Constant Infusion Method Research Articles

Intercompartmental communication between the cerebrospinal and adjacent spaces during intrathecal infusions in an acute ovine in-vivo model

IntroductionThe treatment of hydrocephalus has been a topic of intense research ever since the first clinically successful use of a valved cerebrospinal fluid shunt 72 years ago. While ample studies elucidating different phenomena impacting this treatment exist, there are still gaps to be filled. Specifically, how intracranial, intrathecal, arterial, and venous pressures react and communicate with each other simultaneously.MethodsAn in-vivo sheep trial (n = 6) was conducted to evaluate and quantify the communication existing within the cranio-spinal, arterial, and venous systems (1 kHz sampling frequency). Standardized intrathecal infusion testing was performed using an automated infusion apparatus, including bolus and constant pressure infusions. Bolus infusions entailed six lumbar intrathecal infusions of 2 mL Ringer’s solution. Constant pressure infusions were comprised of six regulated pressure steps of 3.75 mmHg for periods of 7 min each. Mean pressure reactions, pulse amplitude reactions, and outflow resistance were calculated.ResultsAll sheep showed intracranial pressure reactions to acute increases of intrathecal pressure, with four of six sheep showing clear cranio-spinal communication. During bolus infusions, the increases of mean pressure for intrathecal, intracranial, arterial, and venous pressure were 16.6 ± 0.9, 15.4 ± 0.8, 3.9 ± 0.8, and 0.1 ± 0.2 mmHg with corresponding pulse amplitude increases of 2.4 ± 0.3, 1.3 ± 0.3, 1.3 ± 0.3, and 0.2 ± 0.1 mmHg, respectively. During constant pressure infusions, mean increases from baseline were 14.6 ± 3.8, 15.5 ± 4.2, 4.2 ± 8.2, and 3.2 ± 2.4 mmHg with the corresponding pulse amplitude increases of 2.5 ± 3.6, 2.5 ± 3.0, 7.7 ± 4.3, and 0.7 ± 2.0 mmHg for intrathecal, intracranial, arterial, and venous pulse amplitude, respectively. Outflow resistances were calculated as 51.6 ± 7.8 and 77.8 ± 14.5 mmHg/mL/min for the bolus and constant pressure infusion methods, respectively—showing deviations between the two estimation methods.ConclusionsStandardized infusion tests with multi-compartmental pressure recordings in sheep have helped capture distinct reactions between the intrathecal, intracranial, arterial, and venous systems. Volumetric pressure changes in the intrathecal space have been shown to propagate to the intraventricular and arterial systems in our sample, and to the venous side in individual cases. These results represent an important step into achieving a more complete quantitative understanding of how an acute rise in intrathecal pressure can propagate and influence other systems.

Mathematical Analysis of a Mathematical Model of Chemovirotherapy: Effect of Drug Infusion Method.

A mathematical model for the treatment of cancer using chemovirotherapy is developed with the aim of determining the efficacy of three drug infusion methods: constant, single bolus, and periodic treatments. The model is in the form of ODEs and is further extended into DDEs to account for delays as a result of the infection of tumor cells by the virus and chemotherapeutic drug responses. Analysis of the model is carried out for each of the three drug infusion methods. Analytic solutions are determined where possible and stability analysis of both steady state solutions for the ODEs and DDEs is presented. The results indicate that constant and periodic drug infusion methods are more efficient compared to a single bolus injection. Numerical simulations show that with a large virus burst size, irrespective of the drug infusion method, chemovirotherapy is highly effective compared to either treatments. The simulations further show that both delays increase the period within which a tumor can be cleared from body tissue.

Measuring Protein Synthesis Rate In Living Object Using Flooding Dose And Constant Infusion Methods

Constant infusion is a method used for measuring protein synthesis rate in living object which uses low concentration of amino acid tracers. Flooding dose method is another technique used to measure the rate of protein synthesis which uses labelled amino acid together with large amount of unlabelled amino acid. The latter method was firstly developed to solve the problem in determination of precursor pool arise from constant infusion method. The objective of this writing is to compare the results from several researches on measuring protein synthesis rate using flooding dose and constant infusion methods and to obtain the criteria or prerequisite on the type of sample and tracer for each method. The measurements of protein synthesis rate using both methods are presented and results from both methods are compared. Both methods are eligible for measuring protein synthesis rate as long as the true precursor can be measured. The uncertainties in determining the precursor in constant infusion method and the effect of flooding on the stimulation of protein are the limitation on using the two methods. Bolus injection which imitates the flooding dose method but using smaller dose of tracer can be a good alternative in measuring protein synthesis rate.

