Erythropoietin alters the pharmacokinetics of organic anions mainly eliminated by the kidney in rats.

Erythropoietin (EPO) is a cytokine originally used for its effects on the hematopoietic system, and is widely prescribed around the world. In the present study, the effects of EPO administration on p-aminohippurate (PAH, a prototype organic anion) pharmacokinetics and on the renal expression of PAH transporters were evaluated. Male Wistar rats were treated with EPO or saline (control group). After 42 h, PAH was administered, and plasma samples were obtained at different time points to determine PAH levels. PAH levels in renal tissue and urine were also assessed. The renal expression of PAH transporters was evaluated by Western blotting. EPO-treated rats showed an increase in PAH systemic clearance, in its elimination rate constant, and in urinary PAH levels, while PAH in renal tissue was decreased. Moreover, EPO administration increased the expression of the transporters of the organic anions evaluated. The EPO-induced increase in PAH clearance is accounted for by the increase in its renal secretion mediated by the organic anion transporters. The goal of this study is to add important information to the wide knowledge gap that exists regarding drug-drug interactions. Owing to the global use of EPO, these results are useful in terms of translation into clinical practice.

Drug Clearance from Cerebrospinal Fluid Mediated by Organic Anion Transporters 1 (Slc22a6) and 3 (Slc22a8) at Arachnoid Membrane of Rats.

Although arachnoid mater epithelial cells form the blood-arachnoid barrier (BAB), acting as a blood-CSF interface, it has been generally considered that the BAB is impermeable to water-soluble substances and plays a largely passive role. Here, we aimed to clarify the function of transporters at the BAB in regulating CSF clearance of water-soluble organic anion drugs based on quantitative targeted absolute proteomics (QTAP) and in vivo analyses. Protein expression levels of 61 molecules, including 19 ATP-binding-cassette (ABC) transporters and 32 solute-carrier (SLC) transporters, were measured in plasma membrane fraction of rat leptomeninges using QTAP. Thirty-three proteins were detected; others were under the quantification limits. Expression levels of multidrug resistance protein 1 (Mdr1a/P-gp/Abcb1a) and breast cancer resistance protein (Bcrp/Abcg2) were 16.6 and 3.27 fmol/µg protein (51.9- and 9.82-fold greater than in choroid plexus, respectively). Among those organic anion transporters detected only at leptomeninges, not choroid plexus, organic anion transporter 1 (oat1/Slc22a6) showed the greatest expression (2.73 fmol/µg protein). On the other hand, the protein expression level of oat3 at leptomeninges was 6.65 fmol/µg protein, and the difference from choroid plexus was within two-fold. To investigate oat1's role, we injected para-aminohippuric acid (PAH) with or without oat1 inhibitors into cisterna magna (to minimize the contribution of choroid plexus function) of rats. A bulk flow marker, FITC-inulin, was not taken up from CSF up to 15 min, whereas uptake clearance of PAH was 26.5 µL/min. PAH uptake was completely blocked by 3 mM cephalothin (inhibits both oat1 and oat3), while 17% of PAH uptake was inhibited by 0.2 mM cephalothin (selectively inhibits oat3). These results indicate that oat1 and oat3 at the BAB provide a distinct clearance pathway of organic anion drugs from CSF independently of choroid plexus.

Influence of surface charge of gold nanorods on skin penetration.

BACKGROUND/PURPOSE: The skin plays an important role as a protective barrier against toxic environments and also is a route of drug administration. In spite of evidence for and interest in the skin penetration of nanoparticles, no study has examined the effect of nanoparticle surface charge on percutaneous absorption. In this study, we investigated the effect of surface charges of gold nanorods (GNs) on skin penetration. METHODS: Using transmission electron microscopy (TEM) and image analysis, we quantitatively measured the ability of GNs to penetrate the skin. RESULTS: Our results showed that the area density of the electron-dense dots of GNs, which penetrated into the stratum corneum, significantly increased for negatively charged GNs compared to those with a positive charge (P < 0.01). To investigate the percutanoues absorption of charged GNs, in vitro skin permeation studies were carried out using a Franz-type diffusion cell (FDC). The penetration of GNs through the skin was quantified by inductively coupled plasma mass spectrometry. Consistent with TEM observations, our penetration study using an FDC also revealed that negative particles were frequently detected in samples of receptor fluid at 48 h after exposure (P < 0.01). CONCLUSION: Together our results showed that anionic GNs penetrate skin better than cationic GNs.

