Indicator Fractionation Technique Research Articles

Quantitative Estimation of Tissue Blood Flow Rate.

The rate of blood flow through a tissue (F) is a critical parameter for assessing the functional efficiency of a blood vessel network following angiogenesis. This chapter aims to provide the principles behind the estimation of F, how F relates to other commonly used measures of tissue perfusion, and a practical approach for estimating F in laboratory animals, using small readily diffusible and metabolically inert radio-tracers. The methods described require relatively nonspecialized equipment. However, the analytical descriptions apply equally to complementary techniques involving more sophisticated noninvasive imaging.Two techniques are described for the quantitative estimation of F based on measuring the rate of tissue uptake following intravenous administration of radioactive iodo-antipyrine (or other suitable tracer). The Tissue Equilibration Technique is the classical approach and the Indicator Fractionation Technique, which is simpler to perform, is a practical alternative in many cases. The experimental procedures and analytical methods for both techniques are given, as well as guidelines for choosing the most appropriate method.

Iodoamphetamine (IMP) uptake in the brain is increased after experimental cerebral venous hypertension in the rat.

Cerebral venous hypertension (VH) is a pathological condition associated with arteriovenous malformation, and is an important determinant factor for clinical symptom and outcome. Using a rat whole brain VH model created by a carotid-jugular fistula and contralateral jugular vein ligation we measured superior sagittal sinus pressure (SSSP) and cerebral blood flow (CBF) by the 14C-iodoamphetamine (IMP) indicator fractionation technique under isoflurane anesthesia with controlled ventilation. SSSP was significantly elevated in the fistula group (15 +/- 1 mmHg) (mean +/- SEM) compared to the vessel occlusion (5 +/- 1) and sham operated groups (5 +/- 1) (P < 0.05). Mean absolute CBF in the fistula (103 +/- 3 ml/ min/100 g) and vessel occlusion (90 +/- 4) groups were elevated compared to the sham group (77 +/- 3) (P < 0.05). Physiological parameters (mean blood pressure, heart rate, and blood gas analysis) before measurement were not different. In this VH model we found increases in SSSP and IMP uptake in the brain. We assume the mechanism of IMP uptake increase in VH animal may be slow circulation with a possible change of first pass extraction of IMP.

Effects of intraventricular infusion of vascular endothelial growth factor on cerebral blood flow, edema, and infarct volume.

Therapeutic cerebral angiogenesis, utilizing angiogenic factors to enhance collateral vessel formation within the central nervous system, is a potential method for cerebral revascularization. A prior dose-response study determined that intracerebroventricular infusion of vascular endothelial growth factor (VEGF) increases vascular density with minimal associated brain edema at a concentration of 5 microg/ml. The purpose of this study was to assess effects of intracerebroventricular infusion of VEGF (5 microg/ml) on cerebral blood flow, infarct volume, and brain edema after ischemia. Recombinant human VEGF(165) was infused into the right lateral ventricle of rats with an osmotic minipump at a rate of 1 microl/hr for 7 days. Control animals received vehicle only. Ischemia was produced by transient (2 hours) middle cerebral artery occlusion (MCAO). After MCAO, cerebral blood flow was determined with the indicator fractionation technique: infarct volume was assessed with 2,3,5-triphenlytetrazolium chloride staining, and brain edema was determined by measuring brain water content. Cerebral blood flow was not significantly different in animals treated with VEGF compared to controls. There was a significant reduction in total infarct volume after temporary MCAO in VEGF-treated animals compared to controls (163+/-37 mm(3) vs. 309+/-54 mm(3), P<0.05). Brain water content after transient MCAO was also significantly reduced in VEGF-treated animals compared to controls (80.9+/-0.7% vs. 83.3+/-0.6%, P<0.05). Intracerebroventricular infusion of VEGF(165) (5 microg/ml) decreases infarct volume and brain edema after temporary MCAO without a significant increase in cerebral blood flow. These results indicate that VEGF may have a direct neuroprotective effect in cerebral ischemia.

β-Adrenergics enhance brain extraction of levodopa.

