Effect of prostaglandin E1 on low density lipoprotein apo-B-receptor binding in human, rat and swine liver in vitro.

The effect of PGE1 on low density lipoprotein (LDL) apo-B-receptor binding was examined in human, rat and swine liver. Autologous LDL (for humans and swines) and homologous LDL (for rats) were isolated by ultracentrifugation and labelled with 123I using Iodogen followed by purification with dialysis. LDL-concentrations of 0.1-6 micrograms protein/ml were used for direct binding assays investigating the specific binding of labelled LDL in presence of increasing PGE1-concentrations (100 pM to 100 microM). In separate experiments the effect of PGE1 on displacement of specifically bound 123I-LDL by unlabelled ones was studied. The binding capacities estimated by Scatchard analysis were similar for human and rat liver LDL-apo-B-receptor binding, however, swine liver exhibited a significantly (p less than 0.001) lower binding capacity for 123I-LDL. PGE1 significantly (p less than 0.01-0.001) increased the amount of 123I-LDL specifically bound to the liver apo-B-receptors and the binding affinity in all liver preparations of the 3 species in a dose-dependent manner. PGE1 also significantly increased competition of unlabelled LDL for 123I-LDL bound to its specific apo-B-receptors in a dose-dependent manner (p less than 0.01-0.001) with an ED50 of 123 +/- 64 nM for human liver, 901 +/- 102 nM for rat liver obtained during anaesthesia, 74 +/- 23 nM for rat liver obtained after decapitation and 941 +/- 121 nM for swine liver. In human liver iloprost (ED50 = 876 +/- 53 nM) and PGI2 (ED50 = 52 +/- 12 microM) were less effective than PGE1, PGE2 had no effect on LDL-induced competition. It is concluded that PGE1 renders LDL more sensitive for apo-B-receptor binding suggesting a potential hypolipidemic action of PGE1.     

Decrease of the prostaglandin I2 binding capacity in thyroids from patients with Graves' disease

Properties of prostaglandin (PG) I2-binding sites in human thyroids from euthyroid and thyrotoxic patients were investigated. The specific binding of 3H-iloprost (ZK 36374, a chemically stable PGI2-analogue) to normal thyroids approached saturation at a concentration of more than 150 nmoles/L and could be displaced most effectively by unlabeled iloprost (concentration causing the half maximal inhibition, IC-50 = 5.9 +/- 2.9 mumoles/L) and PGI2 (IC-50 = 9.8 +/- 3.1 mumoles/L) and less effectively by unlabeled PGF2 alpha (IC-50 = 847.9 +/- 123.8 mumoles/L) at 4 degrees C. The Scatchard analysis clearly indicated heterogeneity revealing the presence of 0.65 +/- 0.18 pmoles/mg protein at the high-affinity binding sites (equilibrium dissociation constant, Kd = 18.2 +/- 9.1 nmoles/L) and 5.35 +/- 1.6 pmoles/mg protein at the low-affinity binding sites (Kd = 151.3 +/- 43.1 nmoles/L). In contrast, in diffuse colloid struma derived from patients with Graves' disease a single class of binding sites with an apparent binding capacity of 0.18 +/- 0.05 pmoles/mg protein (Kd = 70.4 +/- 27.3 nmoles/L) was indicated. However, in diffuse colloid struma derived from euthyroid patients no difference in the number of binding sites and binding affinity of 3H-iloprost was noted compared to normal thyroids. The data provide evidence for the presence of specific PGI2-binding sites in the human thyroid gland and demonstrate a significant decrease of the receptor density in patients with Graves' disease. It is suggested, that PGI2 has an important role in the regulation of human thyroid function.     