Gap junction permeability: selectivity for anionic and cationic probes.

Gap junction channels formed by different connexins exhibit specific permeability to a variety of larger solutes including second messengers, polypeptides, and small interfering RNAs. Here, we report the permeability of homotypic connexin26 (Cx26), Cx40, Cx43, and Cx45 gap junction channels stably expressed in HeLa cells to solutes with different size and net charge. Channel permeability was determined using simultaneous measurements of junctional conductance and the cell-cell flux of a fluorescent probe. All four connexins allowed passage of both cationic and anionic probes, but the transfer rates were connexin dependent. The negatively charged probes [Lucifer yellow (LY; median axial diameter 9.9 Å, charge -2), carboxyfluorescein (CF; 8.2 Å; -2), and Alexa Fluor350 (AF350, 5.4 Å; -1)] exhibited the following permeability order: Cx43 > Cx45 > Cx26 > Cx40. In contrast, for the positively charged species permeability, the orders were as follows: Cx26 ≈ Cx43 ≈ Cx40 ≈ Cx45 for N,N,N-trimethyl-2-[methyl-(7-nitro-2,1,3-benzoxadiol-4-yl) amino] ethanaminium (NBD-m-TMA; 5.5 Å, +1) and Cx26 ≥ Cx43 ≈ Cx40 > Cx45 for ethidium bromide (10.3 Å, +1). Comparison of probe permeability relative to K(+) revealed that Cx43 and Cx45 exhibited similar permeability for NBD-m-TMA and AF350, indicating weak charge selectivity. However, lesser transfer of CF and LY through Cx45 relative to Cx43 channels suggests stronger size-dependent discrimination of solute. The permeability of NBD-m-TMA for Cx40 and Cx26 channels was approximately three times higher than to anionic AF350 despite the fact that both have similar minor diameters, suggesting charge selectivity. In conclusion, these results confirm that channels formed from individual connexins can discriminate for solutes based on size and charge, suggesting that channel selectivity may be a key factor in cell signaling.

The impact of hepatic uptake on the pharmacokinetics of organic anions.

The disposition of seven marketed and two AstraZeneca acid (organic anion) compounds with a range of volume of distribution at steady state (V(ss)) and clearance have been profiled in rat and dog. Pharmacokinetic (PK) parameters along with liver and muscle tissue levels were collected, and their contributions to total V(ss) were calculated. The physiologically based prediction of V(ss) correlated (all predictions within 2-fold) with the V(ss) obtained from plasma PK analysis. The V(ss) of the acid drugs with atypically high values could be explained by significant sequestering of compound to the liver. A "media loss" in the in vitro hepatocyte assay that monitors loss of compound from the incubation media along with physiologically based PK (PBPK) modeling was assessed for its ability to accurately predict the impact of hepatic uptake on both clearance and V(ss). This methodology significantly improved the prediction of metabolic in vivo clearance compared with standard hepatocyte scaling approaches that do not take into account hepatic uptake. Predictions of V(ss) from the media loss assay also correlate with the measured values from plasma PK analysis. However, hepatic uptake will have little overall impact on half-life, because of the concomitant impact on both Cl and V(ss), as long as hepatic extraction is not high. The methodology described here is particularly useful when there is no allometric relationship between species as a result of interspecies differences in liver uptake. In this situation, the potential use of human hepatocytes combined with PBPK modeling avoids the question of which species pharmacokinetics is most predictive to humans.

