Susan J. Sulakhe

Hepatic α1 and β adrenergic receptors in various animal species

SummaryPlasma membranes were isolated from the livers of various animal species representing the four vertebrate classes: Amphibia, Reptilia, Aves and Mammalia. These liver plasma membranes displayed comparable levels of purity as judged by marker enzyme analysis. The activities of the two marker enzymes, 5′-nucleotidase and γ-glutamyltranspeptidase displayed striking, and quite different, species-dependent differences, with no apparent relationship to phylogeny. α1 and β-adrenergic receptors were characterized in isolated liver plasma membranes by radioligand binding techniques. The hepatic β-adrenergic receptor was found to be expressed in all animals studied; the hepatic α1-adrenergic receptor was absent in Amphibia and Reptilia, co-expressed with the β receptor in Aves, and dominant over the β receptor in Mammalia. These results suggest that, in liver, the β-adrenergic receptor is more primitive while the α1-adrenergic receptor is of a more recent phylogenetic origin. It is proposed that the latter may have evolved in conjunction with hepatic sympathetic innervation.

The activity of γ-glutamyltranspeptidase in regenerating rat liver

γ‐Glutamyltranspeptidase is expressed at low levels in the liver of the male Fischer 344 rat where it exhibits 15‐fold purification and 33% recovery in isolated plasma membranes. While the activity of the enzyme is unaltered in regenerating liver 24 h after partial hepatectomy, it increases steadily thereafter over a period of one week. Seven days after partial hepatectomy the enzyme is maximally activated: 5.6‐fold in liver homogenates and 5.3‐fold in isolated liver plasma membranes. The enzyme declines in activity over the next fourteen days and is expressed at normal levels three weeks after partial hepatectomy. These results demonstrate that the activity of γ‐glutamyltranspeptidase increases in regenerating liver but that the increase is out of phase with the proliferative response.

A characterization of γ-glutamyltranspeptidase in normal, perinatal, premalignant and malignant rat liver

gamma-Glutamyltranspeptidase displays the following order of activity in tissues of the Fischer 344 rat: kidney much greater than small intestine much greater than cerebral cortex = testis greater than lung much greater than liver = heart. The activity of the hepatic enzyme in rats is: 4-fold higher in females than males; 4-fold higher in male Wistar, Sprague-Dawley and Zucker rats than male Fischer 344 rats; increased 10-fold in very old vs young male Fischer 344. The hepatic enzyme displays significant species variation: the mouse and rat liver enzymes are similar and low in activity, while duck, dog, pig and beef enzymes are 7, 13, 86 and 92-fold higher, respectively, in activity than the male Fischer rat liver enzyme. A liver plasma membrane isolation procedure has been devised which selects for the sinusoidal face of the liver parenchymal cell as assessed by marker enzyme analysis: for these plasma membranes the purification of gamma-glutamyltranspeptidase is 21.5 and the recovery is 42% indicating that this is the cellular and subcellular locus of the enzyme in rat liver. The characteristics of the liver plasma membrane from female rats are: pH optimum of 8.0; classical Michaelis-Menten kinetics; Km of 1.43 mM and Vmax of 33.3 nmol X mg-1 X min-1. In Fischer 344 rats, gamma-glutamyltranspeptidase activities are elevated over adult levels in perinatal liver: in fetal liver homogenates and plasma membranes the activities are increased 179 and 109-fold, respectively. The activity peaks just after birth and declines rapidly over the first 15 postnatal days. The activity of the liver enzyme in the male Fischer 344 rat exhibits a progressive increase throughout diethylnitrosamine-induced hepatocarcinogenesis: it is increased 7.8-fold in homogenates and 5.4-fold in plasma membranes at the early premalignant stage; 74-fold in homogenates and 31-fold in plasma membranes at the later hyperplastic nodular premalignant stage; and 174-fold in homogenates and 61-fold in plasma membranes at the hepatoma stage. The gradual drop in purification during hepatocarcinogenesis is associated with the appearance of the enzyme in the blood.

The activity of hepatic γ-glutamyltranspeptidase in various animal species

The activity of gamma-glutamyltranspeptidase was determined in liver homogenates derived from various animals belonging to different phylogenetic groups. Although gamma-glutamyltranspeptidase was present in all livers studied, the activity varied greatly, with the order of activity being: guinea pig much greater than frog much greater than rabbit greater than toad greater than dog congruent to cat = duck greater than hamster = budgerigar = goldfish congruent to rat = mouse. There was no trend with respect to enzyme activity, phylogeny, diet, or habitat.

Modulation of γ-glutamyltranspeptidase activity in rat liver plasma membranes by thyroid hormone

1. 1. In adult male and female rats, liver plasma membrane γ-glutamyltranspeptidase activities were 16-fold higher in the propylthiouracil (PTU)-induced hypothyroid state than in the control euthyroid state; thyroxine (T4replacement resulted in an 80% restoration to control levels. 2. 2. Liver plasma membrane γ-glutamyltranspeptidase activities were 6.7-fold higher in PTU-induced congenitally hypothyroid rats than in control euthyroid rats; T4-replacement reduced enzyme activities to 37% of control levels. 3. 3. In adult rats, in response to the development and recovery from tri-iodothyronine (t3) excess, liver plasma membrane γ-glutamyltranspeptidase activities were inversely related to, and out of phase by 12 hr, to the earlier changes in T3. 4. 4. Liver γ-glutamyltranspeptidase is a thyroid hormone-dependent enzyme.

