Regulation of 21-hydroxylase gene expression.

The steroid 21-hydroxylase (21-OH) gene is selectively expressed in the adrenal cortex and is transcriptionally regulated by ACTH. We examined the role of the 5'-flanking sequences of 21-OH in this regulated expression by analyzing their ability to direct the expression of a human growth hormone (hGH) reporter gene upon transfection into Y1 mouse adrenocortical tumor cells. The 330 bp of 5'-flanking sequences directed basal and hormonally-inducible expression of hGH in Y1 cells, but did not direct expression in I-10 mouse testicular Leydig cells.

Dexamethasone accelerates differentiation of A6 epithelia and increases response to vasopressin.

When seeded heavily on a porous tissue culture dish, A6 cells, derived from the kidney of Xenopus laevis, form a highly differentiated epithelium within 4-6 days. When dexamethasone is added to the culture medium, morphological differentiation is completed by day 2, a time at which the control (untreated) is still a disorganized multilayer of cells. In addition to the morphologically evident monolayer of cuboidal cells, the accelerated differentiation is expressed as high transepithelial electrical resistance, short-circuit current, and adenylate cyclase response to vasopressin.

Hepatic ethanol metabolism is mediated predominantly by catalase-H2O2 in the fasted state.

Methanol and butanol were employed as selective substrates for catalase-H2O2 and alcohol dehydrogenase (ADH), respectively, in the perfused rat liver. As expected, rates of butanol metabolism accounted for over 85% of overall rates of alcohol oxidation indicating that ADH was the predominant pathway of alcohol metabolism in both the fed or fasted state in the absence of added substrate. In the fasted state, however, addition of oleate (1 mM) diminished butanol oxidation 20-25% yet increased rates of methanol oxidation over 4-fold.

Catalase-dependent ethanol oxidation in perfused rat liver. Requirement for fatty-acid-stimulated H2O2 production by peroxisomes.

The purpose of this study was to measure rates of catalase-dependent ethanol uptake and rates of H2O2 generation in perfused rat livers in the presence of fatty acids of varying chain length. Rates of ethanol uptake in livers from fasted rats, perfused in a recirculating system, of about 80 mumol g-1 h-1 were decreased to about 10 mumol g-1 h-1 by the addition of an inhibitor of alcohol dehydrogenase (ADH), 4-methylpyrazole. The medium-chain-length fatty acid, laurate (12:0; 1 mM), increased rates of 4-methylpyrazole-insensitive ethanol uptake maximally to 80-85 mumol g-1 h-1.

An enhancer element and a functional cyclic AMP-dependent protein kinase are required for expression of adrenocortical 21-hydroxylase.

The steroid 21-hydroxylase (21-OHase) gene is selectively expressed at high levels in cells of the adrenal cortex and is transcriptionally regulated by corticotropin (ACTH). In this study, we examined the contribution of cis-acting nucleotide sequences to the regulated expression of the mouse 21-OHase gene. The 5'-flanking sequences of the mouse 21-OHase gene, extending 330 bp upstream from the transcription initiation site, were placed in front of the human growth hormone (hGH) reporter gene, and expression of the fusion gene was measured following transient transfection in Y1 mouse adrenocortical tumor cells.

Antidiuretic hormone moves membranes.

This review focuses on events at the apical plasma membrane of toad urinary bladder and mammalian collecting duct as their permeability to water changes in response to antidiuretic hormone (ADH) and to its withdrawal. The major marker of the permeability change is observed in freeze-fracture electron microscopy of the apical plasma membrane and consists of a dramatic increase in membrane particle aggregates and, in toad bladder but not in collecting duct, in fused vesicles (aggrephores) that contain particle aggregates in their limiting membranes. Withdrawal of ADH is accompanied by endocytosis at the apical membrane, reflecting retrieval of water-permeable, particle aggregate-containing membrane.

Identification of P-450ALC in microsomes from alcohol dehydrogenase-deficient deermice: contribution to ethanol elimination in vivo.