We sought to determine whether beta-adrenergic agonists enhance the brain extraction of L-dopa and L-leucine. Systemic administration of beta-adrenergic agonists increase brain concentrations of L-dopa and other large neutral amino acids (LNAA) in rats and monkeys and may improve symptoms and reduce daily L-dopa requirement in patients with Parkinson's disease. Cerebral blood flow (CBF) using [3H]nicotine and the extraction fraction of 14C-labeled L-dopa or L-leucine were measured simultaneously in various brain regions of conscious rats using the dual-isotope indicator fractionation technique after intraperitoneal administration of isoproterenol (a peripheral nonselective beta-adrenergic agonist), or clenbuterol (a beta2-adrenergic agonist that crosses the blood-brain barrier), or beta-adrenergic agonist preceded by nadolol (a peripheral nonselective beta-adrenergic antagonist), or saline vehicle. Both beta-adrenergic agonists increased regional brain extraction fraction of L-dopa and L-leucine tracers by 35-45%, without altering regional CBF. These changes were accompanied by about a 30% decrease in plasma branched chain LNAA concentrations. Nadolol blocked all these effects. beta-Adrenergic agonists increase the brain extraction of L-dopa and leucine, mainly by peripheral mechanisms that reduce the levels of other competing plasma LNAAs for transport. Thus, beta-adrenergic agonists might be useful in the treatment of patients with Parkinson's disease by enhancing delivery of L-dopa to the brain.

Mevastatin, an HMG-CoA reductase inhibitor, reduces stroke damage and upregulates endothelial nitric oxide synthase in mice.

The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) lower serum cholesterol and decrease the incidence of stroke and cardiovascular disease. There is growing evidence that statins exert some of their beneficial effects independent of cholesterol lowering. Indeed, we have previously demonstrated that chronic simvastatin administration upregulates endothelial nitric oxide synthase (eNOS), resulting in more functional protein, augmentation of cerebral blood flow, and neuroprotection in a murine model of cerebral ischemia. In this report we examined whether another member of the statin family shared these effects and whether eNOS upregulation is sustained with longer treatment. Mevastatin (2 mg/kg or 20 mg/kg per day) was administered to 18- to 22-g male mice for 7, 14, or 28 days before 2-hour middle cerebral artery occlusion with the use of the filament model (n=9 to 12). Neurological deficits and cerebral infarct volumes were assessed at 24 hours. Arterial blood pressure and gases, relative cerebral blood flow, and blood cholesterol levels were monitored in a subset of animals (n=5). Absolute cerebral blood flow was measured by the [(14)C]iodoamphetamine indicator fractionation technique (n=6). eNOS mRNA and protein levels were determined. Mevastatin increased levels of eNOS mRNA and protein, reduced infarct size, and improved neurological deficits in a dose- and time-dependent manner. Greatest protection was seen with 14- and 28-day high-dose treatment (26% and 37% infarct reduction, respectively). Cholesterol levels were reduced only after 28 days of treatment and did not correlate with infarct reduction. Baseline absolute cerebral blood flow was 30% higher after 14-day high-dose treatment. Chronic prophylactic treatment with mevastatin upregulated eNOS and augmented cerebral blood flow. These changes occurred in the absence of changes in serum cholesterol levels, were sustained for up to 1 month of treatment, and resulted in neuroprotection after middle cerebral artery occlusion.

Strain-related differences in susceptibility to transient forebrain ischemia in SV-129 and C57black/6 mice.

