Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.6.1.1 (adenylate cyclase)
19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of the cytosol activator protein obtained from rat reticulocytes (RCAP) were investigated in a heterologous membrane system--partially purified cell membranes from dog renal cortex. RCAP enhanced the response of dog renal cortical adenylate cyclase to bovine parathyroid hormone (1-34) [bPTH (1-34)] from two- to three-fold. RCAP also enhanced the response to 5 microM arginine vasopressin, 10 microM glucagon, and 10 microM isoproterenol. Analysis of double-reciprocal plots of substrate concentration and enzyme activity indicated that bPTH (1-34) alone and together with RCAP increased the Vmax of the adenylate cyclase enzyme and did not alter the apparent Km of the enzyme for MgATP. Membranes from dog renal cortex contain 42K and 39K proteins that are ADP-ribosylated by cholera toxin and pertussis toxin, respectively, and appear to be the stimulatory (Ns) and inhibitory (Ni) guanine nucleotide binding proteins described in many other hormone-responsive membrane preparations. Similar to its effects in rat reticulocytes, RCAP inhibited ADP-ribosylation of Ns and enhanced ADP-ribosylation of Ni. The muscarinic agonist, carbachol, inhibited PTH-responsive adenylate cyclase activity in dog renal cortical membranes and this inhibition was reversed by RCAP. These results indicate that RCAP enhances stimulation of adenylate cyclase by a variety of hormones in a heterologous membrane preparation and supports the hypothesis that RCAP's site of action is common to all adenylate cyclase systems. RCAP may facilitate coupling between Ns and the catalytic unit of adenylate cyclase by a pertussis toxin-like effect to inactivate Ni. The dual effects of RCAP upon ADP-ribosylation of Ni and Ns alpha subunits suggest that a binding site for RCAP may exist at a site of homology between Ns alpha and Ni alpha.
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PMID:Enhancement of parathyroid hormone-responsive renal cortical adenylate cyclase activity by a cytosol protein activator from rat reticulocytes. 350 32

This study describes a method for the separation of distal cell populations based on the sequestration of proximal cells on immunoadsorbent columns (CNBr-activated Sepharose 6MB) bound with three brush-border monoclonal antibodies (S6-Mab). A high yield of isolated cell suspension from rabbit kidney cortex was prepared by mechanical dissociation after perfusion and incubation of the kidneys with 10(-3) M EDTA. The sequestration of the proximal cells was achieved in two sequential chromatographic steps. About 92% of the applied cells were first retained on an S6-Mab column after a 60-min stationary stage and the unbound cells were submitted by direct flow to a second S6-Mab column. In such conditions, 8 X 10(6) cells were recovered when starting with 331 X 10(6) cortical cells. The efficiency of the proximal cell depletion process was confirmed by an 80% decrease in brush-border enzymes, a very low phosphoenolpyruvate carboxykinase activity, and absence of cells bearing long microvilli, as ascertained by electron microscopy. This immunodepleted cell population presented the enzymatical characteristics of cells from the more distal segments. As compared with the initial cell suspension, these cells exhibited higher hexokinase (2.3 times), succinate dehydrogenase (1.5 times), and Na+-K+-ATPase (2.6 times) activities. In addition, adenylate cyclase activities remained sensitive to parathormone, arginine vasopressin, and isoproterenol. The functional capacity of these immunodepleted cells was assessed by an almost complete exclusion of eosin dye, a low Na+ and high K+ intracellular content, and a high respiratory rate of oxygen consumption. In conclusion, this immunoselective process makes it possible to obtain subpopulations of renal cortical cells possessing the main characteristics of the distal, connecting, and collecting cells for physiological and metabolic studies.
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PMID:Indirect immunoselection of late distal cell populations from rabbit kidney cortex. 351 19

