Absolute timing of the photoelectric effect.

Photoemission spectroscopy is central to understanding the inner workings of condensed matter, from simple metals and semiconductors to complex materials such as Mott insulators and superconductors1. Most state-of-the-art knowledge about such solids stems from spectroscopic investigations, and use of subfemtosecond light pulses can provide a time-domain perspective. For example, attosecond (10-18 seconds) metrology allows electron wave packet creation, transport and scattering to be followed on atomic length scales and on attosecond timescales2-7. However, previous studies could not disclose the duration of these processes, because the arrival time of the photons was not known with attosecond precision. Here we show that this main source of ambiguity can be overcome by introducing the atomic chronoscope method, which references all measured timings to the moment of light-pulse arrival and therefore provides absolute timing of the processes under scrutiny. Our proof-of-principle experiment reveals that photoemission from the tungsten conduction band can proceed faster than previously anticipated. By contrast, the duration of electron emanation from core states is correctly described by semiclassical modelling. These findings highlight the necessity of treating the origin, initial excitation and transport of electrons in advanced modelling of the attosecond response of solids, and our absolute data provide a benchmark. Starting from a robustly characterized surface, we then extend attosecond spectroscopy towards isolating the emission properties of atomic adsorbates on surfaces and demonstrate that these act as photoemitters with instantaneous response. We also find that the tungsten core-electron timing remains unchanged by the adsorption of less than one monolayer of dielectric atoms, providing a starting point for the exploration of excitation and charge migration in technologically and biologically relevant adsorbate systems.

[Clinical Comparison of a Trifocal and a Trifocal-Toric Intraocular Lens Based on the Same Diffractive Platform]. / Klinischer Vergleich einer trifokalen und einer trifokal-torischen Intraokularlinse auf gleicher diffraktiver Plattform.

Purpose To evaluate and compare the visual performance of a trifocal and a trifocal-toric intraocular lens (IOL) based on the same diffractive platform. Methods Prospective non-randomized comparative study enrolling 142 eyes of 77 patients (age 40-73 years) undergoing uneventful phacoemulsification lens surgery. Two groups were differentiated according to the implanted IOL: trifocal group, 98 eyes (50 patients) implanted with the trifocal diffractive IOL AT LISA tri 839MP (Carl Zeiss Meditec), and trifocal-toric group, 44 eyes (27 patients) implanted with the trifocal-toric diffractive IOL AT LISA trifocal-toric 939MP (Carl Zeiss Meditec). Visual and refractive changes were evaluated 3 months postoperatively. Results No significant differences between groups were found in postoperative refraction (p ≥ 0.144), monocular and binocular uncorrected intermediate visual acuity (UIVA; p = 0.519 and 0.398, respectively) and binocular uncorrected near visual acuity (UNVA; p = 0.073). In contrast, significantly better monocular and binocular uncorrected distance visual acuity (UDVA; p ≤ 0.002), as well as monocular UNVA (p = 0.005), were found in the trifocal group. A postoperative spherical equivalent within ± 1.00 D was found in 98 % and 100 % of eyes in the trifocal and trifocal-toric groups, respectively. Postoperative binocular UDVA, UIVA and UNVA of 20/30 or better were found in 100, 95 and 100 % of eyes, and in 96.3, 95.8 and 90.9 % of eyes in the trifocal and trifocal-toric groups, respectively. Conclusions The evaluated trifocal and trifocal-toric IOLs both provide a successful restoration of the visual function after cataract surgery, with good levels of refractive precision, as well as UIVA and UNVA.

[Clinical Results After Implantation of a New Diffractive, Multifokal Intraocular Lens with a Reduced Near Add Power (+ 2.75 D)]. / Klinische Ergebnisse nach Implantation einer neuen diffraktiven multifokalen Intraokularlinse mit einer reduzierten Nahaddition (+ 2,75 dpt).