We explored susceptibility to injury after global ischemia in SV-129 and C57Black/6 mice, two commonly used-background strains in genetically engineered mice. Mice (n = 84) were subjected to 15, 30, or 75 minutes of bilateral common carotid artery (BCCA) occlusion followed by reperfusion for 72 hours. BCCA occlusion was performed under halothane or chloral hydrate anesthesia, in one experiment, mean arterial blood pressure and regional cerebral blood flow (laser Doppler flowmetry) were matched by controlled exsanguination. Baseline absolute blood flow measurements were obtained in both strains using a tracer, N-isopropyl-[methyl 1,3-14C]-p-iodoamphetamine, indicator fractionation technique (n = 5 per group). Vascular anatomy of the circle of Willis was visualized by intravascular perfusion of carbon black ink (n = 10 per group). Cerebrovascular reactivity was assessed by measuring the diameter of pial vessels (intravital microscopy) to acetylcholine (ACh) superfusion (0.1 to 10 mmol/L) in a closed cranial window preparation (n = 29). Resting blood flow values did not differ between groups in striatum, cerebellum, and brain-stem regions. SV-129 mice were less susceptible than C57Black/6 mice to ischemic injury (0.0 +/- 0.0 versus 1.3 +/- 0.3 damage in hippocampal CA1 region after 30 minutes of ischemia in SV-129 and C57Black/6, respectively; P < .01). Cellular damage (grade 1 to 3 injury) comparable to 30-minute BCCA occlusion was achieved only after 75 minutes of ischemia in SV-129 mice (1.1 +/- 0.3). Ischemic damage was also significantly less in SV-129 mice after blood pressure and flow were matched during ischemia in halothane-anesthetized SV-129 mice (0.5 +/- 0.3 versus 1.4 +/- 0.2, P < .05), or after chloral hydrate anesthesia (0.4 +/- 0.2 versus 1.5 +/- 0.4, P < .05). Hypoplastic posterior communicating arteries were found in all 10 C57Black/6 mice and may explain the greater susceptibility of these mice to injury after BCCA occlusion. More robust vasodilation to ACh in C57Black/6 mice could also indicate genetic differences in responses to vasoactive substances. C57Black/6 mice exhibit enhanced susceptibility to global cerebral ischemic injury, an incompletely formed circle of Willis, and augmented pial vessel dilation to ACh compared with SV-129 mice. Our findings suggest that strain differences may confound results when genetically engineered mice generated from more than a single background strain are used.

Effects of intra-ischemic blood pressure on outcome from 2-vessel occlusion forebrain ischemia in the rat

Halothane anesthetized Sprague-Dawley rats underwent 10 min of bilateral carotid artery occlusion with mean arterial pressure (MAP) held at 30, 50 or 60 mmHg. Sham rats did not undergo ischemia. A 7-day recovery interval was allowed. Intra-ischemic electroencephalographic (EEG) changes, behavioral function (Days 5–7), and histologic injury (Day 7) were evaluated. Under similar conditions, cerebral blood flow was determined after 10 min ischemia by the [ 3H]nicotine indicator fractionation technique. EEG isoelectricity was observed in 11 of 11.5 of 10, and 2 of 11 rats in the 30 mmHg, 50 mmHg, and 60 mmHg groups respectively. Neither passive avoidance cross-over latencies nor general motor scores were affected by intra-ischemic MAP and no differences from sham performance were observed. The per cent of CA1 neurons counted as dead (left and right hemispheres combined) was significantly affected by intra-ischemic MAP (72, 46 and 28% in the 30 mmHg, 50 mmHg, and 60 mmHg groups, respectively; P < 0.001. A greater than 50% CA1 neuronal mortality rate was present only in those rats exhibiting EEG isoelectricity. However, the number of rats demonstrating greater than a 25% interhemispheric difference in CA1 neuronal loss was greatest in the 50 mmHg group ( P < 0.02). Hippocampal blood flow decreased in association with severity of hypotension 8±1.35±8, and 48 + ml/100g/min (mean ∓ S.E.M. for 30, 50, and 60 mmHg, respectively; P <0.01. Again, however, the greatest variability in blood flow was observed at MAP = 50 mmHg. These results indicate that reduction of MAP to 30 mmHg during bilateral carotid artery occlusion will yield the most consistent hemodynamic and electrophysiologic events corresponding to reduced inter- and intra-animal variability in histologic outcome in this model of near-complete forebrain ischemia.

Importance of freezing time when iodoantipyrine is used for measurement of cerebral blood flow

The goal of the present study was to determine effects of delay of freezing of the brain on distribution of [14C]iodoantipyrine in the brain. Blood flow to parietal cerebral cortex (CBF) and choroid plexus was measured with the indicator fractionation technique and quantitative autoradiography. After injection of iodoantipyrine, each rat was decapitated, and the head was immersed in chlorodifluoromethane (-40 degrees C). The brain was removed from the skull after it was frozen. In some rats, heads were immersed immediately after decapitation, and CBF was markedly heterogeneous. In another group, heads were frozen 3 min after decapitation. In this case, CBF was uniform in its distribution. Average CBF was similar in groups with immediate and delayed freezing (90-104 ml.min-1 x 100 g-1). In contrast, delays in freezing decreased blood flow to choroid plexus from 551 +/- 115 to 261 +/- 48 ml.min-1 x 100 g-1. Our findings indicate that immediate freezing of the brain after decapitation is necessary to prevent diffusion of iodoantipyrine from regions of high blood flow to regions of lower blood flow and underestimations of blood flow in regions with high blood flow.

