CYCLIC AMP STIMULATION OF ELECTROGENIC UPTAKE OF Na+ AND Cl- ACROSS THE GILL EPITHELIUM OF THE CHINESE CRAB ERIOCHEIR SINENSIS

1994 ◽  
Vol 188 (1) ◽  
pp. 159-174 ◽  
Author(s):  
S Riestenpatt ◽  
W Zeiske ◽  
H Onken
Keyword(s):  
Cyclic Amp ◽  
Electromotive Force ◽  
Single Channel ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Eriocheir Sinensis ◽  
Na Channel ◽  
Before And After ◽  
Gill Lamellae ◽  
Stimulation Of

Split gill lamellae (epithelium plus cuticle) of hyperregulating Chinese crabs acclimated to fresh water were mounted in a modified Ussing chamber. Active and electrogenic absorption of sodium and chloride were measured as positive amiloride-sensitive and negative Cl--dependent short-circuit currents (INa, ICl), respectively. Both currents were characterized before and after treatment of the tissue with theophylline or dibutyryl cyclic AMP. Both drugs increased INa and ICl. A simple circuit analysis showed that INa stimulation reflected a marked increase in the transcellular Na+ conductance, whereas the respective electromotive force was unchanged. The Michaelis constant (KNa) for Na+ current saturation was decreased after INa stimulation, indicating an increased affinity of the transport mechanism for its substrate. Consequently, the affinity for the Na+ channel blocker amiloride decreased as expected for a competitive interaction between substrate and inhibitor. Analysis of the amiloride-induced current-noise revealed a marked increase in the number of apical Na+ channels after INa stimulation with theophylline, whereas there was little change in the single-channel current. Stimulation of Cl- absorption was accompanied by a substantial increase in both transcellular conductance and electromotive force, indicating an activation of the apical H+ pump that provides the driving force for active Cl- uptake via apical Cl-/HCO3- exchange and basolateral Cl- channels.

Active NaCl absorption across split lamellae of posterior gills of the chinese crab Eriocheir sinensis: stimulation by eyestalk extract

2000 ◽  
Vol 203 (8) ◽  
pp. 1373-1381 ◽  
Author(s):  
H. Onken ◽  
A. Schobel ◽  
J. Kraft ◽  
M. Putzenlechner
Keyword(s):  
Electromotive Force ◽  
Single Channel ◽  
Short Circuit ◽  
Noise Analysis ◽  
Eriocheir Sinensis ◽  
Dialysis Tubing ◽  
Channel Current ◽  
Dose Dependent ◽  
Cl Conductance ◽  
Stimulation Of

Split lamellae of the posterior gills of freshwater-adapted Chinese crabs (Eriocheir sinensis) were mounted in a modified Ussing-type chamber, and active and electrogenic absorption of Na(+) and Cl(−) were measured as positive (I(Na)) or negative (I(Cl)) short-circuit currents. Haemolymph-side addition of eyestalk extract stimulated I(Cl) by increasing both the transcellular Cl(−) conductance and the electromotive force for Cl(−) absorption. The effect was dose-dependent. Boiling the eyestalk extract did not change its effectiveness. The stimulating factor passed through dialysis tubing, indicating that it has a molecular mass of less than 2 kDa. R(p)cAMPS, a blocker of protein kinase A, reduced the stimulated I(Cl). Eyestalk extract stimulated I(Na) by increasing the transcellular Na(+) conductance at constant electromotive force. Amiloride-induced current-noise analysis revealed that stimulation of I(Na) was accompanied by an increase in the apparent number of open apical Na(+) channels at a slightly reduced single-channel current. In addition to the electrophysiological experiments, whole gills were perfused in the presence and in the absence of putative transport stimulators, and the specific activities of the V-ATPase and the Na(+)/K(+)-ATPase were measured. Eyestalk extract, theophylline or dibutyryl-cyclic AMP stimulated the activity of the V-ATPase, whereas the activity of the Na(+)/K(+)-ATPase was unaffected. The simultaneous presence of R(p)cAMPS prevented the stimulation of V-ATPase by eyestalk extract or theophylline.

