Measurement of carbamazepine and carbamazepine epoxide in the human brain using in vivo microdialysis

We report the first study of carbamazepine and carbamazepine-10,11-epoxide concentrations determined by using intracerebral microdialysis in three patients undergoing depth electrode studies for the evaluation of medically intractable epilepsy. Very small microdialysis catheters, affixed to and inserted with the depth electrodes, sampled drug concentration in the extracellular environment. We perfused artificial extracellular fluid continuously, and varied the perfusion rate to permit estimation of the absolute drug concentration in the extracellular space. Serum samples were obtained simultaneously. The relation between dialysate and extracellular concentration (recovery fraction) depended, in vivo but not in vitro, on the relative lipophilicity of the compounds, suggesting that diffusion of the drug within the brain is a major determinant of microdialysate drug concentration. When this is taken into account, the steady-state extracellular concentrations of these compounds closely mirror their unbound serum concentrations.     

Dose-dependent effects of acute in vivo ethanol exposure on extracellular glutamate concentration in the cerebral cortex of the near-term fetal sheep

The cerebral cortex is a target site of ethanol teratogenesis. L-Glutamate is a major excitatory neurotransmitter that plays an important neurotrophic role in brain development. It has been proposed that optimal function of the glutamate neuronal system is required for normal brain development; overactivation could lead to excitotoxic-induced neuronal injury, whereas underactivation could delay/restrict brain development. The objective of this study was to test the hypothesis that acute in vivo ethanol exposure alters basal glutamate release in the fetal cerebral cortex. The experimental approach involved measuring fetal cortical extracellular glutamate concentration using the technique of in vivo microdialysis. Near-term fetal sheep were chronically instrumented with a microdialysis probe placed in the parasagittal cortex. At 124 +/- 3 days of gestation, the effects of maternal intravenous infusion of 2 g or 4 g ethanol/kg maternal body weight or an equivalent volume of saline, given as four equally divided doses over 5 hr, on fetal cerebral cortical extracellular glutamate concentration were determined. None of the three treatment regimens produced fetal or maternal demise during the time course of the study. There was an ethanol dose-dependent increase, p = 0.005, in extracellular glutamate concentration in the fetal cerebral cortex. This increase was paroxysmal in nature and was not directly related to the fetal blood ethanol concentration. In view of the proposed role for glutamate in neuronal development, this apparent ethanol-induced increase in glutamate release may be important in the pathogenesis of ethanol teratogenesis involving the cerebral cortex.     

In situ assessment of the role of the beta 1-, beta 2- and beta 3-adrenoceptors in the control of lipolysis and nutritive blood flow in human subcutaneous adipose tissue

1. The involvement of beta 1-, beta 2- and beta 3-adrenoceptors in the control of lipolysis and nutritive blood flow was investigated in abdominal subcutaneous adipose tissue of healthy young adults by use of an in situ microdialysis technique. 2. Dialysis probes were infused either with isoprenaline (non-selective beta-adrenoceptor agonist), CGP 12,177 (selective beta 3-adrenoceptor agonist having beta 1-/beta 2-antagonist properties), dobutamine (selective beta 1-adrenoceptor agonist) or terbutaline (selective beta 2-adrenoceptor agonist). The recovery of each probe used for perfusion was calculated by an in vivo calibration method. The local blood flow was estimated through the measurement of the escape of ethanol infused simultaneously with the drugs included in the probe. 3. Isoprenaline infusion at 0.01 microM had a weak effect while higher concentrations of isoprenaline (0.1 and 1 microM) caused a rapid, sustained and concentration-dependent increase of glycerol outflow; the maximum increase was 306 +/- 34% with 1 microM. Isoprenaline also increased the nutritive blood flow in adipose tissue; a significant effect appeared at 0.1 microM isoprenaline and was greater at 1 microM. 4. CGP 12,177 (10 and 100 microM) increased the glycerol concentration in the dialysate (128 +/- 8 and 149 +/- 12%, respectively) and nutritive blood flow. Terbutaline and dobutamine (100 microM) both provoked rapid and similar increases in glycerol outflow (252 +/- 18 and 249 +/- 18%, respectively). Both, terbutaline and dobutamine increased nutritive blood flow. 5. It is concluded that beta 1- and beta 2-adrenoceptor subtypes are both mainly involved in the mobilization of lipids and in the control of nutritive blood flow. beta 3-Adrenoceptors play a weaker role in the control of lipolysis and nutritive blood flow in human subcutaneous abdominal adipose tissue.     