Comparison of bolus injection and constant infusion methods for measuring muscle protein fractional synthesis rate in humans

BackgroundThe use of stable isotope tracer techniques to measure muscle protein fractional synthesis rate (FSR) has been well established and widely used. The most common method that has been utilized so far is a primed constant infusion (CI) method, which requires 3–4h of tracer infusion. However, recently our group has developed a bolus injection (BI) method, which requires an injection of bolus of tracer and can be completed within 1h. In this study, we compared calf (gastrocnemius) muscle protein FSR measured using these two different methods — CI and BI. MethodFSRs were measured in eight people (5 men and 3 women; age: 62.3±6.9years (mean±SD); body weight: 75.4±21.5kg) at basal, postabsorptive state using L-[ring-2H5]-phenylalanine. In the CI protocol, a primed continuous infusion was given for 4h, and muscle biopsies were taken at 120 and 240min; in the BI, a bolus injection of the tracer was given at 0min and biopsies were taken at 5 and 60min. Tracer enrichments in blood and muscle tissue were determined by gas chromatography–mass spectrometry. Data are expressed as mean±SE; t-test, linear regression and Levene Median equal variance test analyses were performed. ResultsCI FSR was 0.066±0.006%/h, whereas BI FSR was 0.058±0.008%/h, p=NS. The linear regression analysis showed a significant relationship between BI and CI, p=0.038. The intra-class correlation coefficient was 0.83. The standard deviation of the differences in the measurements was 0.015%/h. The Levene Median equal variance test demonstrated no difference in variance between the CI and BI measurements (p=0.722). ConclusionNo difference could be detected in calf muscle protein FSR measured by CI and BI methods; the BI method can be used for the measurement of muscle protein FSR in humans.

The Functional DRD3 Ser9Gly Polymorphism (rs6280) Is Pleiotropic, Affecting Reward as Well as Movement

Abnormalities of motivation and behavior in the context of reward are a fundamental component of addiction and mood disorders. Here we test the effect of a functional missense mutation in the dopamine 3 receptor (DRD3) gene (ser9gly, rs6280) on reward-associated dopamine (DA) release in the striatum. Twenty-six healthy controls (HCs) and 10 unmedicated subjects with major depressive disorder (MDD) completed two positron emission tomography (PET) scans with [11C]raclopride using the bolus plus constant infusion method. On one occasion subjects completed a sensorimotor task (control condition) and on another occasion subjects completed a gambling task (reward condition). A linear regression analysis controlling for age, sex, diagnosis, and self-reported anhedonia indicated that during receipt of unpredictable monetary reward the glycine allele was associated with a greater reduction in D2/3 receptor binding (i.e., increased reward-related DA release) in the middle (anterior) caudate (p<0.01) and the ventral striatum (p<0.05). The possible functional effect of the ser9gly polymorphism on DA release is consistent with previous work demonstrating that the glycine allele yields D3 autoreceptors that have a higher affinity for DA and display more robust intracellular signaling. Preclinical evidence indicates that chronic stress and aversive stimulation induce activation of the DA system, raising the possibility that the glycine allele, by virtue of its facilitatory effect on striatal DA release, increases susceptibility to hyperdopaminergic responses that have previously been associated with stress, addiction, and psychosis.