Sex-specific extraction of organic anions by the rat liver.

The capacity for hepatic elimination of some compounds is different in males and females and differential expression of a number of sinusoidal and canalicular transporters exists. However, the specific events underlying the functional differences are not understood. To determine how sex influences sinusoidal and canalicular organic anion transport, bile duct-cannulated livers from mature Sprague-Dawley rats of both sexes were single-pass perfused with saline containing the model organic anions bromosulphophthalein (BSP), carboxyfluorescein (CF), carboxyfluorescein diacetate (CFDA) or 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS). Assay of effluent perfusate anion concentration showed that BSP, but not DIDS, extraction was significantly higher in male versus female rats. At 20 min perfusion with 50 microM BSP the mean effluent concentration was 5.6 and 20.1 microM in, respectively, male and female rats. HPLC confirmed that the effluent perfusate concentration of BSP was higher in female as compared with male rats and was not contributed to by its glutathione conjugate. With 25 microM DIDS, the effluent concentration reached 7.3 (male) and 8.2 microM (female), indicating high extraction efficiency. In contrast to BSP and DIDS, CF extraction was very low (<20%) so that differences between male and females could not be assessed. Biliary BSP and CF excretion were, respectively, 3.5- and 4-fold higher in male rats. Neither sinusoidal efflux nor biliary excretion of CF was sex-dependent with a higher cytoplasmic load of CF (during CFDA perfusion). Our results suggest that differences in sinusoidal uptake are responsible for the sex-specific hepatic excretion of some organic anions.

[The role of ABC transporters in the renal elimination of organic anions and drugs]. / Rola transporterów ABC w nerkowym wydalaniu organicznych anionów i leków.

The ATP-binding cassette (ABC) transporters are a large superfamily of integral membrane proteins that actively transport organic ions across biological membranes. These transporters play important role in the reabsorptive and excretory capacity of the kidney. Nine structurally and functionally related members (MRP1-7, P-gp, BCRP) have been identified, which differ from each other by their localization, expression levels, and substrate specificity. This review discusses current knowledge on the renal characteristics of ABC transporters, with specific focus on their regulation.

Structural properties governing drug-plasma protein binding determined by high-performance liquid chromatography method.

The high-performance liquid chromatography (HPLC) method employing stationary phases immobilized with plasma proteins was used for this study to investigate the structural properties governing drug-plasma protein binding. A set of 65 compounds with a broad range of structural diversity (in terms of volume, hydrogen-bonding, polarity and electrostatic force) were selected for this purpose. The Abraham linear free energy relationship (LFER) analyses of the retention factors on the immobilized HSA (human serum albumin) and AGP (α1-acid glycoprotein) stationary phases showed that McGowan's characteristic molecular volume (V), dipolarity/polarizability (S) and hydrogen bond basicity (B) are the three significant molecular descriptors of solutes determining the interaction with immobilized plasma proteins, whereas excess molar refraction (E) is less important and hydrogen bond acidity (A) is not of statistical significance in both systems, for electrically neutral compounds. It was shown that ionised acids, as carboxylate anions, bind very strongly to the immobilized HSA stationary phase and that ionised bases, as cations bind strongly to the AGP stationary phase. This is the first time that the effect of ionised species on plasma protein binding has been determined quantitatively; the increased binding of acids to HSA is due almost entirely to acids in their ionised form.

Track analysis of the passage of rhodamine-labeled liposomes across porcine jejunal mucus in a microchannel device.