Studies on the stimulation of (Na+-K+) ATPase of neural tissues by catecholamines

Abstract 1. 1. (Na+-K+) ATPase activities of hypothalamic, cortical, cerebellar, caudate nucleus and superior cervical ganglia homogenates of either rat or rabbit were stimulated by catecholamines to a varying degree. 2. 2. This stimulatory effect depended on the method of homogenization of tissue and assay of enzyme activity. 3. 3. Cyclic nucleotides failed to show any stimulatory action. 4. 4. Partially purified (Na+-K+) ATPase preparations were phosphorylated in the presence of ATP by the reaction catalyzed by protein kinase in the presence and absence of cyclic AMP. 5. 5. The phosphorylated product was hydroxylamine-insensitive, alkaline phosphatase-sensitive, acid-resistant and alkali-labile. 6. 6. However, phosphorylation did not stimulate the (Na+-K+) ATPase activity.

The impact of hypothyroidism and thyroxine replacement on the expression of hepatic α1-, α2- and β-adrenergic receptors in rat liver plasma membranes

Abstract 1. 1. Liver plasma membranes were isolated from control, propylthiouracil-induced hypothyroid and thyroxine-replaced rats; relative specific activities of 5′-nucleotidase were found to be similar, 5.6–6.1, demonstrating that comparable purity levels were achieved. 2. 2. Radioligand binding studies indicated that hepatic α1-, α2- and β-adrenergic receptor binding to control liver membranes was 1963.23 ± 59.34, 77.64 ± 2.20 and 111.18 ± 11.04 fmol·mg−1, respectively. 3. 3. Hypothyroidism induced a 67% and 54% decrease, respectively, in hepatic α1- and α2-adrenergic receptor binding with no change in β-adrenergic receptor binding. 4. 4. Thyroxine replacement achieved an 85% and 100% restoration, respectively, in hepatic α1- and α2-adrenergic receptor expression with no effect on the β-adrenergic receptor.

Interactions of Ionophores (A23187 and X-537A) with sarcolemma and sarcoplasmic reticulum

Abstract 1. The effects of the ionophores A23187 and X-537A on various muscle sarcoplasmic reticular and sarcolemmal enzyme activities were studied. 2. Both ionophores (A23187 more potent than X-537A) profoundly stimulated MgCaATPase (not MgATPase or CaATPase) and also stimulated MgCaϱ-nitrophenyl phosphatase and MgCaGTPase. A23187 stimulated sarcolemmal MgCaGTPase in part by increasing the turnover of the phosphoprotein enzyme intermediate. 3. The ionophores (A23187 more effective than X-537A) both blocked calcium influx into membrane vesicles and effected an instantaneous release of bound (uncomplexed) calcium. 4. X-537A inhibited cardiac sarcolemmal (Na+-K+)ATPase. 5. A23187 modestly inhibited adenylate cyclase activity (basal, epi, NaF-stimulated) while X-537A (low concentrations) modestly stimulated guanylate cyclase activity of these membranes.

Calcium transport activities of plasma membranes isolated from the livers of various animal species

1. Plasma membranes of comparable yield and purity were isolated from the livers of various animal species belonging to phylogenetic groups from Amphibia to Mammalia. 2. Calcium transport activity was observed in all liver plasma membranes examined. 3. No phylogenetic pattern of expression of the liver plasma membrane calcium transport system was observed, with the order of activity being: guinea pig greater than rabbit greater than frog greater than chicken = hamster greater than rat = budgerigar = turtle greater than beef cattle greater than mouse = duck. 4. Calcium transport activity was only 9.7 and 8.7% of adult frog levels in plasma membranes isolated from the livers of tadpoles without and with limbs, respectively. 5. Liver plasma membrane calcium transport activity was 25% higher in adult chickens than in day-old chicks. 6. A possible role for thyroid hormone in the development of the liver plasma membrane calcium transport system is discussed.

Diethylnitrosamine-induced increase in γ-glutamyltranspeptidase in rat liver: Its association with thyroid hormone deficiency and its reversal by tri-iodothyronine

1. The nodular phase of hepatic premalignancy was induced in male Fischer 344 rats by the administration of diethylnitrosamine, 200 mg/kg i.p., followed by promotion utilizing the Solt-Farber promoting regime. 2. Relative to the situation in normal non-treated control rats: the activity of gamma-glutamyltranspeptidase was found to be increased 9.42-fold in homogenate and 7.33-fold in plasma membrane fractions prepared from the livers of saline-injected control rats; and 81.37-fold in homogenates and 91.92-fold in plasma membranes prepared from the livers of diethylnitrosamine-injected rats; plasma levels of total T3 and total T4 were found to be decreased 42.06 and 47.45% in saline-injected control rats and 88.7 and 83.2% in diethylnitrosamine-injected rats, respectively. 3. An early pre-nodular phase of hepatic premalignancy was produced in young immature and mature adult male Fischer 344 rats by the administration of diethylnitrosamine, 75 mg/kg, without subsequent application of the promotion regime. 4. Relative to the situation in control rats: the activity of gamma-glutamyltranspeptidase was found to be increased in liver homogenates prepared from diethylnitrosamine-treated rats, 1.62-fold in young immature rats 1.20-fold in mature adult rats; plasma levels of total T3 were found to be reduced in diethylnitrosamine-treated rats, 28% in young immature rats 9% in mature adult rats. 5. Treatment of diethylnitrosamine-injected young immature male Fischer 344 rats at the prenodular phase of hepatic premalignancy with tri-iodothyronine at 0.005 micrograms/kg s.c. daily for 7 days reversed the diethylnitrosamine-induced increase in liver homogenate gamma-glutamyltranspeptidase activity and the decrease in plasma total T3, restoring these parameters to normal levels.(ABSTRACT TRUNCATED AT 250 WORDS)