scholarly journals Acid-Base Transport and Control in Locust Hindgut: Artefacts Caused by Short-Circuit Current

1991 ◽  
Vol 155 (1) ◽  
pp. 455-467
Author(s):  
R. BRENT THOMSON ◽  
N. AUDSLEY ◽  
JOHN E. PHILLIPS
Keyword(s):  
Cyclic Amp ◽  
Acid Secretion ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Salt Bridges ◽  
Electrical Parameters ◽  
Acid Base ◽  
Before And After ◽  
And Control ◽  
Stimulation Of

The commonly used method of passing short-circuit current (Isc) across insect epithelia through Ag-AgCl electrodes, without the use of salt bridges, leads to significant OH− production at the cathode (lumen side) when high currents are applied. The alkalization of the lumen previously reported when cyclic AMP was added to short-circuited locust hindgut is a result of this phenomenon rather than cyclic-AMP-mediated stimulation of acid-base transport in the hindgut. When salt bridges are used to pass short-circuit current across locust hindgut, acid secretion (JH) into the lumen equals alkaline movement (JOH) to the haemocoel side, and JH is similar under both open- and short-circuit conditions. JH is similar (1.5 μequiv cm−2 h−1) in recta and ilea. Addition of cyclic AMP inhibits JH across the rectum by 42–66%, but has no effect on the ileum when salt bridges are used. Electrical parameters (Isc, Vt, Rt) reflecting hindgut Cl− transport (JCL) before and after stimulation with cyclic AMP are the same whether or not salt bridges are used. We found no evidence of any coupling between JCl and JH/JOH.

Metabolic Support of Chloride-dependent Short-circuit Current Across Locust Rectum

1982 ◽  
Vol 99 (1) ◽  
pp. 349-362
Author(s):  
M. CHAMBERLIN ◽  
J. E. PHILLIPS
Keyword(s):  
Cyclic Amp ◽  
Short Circuit ◽  
Malpighian Tubule ◽  
Short Circuit Current ◽  
Metabolic Substrates ◽  
Endogenous Substrates ◽  
Tubule Fluid ◽  
Substrate Source ◽  
Stimulation Of

1. Recta of desert locusts were short-circuited and depleted of endogenous substrates by exposing them to saline containing cyclic AMP but no metabolites. Individual substrates were then added to substrate-depleted recta and the change in short-circuit current (Isc) monitored. 2. Proline or glucose (50 mM) caused by far the largest increase in Isc of all substrates tested. Stimulation of the Isc by proline was not dependent upon external sodium, but did require external chloride. 3. Physiological levels of proline also caused a large increase in Isc, while physiological levels of glucose produced a much smaller stimulation. Over 90% of the proline-dependent Isc stimulation can be produced by adding 15 mM proline solely to the lumen side of the tissue. 4. These results are discussed with regard to rectal oxidative metabolism and availability of metabolic substrates in vivo. High levels of proline in Malpighian tubule fluid are probably the major substrate source for rectal Cl−transport. Note:

Cholinergic modulation of apical Na+ channels in turtle colon: analysis of CDPC-induced fluctuations

1990 ◽  
Vol 259 (4) ◽  
pp. C668-C674 ◽  
Author(s):  
D. J. Wilkinson ◽  
D. C. Dawson
Keyword(s):  
Single Channel ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Rate Coefficients ◽  
Corner Frequency ◽  
Na Channel ◽  
Fluctuation Analysis ◽  
Na Channels ◽  
Na Current