Cerebral Blood Flow with the Indicator Fractionation of [14C]Iodoantipyrine: Effect of Paco2 on Cerebral Venous Appearance Time

The indicator fractionation technique using a diffusible indicator as a tracer for the determination of CBF has been used for numerous investigations of the cerebral circulation and its pathophysiology. The diffusible tracer is "trapped" in the brain based on the proper delay between tracer injection and cessation of the cerebral circulation by decapitation before the appearance of the tracer in the cerebral venous circulation. If this delay is too long, the quantitative assumption of the indicator fractionation technique will not be met, and CBF values will be underestimated. In 13 Sprague-Dawley rats anesthetized with pentobarbital, the appearance of [14C]iodoantipyrine at the torcular was assessed as a function of PaCO2. An inverse linear relationship between PaCO2 (in millimeters of mercury) and cerebral venous appearance, Ta (in seconds), was established with the regression equation Ta = -0.0842.PaCO2 + 12.3 (R2 = 0.70, slope significantly different from zero, p less than 0.001). Ta varied between 5 and 12 s and PaCO2 varied between 84 and 18 mm Hg, respectively. Thus, in low-flow states, the decapitation time may be lengthened to 12 s, whereas in high-flow states, the time must be 5 s to eliminate the possibility of backflux of tracer out of the brain.

Chronic cerebral blood flow changes following experimental subarachnoid hemorrhage in rats.

Experimental subarachnoid hemorrhage was induced in 52 adult male Wistar rats by microsurgical transclival basilar artery puncture. Telencephalic blood flow measured in 24 rats with subarachnoid hemorrhage was compared with that in 23 sham-operated rats and 10 unoperated control rats using the [14C]butanol indicator fractionation technique. Telencephalic blood flow was significantly less in the rats with subarachnoid hemorrhage than in the sham-operated rats 3 (78.7 +/- 6.9 [n = 7] and 112.0 +/- 8.5 [n = 8] ml/100 g/min, respectively; p less than 0.01), 7 (74.9 +/- 5.1 [n = 9] and 112.6 +/- 4.6 [n = 8] ml/100 g/min, p less than 0.001), and 14 (81.9 +/- 6.0 [n = 8] and 104.1 +/- 5.4 [n = 7] ml/100 g/min, p less than 0.01) days after surgery. Telencephalic blood flow in unoperated controls (114.7 +/- 4.9 ml/100 g/min) did not differ significantly from sham-operated rats. Clinically, the 52 rats with subarachnoid hemorrhage were indistinguishable from 32 sham-operated rats. Postmortem examinations in 10 rats used in a preliminary investigation demonstrated significant blood clot in the basal cisterns 2 hours after basilar artery puncture. Intracranial pressure was slightly elevated (2.3 mm Hg over baseline) 30 minutes after the hemorrhage (n = 7), but when measured 3 (n = 3) or 7 (n = 3) days after surgery it had returned to baseline. Histologic examination of the brains from 10 rats subjected to subarachnoid hemorrhage 7 (n = 5) or 14 (n = 5) days before sacrifice revealed no evidence of cerebral ischemia or vasculopathic changes in the cerebral arteries.(ABSTRACT TRUNCATED AT 250 WORDS)

Subarachnoid hemorrhage fails to produce vasculopathy or chronic blood flow changes in rats.

Cerebral blood flow was measured by a [14C]butanol indicator fractionation technique in rats subjected to subarachnoid hemorrhage, in control rats, and in rats given injections of buffered saline into the subarachnoid space (sham hemorrhage). Cerebral blood flow was significantly decreased in both the subarachnoid hemorrhage and sham hemorrhage rats 3 hours after injection. However, blood flow returned to control levels by 24 hours, and measurement for 14 days after subarachnoid hemorrhage failed to show any delayed decrease in cerebral blood flow. Electron microscopic studies of basilar arteries from rats subjected to subarachnoid hemorrhage 72 hours before killing failed to show any of the morphologic changes that have been associated with vasospasm in humans or in higher animal models. Our studies indicate that the rat model of subarachnoid hemorrhage has limited applicability to the study of subarachnoid hemorrhage following ruptured cerebral aneurysms in humans. However, although rats are not a perfect model of this clinical condition, some pathophysiologic changes similar to those observed in human subarachnoid hemorrhage have been demonstrated in this model and deserve further investigation.