Reference values for CSF outflow resistance and intracranial pressure in healthy elderly

The intracranial pressure (ICP) and CSF outflow resistance (R(out)) are essential to describe the dynamics of the CSF system. R(out) affects ICP, pulse amplitudes, CSF absorption, and the compliance of the system. The objective of this study was to determine the reference values in healthy elderly subjects. Elderly people (60-82 years), who considered themselves healthy, were recruited through an advertisement in the local newspaper. All were evaluated with a 3-T MRI. Subjects were eligible if they did not have any psychiatric or neurologic disorder or signs of advanced atherosclerotic disease. CSF resting pressure (ICP) and R(out) were determined by a constant pressure infusion method with the patient in the supine position. The study population consisted of 40 subjects (mean age 70 years; 23 women). The median ICP was 11.6 mm Hg (15.8 cmH(2)O) and the reference interval was ICP 7.8-14.3 mm Hg (10.6-19.4 cmH(2)O) (defined as 5th to 95th percentiles). The median R(out) was 8.6 mm Hg/mL/min. The variation in R(out) was large and not normally distributed. The 90th percentile of R(out) was 17.4 mm Hg/mL/min. This study reports reference values for ICP and R(out) and should be used for comparison when investigating disorders with suspected CSF dynamic disturbances in the elderly. ICP was in the same range as that reported in the young and middle-aged. The upper limit of R(out) was higher than previously believed to be the upper limit of normal for this age group.

Advantage of predictive modeling in tracer studies

Letters to the EditorAdvantage of predictive modeling in tracer studiesStewart RussellStewart RussellMarc and Ruti Bell Vascular Biology Program, Department of Medicine (Cardiology), New York University School of Medicine, New York, New YorkPublished Online:01 Jan 2011https://doi.org/10.1152/ajpendo.00537.2010MoreSectionsPDF (27 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations to the editor: Sorensen et al. (1) compare the production rate of triglyceride, as calculated from a bolus injection decay curve, to that obtained from a primed constant infusion curve of the specific activity of radiolabeled tracer. They note a difference in the two rates and choose the constant infusion as the more accurate method. But is it? The bi-exponential model is a predictive model and uses measurements of the disappearance of tracer from the plasma compartment, over a limited period of time, to predict the plasma concentration of tracer for all time. The integral of this function over all time is the specific activity of the injected dose, which is used to calculate the production rate. On the other hand, the steady-state infusion model relies on measurement of the appearance of tracer in the plasma and therefore is only appropriate once the bi-directional flux of tracer between compartments has reached a steady state. At equilibrium, the primed constant infusion method will give the same production rate as the bolus. That it is different suggests that the time course is long enough to fit a bi-exponential decay curve but not long enough to establish steady state. In the bolus experiment, if tracer loss from the fast compartment is very rapid, its effect is negligible by the end of the experiment, which allows a second, slower decay curve to be detected. But in constant infusion, tracer that is fast to leave but slow to return will make the specific activity of the injected dose appear artificially low until equilibrium is reached. This is the power of the exponential function when used in this context: if the decay data fit a bi-exponential model, then all future information about the transfer of tracer between compartments will be described by the shape of the predicted curves. This two-pool model was introduced in these pages by Sapirstein et al. (2) as a simplified method to avoid possible errors in estimation attendant upon constant infusion methods. The specific activity of the rapid pool, as calculated from Sørensen, et al. solution of the bi-exponential model, is 22% of the total specific activity; if this missing amount is added to the measured specific activity in the constant infusion experiment, then the calculated production rates are the same. The more accurate number is derived from the bi-exponential equation, not the steady-state experiment, and, as the authors correctly conclude, is a strong indicator of rapid recycling of VLDL-TG through the periphery.DISCLOSURESNo conflicts of interest are reported by the author.REFERENCES1. Sørensen LP, Gormsen LC, Nielsen S. VLDL-TG kinetics: a dual isotope study for quantifying VLDL-TG pool size, production rates, and fractional oxidation in humans. Am J Physiol Endocrinol Metab 297: E1324–E1330, 2009.Link | ISI | Google Scholar2. Sapirstein LA, Vidt DG, Mandel MJ, Hanusek G. Volumes of distribution and clearances of intravenously injected creatinine in the dog. Am J Physiol 181: 330–336, 1955.Link | ISI | Google ScholarAUTHOR NOTESAddress for reprint requests and other correspondence: S. Russell, Marc and Ruti Bell Vascular Biology Program, Department of Medicine (Cardiology), NYU School of Medicine, Smilow Bldg., 522 1st Ave., 8th Floor, New York, NY 10016 (e-mail: stewart.[email protected]org). Download PDF Previous Back to Top Next FiguresReferencesRelatedInformation Related ArticlesReply to Russell: VLDL-TG kinetics: how to interpret a dual-isotope study 01 Jan 2011American Journal of Physiology-Endocrinology and Metabolism More from this issue > Volume 300Issue 1January 2011Pages E252-E252 Copyright & PermissionsCopyright © 2011 the American Physiological Societyhttps://doi.org/10.1152/ajpendo.00537.2010PubMed21189361History Published online 1 January 2011 Published in print 1 January 2011 Metrics