Current fluctuation analysis was used to investigate the properties of apical Na+ channels during muscarinic inhibition of active Na+ absorption. A reversible Na+ channel blocker, 6-chloro-3,5-diaminopyrazine-2-carboxamide (CDPC), was used to induce fluctuations in the short-circuit current (I(sc)). Power density spectra of the CDPC-induced fluctuations exhibited a clearly discernible Lorentzian component, characterized by a corner frequency that was linearly related to CDPC concentration between 20 and 100 microM. The on (k'on) and off (k(off)) rate coefficients for the CDPC blocking reaction were k'on = 11.1 +/- 0.8 rad.s-1.microM-1 and k(off) = 744 +/- 53 rad/s, and the microscopic inhibition constant was 67 microM (n = 11). CDPC blocking kinetics were not significantly different after inhibition of Isc by 5 microM serosal carbachol. Single-channel Na+ current (iNa) and the density of open and blocked Na+ channels (N(ob)) were estimated from the fluctuations induced by 40 microM CDPC. Under control conditions, iNa was 0.43 +/- 0.05 pA and N(ob) was 251 +/- 42 X 10(6)/cm2 (n = 10). After exposure to serosal carbachol (2-10 microM) for 60 min, Na+ current and N(ob) were reduced by approximately 50%, but iNa was not changed significantly. These results indicate that muscarinic inhibition of electrogenic Na+ absorption was associated with a reduction in the number of open Na+ channels in the apical membrane. They also suggest that this downregulation of transport involved a coordinated decrease in both apical and basolateral membrane conductances.

Studies on Locust Rectum: II. Identification of Specific Ion Transport Processes Regulated by Corpora Cardiaca and Cyclic-AMP

1980 ◽  
Vol 86 (1) ◽  
pp. 225-236
Author(s):  
J. H. SPRING ◽  
J. E. PHILLIPS
Keyword(s):  
Cyclic Amp ◽  
Ion Transport ◽  
Time Course ◽  
Transport Processes ◽  
Electrogenic Transport ◽  
Corpora Cardiaca ◽  
Before And After ◽  
Net Flux ◽  
Unidirectional Fluxes ◽  
Stimulation Of

Please send reprint requests to J. E. Phillips. 1. The unidirectional fluxes of 36Cl− and 22Na+ across short-circuited locust recta bathed in a simple NaCl saline were followed with time. Unidirectional fluxes and net flux of 22Na+ to the haemocoel side all remained constant for at least 4 h and were unaffected by either corpora cardiaca homogenate (CC) or cAMP. 2. Both CC and cAMP stimulated influx and net flux of 36Cl− to the haemocoel side. Over the whole time course of the experiment, i.e. both before and after stimulation, net Cl− flux approximately equalled the shortcircuit current (ISC). 3. Neither CC nor cAMP caused substantial stimulation of ISC or transepithelial electropotential difference (PD) if all Cl− in the bathing saline was replaced by either sulphate or nitrate or acetate. 4. Acetate saline sustains ISC, PD and transepithalial resistance (R) at higher levels than does simple Cl-saline. 5. Experiments with Cl-free, SO4-salines suggest that alternate electrogenic transport processes can be slowly turned on when Cl− is absent, provided a complex saline which contains several organic constituents, or simple acetate saline, is present.

Electrophysiology and noise analysis of K+-depolarized epithelia of frog skin

1985 ◽  
Vol 249 (5) ◽  
pp. C421-C429 ◽  
Author(s):  
J. Tang ◽  
F. J. Abramcheck ◽  
W. Van Driessche ◽  
S. I. Helman
Keyword(s):  
Frog Skin ◽  
Single Channel ◽  
Apical Membrane ◽  
Short Circuit ◽  
Noise Analysis ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Membrane Resistance ◽  
Na Channel ◽  
Channel Density

Epithelia of frog skin bathed either symmetrically with a sulfate-Ringer solution or bathed asymmetrically and depolarized with a 112 mM K+ basolateral solution (Kb+) were studied with intracellular microelectrode techniques. Kb+ depolarization caused an initial decrease of the short-circuit current (Isc) with a subsequent return of the Isc toward control values in 60-90 min. Whereas basolateral membrane resistance (Rb) and voltage were decreased markedly by high [Kb+], apical membrane electrical resistance (Ra) was decreased also. After 60 min, intracellular voltage averaged -27.3 mV, transcellular fractional resistance (fRa) was 86.8%, and Ra and Rb were decreased to 36.1 and 13.0%, of their control values, respectively. Amiloride-induced noise analysis of the apical membrane Na+ channels revealed that Na+ channel density was increased approximately 72% while single-channel Na+ current was decreased to 39.9% of control, roughly proportional to the decrease of apical membrane voltage (34.0% of control). In control and Kb+-depolarized epithelia, the Na+ channel density exhibited a phenomenon of autoregulation. Inhibition of Na+ entry (by amiloride) caused large increases of Na+ channel density toward saturating values of approximately 520 X 10(6) channels/cm2 in Kb+-depolarized tissues.