PURPOSE: Evaluation of the clinical data 3 months after implantation of a new diffractive multifocal intraocular lens (MIOL) with a reduced near add power of + 2.75 D. METHODS: In a prospective study, patients who underwent cataract surgery or refractive lens exchange with implantation of an MIOL (Tecnis ZKB00, Abbott Medical Optics, Santa Ana, California, USA) were included. Three months postoperative corrected and uncorrected visual acuities at different distances were measured and evaluated. Those patients that underwent bilateral MIOL implantation additionally filled out a questionnaire 3 months postoperatively. RESULTS: Between October 2013 and August 2014, 115 eyes of 62 patients were implanted with the ZKB00 IOL. Mean postoperative refractions were - 0.27 ± 0.44 D for the spherical equivalent, respectively. Mean binocular CDVA was - 0.01 ± 0.3 logMAR with a mean binocular UDVA of 0.06 ± 0.08 logMAR. For near distance in 40 cm, an UNVA of 0.07 ± 0.10 logMAR three months postoperatively was measured. CONCLUSION: The ZKB00 IOL belongs to a group of novel MIOL with an increased intermediate visual performance. Our study shows good visual acuity at all distances, as well as a high rate of satisfaction and subjectively good image quality.

[Functional Results 3 Months after Implantation of an "Extended Range of Vision" Intraocular Lens]. / Funktionelle Ergebnisse 3 Monate nach Implantation einer "Extended-Range-of-Vision"-Intraokularlinse.

PURPOSE: Evaluation of the visual and refractive results 3 months after implantation of a diffractive extended range of vision (ERV) intraocular lens (IOL) during cataract surgery. METHODS: In a prospective multicentre study, patients with a calculated postoperative corneal astigmatism of ≤ 1.5 D received a diffractive ERV IOL (TECNIS Symfony, model ZXR00, Abbott Medical Optics, USA) during cataract surgery. After 3 months, the monocular and binocular corrected and uncorrected far, intermediate and near visual acuity, as well as refraction, were evaluated. RESULTS: 18 patients (36 eyes) with a mean age of 63.34 ± 4.6 years underwent bilateral cataract surgery. After 3 months, the binocular uncorrected distance visual acuity (UDVA) of logMAR was - 0.05 ± 0.11 and the corrected distance visual acuity (CDVA) of logMAR - 0.14 ± 0.05. Binocular uncorrected intermediate (UIVA) and near visual acuity (UNVA) were logMAR - 0.09 ± 0.02 and 0.19 ± 0.09, respectively. A target refraction of ± 0.75 D was reached by 89 % of the patients. CONCLUSION: Implantation of an extended range of vision intraocular lens offers an effective way for visual rehabilitation at far and intermediate distances. Near vision is still in a functional range.

[Refraction after Implantation of Multifocal and Presbyopia-Correcting Intraocular Lenses]. / Richtiges Refraktionieren nach Implantation von Multifokal- und presbyopiekorrigierenden Intraokularlinsen.

Nowadays, further developments in the field of intraocular lenses offer a higher level of spectacle independence for our patients. As light gets scattered on different focal points a wider range of defocus is created. This greater defocus area makes it more difficult for us to determine the objective or subjective refraction. This contribution is concerned with the difficulties of measuring visual acuity in different intraocular lens designs and different measurement distances. Measuring refraction after implantation of a multifocal intraocular lens is a complex procedure and the experience of the examiner plays a crucial role. Retinoscopy, keratometry and the defocus curve are reliable methods for testing, while the auto refractometer, bichromatic testing and the cross-cylinder have limitations.

[Functional Results One Year after Implantation of a Bitoric, Trifocal Intraocular Lens]. / Funktionelle Ergebnisse 1 Jahr nach Implantation einer bitorischen, trifokalen Intraokularlinse.

PURPOSE: An evaluation of the visual and refractive results was undertaken one year after implantation of a trifocal diffractive toric intraocular lens (IOL) during cataract surgery. METHODS: In a prospective study patients with a calculated postoperative corneal astigmatism of ≥ 0.75 D received a diffractive trifocal toric IOL (AT LISA tri toric 939MP, Carl Zeiss Meditech, Jena, Germany) during cataract surgery. One year postoperatively the near, intermediate and distance visual acuity, corrected and uncorrected vision as well as refraction were evaluated. RESULTS: 20 patients (40 eyes) with a median age of 59 ± 11 years of which 15 were female underwent bilateral cataract surgery. One year postoperatively a binocular uncorrected distance visual acuity (UDVA) of 0.10 logMAR ± 0.11 and a corrected distance visual acuity (CDVA) of 0.00 logMAR ± 0.08 could be found. Binocular intermediate visual acuity (UIVA) and near visual acuity (UNVA) were 0.00 logMAR ± 0.05 and 0.09 logMAR ± 0.07, respectively. 100 % of patients were between ± 1.0 D from target refraction. Even 1 year after surgery no patient had an IOL rotation greater than 5°. CONCLUSION: The implantation of a trifocal toric intraocular lens offers an effective way for visual rehabilitation in near, intermediate and far distances with a good rotational stability of the IOL platform.