Human muscle protein turnover—why is it so variable?

We undertook a comprehensive review of the literature to unravel the nature of the variability in the reported rate of human muscle protein synthesis. We analyzed the results from studies that report the protein fractional synthesis rate (FSR) in the vastus lateralis in healthy, nonobese, untrained adults ≤50 yr of age in the postabsorptive state at rest by using the primed, constant tracer amino acid infusion method according to experimental design characteristics. We hypothesized that if the variability is methodological (rather than physiological) in nature, systematic clustering of FSR values would be evident, and outliers would become apparent. Overall, as expected, the mixed muscle protein FSR values were significantly (P < 0.001) greater when the muscle vs. the plasma free amino acid enrichment is used as the surrogate precursor pool enrichment, and the average mixed muscle protein FSR values were significantly greater (P = 0.05) than the myofibrillar/myosin heavy chain FSR values. The within-study variability (i.e., population variance) was somewhat smaller in studies that used plasma amino acid/ketoacid enrichments vs. muscle free amino acid enrichment (∼24 vs. ∼31%), but this was not apparent in all circumstances. Furthermore, the between-study consistency of measured FSR values (i.e., interquartile range) was inversely correlated with the average duration between biopsies. Aside from that, the variation in reported FSR values could not be explained by differences in the experimental design and analytical methods, and none of the most commonly used approaches stood out as clearly superior in terms of consistency of results and/or within-study variability. We conclude that the variability in reported values is in part due to 1) differences in experimental design (e.g., choice of precursor pool) and 2) considerable within-subject variability. The summary of the results from our analysis can be used as guidelines for "normal" average basal FSR values at rest in healthy adults.

Comparison between 3 infusion methods to measure cerebrospinal fluid outflow conductance

There are several infusion methods available to estimate the outflow conductance (Cout) or outflow resistance (Rout=1/Cout) of the CSF system. It has been stated that for unknown reasons, the bolus infusion method estimates a higher C(out) than steady-state infusion methods. The aim of this study was to compare different infusion methods for estimation of Cout. The following 3 different infusion methods were used: the bolus infusion method (Cout bol); the constant flow infusion method, both static (Cout stat) and dynamic (Cout dyn) analyses; and the constant pressure infusion method (Cout cpi). Repeated investigations were performed on an experimental model with well-known characteristics, with and without physiological pressure variations (B-waves, breathing, and so on). All 3 methods were also performed in a randomized order during the same investigation in 20 patients with probable or possible idiopathic normal-pressure hydrocephalus; 6 of these patients had a shunt and 14 did not. Without the presence of physiological pressure variations, the concordance in the experimental model was good between all methods. When they were added, the repeatability was better for the steady-state methods and a significantly higher Cout was found with the bolus method in the region of clinically relevant Cout (p<0.05). The visual fit for the bolus infusion was dependent on subjective assessment by the operator. This experimental finding was confirmed by the clinical results, where significant differences were found in the investigations in patients without shunts between Cout of the visual bolus method and Cout stat, Cout dyn, and Cout cpi (4.58, 4.18, and 6.12 μl/[second×kPa], respectively). This study emphasized the necessity for standardization of Cout measurements. An experienced operator could partly compensate for difficulties in correctly estimating the pressure parameters for the bolus infusion method, but for the general user this study suggests a steady-state method for estimating Cout.

