Effects of A-23187 and verapamil on the active transport enzymes in turtle bladder epithelial cells

1997 ◽  
Vol 272 (5) ◽  
pp. R1379-R1389 ◽  
Author(s):  
S. Sabatini
Keyword(s):  
Direct Action ◽  
Short Circuit ◽  
Calcium Ionophore ◽  
Short Circuit Current ◽  
Sodium Reabsorption ◽  
Proton Secretion ◽  
G Cells ◽  
A 23187 ◽  
Turtle Bladder ◽  
Intact Membrane

These experiments were designed to examine the effects of A-23187 (5 x 10(-4) M) and verapamil (100 microM) on membrane transport, 45Ca fluxes, and adenosine-triphosphatase (ATPase) activities in turtle bladder. In the intact membrane, the calcium ionophore decreased proton secretion and sodium transport [short-circuit current (SCC)] to approximately the same degree (by approximately 55% at 30 min). During the same period of time, verapamil decreased SCC (by approximately 58%), but proton secretion was unaffected. The turtle bladder membrane is composed predominantly of two cell types: 1) the mitochondrial-rich cells (MR cells) thought to be involved in proton (and bicarbonate) secretion containing significant H(+)-ATPase and Ca(2+)-ATPase and 2) the granular cells (G cells), postulated important in sodium reabsorption, having abundant Na(+)-K(+)-ATPase. That Na(+)-K(+)-ATPase activity was unchanged by either a calcium ionophore or a calcium channel blocker suggests that the decrease in SCC noted in the intact membrane is not directly mediated by changes in the sodium "pump." The decrease of H(+)-ATPase in MR cells, which resulted after the A-23187, suggests that it probably exerts a direct action on the proton pump, which decreases hydrogen ion secretion. The increase in ATP-dependent 45Ca transport seen after the ionophore (or the decrease in ATP-independent 45Ca transport after verapamil) most likely reflects increased (or decreased) Ca2+ availability within the cytosol, and the high (or low) cell calcium could decrease the SCC. These results thus suggest that cytosolic Ca2+ reciprocally sets, by different mechanisms, the rate of proton secretion in MR cells and the sodium reabsorption in G cells.

scholarly journals Spontaneous water secretion in T84 cells: effects of STa enterotoxin, bumetanide, VIP, forskolin, and A-23187

2001 ◽  
Vol 281 (3) ◽  
pp. G816-G822 ◽  
Author(s):  
Roxana Toriano ◽  
Arlinet Kierbel ◽  
Marco Antonio Ramirez ◽  
Gerhard Malnic ◽  
Mario Parisi
Keyword(s):  
Short Circuit ◽  
Second Messengers ◽  
Short Circuit Current ◽  
Rt Pcr ◽  
T84 Cells ◽  
Pharmacological Agents ◽  
Heat Stable ◽  
A 23187 ◽  
Water Secretion ◽  
Secretory Apparatus

The regulated Cl− secretory apparatus of T84 cells responds to several pharmacological agents via different second messengers (Ca2+, cAMP, cGMP). However, information about water movements in T84 cells has not been available. In the absence of osmotic or chemical gradient, we observed a net secretory transepithelial volume flux ( J w = −0.16 ± 0.02 μl · min−1 · cm−2) in parallel with moderate short-circuit current values ( I sc = 1.55 ± 0.23 μA/cm2). The secretory J wreversibly reverted to an absorptive value when A-23187 was added to the serosal bath. Vasoactive intestinal polypeptide increased I sc, but, unexpectedly, J w was not affected. Bumetanide, an inhibitor of basolateral Na+-K+-2Cl−cotransporter, completely blocked secretory J wwith no change in I sc. Conversely, serosal forskolin increased I sc, but J w switched from secretory to absorptive values. Escherichia coli heat-stable enterotoxin increased secretory J w and I sc. No difference between the absorptive and secretory unidirectional Cl−fluxes was observed in basal conditions, but after STa stimulation, a significant net secretory Cl− flux developed. We conclude that, under these conditions, the presence of secretory or absorptive J w values cannot be shown by I sc and ion flux studies. Furthermore, RT-PCR experiments indicate that aquaporins were not expressed in T84 cells. The molecular pathway for water secretion appears to be transcellular, moving through the lipid bilayer or, as recently proposed, through water-solute cotransporters.