Release and specific binding of prostaglandins in bovine pineal gland

Incubated bovine pineal glands released prostaglandin E-and prostglandin F-like material (304 +/- 20 and 582 +/- 56 pg/mg dry tissue wt/h, respectively) and the release was increased 2.2 2.9-fold by adding 10(-4)-10(-6)M of norepinephrine to the medium. Binding assays revealed the existence of high affinity binding of 3H-prostaglandin E2 (3H-PGE2) and 3H-prostaglandin F1 alpha (3H-PGF2 alpha) in low speed supernatants of pineal homogenates. Binding was increased by increasing Ca++ concentration in medium up to 2 mM, was heat labile and was depressed following incubation with trypsin. In subcellular fractionation studies maximal 3H-PG binding was found in the 27000 x g pellet. Scatchard analysis of 3H-PGE2 binding revealed the presence of a single population of binding sites with a Kd= 1.2 nM and a binding site concentration of 1-2 pmoles/g protein. A single population of binding sites for 3H-PGF2 alpha was also detected with a Kd= 1.7 nM and a similar binding site concentration. Non-radioactive PGE1 and PGE2 were almost equally effective to compete for 3H-PGE1 binding sites (ED50= 5 and 2 nM, respectively). Unlabeled PGF1 was relatively ineffective to compete for 3H-PGE2 binding (ED50 greater than 1000 nM) but displaced effectively 3H-PGF2 alpha binding (ED20=1.2 nM).     

Modulation of 3H-prostaglandin E2 binding sites in the rabbit gastric mucosa

The binding of 3H-prostaglandin E2 (PGE2) to rabbit gastric mucosa was investigated. Binding depended on incubation time, temperature and pH, and was saturable and reversible. Scatchard plot analysis revealed a single class of binding sites with a dissociation constant (Kd) of 5.33 +/- 0.21 nM and a maximum number of binding sites (Bmax) of 138.1 +/- 3.4 fmol/mg protein. PGE1 and 16,16-dimethyl PGE2 potently competed with 3H-PGE2 for the binding sites of gastric mucosa, whereas PGA2, PGF2 alpha, 6-keto PGF1 alpha and thromboxane B2 were less potent. The gastric mucosa prepared from the rabbits given indomethacin (5 mg/kg s.c. three times) showed a lower Kd (2.47 +/- 0.19 nM) for 3H-PGE2 than that from untreated one. Treatment with a PGE1 analog, misoprostol (320 micrograms/kg s.c. three times) lowered the Bmax to 74.1 +/- 2.4 fmol/mg protein without any significant effect on the Kd value. It is concluded that rabbit gastric mucosa has specific binding sites for 3H-PGE2 which may be modulated by the levels of PGs in vivo.     

Prostaglandin E2 binding site distribution and subtype classification in the rabbit iris-ciliary body.

The distribution and characteristics of specific binding sites for tritium labeled prostaglandin E2 (3H-PGE2) were examined in membrane preparations from rabbit iris-sphincter, iris and ciliary body. The majority of 3H-PGE2 specific binding sites were found in the ciliary body (46%) followed by the iris (37%) and the iris-sphincter muscle (5%). Scatchard analysis of saturable 3H-PGE2 binding sites in the ciliary body indicated a single binding site with a Kd of 2.81 nM and Bmax value of 84 fmoles bound/mg protein. Competition by agonists selective for the EP1, EP2 and EP3 receptor subtypes of the EP (PGE2) prostanoid receptor indicated that the majority of rabbit ciliary body 3H-PGE2 binding sites are of the EP2 subtype. Incomplete displacement of labeled 3H-PGE2 from its binding sites by the EP2 selective agonist 11-deoxy PGE1 suggests the presence of additional EP or non-EP binding sites. There was essentially no binding to EP1 receptor sites as defined by the displacement of 3H-PGE2 by 17-phenyl-trinor PGE2. A weak displacement of 3H-PGE2 by the EP3/EP1 specific agonist, sulprostone, may account for the presence of a small number of EP3 specific binding sites in this tissue. The predominant distribution of PGE2 binding sites in the ciliary body and their identification as EP2 selective, supports recent functional studies where topical application of prostanoids with EP2 but not EP1 or EP3 agonist activity resulted in breakdown of the blood-aqueous barrier.     

Down-regulation of prostaglandin F2 alpha receptors in rat liver during chronic endotoxemia.