Tear volume, turnover rate, and flow rate in ophthalmologically normal horses

To determine tear volume, turnover rate, and flow rate in ophthalmologically normal horses by use of fluorophotometry. 12 mares free of ophthalmic disease. 2 microL of 10% sodium fluorescein was instilled onto 1 eye of each horse, and tear samples were collected via microcapillary tubes from the inferonasal conjunctival culde-sac at 0, 2, 4, 6, 10, 15, and 20 minutes after instillation. Collected tear samples were then measured for fluorescein concentrations with a computerized scanning ocular fluorophotometer. A decay curve plot of concentration changes over time was used to determine tear flow rate and volume through 2 different mathematical treatments of the data (the including method and the excluding method). Fluorescein concentration in tears decreased in a first-order manner. The including method yielded a mean tear volume of 360.09 microL, a turnover rate of 12.22%/min, and a flow rate of 47.77 microL/min. The excluding method yielded values of 233.74 microL, 13.21%/min, and 33.62 microL/min, respectively. Mean +/- SD correlation coefficients for the natural logarithm of the fluorescein concentration versus time were 0.93 +/- 0.12 for the including method and 0.98 +/- 0.03 for the excluding method. The excluding method yielded more accurate results. A tear flow rate of 33.62 microL/min and a tear volume of 233.74 microL imply a complete recycling of the tear volume in approximately 7 minutes and suggest that increased dosing regimens or constant infusion methods for topical administration of ophthalmic drugs may be indicated when treating horses for corneal disease in which high ocular surface concentrations are needed.

EGFR

GFR is the accepted metric for evaluating renal function but measuring it directly (mGFR) can be cumbersome, time-consuming, and costly. The constant infusion method using inulin as the filtration marker, introduced in 1934 by Richards and Smith (1), has undergone many revisions that have now made mGFR easier to perform, although not widely available (2,3). A simple, reliable, unbiased, precise, and accurate estimation of mGFR, without reliance on constant infusions or timed urinary collections, would be welcome; however, as illustrated by the extensive compendium of comparisons of mGFR (using classical inulin clearance methods) to eGFR by Botev et al. (4) in this issue of CJASN , this objective is far from being realized. Although formulaic methods to approximate the values of mGFR from serum creatinine (Scr) levels have been used for many decades, the seminal description of the Modification of Diet in Renal Disease (MDRD) equation (eGFR-MDRD) in 1999 galvanized interest in this common and important clinical issue (5). The often used Cockcroft-Gault equation (6) was never intended to be an approximation of GFR but rather was designed to estimate endogenous creatinine clearance, which is not equivalent to GFR because of the effects of tubular secretion of creatinine (TScr). In normal individuals, this can be compensated for by a correction factor, because TScr is relatively constant (at approximately 22 to 24% of GFR) (7); however, TScr can vary markedly in disease states, particularly those associated with heavy proteinuria (8). Compared with mGFR, the current widely used eGFR-MDRD formulas (original and reexpressed [5,9]) have limitations because of variable degrees of bias, imprecision, and inaccuracy (10). The studies of Botev et al. (4) in this issue of CJASN provide additional quantitative data on the ranges of bias, precision, and accuracy of eGFR-MDRD according to the levels …

Feasibility and Impact of the Measurement of Extracellular Fluid Volume Simultaneous with GFR by 125I-Iothalamate

The feasibility, validity, and possible applications of the assessment of extracellular fluid volume (ECFV) simultaneous with glomerular filtration rate (GFR) were assessed in a series of validation studies using the constant infusion method of (125)I-iothalamate (IOT). In 48 subjects with a broad range of GFR, distribution volume (V(d)) of IOT corresponded well with V(d) bromide (16.71 +/- 3.0 and 16.73 +/- 3.2 l, respectively, not significant), with a strong correlation (r = 0.933, P < 0.01) and without systematic deviations. Reproducibility assessment in 25 healthy male subjects showed coefficients of variation of 8.6% of duplicate measurement of V(d) IOT during strictly standardized (50 mmol Na(+)/d) sodium intake. An increase in dietary sodium intake (200 mmol Na(+)/d) induced a corresponding rise in V(d) IOT of 1.11 +/- 1.5 l (P < 0.01). In 158 healthy prospective kidney donors, the impact of indexing of GFR to ECFV was analyzed. Age, gender, height, and body surface area (BSA) were determinants of GFR. Whereas GFR, GFR/BSA, and GFR/height were gender-dependent, GFR/ECFV was gender-independent and not related to height or BSA. This supports the potential of normalizing GFR by ECFV. Thus, ECFV can be simultaneously assessed with GFR by the constant infusion method using IOT. After appropriate validation, also other GFR tracers could be used for such a simultaneous estimation, providing a valuable resource of data on ECFV in renal studies and, moreover, allowing GFR to be indexed to the body fluid compartment it clears: the ECFV.