Aldosterone response in the turtle bladder is associated with an increase in ATP

1983 ◽  
Vol 245 (4) ◽  
pp. F512-F514
Author(s):  
N. Cortas ◽  
E. Abras ◽  
M. Walser
Keyword(s):  
Perchloric Acid ◽  
Sodium Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Open Circuit ◽  
Potential Difference ◽  
Freshwater Turtles ◽  
Active Sodium ◽  
Active Sodium Transport ◽  
Turtle Bladder

Urinary bladders from freshwater turtles, mounted as sacs, were stripped of their serosa and submucosa. This did not alter conductance. They were maintained in open circuit except for brief observation of short-circuit current (SCC) every 15 min. Potential difference (PD) averaged 68 +/- 14 mV and SCC 485 +/- 100 microA. Acetazolamide 10(-3) M increased SCC by 46 +/- 27 microA. Aldosterone 10(-7) M following acetazolamide resulted in a rise in SCC that began at about 75 min and reached a plateau between 3 and 5 h. SCC rose 127 +/- 15% compared with control bladder halves. ATP measured in perchloric acid extracts 5 h after addition of aldosterone increased by 33% (P less than 0.01) and (ATP)/(ADP) X (Pi) by 81% (P less than 0.01). These results support the view that the stimulatory effects of aldosterone on active sodium transport involve an increase in ATP and (ATP)/(ADP) X (Pi).

Effect of parathyroid hormone on transport by toad and turtle bladder

1987 ◽  
Vol 252 (1) ◽  
pp. R63-R68
Author(s):  
S. Sabatini ◽  
N. A. Kurtzman
Keyword(s):  
Parathyroid Hormone ◽  
Epithelial Cells ◽  
Calcium Uptake ◽  
Short Circuit ◽  
Calcium Ionophore ◽  
Toad Bladder ◽  
Base Line ◽  
Ionophore A23187 ◽  
45Ca Uptake ◽  
Turtle Bladder

We recently demonstrated that parathyroid hormone (PTH) inhibited both vasopressin- and cyclic AMP-stimulated water transport in the toad bladder. This was associated with an increase in calcium uptake by isolated epithelial cells. We postulated that PTH exerts its action on H2O transport by directly stimulating calcium uptake. The current study was designed to compare the effects of PTH and the calcium ionophore, A23187, on H2O and Na transport and H+ secretion in toad and turtle bladders. In toad bladder, PTH and A23187 decreased arginine vasopressin (AVP)-stimulated H2O flow and short-circuit current (SCC) after 60 min serosal incubation. In turtle bladder A23187 decreased SCC to 79.3 +/- 3.6% of base line (P less than 0.05), and significantly decreased RSCC as well. PTH had no effect on SCC or H+ secretion in turtle bladders. Both PTH and A23187 increased 45Ca uptake in toad bladder epithelial cells; only A23187 increased 45Ca uptake in the turtle bladder. The different action of PTH in these two membranes, compared with that of the calcium ionophore, illustrates the selectivity of PTH on membrane transport. PTH increases calcium uptake and decreases transport only in a hormone-sensitive epithelium, whereas the ionophore works in virtually all living membranes. The mode of action of these two agents to increase calcium uptake is, therefore, likely different.

