Mechanism of ion transport by avian salt gland primary cell cultures

1989 ◽  
Vol 256 (6) ◽  
pp. R1184-R1191
Author(s):  
R. J. Lowy ◽  
D. C. Dawson ◽  
S. A. Ernst
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Salt Gland ◽  
Secretory Cells ◽  
Adrenergic Stimulation ◽  
Beta Adrenergic ◽  
Dose Dependent Fashion ◽  
Active Ion Transport ◽  
Net Flux

Confluent sheets formed from primary culture of avian salt gland secretory cells exhibit a short-circuit current (Isc) in response to cholinergic and beta-adrenergic stimulation [Lowy, R. J., D. C. Dawson, and S. A. Ernst. Am J. Physiol. 249 (Cell Physiol. 18): C41-C47, 1985]. To establish the ionic basis for the Isc, transmural fluxes of 22Na and 36Cl were measured. Under short-circuit conditions there was little net flux of either ion in the absence of agonists. Addition of carbachol elevated net serosal-to-mucosal Cl flux to 1.71 mu eq.h-1.cm-2, whereas a smaller increase to 0.85 mu eq.h-1.cm-2 occurred with isoproterenol. Neither agonist altered net Na flux. The stimulated Isc accounted for 70% of the net Cl flux induced by carbachol and nearly 100% of that induced by isoproterenol. Replacement of Cl by gluconate or Na by choline abolished (carbachol) or greatly reduced (isoproterenol) the Isc, which could be restored in a dose-dependent fashion by ion restitution. Active ion transport was preferentially inhibited by basal (vs. apical) addition of ouabain, furosemide, or barium. The results provide evidence that cholinergic and beta-adrenergic agonists elicit active transmural Cl secretion. They further suggest that transport is dependent on the Na+-K+-adenosine-triphosphatase, a Na-Cl cotransport process, and a basal K conductance, all features of a secondary active Cl secretory mechanism.

Primary culture of duck salt gland. II. Neurohormonal stimulation of active transport

1985 ◽  
Vol 249 (1) ◽  
pp. C41-C47 ◽  
Author(s):  
R. J. Lowy ◽  
D. C. Dawson ◽  
S. A. Ernst
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Primary Cultures ◽  
Short Circuit Current ◽  
Salt Gland ◽  
Hormonal Control ◽  
Regulatory Control ◽  
Secretory Cells ◽  
Active Ion Transport ◽  
Secretory Transport

Primary cultures of structurally polarized sheets of avian salt gland secretory cells were mounted in Lucite chambers for transmural electrophysiological analysis. Transmural resistance values increased during the first 3 days of culture to 293 +/- 35 omega X cm2 and then decreased slowly thereafter. There was little short-circuit current (Isc) in the absence of secretagogues. Serosal addition of either carbachol or epinephrine resulted in a Isc consistent with positive charge flow from mucosa to serosa, thus demonstrating that these cell layers were capable of active ion transport in response to either cholinergic or adrenergic neurohormonal stimulation. Serosal ouabain or furosemide abolished the response to either agonist, while theophylline enhanced the response. Receptor specificity for the electrical responses was shown by selective inhibition of carbachol- and epinephrine-induced Isc by atropine and propranolol, respectively. The results demonstrate that these primary epithelial cell cultures are capable of active ion transport and are sensitive to known inhibitors of secretory transport, and suggest that intracellular coupling mechanisms for hormonal control are retained in culture. These cultures should be useful for studying mechanisms of ion secretory transport and their regulatory control.

Beta-adrenergic stimulation of ion transport in primary cultures of avian salt glands

1987 ◽  
Vol 252 (6) ◽  
pp. C670-C676 ◽  
Author(s):  
R. J. Lowy ◽  
S. A. Ernst
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Primary Cultures ◽  
Short Circuit Current ◽  
Salt Gland ◽  
Adrenergic Stimulation ◽  
Salt Glands ◽  
Beta Adrenergic ◽  
Effector Pathway ◽  
Stimulation Of

Adrenergic stimulation of transmural ion transport was identified and characterized in primary cultures of avian salt gland. Adrenergic activation was mediated by beta-receptors since stimulation of the short-circuit current (Isc) was blocked by propranolol but not phentolamine. The Isc's elicited by isoproterenol, epinephrine, and norepinephrine were dose dependent, with respective EC50 values of 1.5 X 10(-8) M, 5.0 X 10(-6) M, and 1.1 X 10(-5) M. The apparent Ki for propranolol inhibition after isoproterenol stimulation was 7.5 X 10(-10) M. 8-Br cyclic AMP (8-Br cAMP) and forskolin-elicited Isc's that were insensitive to propranolol, were potentiated by theophylline, and inhibited by furosemide or ouabain. Isoproterenol also induced an increase in ouabain-sensitive respiration in acutely dispersed cells from salt-stressed juvenile or unstressed adult animals, but not in fully salt-stressed adults. The data indicate that, in addition to the well-established cholinergic receptors, beta-adrenergic receptors can control ion transport in these glands. Furthermore, the results suggest for the first time that an intracellular effector pathway involving cAMP is present.