Efficacy and mechanism of action of a calcium channel blocker after global cerebral ischemia in rats.

Dihydropyridine calcium channel blockers such as nicardipine are under evaluation for treating acute cerebral ischemia because they may increase cerebral blood flow by causing vasodilation and because they may be cytoprotective in part by limiting production of arachidonic acid metabolites. We demonstrated in a previous study that nicardipine improves postischemic neuronal function, as measured by somatosensory evoked potentials, without reducing the extent of light-microscopic CA-1 hippocampal histologic damage. To characterize further the effect of nicardipine on global ischemic injury, we administered the drug beginning 24 hours before 30 minutes of four-vessel ischemia in Wistar rats. We then measured hippocampal ATP, phosphocreatine, and glucose contents immediately and 2 hours after ischemia, and measured learning ability (working and reference errors) on an eight-arm radial maze beginning 30 days after ischemia. To gain insight into the possible mechanism of action, we measured production of arachidonic acid metabolites (eicosanoids: TXB2 and 6-keto-PGF1 alpha) and hemispheric and hippocampal cerebral blood flow by the [14C]butanol indicator fractionation technique immediately and 2 hours after ischemia. Nicardipine was associated with fewer working errors (p less than 0.02) but no difference in reference errors. The drug had no effect on energy metabolites, cerebral blood flow, or eicosanoids immediately after ischemia, but ATP, phosphocreatine, and cerebral blood flow all returned to normal levels significantly more rapidly during reperfusion in treated rats. Nicardipine improves behavioral, electrophysiologic, and mitochondrial function after ischemia without preventing cellular damage and improves postischemic reperfusion. The drug's positive effect appears to occur during reperfusion.

Blood flow in an experimental rat brain tumor by tissue equilibration and indicator fractionation.

The tissue equilibration technique (Kety) was compared with the indicator fractionation technique for the measurement of blood flow to normal brain and an experimental brain tumor in the rat. The tumor was a cloned astrocytic glioma implanted in the cerebral hemisphere of F-344 rats. I-125 Iodoantipyrine, using a rising infusion for one minute, was used for the tissue equilibration technique. C-14 butanol, injected as a bolus 8 seconds before sacrifice, was used for the indicator fractionation technique. Samples were assayed using liquid scintillation counting and the iodoantipyrine results were regressed against the butanol results. For normal tissue R = 0.832, SEE = 0.115 ml/g/min, and Slope = 0.626. For tumor R = 0.796, SEE = 0.070 ml/g/min, and Slope = 0.441. The iodoantipyrine tissue/blood partition coefficient for normal hemisphere (gray and white matter) was 0.861 +/-0.037 (SD) and for tumor was 0.876 +/-0.042. The indicator fractionation technique with C-14 butanol underestimated blood flow in a consistent manner, probably because of incomplete extraction, early washout of activity from tissue and from evaporation of butanol during processing. Our experiments revealed no differences between tumor and normal brain tissue that might invalidate the comparison of iodoantipyrine blood flow results in brain tumors and surrounding normal brain.

1441 PERINATAL REGIONAL CEREBRAL BLOOD FLOW (rCBF): EFFECT OF THEOPHYLLINE

Theophylline (T) is known to produce cerebral vasoconstriction in both animals and humans. It is unclear whether these effects vary regionally, especially in the developing brain. Using the tracer iodo-[C] antipyrine and the indicator-fractionation technique described by Cavazutti and Duffy, we studied rCBF in awake newborn dogs (3 to 9 days old), treated with either saline (n=4) or T (n=3).Catheters were placed in the right common carotid artery and the right external jugular vein while under halothane anesthesia. Following a 4 hour recovery period, either T, 45 mg/kg, or saline was administered by slow IV infusion. After an additional 1 hour equilibration period, the rCBF measurement was performed. pCO2, measured just prior to rCBF determination, was 36 ± 3 mm Hg (mean ± S.D.) in the saline group and 26 ± 6 mm Hg in the T group. T levels in treated animals equaled 38 ± 4 μg/ml.RCBF in the saline group ranged from 6 ml/100 gm/min in sub-cortical white matter to 75 ml/100g/min in the oculomotor n. nucl. T treated dogs, when compared to saline treated dogs, had lower values in all 28 areas of the brain analyzed. Thalamic and midbrain nuclei showed the greatest reductions (35-48%). At the other extreme, medullary and cerebellar structures showed little reduction (7-16%) with T. Though exceptions were noted, the degree of T associated reduction in rCBF generally paralleled reported newborn dog CO2 - CBF sensitivity.