Assessment of cerebrospinal fluid outflow conductance using an adaptive observer—experimental and clinical evaluation

Idiopathic normal pressure hydrocephalus (INPH) patients have a disturbance in the dynamics of the cerebrospinal fluid (CSF) system. The outflow conductance, C, of the CSF system has been suggested to be prognostic for positive outcome after treatment with a CSF shunt. All current methods for estimation of C have drawbacks; these include lack of information on the accuracy and relatively long investigation times. Thus, there is a need for improved methods. To accomplish this, the theoretical framework for a new adaptive observer (OBS) was developed which provides real-time estimation of C. The aim of this study was to evaluate the OBS method and to compare it with the constant pressure infusion (CPI) method. The OBS method was applied to data from infusion investigations performed with the CPI method. These consisted of repeated measurements on an experimental set-up and 30 patients with suspected INPH. There was no significant difference in C between the CPI and the OBS method for the experimental set-up. For the patients there was a significant difference, −0.84 ± 1.25 µl (s kPa)−1, mean ± SD (paired sample t-test, p < 0.05). However, such a difference is within clinically acceptable limits. This encourages further development of this new real-time approach for estimation of the outflow conductance.

The Increase in Human Muscle Protein Synthesis Induced by Food Intake Is Similar When Assessed with the Constant Infusion and Flooding Techniques

Food intake is accompanied by a stimulation of muscle protein synthesis. However, the reported magnitude of the response differs with different methods of measurement. The aim of this study was to assess whether the response to feeding is dependent on the technique used for measurement when length and amount of feeding are controlled. Muscle protein fractional synthesis rates (FSRs) were measured both in the fasting and feeding states in 2 groups of healthy volunteers (n = 8). Two techniques were used to measure FSR: in one group, FSRs were assessed with a primed constant infusion of L-[2H5]phenylalanine, whereas in the other, a flooding amount of the same label was employed. The fasting FSRs assessed with the constant infusion method and estimated using the free amino acid in the tissue fluid to represent the precursor pool for protein synthesis were comparable to those obtained with the flooding method (1.94 +/- 0.15 vs. 1.86 +/- 0.13%/d). The degree of stimulation due to feeding (P < 0.02) did not differ between the constant infusion (+15%) and flooding (+22%) techniques. The stimulatory effect of feeding on muscle FSR was associated with enhanced phosphorylation of the Mr = 70,000 ribosomal protein S6 kinase, suggesting that it may involve activation of translation. This study demonstrates that human muscle FSRs obtained with the constant infusion technique are comparable to those obtained with the flooding method and that, in response to feeding, the 2 techniques give comparable estimates of stimulation.

Response of human muscle protein synthesis to feeding: a comparison of methods

Feeding has been shown to stimulate muscle protein synthesis rates (FSR) in humans. However, while a substantial stimulation (+60–100%) of muscle FSR in response to feeding has been shown in studies using the constant infusion method, a much smaller response (+30%, P=NS) has been detected with the flooding method. The aim of this study was to assess whether the response of muscle protein synthesis to feeding depends on the technique used for measurement. Muscle FSR were measured both in the fasting and feeding states in two groups (n=8) of healthy volunteers (Age 31±2 y), with the same tracer and feeding regimen. However, in one group FSR were assessed with a flooding amount of L-[2H5]phenylalanine (45 mg/kg BW) and in the other one with a primed constant infusion of the same labeled amino acid (6 μmol/kg/h). Fasting FSR obtained with the flooding method were similar to that obtained with the constant infusion method calculated with phenylalanine within the tissue taken as the precursor pool for protein synthesis (1.86±0.13 vs. 1.94±0.15 %/day). The results also show that feeding is associated with a stimulation of FSR, whether assessed by the flooding method (+22%, P<0.01) or constant infusion (+13%, P<0.05). Lower fasting FSR (1.31±0.07 %/day, P<0.01) and a greater stimulation after feeding (+25%, P<0.05) were observed with the constant infusion method when plasma was used as a surrogate precursor pool. This study demonstrates that human muscle FSR measured with the flooding method is comparable to that obtained with the constant infusion, and that, under similar experimental conditions, the two methods give comparable estimates of stimulation from feeding. Supported by NIH grants AG017446 and 5-MO1-RR-10710