Prostaglandin (PG) F2 alpha binding parameters were measured in purified plasma membrane preparations isolated from livers of chronically endotoxin-(ET) treated rats and corresponding controls. Two classes of binding sites were detected in both groups: high affinity, low capacity, with a KD of 44.4 +/- 8.8 nM for saline- and 28.6 +/- 11.3 nM for ET-treated rats (n = 5 for both, p greater than 0.05) and low affinity, high capacity with a KD of 1.12 +/- 0.49 microM for saline- and 1.24 +/- 0.43 microM for ET-treated rats (p greater than 0.05). Bmax values for high affinity sites were 1.01 +/- 0.18 fmol.mg-1 protein for saline- and 1.02 +/- 0.54 (same units) for ET-treated rats (p greater than 0.05). There was a significant difference (p less than 0.01) between the Bmax values for low affinity sites in saline- (675 +/- 332 fmol.mg-1 protein) and ET-treated rats (12 +/- 1, same units). This decrease in the amount of PGF2 alpha low affinity high capacity binding sites may underlie the depression of the PGF2 alpha stimulatory effect on hepatic gluconeogenesis induced by non-lethal, chronic ET treatment of rats, recently described by us (9).     

Specific 3H-PGE1 binding sites in rat ovary in estrous cycle and pregnancy.

Stimulation of cAMP synthesis by prostaglandins E series in the rat ovary is consistent with the presence of a prostaglandin receptor in this tissue. Prostaglandin binding sites with specificity for PGE1 in vitro incubation systems have been demonstrated in rat ovary slices and corpora lutea. The binding of 3H-PGE1 was progressively inhibited with increasing amounts of unlabelled PGE1 and PGE2. PGF2alpha inhibitory effect was markedly smaller than that of PGE. 3H-PGE1 binding to the ovary was higher in 3-day-old rats than in 5-day-old and adult animals, when the highest binding was present in estrus. The specific binding of 3H-PGE1 to rat corpora lutea (CL) decreased on days 11 and 13 of pregnancy and then gradually returned to the level found on day 1 during the second half of gestation. This binding of labelled prostaglandin during pregnancy has been studied in relation to the PGE1 stimulation of cAMP synthesis in rat corpora lutea, but no consistent changes were observed in responsiveness.     

Identification and characterization of prostaglandin-binding components in rat ileum

A specific prostaglandin E2 (PGE2) binding component was identified in the membrane fractions of rat ileum. Two ligands, 3H-PGE2 and 14C-PGE2, competed for the binding sites in crude homogenates of ileum during competition experiments, demonstrating the presence of a limited number of binding sites. There was enhancement in the specific 3H-PGE2 binding to particulate fractions over control when endogenous prostaglandin synthesis was blocked by the administration of indomethacin to rats prior to sacrifice. The specific prostaglandin-binding protein was purified by a combination of [NH4]2SO4 fractionation and Sephacryl S-200 column chromatographic techniques, and shown to be active after these biochemical steps.     

Study of oxytocin receptor: II. oxytocin and prostaglandin F2 alpha receptors in human myometria and amnion-decidua complex during pregnancy and labor

Oxytocin receptors (OXT-R) and prostaglandin F2 alpha receptors (PGF2 alpha-R) in human myometrium, amnion and decidua during pregnancy and at parturition were examined in an effort to clarify their role in the initiation and maintenance of uterine contractions. The number of binding sites for OXT in myometria showed an increase as gestation advance (Ist trimester v.s. at term; 205 +/- 90 v.s. 671 +/- 98 fmol/mg protein, N = 5, p less than 0.01), and a rapid decrease following the onset of labor (254 +/- 60 fmol/mg protein, N = 5, p less than 0.02). On the other hand the number of PGF2 alpha-R, remained unchanged throughout pregnancy and in labor. This myometrial PGF2 alpha binding capacity was approximately 1/20 to 1/30 that of the OXT binding, while binding affinity was almost equal. The OXT-R both in amnion and decidua, which was 1/6 to 1/7 that in myometrium, showed no significant changes throughout pregnancy or after the onset of labor. Binding affinity for each tissue was almost the same and appeared to increase towards term but no statistical significance was detected. Present data confirmed the presence of OXT as well as PGF2 alpha receptors in the three functionally distinct entities of pregnant human uterus; myometrium, amnion, and decidua. Among the components, the OXT binding increased only in the myometrium during pregnancy, suggesting this tissue specifically responds to OXT. In contrast, there was a constant binding in myometria for PGF2 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)     