Evidence for separate cellular origins of sodium and acid-base transport in the turtle bladder

1986 ◽  
Vol 250 (4) ◽  
pp. C609-C616 ◽  
Author(s):  
J. H. Durham ◽  
W. Nagel
Keyword(s):  
Epithelial Cells ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Transport Processes ◽  
Electrical Parameters ◽  
Acid Base ◽  
Intracellular Potential ◽  
Turtle Bladder ◽  
Stimulation Of

Transmembrane electrical parameters of the epithelial cells in short-circuited turtle bladders were measured to determine whether those cells participating in Na reabsorption also participate in electrogenic transepithelial acidification and alkalinization. Amiloride-induced increases in intracellular potential (Vsca), apical fractional resistance (FRa), and concomitant decreases in short-circuit current (Isc) denote the participation of the impaled cells in Na reabsorption. In bladders from postabsorptive turtles, amiloride increased Vsca by -45 mV, increased FRa by 37%, and decreased Isc from 36 to -10 microA/cm2. In bladders from NaHCO3-loaded turtles, amiloride increased Vsca by -21 mV, FRa by 21%, and decreased Isc from 22 to 0 microA/cm2. Neither the subsequent inhibition of the negative acidification current in postabsorptive bladders, nor stimulation of positive alkalinization current in bladders from NaHCO3-loaded turtles was associated with any transmembrane electrical change that could be attributed to changes in those transport processes. It is concluded that the electrogenic luminal acidification and alkalinization processes of the turtle bladder are not produced by, or electrically coupled to, those cells that are involved in Na reabsorption.

Release of cyclooxygenase products from primary cultures of tracheal epithelia of dog and human

1989 ◽  
Vol 257 (6) ◽  
pp. L361-L365 ◽  
Author(s):  
J. H. Widdicombe ◽  
I. F. Ueki ◽  
D. Emery ◽  
D. Margolskee ◽  
J. Yergey ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Gas Chromatography ◽  
Vasoactive Intestinal Peptide ◽  
Short Circuit ◽  
Primary Cultures ◽  
Platelet Activating Factor ◽  
Calcium Ionophore ◽  
Short Circuit Current ◽  
Gas Chromatography Mass Spectrometry ◽  
Pg E2

Release of cyclooxygenase products from primary cultures of dog or human tracheal epithelium was measured by radioimmunoassay. In both species, bradykinin, platelet-activating factor (PAF), and A23187 (a calcium ionophore) caused increases in the rate of release of prostaglandin (PG) E2 and smaller increases in PGF2 alpha, 6-keto-PGF1 alpha, and thromboxane B2 output. Isoproterenol, vasoactive intestinal peptide (VIP), methacholine, and leukotrienes C4 and D4 had no effect on release of these cyclooxygenase products. Gas chromatography-mass spectrometry showed that the ratio of PGE2 to PGD2 released from dog cells by A23187 was 30:1. Short-circuit current across dog cells was stimulated by bradykinin, A23187, PAF, VIP, methacholine, and isoproterenol. Only the responses to bradykinin and A23187 were reduced by pretreatment with indomethacin.

Sch-28080 inhibits bafilomycin-sensitive H+ secretion in turtle bladder independently of luminal [K+]

1993 ◽  
Vol 265 (2) ◽  
pp. F174-F179
Author(s):  
O. F. Kohn ◽  
P. P. Mitchell ◽  
P. R. Steinmetz
Keyword(s):  
Competitive Inhibition ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Atpase Inhibitor ◽  
Turtle Bladder ◽  
High K ◽  
Ph Stat ◽  
Low Levels ◽  
K Concentration ◽  
Removal Studies

To explore the possible contribution of an H-K-adenosine-triphosphatase (H-K-ATPase) to H+ secretion (JH) in the isolated turtle bladder, we measured electrogenic JH (JeH) as short-circuit current and total JH (JTH) by pH stat titration in the presence of ouabain at different ambient K+ concentration ([K+]) and during luminal addition of a known gastric H-K-ATPase inhibitor, Schering (Sch)-28080. JH was not reduced by decreasing ambient [K+] to undetectable or very low levels (< 0.05 mM by atomic absorption) and luminal BaCl2 addition to further reduce local [K+] at the apical membrane. These K(+)-removal studies indicate that H+ transport is not coupled to countertransport of K+. JTH did not exceed JeH at any point: in K(+)-free solutions JTH was 0.73 +/- 0.05, and JeH was 0.95 +/- 0.08 mumol/h; in standard (3.5 mM) K+ solutions JTH was 0.72 +/- 0.05 and JeH 0.98 +/- 0.06 mumol/h; in high (118 mM) K+ solutions JTH was 0.65 +/- 0.07 and JeH 0.94 +/- 0.08 mumol/h. Sch-28080 caused a rapid inhibition of JH, with similar half-maximal inhibitory concentrations (IC50) in K(+)-free, standard [K+], and high [K+] solutions. Bafilomycin inhibited JeH and JTH with an IC50 of approximately 100 nM. The observed non-potassium-competitive inhibition of JH by Sch-28080 and the bafilomycin sensitivity distinguish the H-ATPase of the turtle bladder from the gastric H-K-ATPase. The rapidity of the inhibition by Sch-28080 suggests that it acts at an accessible luminal site of the ATPase.