Nitric oxide and cyclic GMP enhance aqueous humor outflow facility in rabbits

Purpose. Nitric oxide (NO) may control intraocular pressure (IOP)-regulating mechanisms physiologically and in ocular diseases such as glaucoma. The aim of the present study was to clarify whether an increase in aqueous humor outflow facility could explain the IOP-lowering effect of the NO/cyclic GMP pathway we recently described. Methods. Test compounds were administered to anesthetized rabbits (New Zealand White, n = 6) intracamerally (5µl) in the following doses: nitrosocaptopril 12.3µg, captopril 10.9µg, sodium nitroprusside (SNP) 13.1µg and 8-Brc-GMP 22.3µg. Outflow facility (C) was determined by the two-level constant pressure infusion method. Outflow facility, C 1 and C 2, was measured at lower and higher pressure levels, respectively. Results. Outflow facility was increased after treatment with SNP (increase in C in the experimental eye as compared to the control eye C 1 80% and C 2 74%), nitrosocaptopril (C 1 69% and C 2 64%) and 8-Br-cGMP (C 1 35% and C 2 33%). Captopril had no effect on outflow facility (C 1 -12% and C 2 2%). Blood pressure was not affected by the drugs. Conclusions. We conclude that enhancement of outflow facility by nitrosocaptopril, SNP and 8-Br-cGMP, their second messenger derivative, at least partly explains the IOP-lowering effect of NO releasing compounds.

Of flux and flooding: the advantages and problems of different isotopic methods for quantifying protein turnover in vivo : II. Methods based on the incorporation of a tracer.

The most common methods for measuring the incorporation of tracer amino acids into tissue protein are the constant tracer infusion and the flooding dose. The flooding dose is an attractive method for measuring tissue protein synthesis because of its convenience and precision. A primary assumption of the method, that the free amino acid precursor pools are equilibrated with the true precursor pool, aminoacyl-transfer RNA, has recently been validated. When short labelling periods are involved, the large dose of amino acid does not appear to alter protein synthesis. The constant tracer infusion is a satisfactory method from a theoretical point of view, but its use requires the measurement of the protein synthetic precursor pool. The best estimate of the aminoacyl-tRNA precursor pool for the constant infusion method appears to be the acid-soluble tissue pool in muscle and VLDL apolipoprotein B-100 in the liver. The experimental approach chosen for measuring tissue protein synthesis should be dictated by the question being addressed.

Effects of vigabatrin on the GABAergic system as determined by [123I]iomazenil SPECT and GABA MRS.