The ontogeny of the rabbit hepatic glucose transporter

We delineated and characterized the fetal hepatic glucose transporter in the rabbit. Employing the 2-deoxy-D-glucose displaceable 3H-cytochalasin B binding assay we estimated the number and Kd of the GT per mg of liver protein. A gradual increase in the number was observed during development, the fetus (23.8 +/- 2.04 pmoles/mg) expressing a lesser amount when compared to the neonate (59.5 +/- 17 pmoles/mg; p less than 0.05) and adult (142 +/- 11 pmoles/mg; p less than 0.05). On the other hand the affinity of the glucose transporter was higher in the fetus (Kd 287 +/- 81 nM) when compared to either the neonate (988 +/- 222 nM, p less than 0.05) or the adult (706 +/- 101 nM, p less than 0.05). We conclude that the fetal hepatic GT is more efficient secondary to a higher affinity for glucose.     

Regulation of prostaglandin E2 receptors in vivo by dietary fatty acids in peritoneal macrophages from rats

Groups of rats were pretreated with 4-week diets containing 12.5% corn oil or linseed oil. At the end of this period peritoneal macrophages were elicited and isolated. These cells were used for binding experiments with 3H-PGE2 and for estimation of prostaglandin-stimulated cAMP production. Specific binding of 3H-PGE2 was saturable, reversible, protein-dependent, and correlated with stimulation of cAMP production, indicating that specific binding referred to receptor binding. PGE1 and PGI2 were far less effective than PGE2 in competition of binding with 3H-PGE2, indicating receptor selectivity for PGE2. Scatchard analysis of the specific binding data revealed a high affinity component (Kd 17 nM) and low affinity component. The total number of high- and low-affinity binding sites, respective Kd values, and PG stimulation of cAMP production of cells from rats fed the linseed oil diet were comparable to controls. The corn oil diet, however, resulted in a twofold increase in total number of high- and low-affinity binding sites, while respective Kd values were unchanged. This enhancement of binding capacity could be explained by an increased density of binding sites on the cells, and may itself be responsible for the increased sensitivity of the macrophages in this diet group for PG-stimulated cAMP production. The data suggest a regulatory mechanism at the receptor level and are discussed in terms of possible altered bioavailability of arachidonic acid-derived PGE2.     

Effect of drug treatment on liver-slice function following 72-hour hypothermic perfusion

The viability of hypothermically perfused dog liver was evaluated with a tissue-slice technique. After being preserved for 72 hr, slices of liver were incubated at 30 degrees C for as long as 2 hr; then water content, K+/Na+ ratio, and ATP concentration were measured. Dog livers were assigned to the following experimental groups: Group 1 (no preservation; control); Group 2 (livers preserved for 72 hr); Group 3 (donor animals pretreated with 3.5 mg/kg of chlorpromazine (CPZ) and 20 mg/kg of methylprednisolone (MP), and livers preserved for 72 hr); Group 4 (livers pretreated with 2-deoxycoformycin (2-DOC), 50 mg/liter, and preserved for 72 hr); and Group 5 (combination of Group 3 and Group 4 treatments). Livers in Groups 2, 3, and 4 lost K+ during preservation, and the mean K+/Na+ ratio significantly decreased from a control value of 4.2 +/- 0.4 to 1.5-1.9 (P less than 0.05). Group 5 livers did not lose K+; mean K+/Na+ ratio was 3.9 +/- 0.5. Fresh livers (no preservation) rapidly reaccumulated K+ when the tissue slices were incubated for 2 hr at 30 degrees C; mean K+/Na+ ratio was 3.7 +/- 0.5. Tissue slices from Group 2 livers (72 hr preservation), and livers pretreated with CPZ-MP (Group 3) or pretreated with 2-DOC (Group 4) did not significantly reaccumulate K+ at 30 degrees C; mean K+/Na+ ratio was 1.7-2.1. Only slices prepared from liver pretreated with both CPZ-MP and 2-DOC reaccumulated K+; mean K+/Na+ ratio was 4.6 +/- 1.2.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of chronic streptozotocin-induced diabetes on [3H]ouabain binding in the rat left ventricle