HCO3-Cl exchange transport in the adaptive response to alkalosis by turtle bladder

1980 ◽  
Vol 239 (2) ◽  
pp. F167-F174
Author(s):  
L. Cohen
Keyword(s):  
Urinary Bladder ◽  
Adaptive Response ◽  
Intact Animal ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Na Transport ◽  
Acid Base ◽  
Turtle Bladder ◽  
Acid Base Status ◽  
Ph Stat

The isolated turtle urinary bladder acidifies its mucosal (M) solution, and the rate of acidification (JH) is equivalent to the short-circuit current after Na+ transport is abolished by ouabain. When HCO3(-) is present in the serosal solution it is secreted into M in an electroneutral exchange for absorbed Cl-. The rate of HCO3(-) secretion (JHCO3(-)) can be measured by pH stat titration after JH is nullified by an opposing pH gradient. With use of these methods JH and JHCO3 were measured sequentially in bladdes from control animals and animals fed NaHCO3 (alkalosis) or NH4Cl (acidosis). JH in alkalosis (57 +/- 6 micro A) was ot different from control values (53 +/- 7 micro A). JHCO3, however, was nearly 40% higher in alkalosis (1.63 +/- 0.11 vs. 1.17 +/- 0.14 mu mol x h-1 x 8 cm-2). In contrast, JHCO3 in acidosis was similar to control values (0.89 +/- 0.15 mu mol x h-1 x 8 cm-2) but JH was increased. As judged from Cl- fluxes, neither alkalosis nor acidosis altered the electroneutral coupling between HCO3(-) secretion and Cl- absorption. JH and JHCO3 appear to be independent processes in the turtle bladder that are capable of responding independently to physiologic changes in the acid-base status of the intact animal.

Mineralocorticoid activity of 19-nor-DOC and 19-OH-DOC in toad bladder

1981 ◽  
Vol 241 (5) ◽  
pp. E406-E409 ◽  
Author(s):  
R. D. Perrone ◽  
J. H. Schwartz ◽  
H. H. Bengele ◽  
S. L. Dale ◽  
J. C. Melby ◽  
...  
Keyword(s):  
Sodium Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Toad Bladder ◽  
Sodium Reabsorption ◽  
Sodium Retention ◽  
Onset Of Action ◽  
Mineralocorticoid Activity

Adrenal enucleation is followed by a period of increased sodium reabsorption thought to be due to excess mineralocorticoid activity. However, it has not been demonstrated that increased production of any known mineralocorticoid accounts for this antinatriuresis. Recently, 19-hydroxydeoxycorticosterone (19-OH-DOC) was found in incubates of regenerating adrenal capsules 3-4 days postenucleation and 19-nordeoxycorticosterone (19-nor-DOC) was identified in the urine of rats with regenerating adrenals. Because it was possible that these hormones might play a role in the sodium retention after adrenal enucleation, we compared the mineralocorticoid activity of these steroids to aldosterone using the toad bladder. Using short-circuit current as a measure of sodium transport, we found that 19-OH-DOC (10(-8) M) had no significant effect on sodium transport. However, 19-nor-DOC (10(-8) M) increased sodium transport to a degree not different from aldosterone (10(-8) M). Furthermore, the onset of action, duration of activity, and inhibition of effect of 19-nor-DOC by spironolactone were not different from that of aldosterone. We conclude that 19-nor-DOC exhibits a significant effect on sodium transport and thus has the potential to play a role in the sodium retention following adrenal enucleation. Under the conditions of these studies, 19-OH-DOC exhibited no effect on sodium transport.