Proteomic and Metabolomic Characterization of Metabolically Healthy Obesity: A Descriptive Study from a Swedish Cohort.

Method: Associations between different biomarkers (proteomics, lipidomics, and metabolomics) coupled to either MHO or metabolically unhealthy obese (MUO) individuals were analyzed through principal component analysis (PCA). Subjects were identified from a subsample of 416 obese individuals, selected from the Malmö Diet and Cancer study-Cardiovascular arm (MDCS-CV, n = 3,443). They were further divided into MHO (n = 143) and MUO (n = 273) defined by a history of hospitalization, or not, at baseline inclusion, and nonobese subjects (NOC, n = 3,027). Two distinctive principle components (PL2, PP5) were discovered with a significant difference and thus further investigated through their main loadings. Results: MHO individuals had a more metabolically favorable lipid and glucose profile than MUO subjects, that is, lower levels of traditional blood glucose and triglycerides, as well as a trend of lower metabolically unfavorable lipid biomarkers. PL2 (lipidomics, p=0.02) showed stronger associations of triacylglycerides with MUO, whereas phospholipids correlated with MHO. PP5 (proteomics, p=0.01) included interleukin-1 receptor antagonist (IL-1ra) and leptin with positive relations to MUO and galanin that correlated positively to MHO. The group differences in metabolite profiles were to a large extent explained by factors included in the metabolic syndrome. Conclusion: Compared to MUO individuals, corresponding MHO individuals present with a more favorable lipid metabolic profile, accompanied by a downregulation of potentially harmful proteomic biomarkers. This unique and extensive biomarker profiling presents novel data on potentially differentiating traits between these two obese phenotypes.

Reversal of visceral adiposity in candy-diet fed female Wistar rats by the CB1 receptor antagonist rimonabant.

OBJECTIVE: The severity of obesity is often more determined by the distribution of fat depots rather than by body weight itself. Therefore, the effect of rimonabant on fat distribution pattern was investigated in female candy-fed Wistar rats. DESIGN: Female Wistar rats were fed a high fat, high carbohydrate (candy-) diet for 12 weeks. During the last 6 weeks rats were treated with rimonabant. Food intake and body weight development were investigated, as well as effects on total body fat, especially visceral fat and ectopic lipid accumulation in skeletal muscle and liver, determined by in vivo magnetic resonance imaging/magnetic resonance spectroscopy. RESULTS: Candy-diet increased body weight, which was predominantly due to the increased total fat mass with predominance of visceral fat accumulation. Treatment with rimonabant fully reversed the weight gain and fat deposition in the visceral cavity and skeletal muscle, in contrast to pair feeding. In spite of an only transient reduction of food intake, body weight reduction, as well as normalized body fat, reduced visceral fat and intramyocellular lipids were maintained over the treatment period. CONCLUSIONS: We conclude that additional factors other than reduced caloric intake must be responsible for the improvements in these lipid parameters. The complete cluster of results is consistent with increased lipid oxidation caused by rimonabant.

Identification of the Arg-Gly-Asp sequence in laminin A chain as a latent cell-binding site being exposed in fragment P1.

A single RGD-containing sequence present within an epidermal growth factor-like repeat of the short arms of laminin is shown by peptide inhibition to block integrin receptors recognizing a latent cell-binding site of laminin. Based on proteolysis data it is proposed that masking occurs by folding of the globular domain IVa over the cell-binding site in the adjacent rod-like structures of laminin A chain.

Cultures with cryopreserved hepatocytes: applicability for studies of enzyme induction.