An evaluation of errors in the determination of blood flow by the indicator fractionation and tissue equilibration (Kety) methods.

In this report, the effects of various errors and plasma time courses of indicator concentration on the accurate determination of cerebral blood flow (F) are theoretically analyzed for the tissue equilibration and the indicator fractionation techniques. For the indicator fractionation technique, the impact of sample timing and tissue assaying errors and of indicator backflux were examined; for the tissue equilibration method, errors in the value of the partition coefficient (lambda), sample timing, and tissue assaying were considered. The recommended ways to decrease the effects of errors in the indicator fractionation technique are to administer the indicator by an intravenous bolus and to sample the tissue about 10 s thereafter. Possible errors in the assessment of F by the tissue equilibration technique are diminished by using an indicator infusion schedule which yields a continuous rise in arterial concentration and by selecting a 30-s experiment duration. Surprisingly, the impact of sample timing errors is greater on the determination of F with the tissue equilibration method than with the indicator fractionation technique. For the chosen plasma time courses, there is always a backflux error in an indicator fractionation estimation of F, and this error increases as the flow rate increases. Thus, provided the sample timing and tissue assay errors are small and the value of lambda is known, the tissue equilibration method is the more accurate of the two. If lambda is unknown, then the indicator fractionation technique should be used. In many cases, the indicator fractionation method will provide as accurate an estimate of F as will the tissue equilibration method.

Regional cerebral blood flow and glucose metabolism following transient forebrain ischemia.

Progressive brain damage after transient cerebral ischemia may be related to changes in postischemic cerebral blood flow and metabolism. Regional cerebral blood flow (rCBF) and cerebral glucose utilization (rCGU) were measured in adult rats prior to, during (only rCBF), and serially after transient forebrain ischemia. Animals were subjected to 30 minutes of forebrain ischemia by occluding both common carotid arteries 24 hours after cauterizing the vertebral arteries. Regional CBF was measured by the indicator-fractionation technique using 4-iodo-[14C]-antipyrine. Regional CGU was measured by the 2-[14C]deoxyglucose method. The results were correlated with the distribution and progression of ischemic neuronal damage in animals subjected to an identical ischemic insult. Cerebral blood flow to forebrain after 30 minutes of moderate to severe ischemia (less than 10% control CBF) was characterized by 5 to 15 minutes of hyperemia; rCBF then fell below normal and remained low for as long as 24 hours. Post-ischemic glucose utilization in the forebrain, except in the hippocampus, was depressed below control values at 1 hour and either remained low (neocortex, striatum) or gradually rose to normal (white matter) by 48 hours. In the hippocampus, glucose utilization equaled the control value at 1 hour and fell below control between 24 and 48 hours. The appearance of moderate to severe morphological damage in striatum and hippocampus coincided with a late rise of rCBF above normal and with a fall of rCGU; the late depression of rCGU was usually preceded by a period during which metabolism was increased relative to adjacent tissue. Further refinement of these studies may help identify salvageable brain after ischemia and define ways to manipulate CBF and metabolism in the treatment of stroke.

Regulation of local cerebral blood flow in normal and hypoxic newborn dogs.