In the present study, we were particularly interested in distinguishing specific patterns of structural and functional proteins in the collecting duct system of neonatal and adult kidneys and in cultured renal collecting duct epithelia in order to ascertain the degree of differentiation in the cultures. We studied the distribution of specific renal collecting duct cell markers using morphological, immunohistochemical and biochemical procedures. Cultured renal collecting duct epithelium undergoes maturation in vitro. Examples of morphological differentiation include the appearance of cilia and microvilli at the apical cell pole, and a basement membrane at the basal aspect of the epithelium. Tight junctions with five to seven strands separate the wide intercellular spaces from the apical cell surface. Physiological maturation from a 'leaky' to a 'tight' epithelium is evident from the acquisition of the alpha-subunit of Na/K-ATPase and the development of a high transepithelial potential difference and resistance. Biochemical differentiation is revealed by the expression of specific proteins. The simple-epithelium cytokeratins, PKK1 and PKK2, which are typical intracellular-matrix proteins of mature collecting duct epithelium, maintain the same distribution in cell culture as in neonatal and adult kidneys. An indicator of maturation in vitro is the expression of the collecting duct-specific proteins, PCD2 and PCD3. Newly developed monoclonal antibodies against these antigens reacted similarly with cultured cells and cells of the mature collecting duct system, but they did not label the embryonic ampullae in the cortex of neonatal rabbit kidneys. In contrast, a third collecting duct-specific protein, PCD1, is not expressed by the cultured cells, which indicates the retention of an embryonic characteristic in vitro. Embryonic collecting duct ampullae of the neonatal kidney in situ contain laminin during their development. Laminin is, however, absent in cultured collecting duct epithelium. Biochemical stimulation of the adenylate cyclase system by arginine vasopressin resulted in a twofold stimulation of the enzyme activity. This degree of stimulation is similar to that found in maturing kidneys of neonatal rabbits and indicates another embryonic feature of the cultures.
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PMID:Differentiation properties of renal collecting duct cells in culture. 355 30

Two groups of Sprague-Dawley rats, Harlan (H) and Charles River (CR), were discovered in that the medullary thick ascending limb (MAL) had a profoundly different adenylate cyclase response to arginine vasopressin (AVP). Using these two groups of rats, we studied the correlation between AVP action on the MAL and maximal urinary concentration. AVP (10(-6) M) significantly stimulated adenylate cyclase in MAL of H rats (7.4 +/- 0.9 to 43.8 +/- 4.6 fmol cAMP formed X 30 min-1 X mm-1, P less than 0.001) but not in CR rats (10.3 +/- 1.4 to 12.7 +/- 2.0 fmol cAMP formed X 30 min-1 X mm-1, NS). In contrast, AVP significantly stimulated adenylate cyclase of cortical, outer and inner medullary collecting tubules from both H and CR rats. Glucagon (10(-6) M) significantly stimulated adenylate cyclase of MAL from both H and CR rats. After 48 h of fluid deprivation, urinary osmolality was significantly higher (P less than 0.001) in the H (4,504 +/- 399 mosmol/kg H2O, n = 14) than CR (2,840 +/- 176 mosmol/kg H2O, n = rats. This observation was not attributable to differences in creatinine clearance (CR, 1.30 +/- 0.24; H, 1.24 +/- 0.03 ml/min, NS, n = 4) or plasma AVP (CR, 12.75 +/- 1.44; H, 12.38 +/- 1.17 pg/ml, NS, n = 6) levels. These results therefore suggest that the action of AVP on the MAL, in addition to the effect on collecting tubules, is involved in maximal urinary concentration in rats.
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PMID:Role of arginine vasopressin in medullary thick ascending limb on maximal urinary concentration. 374 Feb 73

Rabbit nephron segments of proximal convoluted tubules (PCT); proximal straight tubules (PST); cortical and medullary thick ascending limbs of Henle's loop (CAL, MAL); and cortical, outer medullary, and inner medullary collecting tubules (CCT, OMCT, IMCT) were individually microdissected and grown in monolayer culture in hormone supplemented, defined media. Factors favoring a rapid onset of proliferation included young donor age, distal tubule origin, and the addition of 3% fetal calf serum to the medium. All primary cultures had polarized morphology with apical microvilli facing the medium and basement membrane-like material adjacent to the dish. Differentiated properties characteristic of the tubular epithelium of origin retained in cultures included ultrastructural characteristics and cytochemically demonstrable marker enzyme proportions. PCT and PST were rich in alkaline phosphatase; CAL stained strongly for NaK-ATPase; CCT contained two cell populations with regard to cytochrome oxidase reaction. A CCT-specific anti-keratin antibody (aLEA) was immunolocalized in CCT cultures, and a PST cytokeratin antibody stained PST cultures. The biochemical response of adenylate cyclase to putative stimulating agents was the same in primary cultures as in freshly isolated tubules. In PCT and PST adenylate cyclase activity was stimulated by parathyroid hormone (PTH) but not by arginine vasopressin (AVP); CAL and MAL adenylate cyclase was stimulated by neither PTH nor AVP; CCT, OMCT, and IMCT adenylate cyclase was stimulated by AVP but not by PTH. NaF stimulated adenylate cyclase activity in every cultured segment. It is concluded that primary cultures of individually microdissected rabbit PCT, PST, CAL, MAL, CCT, OMCT, and IMCT retain differentiated characteristics with regard to ultrastructure, marker enzymes, cytoskeletal proteins, and hormone response of adenylate cyclase and provide a new system for studying normal and abnormal functions of the heterogeneous tubular epithelia in the kidney.
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PMID:Retention of differentiated characteristics by cultures of defined rabbit kidney epithelia. 381 2