The use of hepatocyte cultures is well established for the study of drug-drug interactions. However, the major hindrance for the use of human hepatocyte cultures is that human hepatocytes are only occasionally available. This problem could be overcome by cryopreservation. Although cryopreserved hepatocytes have been recommended for short term applications in suspension, studies on induction of enzyme activity, requiring a more prolonged maintenance of cryopreserved hepatocytes in culture, represent a new field of research. In the present study, we established a technique that allows preparation of rat hepatocyte co-cultures, using cryopreserved hepatocytes. After incubation with phenobarbital (0.75 mM; 72 h) induction factors for the isoenzyme-dependent regio and stereoselective testosterone hydroxylations were 1.6, 2.2, 1.0, 2.1, 5.6, 2.4, 3.6, 4.5 and 0.9 for 2alpha-, 2beta-, 6alpha-, 6beta-, 7alpha-, 15beta-, 16alpha- and 16beta-hydroxytestosterone and 4-androsten-3,17 dione. Regarding induction factors of less than 2-fold, as questionable these induction factors were similar to those of cultures with freshly isolated hepatocytes and the induction pattern of the individual hydroxylation products was similar to the in vivo situation. In addition 3-methylcholanthrene (5 microM; 72 h) induced exclusively the formation of 7alpha-hydroxytestosterone (6.6-fold) in cultures with cryopreserved hepatocytes. This specificity also correlates to that obtained in rats. Although these induction factors were clearly satisfactory in cryopreserved cultures, the absolute activities of the main testosterone hydroxylation products were reduced when compared to fresh cultures. For instance, 6beta-hydroxytestosterone, the main metabolite in solvent controls was reduced to 79%, 7alpha-hydroxytestosterone, the main metabolite after induction with 3-MC, was reduced to 66% and 16beta-hydroxytestosterone, the main metabolite after induction with PB, was reduced to 52%. Similarly, EROD activity after induction with 3-methylcholanthrene in cryopreserved cultures was reduced to 62%, compared with that in fresh cultures. Although further optimization and validation is required, the data show that cytochrome P450 activities can clearly be induced in co-cultures of cryopreserved hepatocytes, in a fashion which for the investigated inducers, is similar to that in cultures from freshly isolated hepatocytes and similar to the in vivo situation.

Gene expression profiling in skeletal muscle of Zucker diabetic fatty rats: implications for a role of stearoyl-CoA desaturase 1 in insulin resistance.

AIMS/HYPOTHESIS: Insulin resistance in skeletal muscle is a hallmark of type 2 diabetes. Therefore, we sought to identify and validate genes involved in the development of insulin resistance in skeletal muscle. MATERIALS: Differentially regulated genes in skeletal muscle of male obese insulin-resistant, and lean insulin-sensitive Zucker diabetic fatty (ZDF) rats were determined using Affymetrix microarrays. Based on these data, various aspects of glucose disposal, insulin signalling and fatty acid composition were analysed in a muscle cell line overexpressing stearoyl-CoA desaturase 1 (SCD1). RESULTS: Gene expression profiling in insulin-resistant skeletal muscle revealed the most pronounced changes in gene expression for genes involved in lipid metabolism. Among these, Scd1 showed increased expression in insulin-resistant animals, correlating with increased amounts of palmitoleoyl-CoA. This was further investigated in a muscle cell line that overexpressed SCD1 and accumulated lipids, revealing impairments of glucose uptake and of different steps of the insulin signalling cascade. We also observed differential effects of high-glucose and fatty acid treatment on glucose uptake and long-chain fatty acyl-CoA profiles, and in particular an accumulation of palmitoleoyl-CoA in cells overexpressing SCD1. CONCLUSIONS/INTERPRETATION: Insulin-resistant skeletal muscle of ZDF rats is characterised by a specific gene expression profile with increased levels of Scd1. An insulin-resistant phenotype similar to that obtained by treatment with palmitate and high glucose can be induced in vitro by overexpression of SCD1 in muscle cells. This supports the hypothesis that elevated SCD1 expression is a possible cause of insulin resistance and type 2 diabetes.

Assembly of apoferritin from horse spleen: comparison of the protein in its native and reassembled state.

The in vitro selfassembly of apoferritin after previous dissociation and unfolding in 7.2M guanidinium chloride, pH 3.5, yields up to 80% of a protein complex exhibiting the molecular mass of the native icositetramer of greater than or equal to 450 kDa. After removal of high molecular mass byproducts, the final reassembly product proves to be indistinguishable from native apoferritin with respect to its functional and conformational properties. These refer to the intrinsic fluorescence and to the far and near UV circular dichroism. The unfolding transitions of the native and reassembled protein in aqueous guanidinium chloride or at acid pH coincide within the range of error. The reassembled protein is also able to catalyze the oxidation of Fe(II). Higher polymers of the apoferritin complex represent most of the residual 20% of the reconstituted protein. They are stabilized by non-covalent (preferentially hydrophobic) interactions, and may be disassembled to the icositetramer by preferential solvation of the protein in the presence of less than or equal to 50% (v/v) ethylene glycol. The change in fluorescence emission accompanying polymerization reflects altered surface properties of the apoferritin subunits compatible with those reported for the ferritin----hemosiderin transition.