Analysis of ceftriaxone and ceftazidime distribution in cerebrospinal fluid of and cerebral extracellular space in awake rats by in vivo microdialysis

In vivo microdialysis was used to estimate the extracellular concentrations of ceftazidime and ceftriaxone, two expanded-spectrum cephalosporins commonly used in the treatment of bacterial meningitis, in two brain regions (the right corpus striatum and the left lateral ventricle_ of awake, freely moving rats. Antibiotics were administered by constant intravenous infusion at 18 mg/h until steady-state levels were reached. Ceftriaxone levels measured at the steady state in the extracellular space of the corpus striatum (0.80 +/- 0.17 micrograms/ml) were statistically equivalent to those obtained in the cerebrospinal fluid of the lateral ventricle (0.71 +/- 0.15 micrograms/ml). The ratios of these levels in the brain to the steady-state levels in plasma were 0.5 +/- 0.1% for both regions. The postinfusion concentrations of ceftriaxone in the brain declined monoexponentially, with an elimination half-life similar to that obtained in plasma. However, the mean antibiotic concentration of ceftazidime in the striatum (2.2 +/- 0.4 micrograms/ml) was lower (P < 0.001) than that in the lateral ventricle (3.8 +/- 0.5% and 4.0 +/- 1.8%, respectively) were higher than those obtained with ceftriaxone. Moreover, the half-life of ceftazidime elimination from plasma was lower than that obtained in the two brain regions. It was concluded that the in vivo microdialysis technique yields useful data on antibiotic distribution in the extracellular space of the brain, that the distribution may not be homogeneous, and that the decay of postinfusion concentrations in the brain may be different from the decay of postinfusion concentrations in plasma.     

Rifampin concentrations in various compartments of the human brain: a novel method for determining drug levels in the cerebral extracellular space

Antimicrobial therapy for brain infections is notoriously difficult because of the limited extent of knowledge about drug penetration into the brain. Therefore, we determined the penetration of rifampin into various compartments of the human brain, including the cerebral extracellular space (CES). Patients undergoing craniotomy for resection of primary brain tumors were given a standard dose of 600 mg of rifampin intravenously before the operation. A microdialysis probe (10 by 0.5 mm) was inserted into the cortex distantly from the resection and was perfused with two different rifampin solutions. Rifampin concentrations in the CES were calculated by the no-net-flux method. Intraoperatively, samples were taken from brain tumor tissue, perifocal tissue, and normal brain tissue in the case of pole resections. Rifampin concentrations in the various samples were determined by using a bioassay with Sarcinea lutea. In the various compartments, rifampin concentrations were highest within tumors (1.37 +/- 1.34 microg/ml; n = 8), followed by the perifocal region (0.62 +/- 0.67 microg/ml; n = 8), the CES (0.32 +/- 0.11 microg/ml; n = 6), and normal brain tissue (0.29 +/- 0.15 microg/ml; n = 7). Rifampin concentrations in brain tumors do not adequately reflect concentrations in normal brain tissue or in the CES. Rifampin concentrations in the CES, as determined by microdialysis, are the most reproducible, and the least scattered, of the values for all compartments evaluated. Rifampin concentrations in all compartments exceed the MIC for staphylococci and streptococci. However, CES concentrations may be below the MICs for some mycobacterial strains.     