AIM: To investigate how surface charge and hydrophilicity affect the mucopermeation of liposomes across intestinal mucus. METHODOLOGY: Rhodamine-labeled liposomes (∼120-130 nm) with different surface charges were investigated for their capacity to flux across fresh porcine jejunal mucus in a microchannel device. Fluorescent microscopy and tracking analysis were used to measure liposome movement, while fluorescence lifetime imaging microscopy was utilized to determine mucus pH. RESULTS: Mucopermeation was dependent on hydrophilicity and surface charge - anionic liposomes permeated more than cationic. The most cationic liposomal prototype agglomerated mucus. Presence of Na+, K+ and Mg2+ increased both speed and straightness of the pathways for all prototypes. Cationic but not anionic liposomes caused acidification (pH 2.5). CONCLUSION: Acidification caused by cationic liposomes explains their ability to interfere with mucus stability. Surface charge of liposomes strongly influences mucopermeation capability.

Synthesis of a quaternary amine anion exchange resin and study [of] its adsorption behaviour for chromate oxyanions.

Glycidyl methacrylate/N,N'-methylene bis-acrylamide (GMA/MBA) was prepared and allowed to react with tetraethylenepentamine (TEP) to give glycidyl methacrylate amine resin (RPA) followed by treatment with glycidyl trimethylammonium chloride (GTA) to give glycidyl methacrylate resin bearing quaternary ammonium chloride moieties (RQA). Zeta potential measurements showed that RQA particles are positively charged over pH 2-10 indicating the strong basic nature of the quaternary amine sites. The effect of pH on the recovery of chromate by RPA and RQA was examined. The results indicated that RQA is an efficient sorbent for chromate from both acidic and basic media. The repeated use of RQA was tested through stripping the adsorbed chromate using a mixture of 0.05 NaOH and 2 M NaCl in the case of the uptake from acidic media and using 2 M NaCl solution in the case of alkaline solutions.

Anion transport through the contraluminal cell membrane of renal proximal tubule. The influence of hydrophobicity and molecular charge distribution on the inhibitory activity of organic anions.

Three different mechanisms of anion transport have been identified for the contraluminal membrane in the proximal tubule of the rat kidney. These mechanisms are specific for the transport of sulfate, dicarboxylate and p-aminohippurate anions. Sulfate transport is inhibited by bivalent organic anions with a distance between the charges of less than 7 A. The sulfate system acts in two modes: in a planar mode for anions with flat charged residues such as COO- and a charge separation of 3-4 A or in a bulky mode for groups such as SO3H- and a charge separation of 4-7 A. Monovalent anions can be accepted if there is a hydrophobic core next to the negative charges. Dicarboxylate transport is inhibited exclusively by anions with two charge centers located within 5 to 9 A, one of those possibly being a partial charge of -0.5 elementary charges. p-Aminohippurate transport is inhibited by monovalent anions, if these have a hydrophobic domain with a minimal length of about 4 A. Bivalent anions inhibit, if they have a charge distance of 6-10 A; both charges can be partial charges of about -0.5 elementary charges. Longer bivalent anions can be effective provided they have a sufficiently large hydrophobic domain. For the sulfate and p-aminohippurate systems it is found that anions with high acidity yield good inhibition. The overlapping specificities of the three systems with respect to charge distance and hydrophobicity allow them to accept a large variety of organic anions.

Anion permeability of the olfactory receptive membrane.

The ionic mechanism of the electropositive olfactory receptor potential was studied in the bullfrog and the swamp frog. The positive receptor potential strikingly decreased in amplitude in chloride-free solution. When the olfactory epithelium was immersed in high-KCl-Ringer's solution and then in Cl-free, high-K solution, the polarity of the positive potential could be reversed. This is supposed to be due to the exit of the increased internal chloride ion. From the above two experiments it is concluded that the positive olfactory receptor potential depends primarily upon the influx of the chloride ion through the olfactory receptive membrane. Some contribution by potassium and possibly other ions may occur. The ability of other anions to substitute for chloride was examined. It was found that only Br-, F-, and HCO2- could penetrate the olfactory receptive membrane. The sieve hypothesis in the inhibitory post-synaptic membrane (Coombs, Eccles, and Fatt, 1955) is not applicable to the olfactory receptive membrane on the basis of the size of hydrated ions, but it may be applicable on the basis of the sizes of naked ions.