Bombesin stimulation of gastrin release from canine gastrin cells in primary culture

1987 ◽  
Vol 252 (3) ◽  
pp. G413-G420 ◽  
Author(s):  
A. S. Giraud ◽  
A. H. Soll ◽  
F. Cuttitta ◽  
J. H. Walsh
Keyword(s):  
Direct Action ◽  
Somatostatin Receptors ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Gastrin Release ◽  
Short Term ◽  
Gastrin Cells ◽  
G Cells ◽  
Short Term Culture

A method is described for the isolation and short-term culture of canine antral gastrin (G) cells. Tissue was dispersed by enzymes and G cells enriched by elutriation and cultured for 40 h. These cultures contained 12% G cells and less than 2% somatostatin- or serotonin-containing cells. Bombesin (0.001–100 pM) potently stimulated gastrin release from cell cultures in a linear fashion over 2 h. The bombesin-specific monoclonal antibody 2A11 dose-dependently blocked bombesin stimulation. Somatostatin (0.001–1,000 nM) inhibited bombesin-stimulated gastrin release. Antibody to somatostatin (Mab S8) prevented the inhibition by exogenous somatostatin but did not alter bombesin-stimulated or basal gastrin release. The substance P (SP) analogue spantide (1 nM-1 microM) did not inhibit bombesin-stimulated gastrin release. Postreceptor activation of adenylate cyclase by forskolin and of protein kinase C by the phorbol ester, beta-TPA, caused gastrin release. The calcium ionophore A23187 also released gastrin in a dose-dependent fashion. This methodology allows enrichment and short-term culture of antral G cells; these cells have stimulatory bombesin and inhibitory somatostatin receptors, suggesting that these peptides have a direct action on antral G cells. Furthermore, G cells are activated by cAMP and calcium/phosphatidylinositol-dependent mechanisms.

Neuropeptide Y inhibits chloride secretion in the shark rectal gland

1993 ◽  
Vol 265 (2) ◽  
pp. R439-R446 ◽  
Author(s):  
P. Silva ◽  
F. H. Epstein ◽  
K. J. Karnaky ◽  
S. Reichlin ◽  
J. N. Forrest
Keyword(s):  
Neuropeptide Y ◽  
Direct Action ◽  
Short Circuit ◽  
Primary Cultures ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Rectal Gland ◽  
Maximal Effect ◽  
Gland Cells ◽  
Shark Rectal Gland

We studied the effects of the 36-amino acid peptide, neuropeptide Y (NPY), on salt secretion by the rectal gland of Squalus acanthias. We used three preparations: whole isolated perfused glands, freshly prepared separated rectal gland tubules, and confluent monolayers of cultured rectal gland cells. In perfused glands NPY inhibited secretion stimulated by vasoactive intestinal peptide (VIP), forskolin, or adenosine 3',5'-cyclic monophosphate (cAMP) and theophylline. Maximal inhibition of 63 +/- 3.4% was seen at 3 x 10(-8) M NPY, with half-maximal effect at 3 x 10(-9) M. NPY did not inhibit the basal activity of rectal gland adenylate cyclase or that stimulated by VIP. The inhibitory action of NPY was not prevented by procaine, nifedipine, or diltiazem, suggesting that it was not secondary to the release of somatostatin or other unknown neurotransmitters from rectal gland nerves. In confirmation, somatostatin was not detected in the venous effluent after administration of NPY. NPY also inhibited transport-related oxygen consumption in separated rectal gland tubules and inhibited short-circuit current generated by confluent monolayers of primary cultures of rectal gland cells. The results indicate that NPY inhibits chloride secretion by a direct action on cells of the shark rectal gland at a site distal to the generation of cAMP.

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