Microdialysis probe for transcutaneous monitoring of ethanol and glucose in humans

A heated (42 degrees C) microdialysis probe and its application for continuous transcutaneous sampling of ethanol and glucose through cellophane-stripped forearm skin are described. Ethanol and glucose concentration in the dialysate were measured on-line with continuous-flow analysis and compared with blood values in human volunteers after ethanol consumption (n = 4) and oral glucose testing (n = 5), respectively. For ethanol and glucose, the dialysate and blood concentrations were linearly related in each subject (r > or = 0.91, P < 0.005), although the dialysate-to-blood ratio varied among subjects. The recovery in vivo was 22.4 +/- 22.7 and 4.7 +/- 2.3% (SD) of the recovery in vitro for ethanol and glucose, respectively. The dialysate glucose concentration was independent of blood flow. When the probe temperature was increased from 32 to 42 degrees C, the dialysate-to-blood glucose ratio increased, with 2.4 +/- 1.4%/degrees C (SD) in fasting subjects (n = 4), which was similar to an increase of 2.1 +/- 0.045%/degree C in dialysate-to-medium ratio in vitro. The present approach for transcutaneous sampling may possibly be used for other substances of low molecular weight as well.     

Increase in extracellular glutamate caused by reduced cerebral perfusion pressure and seizures after human traumatic brain injury: a microdialysis study

OBJECT: To determine the extent and duration of change in extracellular glutamate levels after human traumatic brain injury (TBI), 17 severely brain injured adults underwent implantation of a cerebral microdialysis probe and systematic sampling was conducted for 1 to 9 days postinjury. METHODS: A total of 772 hourly microdialysis samples were obtained in 17 patients (median Glasgow Coma Scale score 5+/-2.5, mean age 39.4+/-20.4 years). The mean (+/-standard deviation) glutamate levels in the dialysate were evaluated for 9 days, during which the mean peak concentration reached 25.4+/-13.7 microM on postinjury Day 3. In each patient transient elevations in glutamate were seen each day. However, these elevations were most commonly seen on Day 3. In all patients there was a mean of 4.5+/-2.5 transient elevations in glutamate lasting a mean duration of 4.4+/-4.9 hours. These increases were seen in conjunction with seizure activity. However, in many seizure-free patients the increase in extracellular glutamate occurred when cerebral perfusion pressure was less than 70 mm Hg (p < 0.001). Given the potential injury-induced uncoupling of cerebral blood flow and metabolism after TBI, these increases in extracellular glutamate may reflect a degree of enhanced cellular crisis, which in severe head injury in humans appears to last up to 9 days. CONCLUSIONS: Extracellular neurochemical measurements of excitatory amino acids may provide a marker for secondary insults that can compound human TBI.     

Gene therapeutic strategies for neuroprotection: implications for Parkinson''s disease

The model used for calculating perfusion by MRI techniques that use endogenous water as a tracer assumes that arterial water is a freely diffusible tracer. Evidence shows that this assumption is not valid in the brain at high blood flow rates, at which movement of water into and out of the microvasculature becomes limited by diffusion across the blood-brain barrier. In this work, the arterial spin-labeling technique is used to show that fraction of arterial water that is dependent on blood flow rate remains in the vasculature and does not exchange with brain tissue water. By using perfusion measurements without and with magnetization transfer (MT) effects, one can distinguish arterial label that exchanges into tissue because blood has much smaller MT than brain tissue. Using this technique, the extraction fraction for water is measured in the rat brain at various cerebral blood flow rates. At high flow rates (approximately 5 ml/g/min), the extraction fraction for water is found to be about 45% in rat brain. Disruption of the blood-brain barrier with D-mannitol caused an increase in the extraction fraction for water. It was possible to form an image related to the extraction fraction for water. The ability to estimate the amount of vascular water exchanging with tissue water by MRI may represent a noninvasive approach to detect the integrity of the blood-brain barrier.     