Vasoactive intestinal peptide stimulates ion transport in avian salt gland

1987 ◽  
Vol 253 (6) ◽  
pp. R801-R808 ◽  
Author(s):  
R. J. Lowy ◽  
J. H. Schreiber ◽  
S. A. Ernst
Keyword(s):  
Ion Transport ◽  
Vasoactive Intestinal Peptide ◽  
Short Circuit ◽  
Primary Cultures ◽  
Short Circuit Current ◽  
Salt Gland ◽  
Nerve Fibers ◽  
Secretory Cells ◽  
Isolated Cells ◽  
Salt Glands

Avian salt glands are considered to be under the control of cholinergic nerve fibers. Here we report evidence that vasoactive intestinal peptide (VIP) also regulates ion transport. Nerve fibers stained immunocytochemically with anti-VIP were distributed throughout the tissue within the peritubular connective tissue and were in close proximity to the secretory tubules. VIP applied to primary cultures of the secretory cells elicited active ion transport as assayed by short-circuit current (Isc) analysis. The mucosal-to-serosal positive Isc was produced in a dose-dependent fashion [(EC50) = 3.1 X 10(-9) M], was potentiated by theophylline, and was inhibited by either ouabain or furosemide. This Isc was independent of activation by cholinergic agonists. VIP also increased ouabain-sensitive respiration 14-18% in acutely isolated cells from salt-stressed and unstressed animals. These data demonstrate for the first time that VIP is present in the avian salt gland and can act as a secretagogue by directly affecting the secretory cells. In addition, the results provide evidence for direct control of ion transport by an adenosine 3',5'-cyclic monophosphate-linked neurohormone in both adult unstressed and fully salt-stressed animals.

Basolateral Cl− uptake mechanisms in Xenopus laevis lung epithelium

2010 ◽  
Vol 299 (1) ◽  
pp. R92-R100 ◽  
Author(s):  
Jens Berger ◽  
Martin Hardt ◽  
Wolfgang G. Clauss ◽  
Martin Fronius
Keyword(s):  
Xenopus Laevis ◽  
Ion Transport ◽  
Anion Exchanger ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Short Circuit Current ◽  
Transport Processes ◽  
Lung Epithelium ◽  
Active Ion Transport ◽  
Uptake Mechanisms

A thin liquid layer covers the lungs of air-breathing vertebrates. Active ion transport processes via the pulmonary epithelial cells regulate the maintenance of this layer. This study focuses on basolateral Cl− uptake mechanisms in native lungs of Xenopus laevis and the involvement of the Na+/K+/2 Cl− cotransporter (NKCC) and HCO3−/Cl− anion exchanger (AE), in particular. Western blot analysis and immunofluorescence staining revealed the expression of the NKCC protein in the Xenopus lung. Ussing chamber experiments demonstrated that the NKCC inhibitors (bumetanide and furosemide) were ineffective at blocking the cotransporter under basal conditions, as well as under pharmacologically stimulated Cl−-secreting conditions (forskolin and chlorzoxazone application). However, functional evidence for the NKCC was detected by generating a transepithelial Cl− gradient. Further, we were interested in the involvement of the HCO3−/Cl− anion exchanger to transepithelial ion transport processes. Basolateral application of DIDS, an inhibitor of the AE, resulted in a significantly decreased the short-circuit current (ISC). The effect of DIDS was diminished by acetazolamide and reduced by increased external HCO3− concentrations. Cl− secretion induced by forskolin was decreased by DIDS, but this effect was abolished in the presence of HCO3−. These experiments indicate that the AE at least partially contributes to Cl− secretion. Taken together, our data show that in Xenopus lung epithelia, the AE, rather than the NKCC, is involved in basolateral Cl− uptake, which contrasts with the common model for Cl− secretion in pulmonary epithelia.