Energetics of coupled active transport of sodium and chloride.

A Clark electrode was used to measure oxygen consumption by the gall bladder, in which there is a direct and one-to-one linkage between active Na and active Cl transport. O2 uptake was reversibly depressed when Cl in the mucosal bathing solution was replaced by a poorly transported anion, such as sulfate. This effect of Cl was abolished by ouabain or in Na-free solutions. When the anion was chloride, treatment with ouabain or replacement of Na by a poorly transported cation depressed QO2 more than did replacement of Cl. However, ouabain or removal of Na also depressed QO2 in Na2SO4 solutions, in which salt transport is minimal. It is concluded that oxygen uptake in the gall bladder consists of three fractions: 9% requires both Na and Cl, is inhibited by ouabain, and is linked to the NaCl pump; 36% requires Na but not Cl, is inhibited by ouabain, and possibly is linked to the cellular K uptake mechanism; and 55% represents basal uptake. If the extra oxygen uptake observed during transport supplies all the energy for transport, then 25 Na + 25 Cl ions are transported actively per O2 consumed; i.e., twice as many ions as in epithelia which transport only Na actively. This extra uptake is more than sufficient to supply the energy for overcoming internal membrane resistance under the experimental conditions used.

Anion permeation in an apical membrane chloride channel of a secretory epithelial cell.

Single channel currents though apical membrane Cl channels of the secretory epithelial cell line T84 were measured to determine the anionic selectivity and concentration dependence of permeation. The current-voltage relation was rectified with single channel conductance increasing at positive potentials. At 0 mV the single channel conductance was 41 +/- 2 pS. Permeability, determined from reversal potentials, was optimal for anions with diameters between 0.4 and 0.5 nm. Anions of larger diameter had low permeability, consistent with a minimum pore diameter of 0.55 nm. Permeability for anions of similar size was largest for those ions with a more symmetrical charge distribution. Both HCO3 and H2PO4 had lower permeability than the similar-sized symmetrical anions, NO3 and ClO4. The permeability sequence was SCN greater than I approximately NO3 approximately ClO4 greater than Br greater than Cl greater than PF6 greater than HCO3 approximately F much greater than H2PO4. Highly permeant anions had lower relative single channel conductance, consistent with longer times of residence in the channel for these ions. The conductance sequence for anion efflux was NO3 greater than SCN approximately ClO4 greater than Cl approximately I approximately Br greater than PF6 greater than F approximately HCO3 much greater than H2PO4. At high internal concentrations, anions with low permeability and conductance reduced Cl influx consistent with block of the pore. The dependence of current on Cl concentration indicated that Cl can also occupy the channel long enough to limit current flow. Interaction of Cl and SCN within the conduction pathway is supported by the presence of a minimum in the conductance vs. mole fraction relation. These results indicate that this 40-pS Cl channel behaves as a multi-ion pathway in which other permeant anions could alter Cl flow across the apical membrane.

Effect of enzyme digestion on anionic sites and charge-selective permeability of dermo-epidermal junction.

To study components of anionic sites on the lamina densa of the dermo-epidermal junction (DEJ) and to assess the effect of removal of sialic acid or glycosaminoglycans on its charge-selective permeability, epidermal sheets, whose dermis had been removed by treatment with dithiothreitol, were digested with heparitinase, chondroitinase ABC, hyaluronidase, or neuraminidase. They were then stained with polyethyleneimine for demonstration of the anionic sites or incubated in a medium containing native anionic ferritin for tracer experiments. The anionic sites were completely removed after heparitinase digestion. Although the numerical density of the sites was not altered, their electron density was decreased after chondroitinase ABC digestion. The other enzymes had no effect on the sites. In the tracer experiments, heparitinase or neuraminidase increased the number of tracer molecules penetrating into the lamina lucida of the epidermal sheet, while the other enzymes had no effect on it. These data indicate that heparan sulfate, which is a main component of the anionic sites, plays an important role in the charge-selective permeability of the DEJ, whereas chondroitin sulfate, which seems to be contained in the sites, does not, probably because of its small amount. These data also indicate that sialic acid, which is not a main component of the anionic sites demonstrated with the cationic probe, has a role in the permeability function.