Increased resting and exercise-induced oxidative stress in young IDDM men

The effects of local application of the 5-HT3 receptor agonist, 1-(m-chlorophenyl)-biguanide (CPBG), and i.p. administration of ethanol on the extracellular levels of dopamine (DA) in the ventral tegmental area (VTA) were studied using in vivo microdialysis. Adult female Wistar rats were implanted with microdialysis probes in the VTA at least 24 h before each experiment. Stable extracellular levels of DA (101 +/- 9 fmol/20 min) were established before initiating the experiments. Application of 10-250 microM CPBG through the microdialysis probe dose-dependently enhanced the extracellular concentrations of DA but did not alter the levels of either 3,4-dihydroxyphenylacetic acid or homovanillic acid in the dialysate. The effects of CPBG were reversible and dependent upon Ca2+. Co-perfusion with the 5-HT3 receptor antagonist, 3-tropanyl-indole-3-carboxylate (ICS 205-930), inhibited the effects of CPBG on enhancing extracellular DA levels. The i.p. administration of 2 g/kg ethanol significantly (p < 0.005) enhanced the levels of DA to 150% of baseline values; this ethanol-induced increase was prevented by local perfusion with 100 microM ICS 205-930. These results suggest that 5-HT3 receptors in the VTA are involved in regulating the somatodendritic release of DA and in mediating the stimulatory effects of ethanol on this neuronal system.     

Quantitative microdialysis of neuropeptide Y

The feasibility of using the difference method of quantitative microdialysis to measure neuropeptide Y (NPY) was evaluated in vitro and in vivo. The accuracy of this method was tested in vitro under steady-state conditions for 3 test solutions containing known concentrations of NPY. The estimated concentrations of NPY were 1.2 +/- 0.6, 3.7 +/- 0.9, and 15.1 +/- 0.7 pg/microliter (mean +/- SEM) in agreement with the actual concentrations of NPY in the test solutions which were 1.1 +/- 0.8, 4.6 +/- 0.6, and 14.6 +/- 0.5 pg/microliter (mean +/- SEM of solution samples), respectively. The responsiveness of the estimated NPYext measure to changes in the external concentration of NPY was also evaluated in vitro. An accurate estimate of NPYext was obtained within the first sampling period (within 15 min) after a 2-3-fold increase in the test solution concentration of NPY and within 2-3 sampling periods (15-45 min) in response to a 2-3-fold decrease in the test solution concentration of NPY. In vivo, the estimated basal concentration of NPY in dialysis samples from probes in the medial basal hypothalamus of anesthetized female rats (n = 4) was 4.0 +/- 1.6 pg/microliters and increased to 9.5 +/- 0.3 pg/microliter during K+ stimulation. Relative recovery was 22% in vivo under steady-state conditions and ranged from 14% to 30% during dynamic conditions. These results demonstrate that the difference method of quantitative microdialysis accurately estimates picomolar concentrations of NPY in vitro, and is sufficiently sensitive to detect basal and increasing concentrations of NPY in vivo.     

Role of phosphodiesterase III in the antilipolytic effect of insulin in vivo

The effect of three types of phosphodiesterase (PDE) inhibitors on in vivo antilipolysis was investigated in healthy subjects using a 2-h euglycemic, hyperinsulinemic (40 mU.m-2.min) clamp together with microdialysis of abdominal subcutaneous adipose tissue. During hyperinsulinemia (approximately 330 pmol/l), the circulating glycerol concentration was reduced to approximately 50% of the basal level of 53.2 +/- 3.6 mumol/l, indicating an antilipolytic effect. The decrease in adipose tissue dialysate glycerol, which mirrors the change in interstitial glycerol concentration, was about 40% during hyperinsulinemia when Ringer's solution alone was perfused. Local perfusion with a selective PDE IV inhibitor, rolipram (10(-4) mol/l), did not influence the insulin-induced decrease in dialysate glycerol (F = 0.8 vs. perfusion with Ringer's solution by two-factor analysis of variance [ANOVA]), although rolipram increased the dialysate glycerol level by 144 +/- 7% of the baseline value. However, local perfusion with a selective PDE III inhibitor, amrinone (10(-3) mol/l), or a nonselective PDE inhibitor, theophylline (10(-2) mol/l), abolished the ability of insulin to lower dialysate glycerol (F = 16.5, P < 0.01 and F = 8.5, P < 0.01, respectively, as compared with perfusion with Ringer's solution). The findings could not be explained by changes in the local blood flow (as measured by a microdialysis--ethanol escape technique), which was not affected by hyperinsulinemia in the presence or the absence of PDE inhibitors in the dialysis solvent. We conclude that PDEs play an important role in mediating the antilipolytic effect of insulin in vivo and that PDE III is the dominant isoenzyme modulating this effect.     