Earlier results from our laboratory have revealed that the inotropic response to ouabain was depressed in chronically diabetic rat heart (1). In this study we examined the effect of chronic streptozotocin induced diabetes (3 months) on [3H]ouabain binding in the rat heart. Scatchard analysis of [3H]ouabain binding to membrane preparations of rat left ventricle revealed two classes of binding sites; a high affinity/low capacity and a low affinity/high capacity binding site. The maximum number of binding sites of the high and low affinity binding sites in membrane preparations obtained from the chronically diabetic rats was significantly (p less than 0.05) reduced to 60.4 and 48.8% of controls, respectively. The dissociation constant of the high and the low affinity binding/sites in the chronically diabetic rat heart, compared to controls, was significantly (p less than 0.05) increased and decreased, respectively. These results suggest that the decreased inotropic response of ouabain in the intact tissue obtained from chronically diabetic rats (1) may be related to a decreased number of ouabain binding sites.     

Identification of specific vasopressin binding sites in cell membranes of the adenohypophysis, liver, kidney and adrenal glands of the rat

3H vasopressin specifically binds to the binding sites in liver, kidney and adenohypophysis with Bmax = 11.8 +/- 5.6, 1.7 +/- 0.5 and 4.9 +/- 1.2 pmol/g tissue and Kd = 1.5 +/- 0.5, 0.66 +/- 0.21 and 0.84 +/- 0.21 nM, correspondingly. Specific binding increases in the presence of Mg2+ and Ni2+ and decreases at high temperature (37 degrees). The presence of high affinity binding sites for 3H vasopressin was shown in the rat adrenals, binding was saturable and reversible. Concentration of vasopressin binding sites in adrenals is 6-8 fold less than in adenohypophysis. It is supposed that vasopressin receptors in adrenals may participate in the regulation of corticosteroid secretion.     

Regulation of liver lipase. I. Evidence for several regulatory sites, studied in corticotrophin-treated rats

The activity of liver lipase, an enzyme that can be released from the liver by heparin, varies under several hormonal conditions. The site(s) at which regulation of the enzyme activity may occur was investigated in vitro. As a model, rats were used which had been treated with a corticotrophin analogue, to induce hypercortisolism, a condition in which liver lipase activity is lowered. Lipases isolated from heparin-containing perfusates of livers from ACTH or control rats were identical with respect to heat stability and specific activity as determined by immunotitration and binding to isolated non-parenchymal liver cells, indicating that the enzyme structure was not affected by the treatment. The secretion of liver lipase by isolated parenchymal liver cells was studied. During incubation of parenchymal cells derived from ACTH rats, less enzyme activity was found to be secreted when compared with hepatocytes isolated from control rats (ACTH rats, 2.30 +/- 0.2 mU/10(6) cells; control rats, 3.3 +/- 0.3 mU/10(6) cells). Liver lipase partially purified from control rats could be bound specifically to saturation by non-parenchymal cells, isolated from ACTH or control rats. Non-parenchymal cells from ACTH rats bound less lipase activity (29 mU/mg cell protein) than cells from control rats (50 mU/mg cell protein). This reduction in binding capacity seems to be due to a diminished number of binding sites, since the affinity based on Scatchard analysis and half-maximal binding was not different. These results suggest that the lowered liver lipase activity found during hypercortisolism may be due to an impaired synthesis and/or secretion of the enzyme by the parenchymal cells and to a reduced binding capacity of the non-parenchymal cells for liver lipase.     

High affinity binding of [3H] cocaine to rat liver microsomes

[3H]Cocaine bound reversibly, with high affinity (KD 2.3 +/- 1.1 nM) and stereospecificity to rat liver microsomes. Little binding was detected in the lysosomal, mitochondrial and nuclear fractions. The binding kinetics were slow (T1/2 for association, 6 min and for dissociation 17 min), and the kinetically calculated KD was 2 nM. Induction of mixed function oxidases by phenobarbital did not produce significant change in [3H]cocaine binding. On the other hand, chronic administration of cocaine reduced [3H]cocaine binding drastically. Neither treatment affected the affinity of the liver binding protein for cocaine. Microsomes from mouse and human livers had less cocaine-binding protein and lower affinity for cocaine than those from rat liver. Binding of [3H]cocaine to rat liver microsomes was insensitive to monovalent cations and greater than 10 fold less sensitive to biogenic amines than the cocaine receptor in rat striatum. However, the liver protein had higher affinity for cocaine and metabolites except for norcocaine. Amine uptake inhibitors displaced [3H]cocaine binding to liver with a different rank order of potency than their displacement of [3H]cocaine binding to striatum. This high affinity [3H]cocaine binding protein in liver is not likely to be a monooxygenase, but may have a role in cocaine-induced hepatotoxicity.     

