 mg/kg). In the controls, the H[14C]O3- curve was very similar to the [3H]OH curve. The dilution curves were all linear transformations of each other, indicating that HCO3-, as had previously been shown for the other diffusible tracers, undergoes delayed-wave flow-limited distribution. The distribution space for H[14C]O3- in the control situation includes the blood plasma and interstitial spaces, the erythrocyte interior modified by a Donnan equilibrium, and the available liver cellular space. The calculated HCO3- concentration in the liver cells was somewhat lower than that in the plasma space; the difference implied a cellular pH lower than that of plasma by approximately 0.08 pH units. When the carbonic anhydrases were inhibited with acetazolamide, the dilution curve for H[14C]O3- changed radically, approaching that for [36Cl-], which does not enter the liver cells. The change indicates that although HCO3-, like Cl-, is rapidly exchanged between plasma and erythrocytes, it also does not readily penetrate hepatocytes unless previously transformed to carbon dioxide by the carbonic anhydrases.

Xenon handling in the liver: red cell capacity effect.

Xenon, despite its lack of chemical reactivity, associates preferentially with red cells in blood. To characterize the effect of this and the nature of xenon-tissue interaction in the liver, multiple indicator dilution studies were performed in the anesthetized normal dog through portal vein injection and hepatic vein collection of anaerobic blood samples. Two experimental runs were carried out in each animal, one at the prevailing hematocrit and the other at reduced hematocrit after bleeding and replacement with dextran. For comparison, the injection mixtures contained labeled red blood cells (a vascular reference), sucrose (an interstitial space reference), and labeled water (which freely enters liver cells), as well as labeled xenon. At the higher hematocrit, the labeled xenon curves generally rose earlier, peaked higher, and decayed more quickly than the labeled water curve; at the lower hematocrit, the xenon curve was delayed and diminished in magnitude in relation to the labeled water curves. Analysis of the curve shapes indicated that xenon, like labeled sucrose and water, underwent delayed wave flow-limited distribution. With knowledge of the red cell plasma partition coefficient (2.89 ml/ml), it was possible to both account for the change in form of the xenon curves with hematocrit and to use the data to estimate the liver cell tissue plasma xenon partition coefficient. Values averaged 1.93 ml/ml liver space, or 1.79 ml/g, and did not change significantly from first to second runs. Theoretical analysis indicated that flow cannot be estimated from xenon downslopes.

Metaiodobenzylguanidine [131I] scintigraphy detects impaired myocardial sympathetic neuronal transport function of canine mechanical-overload heart failure.

In heart failure secondary to chronic mechanical overload, cardiac sympathetic neurons demonstrate depressed catecholamine synthetic and transport function. To assess the potential of sympathetic neuronal imaging for detection of depressed transport function, serial scintigrams were acquired after the intravenous administration of metaiodobenzylguanidine [131I] to 13 normal dogs, 3 autotransplanted (denervated) dogs, 5 dogs with left ventricular failure, and 5 dogs with compensated left ventricular hypertrophy due to a surgical arteriovenous shunt. Nine dogs were killed at 14 hours postinjection for determination of metaiodobenzylguanidine [131I] and endogenous norepinephrine content in left atrium, left ventricle, liver, and spleen. By 4 hours postinjection, autotransplanted dogs had a 39% reduction in mean left ventricular tracer accumulation, reflecting an absent intraneuronal tracer pool. Failure dogs demonstrated an accelerated early mean left ventricular tracer efflux rate (26.0%/hour versus 13.7%/hour in normals), reflecting a disproportionately increased extraneuronal tracer pool. They also showed reduced late left ventricular and left atrial concentrations of tracer, consistent with a reduced intraneuronal tracer pool. By contrast, compensated hypertrophy dogs demonstrated a normal early mean left ventricular tracer efflux rate (16.4%/hour) and essentially normal late left ventricular and left atrial concentrations of tracer. Metaiodobenzylguanidine [131I] scintigraphic findings reflect the integrity of the cardiac sympathetic neuronal transport system in canine mechanical-overload heart failure. Metaiodobenzylguanidine [123I] scintigraphy should be explored as a means of early detection of mechanical-overload heart failure in patients.

Lack of liver vascular response to carotid occlusion in mildly acidotic dogs.