Confluent monolayers of pig renal papillary collecting tubule (RPCT) cells were formed on Millipore filters coated with collagen. They were clamped in Ussing-type chambers and used to measure short-circuit current (SCC). The monolayers had low potentials (0.1 mV) with the basolateral side positive. Small inward currents flowed under short-circuit conditions. Increases in SCC were obtained following addition of a number of agents. Receptors associated with SCC changes were disposed as follows: for kinins (e.g., lysyl-bradykinin) they were present on both sides of the tissue, while those for arginine vasopressin and norepinephrine were present on the basolateral side only. Epithelia responded to PGE2 added to the apical or basolateral face of the tissue; application to one side prevented the response from the contralateral side. The tissues also responded to forskolin, an activator of adenylate cyclase, with a sustained inward current that was sensitive to furosemide. Similar sustained inward currents were recorded following exposure to 8-bromoadenosine-3',5'-cyclic monophosphate (BrcAMP). Responses to kinins were attenuated by inhibition of fatty acid cyclooxygenase with either indomethacin or piroxicam or by replacing chloride with impermeant ions. If the SCC was first increased with forskolin, BrcAMP, or norepinephrine, the kinin effects on SCC were either abolished or reversed. It is concluded that kinin can cause chloride secretion in RPCT monolayers, possibly via a prostaglandin or a prostaglandin-adenylate cyclase mechanism. Secondary effects of kinin, exposed by first raising tissue cAMP levels, are not precluded.
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PMID:Kinin effects on electrogenic ion transport in primary cultures of pig renal papillary collecting tubule cells. 386 51

To understand the molecular mechanism of action of the novel class of diuretic agents, the antidiuretic hormone antagonists ["aquaretics" (specific water-losing activity as caused by vasopressin antagonists, as distinguished from the saluresis of conventional diuretics)], in the dog studies were made of the properties of the vasopressin-responsive adenylate cyclase system and the antagonist potencies of the vasopressin analogs [1-(beta-mercapto-beta, beta-cyclopentamethylenepropionic acid), 2-(O-ethyl)tyrosine,4-valine,8-arginine]vasopressin; [1-(beta-mercapto-beta-mercapto-beta, beta-cyclopentamethylenepropionic acid), 2-D-phenylalanine,4-valine,8-arginine]vasopressin; and [1-(beta-mercapto-beta, beta-cyclopenta-methylenepropionic acid), 2-D-(O-ethyl)tyrosine,4-valine,8-arginine]vasopressin (SK&F 100398, 101071 and 101498, respectively) using plasma membranes prepared from cortex, medulla and papilla of dog kidney. It was observed that the greatest sensitivity for vasopressin was in the papilla (concentration of 8-arginine vasopressin required for 50% activation of adenylate cyclase [Kact] was 2.0 X 10(-9)M, 1.1 X 10(-9)M and 5.1 X 10(-10) M in the cortex, medulla and papilla, respectively). The addition of 10(-5)M GTP did not alter the Kact of the cortex but enhanced 10-fold the vasopressin sensitivity of the papilla to 5.2 X 10(-11) M. The vasopressin analogs were competitive antagonists of vasopressin-stimulated adenylate cyclase of cortex and papilla with the greatest potency for the papillary enzyme (Ki in papilla was 3.6 X 10(-9)M, 4.6 X 10(-9)M and 1.0 X 10(-9)M for SK&F 100398, 101071 and 101498, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Antidiuretic hormone antagonists and aquaresis in dogs: different vasopressin sensitivity and antagonist potency in renal cortex and papilla. 396 88