Modification of carboxyl of Na+ channel inhibits aldosterone action on Na+ transport

1983 ◽  
Vol 245 (6) ◽  
pp. F726-F734 ◽  
Author(s):  
J. Kipnowski ◽  
C. S. Park ◽  
D. D. Fanestil
Keyword(s):  
Amphotericin B ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Na Channel ◽  
Dependent Manner ◽  
Na Channels ◽  
Na Transport ◽  
Maximal Increase ◽  
Osmotic Water ◽  
Stimulation Of

We investigated the effect of the carboxyl-selective reagent N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) on aldosterone stimulation of Na+ transport in the urinary bladder of the toad. Na+ transport, measured as the short-circuit current (SCC), was irreversibly inhibited by EEDQ in a dose- and time-dependent manner prior to addition of aldosterone. The greater the percentage inhibition by EEDQ (X), the smaller was the maximal increase of SCC after aldosterone (Y). This relationship gave the regression equation Y = 128.41 - 1.73X, r = -0.99 (n = 35). Evidence that the inhibition of SCC produced by EEDQ was limited to effects at the mucosal membrane was attested by the following: 1) EEDQ did not alter the stimulation by aldosterone of the osmotic water flow response to antidiuretic hormone; 2) whereas inhibition of protein synthesis by cycloheximide prevented this effect of aldosterone; 3) amphotericin B fully restored SCC previously inhibited by EEDQ to the level produced in tissues not inhibited by EEDQ; 4) comparison of the effects of amiloride vs. EEDQ pretreatment on the SCC response to aldosterone and amphotericin B revealed nearly identical characteristics; 5) in contrast, amphotericin B stimulation of SCC was limited when Na+ transport was limited by antimycin A (an inhibitor of energy production) or by ouabain. The findings fail to provide positive evidence for the hypothesis that aldosterone induces the synthesis of new Na+ channels but are consistent with hormonal activation of previously existing but nonfunctioning Na+ channels.

Regulation of electrogenic Na+ transport across leech skin

1995 ◽  
Vol 268 (3) ◽  
pp. R605-R613 ◽  
Author(s):  
W. M. Weber ◽  
U. Blank ◽  
W. Clauss
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Na Channel ◽  
Monovalent Cations ◽  
Serosal Side ◽  
Na Transport ◽  
Permeability Ratio ◽  
Cyclic Monophosphate ◽  
Apical Side ◽  
Stimulation Of

The dorsal integument of the medical leech Hirudo medicinalis exhibits a marked amiloride-sensitive Na+ absorption. With 20 mM Na+ in the apical solution, the transepithelial short-circuit current (Isc) was approximately 40% higher than with 115 mM Na+, whereas the transepithelial potential (VT) with 20 mM Na+ was -35.7 +/- 4.5 and -20.6 +/- 2.6 mV with 115 mM Na+. Amiloride (100 microM) inhibition at 20 mM apical Na+ was also significantly larger than with 115 mM Na+ in the solution. Benzamil (100 microM) showed additional inhibition after amiloride. Large transient overshooting currents occurred only when 115 mM Na+ was added after some minutes of Na(+)-free apical solution. Addition of adenosine 3',5'-cyclic monophosphate (cAMP) to the serosal side in the presence of 115 mM apical Na+ nearly doubled Isc. This cAMP effect was reduced to only 20% in the presence of 20 mM Na+. Guanosine 3',5'-cyclic monophosphate (cGMP) slightly increased Isc, whereas ATP showed biphasic potency. Removal of calcium from the apical side resulted in a large stimulation of amiloride-sensitive Isc only in the presence of 115 mM Na+. When currents were activated with cAMP, a deprivation of Ca2+ modestly reduced the amiloride-sensitive Isc. The Na+ channel of leech integument was found highly selective for Na+ over other monovalent cations. The permeability ratio for Na+ over K+ was approximately 30:1; the selectivity relationship for the investigated cations was Na+ > Li+ > NH4+ > K+ approximately Cs+ approximately Rb+.