Effects of two inhibitors of anion transport on bone resorption in organ culture.

The present investigation was undertaken to examine the effects of 4-acetamido-4'-isothiocyanostilbene-2,2-disulfonic acid (SITS) and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), known amino-reactive and selective inhibitors of anion exchange across the plasma membrane, on bone resorption in organ cultures of fetal rat long bones. Cultures were treated with SITS and DIDS under control unstimulated conditions or with PTH. Both SITS and DIDS were found to be potent inhibitors of 45Ca release from previously labeled fetal rat long bones. Both control resorption and the response to PTH were inhibited in a dose-related fashion. SITS and DIDS also inhibited the incorporation of [3H]thymidine in bone. The effects on resorption and [3H]thymidine incorporation were reversible when the drugs were withdrawn. These findings indicate that SITS and DIDS are potent inhibitors of bone resorption which may act by blocking the anion exchange, Cl-/HCO3-.

Microperfusion studies on the permeability of retinal vessels. A new model demonstrating organic anion transport and a reabsorptive fluid flux.

We developed an experimental model to study the permeability of individual retinal vessels in vitro using microperfusion techniques adapted from kidney tubule studies. The retinal vessels were isolated by freehand dissection and mounted on a microperfusion apparatus. When inulin was perfused luminally, it was diluted to 80.2 +/- 2.3% of its initial concentration. However, no radioactive leak into the bath side was observed, suggesting that the dilution was due to fluid flux from bath to lumen. The dilution of fluorescein (81.9 +/- 3.8%) was in the same range as that of inulin, the reference marker. The extremely low lumen-to-bath fluorescein flux, 0.5 +/- 0.9 X 10(-12) mol/min/mm, increased by 68% when probenecid was added to the perfusate and by 210% when probenecid was placed in the bath. The effect was concentration-dependent. When placed in the bath, fluorescein moved rapidly across the retinal vessel walls, accumulating in the lumen to concentrations 40 times higher than in the bath. This movement from bath to lumen, which was much higher (13.6 +/- 0.3 X 10(-12) mol/min/mm) than the lumen-to-bath fluorescein flux for the same fluorescein concentration, decreased by adding probenecid to the bath. The kinetics of this unidirectional movement of fluorescein were consistent with a saturable active transport process. The fluid flux from bath to lumen across the retinal vessels, which was 6.3 +/- 1.0 nl/min/mm for perfusion rates of 6.6 +/- 0.2 nl/min, was temperature-dependent and was coupled to the fluorescein transport. Fluorescein stimulated the fluid flux by 17% when added to the perfusate and by 60% when added to the bath, and this effect could be reversed by probenecid. Our results showed an active transport of fluorescein in the rabbit retinal vessels coupled with net fluid flux from outside the vessels into the lumen.

Effects of sulphonylureas on the volume-sensitive anion channel in rat pancreatic beta-cells.