The effects of 1-desamino-8-D-arginine vasopressin (dDAVP) on the handling of water and electrolytes by the juxtamedullary nephrons were studied on rats with reduced circulating levels of antidiuretic hormone (ADH), parathyroid hormone, calcitonin, and glucagon, all of which stimulate the adenylate cyclase system of the thick ascending limb and the distal tubule. In such hormone-deprived rats and in hormone-deprived + dDAVP rats, the concentration of Na, Cl, and total solutes was lower in the ascending than in the descending limbs, whereas the inulin concentration was similar at both sites. dDAVP did not alter the fraction of NaCl remaining in the thin limbs, but tended to reduce that of Mg and Ca. On the other hand, dDAVP significantly increased the fraction of filtered K remaining from 65.8 +/- 5.2 to 107.3 +/- 15.8%. A direct correlation was observed between the fraction of filtered K remaining at the tip of the juxtamedullary loops and the fractional excretion rate of K in urine. Since dDAVP enhances distal K net secretion, as previously shown in our laboratory, these results indicate that the medullary recycling of K from nephron terminal segments to Henle's loop of juxtamedullary nephrons is stimulated by this peptide.
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PMID:Effects of dDAVP on rat juxtamedullary nephrons: stimulation of medullary K recycling. 402 56

Distal nephron epithelia of defined anatomical origin were microdissected from rabbit kidneys and individually explanted into an in vitro culture system. The 7 day monolayers grown from four different nephron epithelia were studied for the presence and amount of adenylate cyclase reaction product. In each case basal adenylate cyclase was compared with the enzyme reaction product after stimulation by arginine vasopressin, calcitonin, parathyroid hormone (PTH) and isoproterenol. In cortical collecting tubule cultures, the reaction was stimulated by vasopressin greater than isoproterenol greater than calcitonin. PTH had no effect. In cortical thick ascending loop of Henle cells, the stimulation was by calcitonin greater than vasopressin = PTH. Isoproterenol had no effect. In medullary ascending loop epithelia, stimulation was by vasopressin = calcitonin. Neither isoproterenol nor PTH had an effect. These observations indicate that adenylate cyclase is histochemically demonstrable in cultivated cells from rabbit distal nephron segments and that the enzyme activation by hormones is differential according to the epithelium of origin.
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PMID:Histochemical localization of hormone sensitive adenylate cyclase in defined nephron epithelia in culture. 403 Apr 8

To evaluate the hypothesis that prostaglandins (PGs) inhibit vasopressin stimulation of adenylate cyclase in the collecting tubule, we have studied the interactions of vasopressin, PGs, and intracellular cAMP in rat renal papillary collecting tubule (RPCT) cells in cell culture. Inhibition of PGE2 synthesis with acetylsalicylic acid (ASA) did not potentiate arginine vasopressin (AVP)-stimulated intracellular cAMP. Augmentation of prostanoid synthesis with arachidonic acid or exogenous addition of PGE2 did not decrease AVP-stimulated cAMP in the RPCT cells. Arachidonic acid or PGE2, used alone, increased cAMP and ASA reduced cAMP, consistent with the presence of a PG-sensitive adenylate cyclase. Six-hour incubations of RPCT cells produced clear evidence of homologous desensitization of the PGE2 receptor, after exposure to either PGE2 or arachidonic acid, but not heterologous desensitization of the AVP receptor linked to cAMP synthesis. Preincubation of the RPCT cells with AVP or 1-desamino-8-D-arginine vasopressin (dDAVP) induced homologous desensitization of AVP- or dDAVP-stimulated cAMP. Three-day incubations with dDAVP or AVP apparently induced cyclooxygenase activity since PGE2 synthesis increased in response to arachidonic acid and recovery of cyclooxygenase activity after ASA was enhanced by dDAVP or AVP. In conclusion, our data on AVP-stimulated cAMP in cultured RPCT cells do not support the hypothesis that PGE2 inhibits AVP-dependent collecting tubule cAMP.
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PMID:Interactions of vasopressin, prostaglandins, and cAMP in rat renal papillary collecting tubule cells in culture. 608 89


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