Mechanism of the self-assembly of apoferritin from horse spleen. Cross-linking and spectroscopic analysis.

Apoferritin from horse spleen can be reversibly dissociated at pH2 or in 7.2 M G-HCl (pH 3.5). Reconstitution of the native icositetramer in 0.1 M TEA buffer (pH 7.9) in the presence of 1 mM EDTA and 3 mM dithioerythritol leads to yields higher than 80%. To monitor the kinetic mechanism, intrinsic fluorescence, far-UV circular dichroism, and covalent cross-linking with glutaraldehyde were applied. The overall mechanism of assembly is characterized by a sequence of concentration-dependent association reactions involving "structured monomers" and a dimeric intermediate as the most prominent species, apart from trimers and dodecamers. The parallel decrease in monomers, dimers and trimers indicates that association equilibria precede the formation of the final assembly product. The assembly reaction is accompanied by characteristic changes in fluorescence emission and dichroic absorption. To a first approximation, renaturation and reassociation may be quantitatively described by one single rate-determining second-order process, subsequent to fast folding steps at the monomer level.

Mechanism and specificity of reconstitution of dimeric lactate dehydrogenase from Limulus polyphemus.

D-Lactate dehydrogenase (EC 1.1.1.28) from Limulus polyphemus is a homodimer which is composed of identical subunits of Mr = 35 000. The enzyme may be reversibly denatured and dissociated at acid pH or in 6M guanidine X HCl. The sigmoidal time course of reactivation obeys a consecutive uni-bimolecular mechanism with k1 = 6 X 10(-4) S-1 and k2 = 1.3 X 10(-4) M-1 S-1 (20 degrees C) as first- and second-order rate constants. Cross-linking experiments with glutaraldehyde prove that reactivation and dimer formation run parallel. Joint "synchronous" reconstitution of the enzyme with dimeric porcine mitochondrial malate dehydrogenase (after denaturation in 6M guanidine X HCl) does not yield active hybrids. The unchanged kinetics of reactivation in the absence and presence of the prospective partner of hybridization prove that inactive hybrid intermediates may also be excluded. The absence of hybrids upon synchronous reconstitution of the two closely related dimeric NAD-dependent dehydrogenases clearly suggests that the assembly of nascent oligomeric proteins must be highly specific.

Localization of a major nidogen-binding site to domain III of laminin B2 chain.

The large pepsin fragments P1 and P1X, which comprise most of the rod-like domains III of the three short arms of laminin from the mouse Engelbreth-Holm-Swarm tumor, possess full binding activity for nidogen in radioligand assays. Partial reduction (70-80%) of disulfide bonds in P1 did not reduce binding activity and allowed the separation of domain III segments originating from the A, B1 and B2 chains of laminin as demonstrated by sequence analysis. Only the B2 chain segment consisting of seven cysteine-rich repeats with similarity to epidermal growth factor showed substantial nidogen-binding activity. Further degradation of this component to an active 28-kDa fragment was achieved by a second pepsin digestion of partially reduced P1. This indicates that a major binding structure for nidogen is located within three or four cysteine-rich repeats occupying sequence positions 755 to about 920 in the B2 chain. The data also show that fragments P1 and P1X differ by the absence or presence of a large portion, domain IIIb, of the laminin A chain but are indistinguishable in nidogen binding.

Leptin impairs metabolic actions of insulin in isolated rat adipocytes.