Adenosine: a mediator of the sleep-inducing effects of prolonged wakefulness

Both subjective and electroencephalographic arousal diminish as a function of the duration of prior wakefulness. Data reported here suggest that the major criteria for a neural sleep factor mediating the somnogenic effects of prolonged wakefulness are satisfied by adenosine, a neuromodulator whose extracellular concentration increases with brain metabolism and which, in vitro, inhibits basal forebrain cholinergic neurons. In vivo microdialysis measurements in freely behaving cats showed that adenosine extracellular concentrations in the basal forebrain cholinergic region increased during spontaneous wakefulness as contrasted with slow wave sleep; exhibited progressive increases during sustained, prolonged wakefulness; and declined slowly during recovery sleep. Furthermore, the sleep-wakefulness profile occurring after prolonged wakefulness was mimicked by increased extracellular adenosine induced by microdialysis perfusion of an adenosine transport inhibitor in the cholinergic basal forebrain but not by perfusion in a control noncholinergic region.     

Determination of optimum retrograde cerebral perfusion conditions

Retrograde cerebral perfusion through a superior vena caval cannula is a new technique used to protect the brain during operations on the aortic arch. We measured cerebral tissue blood flow, oxygen consumption, and cerebrospinal fluid pressure under various perfusion conditions in hypothermic (20 degrees C) mongrel dogs (n = 18, 12.8 +/- 0.6 kg) to determine the optimum conditions for retrograde cerebral perfusion. Retrograde cerebral perfusion was performed by infusion via the superior vena caval cannula and drainage via the ascending aortic cannula while the inferior vena cava and azygos vein were clamped. Retrograde cerebral perfusion was performed as the external jugular venous pressure was changed from 15 to 35 mm Hg in increments of 5 mm Hg. Cerebral tissue blood flow was measured by the hydrogen clearance method. Hypothermic retrograde cerebral perfusion with an external jugular venous pressure of 25 mm Hg provided about half the cerebral tissue blood flow of hypothermic (20 degrees C) cardiopulmonary bypass with a flow rate of 1000 ml/min (13.7 +/- 7.9 versus 32.7 +/- 8.5 ml/min per 100 gm). It decreased significantly as the external jugular venous pressure was decreased from 25 to 15 mm Hg but did not increase significantly as the external jugular venous pressure was increased from 25 to 35 mm Hg. Whole-body oxygen consumption during hypothermic retrograde cerebral perfusion with an external jugular venous pressure of 25 mm Hg was one quarter of that during hypothermic cardiopulmonary bypass (3.4 +/- 0.7 versus 12.7 +/- 5.6 ml/min) and varied in proportion to external jugular venous pressure. The cerebrospinal fluid pressure was a little lower than the external jugular venous pressure (19.2 +/- 4.5 mm Hg versus 24.8 +/- 2.4 mm Hg) but also varied with the external jugular venous pressure. The cerebrospinal fluid pressure remained lower than 25 mm Hg so long as the external jugular venous pressure remained lower than 25 mm Hg. High external jugular venous pressure was associated with high intracranial pressure, which restricts cerebral tissue blood flow and may cause brain edema. We believe that a venous pressure of 25 mm Hg is the optimum condition for retrograde cerebral perfusion.     

The ex vivo human placental transfer of the anti-HIV nucleoside inhibitor abacavir and the protease inhibitor amprenavir

OBJECTIVE: The transfer of abacavir, a new nucleoside inhibitor, and amprenavir, a new protease inhibitor, used for the treatment of human immunodeficiency virus, has been studied in the ex vivo human placental model. METHODS: The ex vivo human placental model used C14 antipyrine to determine the transport fraction and clearance index of these compounds at both the peak and trough serum concentrations. The clearance index accumulation and tissue concentrations were determined for each drug by high pressure liquid chromatography. RESULTS: The clearance index of abacavir was 0.47 +/- 0.19 and 0.50 +/- 0.07 at peak and trough concentrations, respectively. The clearance index of amprenavir was 0.38 +/- 0.09 and 0.14 +/- 0.08 at peak and trough concentrations, respectively. There was no unusual accumulation of either drug in the media or tissue when the perfusion system was closed. CONCLUSION: Abacavir is the first nucleoside compound studied in the perfusion system with a high clearance index. The transfer of the protease inhibitor amprenavir had a clearance index 2.75 times greater than the clearance index of ritonavir at peak concentration determined in a previous study. At trough concentration the clearance index was much less than at the peak concentration. A similar result was found with ritonavir.     