Local cerebral blood flow (LCBF) was measured autoradiographically in newborn puppies by an indicator fractionation technique using 4-iodo-[14C]antipyrine as the diffusible indicator. Measurements were obtained in unanesthetized, normotensive animals, and the sensitivity of blood flow to hypercapnia and acute hypoxia was determined in 32 brain structures. LCBF in normal and hypoxic puppies was correlated with local cerebral glucose utilization (LCGU) obtained under the same experimental conditions (Duffy et al, 1982). In normocapnic (PaCO2 33 mm Hg) control animals, highest rates of blood flow were found in gray matter nuclei of the brainstem, in the medulla oblongata, and in the posterolateral nucleus of the thalamus (50 to 77 ml/100 gm/min); far lower flows were recorded among white matter structures (5 to 11 ml/100 gm/min). The vasodilatory response to both hypercapnia and hypoxia was greatest among brainstem gray matter structures, intermediate among cortical and diencephalic gray matter structures, and least in white matter. When LCBF was plotted as a function of LCGU for control animals, a positive linear correlation was obtained for all structures (p less than 0.001), implying that in newborns, as in adults, cerebral blood flow and metabolism are physiologically coupled. In hypoxic puppies, no consistent relationship between LCGU and LCBF could be demonstrated; however, there was suggestion that the two measurements correlated inversely, presumably reflecting enhanced anaerobic glycolysis in structures (e.g., hemispheric white matter) that were not adequately protected by compensatory hyperemia. White matter damage, a frequent complication of perinatal hypoxia-asphyxia, may be a consequence in part of the limited capacity of white matter to vasodilate in response to te chemical "signals" of hypercapnia and lactic acidosis.

Measurement of 42K concentration in the mucosal portion of the intestine without tissue samples: an approach to the measurement of mucosal blood flow in the dog.

1. The concentration of 42K within the jejunal mucosa was measured by intraluminal miniatrue Geiger-Müller tubes shieleded with platinum foil. 2. Isotope concentration measured by tissue assay of jejunal mucosa gave similar results to that obtained by the Geiger-Müller tubes. 3. Isotope measurement by shielded Geiger-Müller tubes obviates the need for tissue samples and permits repeat measurement in the same animal. This should find physiological application in measuring the mucosal portion of total intestinal blood flow by the indicator fractionation technique.

The distribution of blood flow along the small intestine of the dog.

The presence of a blood-flow gradient along the small intestine is disputed. Geber (1), using electromagnetic flowmeters in dogs, and Steiner and Mueller (2) using the indicator fractionation technique in rats demonstrated that blood flow per unit weight was highest in the duodenum and then decreased progressively along the jejunum, ileum, and colon. On the other hand, Grim and Lindseth (3) using direct collection of venous outflow isolated loops of intestine in the dog, and Delaney and Custer (4) using the indicator fractionation technique in dogs did not observe a blood flow gradient. The negative view has also been supported by a more recent study of Goodhead (5) using the indicator fractionation technique in the dog. This paper presents evidence that a blood flow gradient does exist in the small intestine of the dog when it is studied under basal conditions of anesthesia. Materials and methods. Eighteen mongrel dogs weighting 15-30 kg were given water but no food for 24 hr in the preparation period. General anesthesia was then induced and maintained with intravenous pentobarbitone sodium (30 mg/kg). Blood pressure and heart rate were monitored from a femoral artery catheter and body temperature was kept at 37.5°. Care was taken to perform all blood flow measurements when these functions were stable. Intestinal blood flow was measured by the indicator-fractionation technique with 42K (15 μCi/Kg) as the indicator. The 42K was injected into the inferior vena cava and tissue 42K content 100 sec after the injection was used for all blood flow calculations (5-8). The cardiac output was measured by the conventional dye dilution technique (9) immediately prior to the injection of the 42K. Cardiac arrest was produced by a rapid injection of saturated magnesium sulphate into the inferior vena cava and this enabled removal of 3-5 cm sleeves of intestine for 42K assay from the third part of the duodenum, the jejunum (10 cm distal to the ligament of Trietz), and the distal ileum (30 cm proximal to the ileo-caecal valve).

Effect of secretogogues on mucosal blood flow in the antrum and corpus of the stomach.

Gastric mucosal blood flow in the rat was determined by the indicator fractionation technique of Sapirstein using Rb86Cl. Antral and corpus flow rates were determined in response to increasing doses of pentagastrin and histamine and the effect of vagotomy was determined. With progressive stimulation the antral:corpus flow ratio increased with both stimulants; however, vagotomy and noradrenaline reduced blood flow in both areas of mucosa. It is concluded that the antrum, as well as the corpus, actively responds to secretory stimulants by an increase in mucosal blood flow and support is provided for the theory that the antrum plays an active role in the control of parietal cell blood flow and secretion.