The effect of mild acidosis on the reduction in liver vascular volume provoked by reflex sympathetic activation was ascertained in dogs by use of the multiple indicator-dilution technique. Portal vein-hepatic vein dilution patterns were obtained following injection of a mixture containing 51Cr-labeled red blood cells (a vascular reference), 14C-labeled sucrose (an interstitial reference), and 3H-labeled water (a cellular reference). Liver vascular, interstitial, and cellular water spaces were measured in normal and mildly acidotic dogs under basal conditions and during bilateral carotid artery occlusion. Carotid occlusion resulted in a large increase in arterial blood pressure in normal and a lesser increase in acidotic dogs, but the increase in portal and hepatic vein norepinephrine and decrease in liver vascular and interstitial volumes observed in normal dogs did not occur in the acidotic animals; no change in labeled water accessible liver cellular spaces was perceptible. The data indicate that mild acidosis abolishes the ordinarily expected reduction in liver vascular volumes on sympathetic activation in the intact dog but not the arterial pressor response.

Decreased basal cardiac interstitial norepinephrine release after neuronal uptake inhibition in dogs.

The effect of neuronal uptake inhibition on basal interstitial release of norepinephrine in the canine heart was examined by use of the multiple tracer dilution-bulk balance technique. A kinetic model incorporating the effects of flow, capillary permeability-surface product for norepinephrine, the interstitial uptake rate constant for neurotransmitter, and plasma norepinephrine input and output values was used to estimate rates of uptake from and release of norepinephrine into the interstitial space. The intravenous injection of the neuronal uptake inhibitor desipramine in anesthetized dogs under basal conditions reduced interstitial uptake of tracer norepinephrine in the heart, without significant changes in plasma concentration of norepinephrine in aorta and coronary sinus. The lack of change in the arteriovenous balance for norepinephrine across the heart, in the face of the lowered uptake for this amine, suggested that the liberation of norepinephrine by cardiac sympathetic fibers was reduced. Analysis of the data with the norepinephrine tracer kinetic-bulk model showed that, after desipramine, the interstitial release of norepinephrine was reduced to the same extent as uptake was diminished. As a result, the concentration of norepinephrine in the extracellular space of the heart did not increase significantly. The findings indicate the presence of a presynaptic neuronal feedback inhibition of release, which serves to fine tune the myocardial interstitial concentration of norepinephrine in the basal state; with this, after desipramine, both norepinephrine uptake and release are correspondingly diminished.

Plasma expansion effect on cardiac capillary and adrenergic exchange in intact dogs.

The effect of plasma volume expansion on transcapillary exchange and norepinephrine release in the heart was examined in pentobarbital sodium-anesthetized dogs by use of the multiple indicator-dilution technique. Animals were studied under basal conditions and following infusion of the plasma expander, dextran. Catheters were placed in coronary artery and coronary sinus in a closed-chest preparation. Labeled albumin, sucrose, and norepinephrine were injected into the coronary artery and outflow-dilution curves were secured. Analysis of these provided parameters reflecting coronary flow and permeability-surface product, and a norepinephrine tracer kinetic-bulk model provided simultaneous estimates of the rate of norepinephrine release into the myocardial interstitial space. The infusion of dextran resulted in a large increase in coronary flow without significant changes in myocardial norepinephrine release; at the same time the permeability-surface product values increased, amplifying the capacity of the higher flow to deliver substrates to sarcolemmal cells. The findings indicate that plasma volume expansion increases transcapillary exchange in the heart without activating the cardiac sympathetic system.

Tracer norepinephrine kinetics in coronary circulation of patients with heart failure secondary to chronic pressure and volume overload.

Controversy exists over the nature of the abnormality in cardiac sympathetic nerves in heart failure. In the cardiomyopathy of the Syrian hamster, reduction in tissue stores and increased turnover of norepinephrine is clearly associated with excessive sympathetic stimulation but in animal models and humans with heart failure secondary to mechanical overload there is evidence for depression of neuronal uptake. Because norepinephrine is both released and taken up by sympathetic fibers it is impossible to assess norepinephrine kinetics in an intact heart without separating these two functions. A technique for doing so has recently been developed in normal dogs and we therefore acquired similar data in humans with heart failure secondary to chronic pressure and volume overload. The technique involves the combination of transient norepinephrine tracer coronary sinus outflow in relation to intravascular and interstitial references after simultaneous injection into the left coronary artery and the measurement of endogenous norepinephrine concentrations in artery and coronary sinus. We found a marked reduction in cardiac norepinephrine release and uptake in a group of patients with clinical left ventricular failure secondary to mechanical overload, relative to a group of patients with no failure. Norepinephrine balance and overflow across the heart were not significantly different. We conclude that there is hypofunction of the cardiac sympathetic nerves in heart failure secondary to mechanical overload and that traditional methods are inadequate in assessing cardiac norepinephrine kinetics when there are simultaneous changes in neuronal uptake and release.