Isozyme 3a of rabbit hepatic cytochrome P-450, also termed P-450ALC, was previously isolated and characterized and was shown to be induced 3- to 5-fold by exposure to ethanol. In the present study, antibody against rabbit P-450ALC was used to identify a homologous protein in alcohol dehydrogenase-negative (ADH-) and -positive (ADH+) deermice, Peromyscus maniculatus. The antibody reacts with a single protein having an apparent molecular weight of 52,000 on immunoblots of hepatic microsomes from untreated and ethanol-treated deermice from both strains.

Involvement of hormones in the swift increase in alcohol metabolism.

The purpose of this study was to determine the time course of changes in blood levels of various hormones in C57BL/6J mice during exposure to ethanol vapor. Groups of adult male mice were given 2.0 g per kg ethanol intraperitoneally or as continuous vapor for four hours and rates of ethanol elimination were measured.

Apical membrane endocytosis via coated pits is stimulated by removal of antidiuretic hormone from isolated, perfused rabbit cortical collecting tubule.

Antidiuretic hormone increases the water permeability of the cortical collecting tubule and causes the appearance of intramembrane particle aggregates in the apical plasma membrane of principal cells. Particle aggregates are located in apical membrane coated pits during stimulation of collecting ducts with ADH in situ. Removal of ADH causes a rapid decline in water permeability.

Identification of specific apical membrane polypeptides associated with the antidiuretic hormone-elicited water permeability increase in the toad urinary bladder.

Antidiuretic hormone (ADH) increases the water permeability of the toad urinary bladder. The increase occurs in the apical plasma membrane of granular cells that line the urinary surface of the bladder and is produced by the insertion of water permeability units that have been identified by freeze-fracture electron microscopy as intramembrane particle aggregates. Under water-impermeable conditions, particle aggregates reside in intracellular vesicles called "aggrephores.

Inhibition of catalase-dependent ethanol metabolism in alcohol dehydrogenase-deficient deermice by fructose.

Hepatic microsomal fractions from ADH (alcohol dehydrogenase)-negative deermice incubated with an NADPH-generating system metabolized butanol and ethanol at rates around 10 nmol/min per mg. In contrast, cytosolic catalase from ADH-negative deermouse liver oxidized ethanol, but not butanol, when incubated with an H2O2-generating system. Thus butanol is oxidized by cytochrome P-450 in microsomal fractions, but not by cytosolic catalase, in tissues from ADH-negative deermice.

Implications of tooth root hypercementosis in a Barbados slave skeletal collection.

A 17th- to 19th-century cemetery sample of 104 slaves from Newton Plantation (Barbados) shows uniquely high hypercementosis prevalence, as well as unexpectedly high and variable skeletal lead content. A variety of biological and archeological factors indicates that individuals with lower amounts of these anomalies (relative to age at death) were probably African-born, first-generation slaves. The hypercementosis is related to the progression of periodontal disease as assessed from alveolar bone.

Rates of H2O2 generation from peroxisomal beta-oxidation are sufficient to account for fatty acid-stimulated ethanol metabolism in perfused rat liver.

Fatty acids generate H2O2 via peroxisomal beta-oxidation and increase ethanol metabolism markedly in a system that involves catalase-H2O2. The present studies were conducted to understand why fatty acid-stimulated ethanol metabolism occurs much faster than rates of H2O2 generation reported previously in perfused rat liver. A new method was developed to measure rates of H2O2 generation based on the fact that methanol is oxidized only by catalase in rat liver.

Precursors of ribose 5-phosphate suppress expression of glucose-regulated proteins in LLC-PK1 cells.

Withdrawal of glucose from the medium bathing mammalian cells in culture results in cessation of growth and induces the synthesis of two stress proteins (Mr approximately 94-100 kDa and 78-80 kDa) that have been termed glucose-regulated proteins (GRPs). In LLC-PK1 cells, proteins of the same molecular weights, assayed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and autoradiography, are fully induced by 24 h in glucose-free medium. The GRPs from LLC-PK1 cells cross-react with antibodies to GRPs of nonpolar cells, confirming their identification.

Isolation, growth, and characterization of a gluconeogenic strain of renal cells.