Bioelectric properties and ion flow across excised human bronchi

1984 ◽  
Vol 56 (4) ◽  
pp. 868-877 ◽  
Author(s):  
M. Knowles ◽  
G. Murray ◽  
J. Shallal ◽  
F. Askin ◽  
V. Ranga ◽  
...  
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Maximal Effect ◽  
Na Transport ◽  
Adrenergic Agents ◽  
Ion Flow ◽  
Cl Secretion ◽  
Active Ion Transport ◽  
Human Bronchi

Bioelectric properties and ion transport of excised human segmental/subsegmental bronchi were measured in specimens from 40 patients. Transepithelial electric potential difference (PD), short-circuit current (Isc), and conductance (G), averaged 5.8 mV (lumen negative), 51 microA X cm-2, and 9 mS X cm-2, respectively. Na+ was absorbed from lumen to interstitium under open- and short-circuit conditions. Cl- flows were symmetrical under short-circuit conditions. Isc was abolished by 10(-4) M ouabain. Amiloride inhibited Isc (the concentration necessary to achieve 50% of the maximal effect = 7 X 10(-7) M) and abolished net Na+ transport. PD and Isc were not reduced to zero by amiloride because a net Cl- secretion was induced that reflected a reduction in Cl- flow in the absorptive direction (Jm----sCl-). Acetylcholine (10(-4) M) induced an electrically silent, matched flow of Na+ (1.7 mueq X cm-1 X h-1) and Cl- (1.9 mueq X cm-12 X h-1) toward the lumen. This response was blocked by atropine. Phenylephrine (10(-5) M) did not affect bioelectric properties or unidirectional ion flows, whereas isoproterenol (10(-5) M) induced a small increase in Isc (10%) without changing net ion flows significantly. We conclude that 1) Na+ absorption is the major active ion transport across excised human bronchi, 2) Na+ absorption is both amiloride and ouabain sensitive, 3) Cl- secretion can be induced by inhibition of the entry of luminal Na+ into the epithelia, and 4) cholinergic more than adrenergic agents modulate basal ion flow, probably by affecting gland output.

Regulation of Na-Cl absorption in rabbit proximal colon in vitro

1987 ◽  
Vol 252 (1) ◽  
pp. G45-G51 ◽  
Author(s):  
J. H. Sellin ◽  
R. De Soignie
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Proximal Colon ◽  
Beta Agonist ◽  
Disulfonic Acid ◽  
Low Concentrations ◽  
Cotransport System ◽  
Net Flux

Ion transport in rabbit proximal colon (PC) in vitro is dominated by a Na-Cl cotransport system stimulated by epinephrine. To further characterize the regulation of Na-Cl transport, we tested the effects of specific adrenergic agonists on ion fluxes under short-circuit conditions. Additionally, we tested the effects of the transport inhibitors bumetanide, furosemide, and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS). Basal Na and Cl absorption were essentially nil [Na net flux (JNanet) = 0.3 +/- 0.4, and Cl net flux (JClnet) = -0.5 +/- 0.5 mu eq X cm-2 X h-1, means +/- SE]. The alpha 2-agonist clonidine significantly increased net Na and Cl absorption (delta JNanet = 3.0 +/- 0.6 mu eq X cm-2 X h-1, delta JClnet = 2.0 +/- 0.4 mu eq X cm-2 X h-1) with a minimal change in short-circuit current (delta Isc = 0.1 +/- 0.1 mu eq X cm-2 X h-1). The alpha 1-agonist phenylephrine and the beta-agonist isoproterenol did not alter ion transport. The alpha 2-blocker yohimbine (YOH) had a complex, concentration-dependent effect. At low concentrations (10(-6)-10(-8) M) YOH effectively inhibited epinephrine-stimulated cotransport. Compared with 10(-8)M YOH, 10(-6) YOH blocked 90% of the epinephrine-induced increases in Na and Cl absorption.(ABSTRACT TRUNCATED AT 250 WORDS)

Chloride transport in glands of frog skin

1983 ◽  
Vol 244 (3) ◽  
pp. C221-C226 ◽  
Author(s):  
I. G. Thompson ◽  
J. W. Mills
Keyword(s):  
Frog Skin ◽  
Chloride Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Serosal Side ◽  
Adrenergic Stimulation ◽  
Beta Adrenergic ◽  
Skin Glands ◽  
Frog Skin Glands ◽  
Beta Adrenergic Agonist