Localization of putative gonadotrophin releasing hormone receptor protein in the anterior pituitary

Specific binding of a fully biologically active 125I-gonadotrophin releasing hormone (GnRH) to isolated anterior pituitary cells is time dependent, saturable and the concentration dependent binding curves exhibit positive cooperativity. Binding to intact or solubilized plasma membranes and an affinity purified GnRH receptor protein reveals in all instances multiple high affinity binding sites. Thus, GnRH receptor protein appears to be an intrinsic constituent of the cell membrane, and perhaps, other membranous organelles. To investigate the latter, the binding of 125I-GnRH to various subcellular fractions was studied and its affinity and time requirements determined. GnRH binding to plasma membranes and secretory granules was to multiple high affinity sites, while that to nuclei and microsomes was to a single high affinity site. Binding was 1.83 +/- 0.07, 0.78 +/- 0.04, 0.31 +/- 0.03 and 0.27 +/- 0.03 fmol micrograms-1 protein for isolated plasma membranes, secretory granules, microsomes and nuclei, respectively, after 30 min incubation with 10(-9) M GnRH. The magnitude of binding to microsomes did not change during the incubation period. It did not show any decrease (p greater than 0.05) in isolated nuclei and plasma membranes, except for the 24 h time period, when a significant drop (p less than 0.001) was seen. Binding to the secretory granule fraction culminated at 15 min and then decreased (p less than 0.001) steadily to a non-detectable level at 24 h. Thus GnRH receptor protein or its portion may be an integral part of some membranous particles in the anterior pituitary cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Thionamides and arsenite inhibit specific T3 binding to the hepatic nuclear receptor

Methimazole (MMI) and propylthiouracil (PTU) are widely used for the treatment of Graves' disease. However, no studies have been reported on the action of these drugs on binding of L-triiodothyronine (T3) to the nuclear receptor. T3 receptors of rat liver nuclei, prepared by differential centrifugation, were extracted with 0.4 M KCl and 5 mM dithiothreitol (DTT). In the assessment of T3 binding to the DTT-reduced receptor, the hepatic nuclear extract was chromatographed on Superose 6 to remove DTT and isolate proteins of relative mass approximately 50,000 (chromatographed nuclear receptors (CNRs)), prior to the addition of [125I]T3 of high specific activity (3300 microCi/micrograms; 1 Ci = 37 GBq). MMI or PTU at 2 mM reduced specific T3 binding to CNR by 84% and 85%, respectively. The inhibitory effects of these reagents and 2 mM sodium arsenite (which complexes dithiols) were additive. Scatchard analyses indicated that neither MMI nor PTU (at 2 mM) significantly altered the affinity constant (Ka) (from 2.41 x 10(9) to 1.74 x 10(9) M-1 for PTU and 1.79 x 10(9) M-1 for MMI), while they both decreased (p less than 0.02) maximal binding capacity (from 0.36 +/- 0.02 to 0.19 +/- 0.02 pmol/mg protein for MMI and 0.17 +/- 0.02 pmol/mg protein for PTU). Dose-response curves showed that 50% inhibition was attained at 0.6 mM PTU or 1.0 mM MMI with approximately 25% inhibition by both at 0.1 mM. Artefactual binding effects by MMI and PTU on [125I]T3 were excluded by chromatography experiments. Similar results were obtained using nuclear receptors prepared from livers of hyperthyroid rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of vasopressin administration and deficiency upon 3H-AVP binding sites in the CNS and periphery during development.