To evaluate effects of vigabatrin (VGB) by using [123I]iomazenil single-photon emission computed tomography (SPECT) to estimate central gamma-aminobutyric acid (GABA(A))/benzodiazepine receptors (BZRs), and magnetic resonance spectroscopy (MRS) to assess tissue GABA levels. Six patients with partial seizures had both SPECT and MRS before and 25-84 days after starting VGB (3 g p.o., q.d.). SPECT was acquired by using the constant-infusion method and, after nonuniform attenuation correction, coregistered with T1-weighted MR Imaging (MRI) A volume of interest (VOI) of 3 x 2 x 2 cc over the occipital cortex, used for MRS acquisition, was positioned on both MRI and coregistered SPECT. Occipital activity was divided by either total plasma activity or plasma [123I]iomazenil concentration to estimate BZR distribution volume (V(T)-p and V'(T), respectively). Wilcoxon's test was used for VOI differences in GABA levels, BZR V(T)-p or V'(T). SPM96 (either no global normalization or proportional scaling) was used to compare BZR V(T)-p changes in the patients with and without VGB with test-retest data in eight healthy age-matched controls. Occipital GABA levels were increased threefold (without VGB, 1.1+/-0.1 micromol/g; with VGB, 2.9+/-0.5 micromol/g; p = 0.027). BZR distribution volumes showed no change, when estimated by either V(T)-p (without VGB, 6.00+/-0.91 ml/g; with VGB, 5.86+/-0.44 ml/g; p = 0.92) or V(T) (without VGB, 41.1+/-11.2 ml/g; with VGB, 41.2+/-9.9 ml/g; p = 0.75). No significant changes were detected by SPM96. A clinically effective dose of VGB caused a threefold increase in tissue GABA levels but was not associated with a substantial BZR downregulation.

Regional cerebral blood flow, white matter abnormalities, and cerebrospinal fluid hydrodynamics in patients with idiopathic adult hydrocephalus syndrome.

OBJECTIVES--(1) to evaluate regional cerebral blood flow (rCBF) with single photon emission computed tomography and 99mTc-hexamethylpropyleneamine oxime in patients with the idiopathic adult hydrocephalus syndrome (IAHS); (2) to examine regional cerebral blood flow (rCBF), gait, and psychometric functions before and after CSF removal (CSF tap test); (3) to assess abnormalities in subcortical white matter by MRI. METHODS--Thirty one patients fulfilling the criteria for IAHS (according to history and clinical and neuroradiological examination) were studied. Quantified gait measurements, psychometric testing, and rCBF before and after removal of CSF were obtained. Pressure of CSF and CSF outflow conductance were investigated with a constant pressure infusion method. Brain MRI was used to quantify the severity of white matter lesions and periventricular hyperintensities. In IAHS a widespread rCBF hypoperfusion pattern was depicted, with a caudal frontal and temporal grey matter and subcortical white matter reduction of rCBF as the dominant feature. Removal of CSF was not accompanied by a concomitant increase in rCBF. Significant white matter lesions were detected only in a minority of patients by MRI. An altered CSF hydrodynamic state with a higher CSF pressure and lower conductance was confirmed. IAHS is characterised by an abnormal CSF hydrodynamic state, associated with a widespread rCBF reduction with preference for subcortical white matter and frontal-temporal cortical regions. Furthermore in most patients MRI did not show white matter changes suggestive of a coexistent subcortical arteriosclerotic encephalopathy. At least in the idiopathic group of patients with AHS, measurements of rCBF before and after temporary relief of the CSF hydrodynamic disturbance will not provide additional information that would be helpful in the preoperative evaluation but is suggestive of a preserved autoregulation of rCBF.

Enteral bioavailability of human granulocyte colony stimulating factor conjugated with poly(ethylene glycol).

The focus of this paper is to demonstrate that pegylation of a therapeutic protein, recombinant human granulocyte colony stimulating factor (PEG-G-CSF), results in an increase in stability and in retention of in vivo bioactivity when administered by the intraduodenal route and may, therefore, be a suitable form of the protein for inclusion in an oral delivery formulation. The ability of PEG-G-CSF to elicit a therapeutic response from the enteral route was investigated by two methods of intraduodenal dosing in an in vivo model to determine the optimal dosing method: by slow, constant infusion, or by a single bolus administration. Circulating levels of the proteins confirmed that PEG-G-CSF was delivered into the systemic circulation from the enteral route and that biological activity was retained. Bioavailability from the enteral route by the constant infusion method was calculated from the intravenous administration of the proteins to be between 1.8 and 3.5% while un-modified G-CSF failed to elicit a quantifiable response by this method. Bolus administration of PEG-G-CSF also resulted in biological activity although responses were short lived and significantly lower than with the pegylated formulation. The possible mechanisms of enteral delivery of PEG-G-CSF are discussed. Our results indicate that oral delivery of pegylated G-CSF may be possible and in fact, preferable to using the un-modified form of the therapeutic.