The effects of beta-adrenergic stimulation on the bidirectional fluxes of Na+ and Cl- across the frog skin glands were determined. Isoproterenol elicited net serosal-to-mucosal fluxes of both Na+ (JNanet) and Cl- (JClnet) equal to 0.19 +/- 0.05 (SE) and 0.57 +/- 0.05 mueq X cm-2 X h-1, respectively. The residual current (JClnet - JNanet) of 0.38 +/- 0.05 mueq X cm-2 X h-1 closely approximates the isoproterenol-induced short-circuit current of 0.30 +/- 0.04 mueq X cm-2 X h-1. Furosemide added to the serosal side prior to isoproterenol inhibited the isoproterenol-induced net fluxes of both Na+ and Cl-. The addition of dibutyryl cAMP and 3-isobutyl-1-methylxanthine to the serosal side mimicked the action of isoproterenol by stimulating glandular short-circuit current. We conclude that an active Cl(-)-transport mechanism resides in the frog skin glands and is 1) stimulated by a beta-adrenergic agonist (its action is mimicked by cAMP) and 2) inhibited by the loop diuretic furosemide.

Phorbol ester and okadaic acid regulation of Na-2Cl-K cotransport in rabbit tracheal epithelial cells

1996 ◽  
Vol 271 (1) ◽  
pp. C338-C346 ◽  
Author(s):  
C. M. Liedtke ◽  
L. Thomas
Keyword(s):  
Epithelial Cells ◽  
Okadaic Acid ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Term Treatment ◽  
Secretory Cells ◽  
Adrenergic Stimulation ◽  
Short Term Treatment ◽  
Tracheal Epithelial Cells ◽  
Tracheal Epithelial

We evaluated a role for protein kinase C (PKC) in the regulation of rabbit tracheal epithelial Na-Cl(K) cotransport. Short-term treatment with phorbol 12-myristate 13-acetate (PMA) dose dependently increased bumetanide-sensitive Na and Cl efflux and elevated staurosporine- and bumetanide-sensitive Na, Cl, and K uptake. PMA and the alpha 2A-adrenergic agonist guanabenz both induced contransport with a stoichiometry of 2 Cl:1 Na and 2 Cl:1 Rb and elevated staurosporine-sensitive PKC activity in cytosolic and particulate fractions. Prolonged PMA treatment did not sustain bumetanide-sensitive 2 Cl:1 Na and 2 Cl:1 Rb transport but did block stimulation of bumetanide-sensitive transport by PMA or guanabenz and elevation of PKC activity by PMA and guanabenz in a particulate fraction. Cells treated with okadaic acid exhibited a staurosporine- and bumetanide-sensitive 2 Cl:1 Na and 2 Cl:1 Rb uptake. In cultured monolayers, basolateral perfusion with epinephrine, isoproterenol, or PMA increased short-circuit current (Isc). Basolateral application of bumetanide reduced elevated Isc to baseline levels, indicating a role for Cl secretory cells in a reconstituted tracheal epithelium. Pretreatment of transmonolayer cultures with PMA diminished the stimulatory response to epinephrine. These results indicate that, in rabbit tracheal epithelial cells, alpha-adrenergic stimulation activated Na-2Cl-K cotransport and that PKC is a critical effector in this process.

Changes in midgut active ion transport and metabolism during larval-larval molting in the tobacco hornworm (Manduca sexta)

1997 ◽  
Vol 200 (3) ◽  
pp. 643-648 ◽  
Author(s):  
M Chamberlin ◽  
C Gibellato ◽  
R Noecker ◽  
E Dankoski
Keyword(s):  
Ion Transport ◽  
Manduca Sexta ◽  
Aerobic Capacity ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Tobacco Hornworm ◽  
Maximal Aerobic Capacity ◽  
Posterior Midgut ◽  
Molting Process ◽  
Active Ion Transport

Ion transport and metabolism in the posterior midgut before, during and after the molt to the fifth instar of the tobacco hornworm Manduca sexta were investigated. In situ measurements reveal that the transepithelial potential difference of the posterior midgut falls during the molting process. This finding was confirmed by in vitro experiments in which it was demonstrated that both the transepithelial potential and the short-circuit current are lower in molting fourth instars compared with feeding fourth instars. The short-circuit current increases after ecdysis, with a maximal rate being achieved approximately 4 h after the molt. Resumption of feeding after the molt is not necessary to initiate this increase in active ion transport. The metabolic organization of the tissue also changes during the molting process. The maximal activities of glycolytic enzymes and 3-hydroxyacyl-CoA dehydrogenase, an enzyme of lipid ss-oxidation, decrease during the molting process and increase after ecdysis. Although citrate synthase activity, an index of maximal aerobic capacity, decreases during the molt and increases again after ecdysis, tissue respiration is the same in feeding fourth instars and molting larvae. This result indicates that a greater percentage of maximal aerobic capacity is used during molting and that energy may be diverted to cell proliferation and differentiation and away from the support of active ion transport at this time.

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