Arginine8-vasopressin (AVP, 40 micrograms/100 g b.wt., SC) was administered to male Long-Evans (LE) pups from day 1 to 7 of life and the pups were sacrificed on day 8 or 60. 3H-AVP binding was performed on membranes prepared from the liver, kidney, and septum. No significant changes were observed in the kidney or septum of animals 8 or 60 days old. However, the chronic AVP treatment did result in a significant increase in the density of 3H-AVP binding sites in the liver when compared to control day 8 pups (control 44 +/- 2 vs. AVP 56 +/- 3 fmol/mg protein), with no change in affinity. This effect was maintained into adulthood, as the day 60 AVP-treated LE rats also showed a significant increase in liver 3H-AVP binding sites compared to control (control 186 +/- 9 vs. AVP 239 +/- 14 fmol/mg protein), with no change in affinity. A comparison of 3H-AVP binding sites in 8-day-old LE, heterozygous Brattleboro (HET-BB), and homozygous Brattleboro rats (HOM-BB) was performed to assess the effect of complete (HOM-BB) and partial (HET-BB) VP deficiency on binding sites in the CNS and periphery. The liver again was the only tissue in which a change in 3H-AVP binding characteristics was noted. The HOM-BB rat (Bmax 144 +/- 6 fmol/mg protein) displayed a significant increase in AVP binding sites from the LE rat (Bmax 100 +/- 7 fmol/mg protein), while the 3H-AVP binding sites in the HET-BB rat liver (Bmax 69.8 +/- 9 fmol/mg protein) were significantly lower than LE rats. Thus hepatic AVP receptors appear most sensitive to the presence or absence of vasopressin during the early postnatal period.     

Effects of method of preservation on functions of livers from fed and fasted rabbits

Livers from fed, fasted (48 h) and glucose-fed rabbits were preserved for 24 and 48 h by either simple cold storage (CS) or continuous machine perfusion (MP) with the University of Wisconsin preservation solutions. After preservation liver functions were measured by isolated perfusion of the liver (at 37 degrees C) for 2 h. Fasting caused an 85% reduction in the concentration of glycogen in the liver but no change in ATP or glutathione. Glucose feeding suppressed the loss of glycogen (39% loss). After 24 h preservation by CS livers from fed or fasted animals were similar including bile production (6.2 +/- 0.5 and 5.6 +/- 0.4 ml/2 h, 100 g, respectively), hepatocellular injury (LDH release = 965 +/- 100 and 1049 +/- 284 U/liter), and concentrations of ATP (1.17 +/- 0.15 and 1.18 +/- 0.04 mumol/g, glutathione (1.94 +/- 0.51 and 2.35 +/- 0.26 mumol/g, respectively), and K:Na ratio (6.7 +/- 1.0 and 7.7 +/- 0.5, respectively). After 48 h CS livers from fed animals were superior to livers from fasted animals including significantly more bile production (5.0 +/- 0.9 vs 2.0 +/- 0.3 ml/2 h, 100 g), less LDH release (1123 +/- 98 vs 3701 +/- 562 U/liter), higher concentration of ATP (0.50 +/- 0.16 vs 0.33 +/- 0.07 mumol/g) and glutathione (0.93 +/- 0.14 vs 0.30 +/- 0.13 mumol/g), and a larger K:Na ratio (7.4 vs 1.5). Livers from fed animals were also better preserved than livers from fasted animals when the method was machine perfusion. The decrease in liver functions in livers from fasted animals preserved for 48 h by CS or MP was prevented by feeding glucose. Glucose feeding increased bile formation after 48 h CS preservation from 2.0 +/- 0.3 (fasted) to 6.9 +/- 1.2 ml/2 h, 100 g; LDH release was reduced from 3701 +/- 562 (fasted) to 1450 +/- 154 U/liter; ATP was increased from 0.33 +/- 0.07 (fasted) to 1.63 +/- 0.18 mumol/g; glutathione was increased from 0.30 +/- 0.01 (fasted) to 2.17 +/- 0.30 mumol g; and K:Na ratio was increased from 1.5 +/- 0.9 to 5.3 +/- 1.0. This study shows that the nutritional status of the donor can affect the quality of liver preservation. The improvement in preservation by feeding rabbits only glucose suggests that glycogen is an important metabolite for successful liver preservation. Glycogen may be a source for ATP synthesis during the early period of reperfusion of preserved livers.