LLC-PK1 cells, derived from pig kidney, retain several properties of the proximal tubule, but are incapable of gluconeogenesis, due to the lack of fructose-1,6-bisphosphatase (FBPase) [Am. J. Physiol.

Isolation and characterization of specialized regions of toad urinary bladder apical plasma membrane involved in the water permeability response to antidiuretic hormone.

Antidiuretic hormone (ADH) increases the apical (external facing) membrane water permeability of granular cells that line the toad urinary bladder. In response to ADH, cytoplasmic vesicles called aggrephores fuse with the apical plasma membrane and insert particle aggregates which are visualized by freeze-fracture electron microscopy. Aggrephores contain particle aggregates within their limiting membranes.

Fatty acids supply H2O2 at high rates for the oxidation of ethanol by catalase.

Fatty acids are physiological substrates for H2O2 production via peroxisomal beta-oxidation and have been shown to increase ethanol metabolism markedly in a system that involves catalase-H2O2. To try to understand why fatty acid-stimulated ethanol uptake occurs much faster than rates of H2O2 generation reported previously, studies were conducted to measure these parameters under identical conditions in the perfused rat liver. New methods were developed to measure H2O2 generation in a recirculating perfusion system based on the fact that methanol is oxidized only by catalase in rat liver.

Catalase-dependent ethanol metabolism in vivo in deermice lacking alcohol dehydrogenase.

Pathways of ethanol elimination in alcohol dehydrogenase (ADH)-positive and -negative deermice were studied using the catalase inhibitor, 3-amino-1,2,4-triazole. To verify that aminotriazole inhibited catalase effectively, the characteristic decrease in catalase-H2O2 which occurs in saline-treated controls when ethanol is peroxidized was monitored at 660-640 nm in perfused deermouse livers. Following 1.

Mechanism of zone-specific hepatic steatosis caused by valproate: inhibition of ketogenesis in periportal regions of the liver lobule.

Microvacuolar steatosis in periportal regions of the liver lobule was produced by injection of fasted rats with a single dose of valproate (500 mg/kg, subcutaneously). In livers perfused in the absence of exogenous fatty acids, ketone body (acetoacetate + beta-hydroxybutyrate) production was decreased by valproate (500 microM) maximally by 67%. Concomitantly, NADH fluorescence detected from the liver surface declined about 30% with a time course similar to that of the inhibition of ketogenesis.

Gangliosides modulate sodium transport in cultured toad kidney epithelia.

Cultured A6 epithelial cells from toad kidney form confluent monolayers with tight junctions separating the apical and basolateral membranes. These two membrane domains have distinct compositions and functions. Thus, sodium is actively transported across the epithelia from the apical to basolateral surface via amiloride-inhibitable sodium channels located in the apical membrane.

Adenosine 3',5'-cyclic monophosphate stimulates chloride secretion in A6 epithelia.

Basal and aldosterone-stimulated short-circuit current (Isc) of A6 epithelia are known to be equivalent to net apical to basal Na flux and are completely inhibited by 0.05 mM amiloride added to the solution bathing the apical surface of the epithelium. In the absence of amiloride, the Isc stimulated by adenosine 3',5'-cyclic monophosphate (cAMP) is also equivalent to net apical to basal Na flux.

Transepithelial water flow regulates apical membrane retrieval in antidiuretic hormone-stimulated toad urinary bladder.

Antidiuretic hormone (ADH) increases the osmotic water permeability (Posm) of toad urinary bladder. This increase is believed to be produced by fusion of intracellular vesicles called aggrephores with the granular cell apical plasma membrane. Aggrephores contain intramembrane particle aggregates postulated to be water channels.

Fluorescent markers to study membrane retrieval in antidiuretic hormone-treated toad urinary bladder.

Antidiuretic hormone (ADH) stimulation of toad urinary bladder causes fusion of intracellular vesicles called aggrephores with the apical plasma membrane of granular cells. Aggrephores contain intramembrane particle aggregates whose appearance in the apical membrane is believed to produce a large increase in its water permeability. ADH removal (ADH washout) is thought to cause the retrieval of aggrephores into granular cell cytoplasm.