Construction and calibration of a low-cost bandage pressure monitor

Pregnant sheep with a microdialysis probe implanted in the fetal cerebral cortex were used to determine if nitrate and nitrite anions (nitrate/nitrite) could be quantitated in the microdialysate as an indirect index of in vivo nitric oxide formation. Pregnant ewes (term, about 147 days) were surgically instrumented at gestational day (GD) 90 (n = 3; preterm) and GD 121 (n = 3; nearterm). Three days later, following an overnight probe equilibration period, five dialysate samples were collected continuously on ice at 1-h intervals (infusion rate of 1 (microl/min). The nitrate/nitrite concentration was determined by reducing a 10-microl aliquot of each dialysate fraction with hot acidic vanadium followed by chemiluminescence quantitation of the nitric oxide product. The lower limit of quantitative sensitivity of the method is 25 picomoles. Nitrate/nitrite concentration was 16.6+/-7.3 microM for the preterm fetus and 19.7+/-1.9 microM for the nearterm fetus. The data demonstrate that nitrate/nitrite, as an index of in vivo nitric oxide formation, can be quantitated in microdialysate samples collected from the intact fetal sheep cerebral cortex.     

An immunologic approach to induction of epidermal growth factor deficiency: induction and characterization of autoantibodies to epidermal growth factor in rats

The role of glutamate as a possible mediator of neurodegeneration is well described, and the homeostasis of extracellular glutamate is considered of major importance when addressing the pathogenesis of excitatory neurodegeneration. Applying the 'indicator diffusion' method to the microdialysis technique, we present a method that is suitable for the in vivo investigation of the capacity of cellular uptake of glutamate. Using 14C-mannitol as reference, we measured the cellular extraction and the cell membrane permeability of the test substance 3H-D-aspartate in the corpus striatum of the rat brain. The cellular extraction fraction of 3H-D-aspartate was 0.29, and the cell membrane permeability 2.24 x 10(-4) cm/s. In the presence of the glutamate-uptake blocker DL-threo-beta-hydroxyaspartate (THA) the extraction of 3H-D-aspartate was completely abolished, indicating that extraction of 3H-D-aspartate was due to cellular uptake by glutamate transporters. The cell membrane permeability towards 3H-D-aspartate was reduced by approximately 98% due to THA, indicating that the cell membranes per se are highly resistant to diffusion of 3H-D-aspartate. It is concluded that the present method can be used in studying the capacity of the glutamate transporters in vivo.     

Comparison of three different experimental methods for the assessment of peripheral compartment pharmacokinetics in humans

In many cases the concentration reached in a peripheral effect compartment rather than in plasma determines the clinical outcome of therapy. Therefore, several experimental approaches have been developed for direct assessment of drug kinetics in peripheral compartments. Particularly saliva sampling, skin blister fluid sampling, and in vivo microdialysis are frequently employed for measuring peripheral drug concentrations. However, data derived from these techniques have never been directly compared. In the present study, the tissue kinetics of theophylline were measured following single dose administration simultaneously in cantharides induced skin blisters, saliva and microdialysates of subcutaneous- and skeletal muscle- tissue and compared to plasma concentrations. Theophylline was administered to 9 healthy volunteers as an i.v. infusion of 240 mg. Mean ratio (AUCsaliva/AUCplasma) was 0.63 +/- 0.05, mean ratio (AUCblister/AUCplasma) was 0.69 +/- 0.12, mean ratio (AUCmuscle/AUCplasma) was 0.41 +/- 0.10, mean ratio (AUCsubcutaneous/AUCplasma) was 0.34 +/- 0.07. The time course of the concentration(peripheral)/concentration(plasma)-ratios showed that tissue concentrations obtained by microdialysis were closely correlated to free plasma levels, whereas saliva- and cantharides blister data overestimated the corresponding free plasma concentrations. It is concluded that microdialysis represents a reliable technique for the measurement of unbound peripheral compartment concentrations and is superior to saliva- and skin blister concentration measurements.     