1. The volume-sensitive anion conductance in rat pancreatic beta-cells was studied directly using the conventional whole-cell and perforated patch recording techniques, and indirectly by measuring 3H-taurine efflux from pre-loaded, perifused islets. 2. Using the conventional whole-cell recording configuration, activation of the outwardly-rectifying, DIDS-sensitive conductance was induced by glibenclamide (10 microM) but not by tolbutamide (100 microM) nor by meglitinide (20 microM). A high concentration of glibenclamide (100 microM) caused a voltage-and time-dependent inhibition of the conductance. Tolbutamide had a modest inhibitory effect on swelling-induced inward currents. 3. In perforated patch recordings, glibenclamide, tolbutamide and meglitinide were all without effect on the conductance, although activation could be induced under these conditions by exposure to a hypotonic bath solution. 4. The rate of efflux of 3H-taurine, a marker for activity of the volume-sensitive anion channel, from preloaded, perifused islets was markedly stimulated by exposure to a hypotonic solution. However, glibenclamide and tolbutamide were both without effect. 5. Electrical activity of beta-cells in response to glibenclamide or tolbutamide was not inhibited by 4,4'-dithiocyanatostilbene-2,2'-disulphonic acid (DIDS), an inhibitor of the volume-sensitive anion channel. 6. It is concluded that activity of the volume-sensitive anion conductance in rat pancreatic beta-cells is not modulated by the sulphonylurea receptor. The activation of the conductance by glibenclamide in whole-cell recordings could be the result of a non-specific interaction of the drug with plasma membrane lipids.

Pharmacological characterization of a 7-benzylidenenaltrexone-preferring opioid receptor in porcine ileal submucosa.

In the intestine, opioids produce antidiarrhoeal and constipating actions that are mediated by enteric neurones. Through interactions with opioid receptors (ORs) on submucosal neurones, opioids suppress active ion transport evoked by transmural electrical stimulation (TES) in mucosa-submucosa sheets from the porcine ileum. In this study, we examined the pharmacological characteristics of the previously described OR, which is sensitive to the delta1-OR antagonist 7-benzylidenenaltrexone and modulates neurogenic transepithelial ion transport in this tissue preparation. Increases in short-circuit current (Isc, a measure of active anion transport) evoked by TES in ileal mucosa-submucosa sheets were inhibited by opioid agonists possessing high selectivity for either delta- or micro-ORs including [d-Pen2,5]enkephalin (DPDPE), [d-Ala2, Glu4]deltorphin II, and [d-Ala2, N-Me-Phe4, Gly5-ol]enkephalin (DAMGO). As determined by the Schild analysis, the actions of these agonists were competitively inhibited by 7-benzylidenenaltrexone. The nonequilibrium micro-OR antagonist beta-funaltrexamine inhibited the actions of DAMGO only at a high concentration (1 microm) but did not alter DPDPE or deltorphin II action. At concentrations up to 10 microm, the nonequilibrium delta-OR antagonist naltrindole 5'-isothiocyanate did not alter the actions of delta- or micro-OR agonists. Radioligand binding analyses of neuronal homogenates from the ileal submucosa revealed that the nonselective OR ligand [3H]diprenorphine bound to two populations of specific binding sites. One of these sites possessed binding characteristics similar to the delta-OR. In summary, neurogenic ion transport in the porcine intestine is modulated by an OR which shares pharmacological characteristics of both micro- and delta-ORs and may represent a novel receptor entity.

Effect of nutritional status on organic anion clearance by the swine liver.

Hepatic dysfunction follows a wide range of insults. Impaired excretion of organic dyes such as bilirubin often occurs before other obvious clinical defects in metabolic processes. Indocyanine green (ICG) is excreted through pathways similar to those of bilirubin. To determine the effectiveness of ICG as a marker of hepatic dysfunction related to clinical malnutrition, pigs received 5 mg/kg ICG with simultaneous sampling from the hepatic vein, pulmonary artery, and aorta over 3 hours. Group I remained well nourished, group II was fasted to a weight loss equal to 20% of initial body weight, and group III was fasted to a 20% weight loss and then refed until the animals regained their initial weight. Both systemic and intrinsic hepatic clearance were depressed significantly with fasting but returned above baseline after refeeding. No significant difference appeared between systemic and intrinsic hepatic clearance. Extraction ratios were low in all groups. In outbred swine, ICG clearance reflects the function of hepatic organic anion excretion in vivo, and venous sampling reflects intrinsic hepatic clearance. The impairment of the carrier-mediated transport system is reversible with refeeding.