Leptin is an adipocyte hormone involved in the regulation of energy homeostasis. Generally accepted biological effects of leptin are inhibition of food intake and stimulation of metabolic rate in ob/ob mice that are defective in the leptin gene. In contrast to these centrally mediated effects of leptin, we are reporting here on leptin effects on isolated rat adipocytes. Leptin impairs several metabolic actions of insulin, i.e. stimulation of glucose transport, glycogen synthase, lipogenesis, inhibition of isoproterenol-induced lipolysis, and protein kinase A activation, as well as stimulation of protein synthesis. Insulin effects were reduced by leptin (2 nM) with a half-life of about 8 h. At low leptin concentrations (<1 nM), the insulin sensitivity was reduced leading to a shift to the right in the dose-response curve. At higher concentrations the responsiveness was diminished, resulting in nearly complete inhibition of insulin effects at >30 nM leptin. The IC50 value of leptin was 3.1 +/- 1 nM after 15 h of preincubation of adipocytes in primary culture. The natural splice variant des-Gln49-leptin exhibited a significantly lower potency. Adipocytes regained full insulin sensitivity within a few hours after leptin removal. The stimulation of glucose transport by vanadate was not affected by leptin. These data show specific and potent impairment of insulin action by leptin in the physiological concentration range of both leptin and insulin, which may be related to the pathophysiology of insulin resistance in both non-insulin-dependent diabetes mellitus and obesity.

Self-assembly of apoferritin from horse spleen after reversible chemical modification with 2,3-dimethylmaleic anhydride.

Apoferritin from horse spleen is composed of 24 subunits that undergo partial dissociation after chemical modification with 2,3-dimethylmaleic anhydride (DMMA), yielding dimeric, trimeric, and tetrameric intermediates, stable at pH 8.5 and 0 degrees C. Deacylation at neutral pH and elevated temperature provides a means to initiate reassembly by appropriate shifts of the solvent conditions. In order to monitor the pathway of self-assembly, starting from different intermediates of dissociation, dimers, trimers, and tetramers were isolated and investigated with respect to their capacity to accomplish reassociation. Intrinsic protein fluorescence, gel permeation chromatography, and analytical ultracentrifugation were applied to characterize the intermediate and final stages of association. The assembly of both the dimer and trimer yields greater than 85% of the native tetracosamer; the overall rate, starting from the dimer, exceeds the one starting from the trimer. Under comparable conditions, the tetramer exhibits only partial reassociation via the dimer and monomer; the corresponding dissociation reaction determines the observed slower rate. Significant assembly intermediates are "structured monomers", dimers, trimers, and dodecamers. Polymerization of the dimer via the tetramer, octamer, etc., does not occur on the pathway of assembly. The results confirm the assembly scheme proposed previously on the basis of cross-linking and spectroscopic experiments [Gerl, M., & Jaenicke, R. (1987) Eur. Biophys. J. 15, 103-109]. Comparison of structural models involving the different subunit interactions responsible for the sequential association supports the monomer----dimer----trimer----hexamer----dodecamer----tetracosamer mechanism of apoferritin self-assembly.

A radioimmunoassay for the N-terminal propeptide of rat procollagen type III. Application to the study of the uptake of the N-terminal propeptide of procollagen type III in isolated perfused rat liver.

Antibodies against a synthetic peptide representing the 14 C-terminal amino acids of the N-terminal propeptide of rat and bovine procollagen type III were raised in rabbits and used to develop a radioimmunoassay. N-Terminal propeptide of procollagen type III, purified from calf skin, served as standard and tracer material. The IC50 of the standard inhibition curve was 2.1 micrograms/l, the lower limit of detection about 0.4 microgram/l, interassay variation was 8.5% and the intraassay variation 6.6% in typical experiments. Three peaks of antigenicity were detected in rat serum after gel chromatography. One peak coeluted with purified N-terminal propeptide of procollagen type III, one peak contained material approximately twice this size, and one peak eluted close to the void volume of the column. The antigen concentration in rat serum decreased in an age dependent manner. Rat, bovine, sheep and minipig serum antigen was sufficiently crossreactive to allow the application of the assay to these species, whereas human, goat and guinea pig samples were not. The degradation product Col 1, causing non-parallel inhibition in commercially available assays for human samples was not recognized since it does not contain the epitope represented by the synthetic peptide. The assay was used to study the half-life of bovine and endogenous N-terminal propeptide of procollagen type III in isolated perfused rat liver. [125I]-labeled antigen was cleared rapidly from the perfusate (t1/2 less than 5 min). The bovine antigen was removed from the perfusate with a half-life of 15 +/- 4 min. Endogenous propeptide was perfused for 120 min with little change in concentration.(ABSTRACT TRUNCATED AT 250 WORDS)