Transport of alpha-aminoisobutyric acid across the blood-brain barrier studied with in situ perfusion of rat brain

Transport of alpha-aminoisobutyric acid (AIB) from blood to brain in pentobarbital-anesthetized rats was examined using in situ perfusion. In situ perfusion with washed sheep red blood cells allowed the precise control of the composition of the perfusate that was necessary for a detailed examination of the transport of AIB. Retrograde perfusion at 4 ml/min through the left external carotid artery with oxygenated, artificial blood (hematocrit = 0.3) maintained a normal electroencephelogram during a 10 min experiment. The perfusate cerebral blood flow, at a value of 1.2 +/- 0.1 ml/g/min, and the perfusate cerebral plasma volume, at a value of 5.4 +/- 1.9 microliter/g, in the left frontal cortex were within the range of reported in vivo values. The in situ PS product for AIB (3.8 +/- 0.4 microliter/g/min) was higher than the value observed in vivo. AIB uptake was reduced to the in vivo value by 2 mM phenylalanine (1.3 +/- 0.3 microliter/g/min) and equally well by a mixture of neutral amino acids at their normal plasma concentrations but was unaffected by 2 mM methyl-AIB or removal of sodium from the perfusate. A kinetic analysis showed that the apparent Ki for phenylalanine inhibition of AIB transport was 19.8 +/- 4.9 microM. Thus, although AIB has affinity for the large neutral amino acid carrier in the blood-brain barrier, brain uptake by this mechanism in vivo is negligible due to competition by other amino acids in the plasma.     

High resolution renal diffusion imaging using a modified steady-state free precession sequence

A modified steady-state free precession (SSFP) diffusion sequence is proposed for high resolution renal imaging. A pair of bipolar diffusion gradients was used to minimize the errors in measured apparent diffusion coefficient (ADC) caused by variations in T1, T2, and RF flip angle that have been observed with previously employed SSFP diffusion sequences. Motion sensitivity was reduced by the use of compensated gradients, frame-by-frame averaging, and a repetition time of 22 ms, which for a single-acquisition 128 x 128 image requires only 3 s. High resolution was achieved by signal averaging. The modified sequence was applied to in vivo diffusion measurements. In six normal rat kidneys the ADCs (mean +/- SD; x 10(-3) mm2/s) of the cortex, outer medulla, and inner medulla were 2.28 +/- 0.05, 2.38 +/- 0.10, and 2.95 +/- 0.05, respectively. The technique requires relatively large gradients to achieve adequate diffusion weighting.     

Percutaneous penetration of 2-phenoxyethanol through rat and human skin

2-Phenoxyethanol applied in methanol was absorbed (64 +/- 4.4% at 24 hr) through unoccluded rat skin in vitro in the static diffusion cell with ethanol/water as receptor fluid. By comparison (43 +/- 3.7% in 24 hr) was absorbed in the flow-through diffusion system with tissue culture medium as receptor fluid. 2-Phenoxyethanol applied in methanol was absorbed (59.3 +/- 7.0% at 6 hr) through unoccluded human skin in vitro in the flow-through diffusion cell with tissue culture medium. With both unoccluded cells, 2-phenoxyethanol was lost by evaporation but occlusion of the static cell reduced evaporation and increased total absorption to 98.8 +/- 7.0%. Skin, post mitochondrial fraction, metabolized phenoxyethanol to phenoxyacetic acid at 5% of the rate for liver. Metabolism was inhibited by 1 mM pyrazole, suggesting involvement of alcohol dehydrogenase. However, first-pass metabolism of phenoxyethanol to phenoxyacetic acid was not detected during percutaneous penetration through viable rat skin in the flow-through system. First-pass metabolism in the skin does not therefore have an influence on systemic availability of dermally absorbed phenoxyethanol. These measures of phenoxyethanol absorption through rat and human skin in vitro agree well with those obtained previously in vivo.