Active and electrogenic absorption of Na + and Cl- across posterior gills of Eriocheir sinensis: influence of short-term osmotic variations

1996 ◽  
Vol 199 (4) ◽  
pp. 901-910 ◽  
Author(s):  
H Onken
Keyword(s):  
Electromotive Force ◽  
Marked Decrease ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Eriocheir Sinensis ◽  
Induced Current ◽  
Na Current ◽  
Short Term ◽  
External Barrier ◽  
Short Circuit Currents

Split lamellae of posterior gills of Chinese crabs (Eriocheir sinensis) acclimated to fresh water were mounted in a modified Ussing-type chamber, and the transepithelial short-circuit current and conductance were measured. The epithelium shows independent active and electrogenic absorption mechanisms for Na+ and Cl- that can be measured as positive and negative short-circuit currents, respectively, in the absence of the counter ion. Increasing the osmolarity of the haemolymph-side saline by addition of sucrose resulted in a marked decrease in active uptake of both Na+ and Cl-. In contrast, increasing the internal osmolarity by addition of urea or moderately decreasing the haemolymph-side osmolarity resulted in a marked increase in Na+ as well as Cl- transport. Circuit analysis revealed that Na+ current changes are mostly due to alterations in the apical amiloride-sensitive Na+ conductance, while Cl- current changes are caused not only by alterations in the transcellular conductance but also by changes in the electromotive force for Cl- absorption. Osmotic perturbations in the external bath induced current changes in the same directions, but the magnitudes of the effects were smaller than those after internal osmotic variations, indicating that the external barrier has a lower water permeability than the internal barrier. Short-term osmotic perturbations did not significantly affect the leak conductance, which is not associated with active transport and which may mostly reflect the paracellular conductance.

Studies on Locust Rectum: I. Stimulants of Electrogenic Ion Transport

1980 ◽  
Vol 86 (1) ◽  
pp. 211-223
Author(s):  
J. H. SPRING ◽  
J. E. PHILLIPS
Keyword(s):  
Steady State ◽  
Cyclic Amp ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Lag Time ◽  
Camp Concentration ◽  
Maximum Increase ◽  
Electrogenic Ion Transport ◽  
The Relationship ◽  
Stimulation Of

1. Homogenates of whole corpora cardiaca (CC) cause increases in the short-circuit current (Isc) and transepithelial electropotential difference (PD) across locust recta of 3-fold and 1.7-fold respectively, in comparison with the values for unstimulated steady-state recta. Maximum stimulation restores rectal ISC and PD to levels observed immediately after removing this organ from animals. 2. Cyclic-AMP causes a similar maximum increase in ISC and PD; however, the response exhibits a much shorter lag time and a faster rate of rise than is observed for stimulation with CC. 3. The addition of CC to the haemocoel side of everted rectal sacs caused whole tissue levels of cAMP in this organ to increase 3-fold. 4. The relationship between the logarithm of CC or cAMP concentration and the increase in ISC is linear, and the decline in ΔISC with time is also dosedependent. 5. Small maximum increases in ISC are caused by homogenates of ventral ganglia, whole brain and rectal tissue, but the concentration of the stimulatory activity in these locust tissues is clearly three orders of magnitude lower than in CC. 6. Inhibitors of HCO3—/H+ and Cl− transport in vertebrate systems, acetazolamide and thiocyanate, do not inhibit the stimulation of recta by CC or cAMP.

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