Toad Urinary Bladder Epithelium Research Articles

Microelectrode studies in toad urinary bladder epithelium. effects of Na concentration changes in the mucosal solution on equivalent electromotive forces

Microelectrode studies in toad urinary bladder epithelium. effects of Na concentration changes in the mucosal solution on equivalent electromotive forces

Cellular and shunt conductances of toad bladder epithelium.

Toad urinary bladders were mounted in Ussing-type chambers and volt-age-clamped. At nonzero voltages only, small fluctuations in current, delta I, and therefore in tissue conductance, delta Gt, were detected. These fluctuations were caused by the smooth muscle of the underlying tissue which could be monitored continuously and simultaneously with the current, I. Inhibition of the smooth muscle contraction with verapamil (2 X 10(-5) M) abolished the fluctuations in I and Gt. Amiloride (10(-4) M) had no significant effect on the magnitude of delta Gt, oxytocin increased Gt without affecting delta Gt, and mucosal hypertonicity produced by mannitol increased delta Gt. These results are consistent with the hypothesis that two parallel pathways exist for passive current flow across the toad urinary bladder: one, the cellular pathway, was not affected by smooth muscle activity; the other, the paracellular pathway, was the route whose conductance was altered by the action of the smooth muscle. Thus the relationship between the cellular and shunt conductances of the epithelium of the toad urinary bladder, under a variety of conditions, can be investigated by utilizing the effects of the movement of the smooth muscle.

Dual effects of amphotericin B on ion permeation in toad urinary bladder epithelium.

The mechanisms of action of amphotericin B on the electrical properties of the toad urinary bladder epithelium were studied with microelectrode techniques. Cell membrane and transepithelial electrical potentials and resistances were measured in the absence and in the presence of the drug during exposure to bathing mediums of different ionic compositions. As observed previously by other investigators, amphotericin B produces a dramatic decrease of transepithelial electrical resistance (Rt) and an increase of the rate of sodium transport. Our results indicate that the effect of the drug on Rt depends in part on an increase in Na conductance across the luminal cell membrane (amiloride-insensitive), but is caused mainly by an increase of ionic conductances (with the sequence GK greater than GNa greater than G choline greater than GCI) at a site in parallel with the impaled cells (i.e., across a cell type not investigated by the microelectrode measurements or across the paracellular pathway.

Neurohypophyseal peptide action on andenylate cyclase and hydro-osmotic properties of toad urinary bladder epithelium

The relationship between increase in water permeability in intact urinary bladder of toad, Bufo marinus, and activation of its membrane-bound adenylate cyclase preparation in response to neurohypophyseal peptides was examined. All of the peptides tested produced a maximal hydro-osmotic response in the intact bladder; in the adenylate cyclase assay the intrinsic activities of peptides varied greatly. While there is a positive qualitative correlation ( r s = 0.85, P < 0.01) between the apparent affinity parameters (p D 2) of the peptides obtained in the adenylate cyclase and hydro-osmotic assays, there is a difference in peptide sensitivity between the two assays of one to four orders of magnitude. The magnitude of this difference increases as the intrinsic activity of the peptides increases. The differences in peptide affinity observed in the two assay systems as well as the lack of correlation between intrinsic activities as measured in subcellular and membrane preparations can in part be explained on the basis that maximal hormonal response in intact tissue may require only a small franction (5% or less) of the cyclic AMP produced in response to maximally stimulating concentrations of hormone or analog. On this basis, the greater the potency of a peptide in generating cyclic AMP, the greater will be the discrepancy between the peptide concentration required to produce half-maximal responsens in the two assays.

Effects of changes in the composition of the mucosal solution on the electrical properties of the toad urinary bladder epithelium.

By the use of microelectrode techniques, the potential profile and the electrical resistances of the cellular and shunt pathways across the toad urinary bladder epithelium were measured under control conditions and after exposing the mucosal side to solutions of low and high NaCl concentrations and osmolalities. The resistance of the shunt pathway increases at low NaCl concentration (even if the osmolality is kept constant), and decreases at high NaCl concentration (by a nonspecific osmotic mechanism). The inverse relationship between mucosal NaCl concentration and shunt resistance suggests a regulatory mechanism of net sodium transport by reduction of the passive blood-to-urine sodium flux at low urinary sodium concentrations. In addition, the transepithelial potential and the potentials at both cell borders fall in both low and high mucosal NaCl, and the magnitude of these changes is such that they cannot be explained by changes in the shunt pathway alone.

Effects of changes in the composition of the serosal solution on the electrical properties of the toad urinary bladder epithelium.

1. The potential profile and the cellular and paracellular transepithelial resistances of the toad urinary bladder were measured, by means of micro-electrode techniques, as functions of the osmolality of the serosal solution. 2. Reductions in serosal osmolality (that increase the rate of active sodium transport) produced proportional decreases in the electrical resistances of the apical and basal-lateral cell membranes, while the changes in resistance of the paracellular pathway were more complex. The apical membrane potential increased. 3. Increases in serosal osmolality (that decrease sodium transport) produced increases in the electrical resistances of both cell membranes, and moderate reduction in the paracellular resistance. The polarity of the apical membrane potential reversed. 4. These results indicate that reductions in serosal solution osmolality stimulate sodium transport by increasing both the sodium permeability of the luminal cell membrane (thus increasing sodium entry), and the electromotive force generated at the serosal border of the cell, thus enhancing the rate of sodium pumping. Conversely, increases in osmolality reduced sodium transport by reducing both the sodium permeability of the luminal membrane and the serosal membrane electromotive force.

Dependence of serosal membrane potential on mucosal membrane potential in toad urinary bladder

Ussing's model for sodium transport across epithelia assumes the existence of two barriers in series; the outer barrier behaves as a sodium electrode and the inner barrier as a potassium electrode (Koefoed-Johnsen and Ussing, 1958); furthermore, these membranes are supposed to be independent of one another. Although experimental determinations of the changes in transepithelial potential across frog skin and toad urinary bladder after ionic substitutions agree with this view (Koefoed-Johnsen and Ussing, 1958; Gatzy and Clarkson, 1965; Leb et al., 1965), microelectrode experiments have shown that, in the steady state, unilateral changes in the composition of the bathing solution alter the potential difference across both barriers in frog skin (Cereijido and Curran, 1965) and Amphiuma distal tubule (Wiederholt and Giebisch, 1974). However, an alternative explanation for these observations would be that significant alterations in intracellular ionic activities took place. Accordingly, it has been proposed that fast changes in the composition of the outer bathing solution in frog skin or toad urinary bladder produce changes in the transepithelial potential by altering only the potential drop across the outer barrier (Lindemann and Gebhardt, 1973). The present experiments were designed to study the alterations in potential profile across the toad urinary bladder epithelium during changes of the composition of the mucosal bathing solution. Urinary bladders from Colombian toads were studied as previously described (Reuss and Finn, 1974). The solutions employed were standard amphibian Ringer (in millimoles per liter, NaCl 109, NaHCO3 2.4, CaC12 0.9, KCI 2.5, glucose 5.5), potassium-Ringer (equimolar K-for-Na substitution), or Ringer plus 10-5 or 10-4 M amiloride. Short current pulses were passed transepithelially in order to measure the total transepithelial resistance and the ratio of the apical to basal-lateral membrane resistances. The mucosal solution was changed by gravity superfusion through a glass pipette with the microelectrode in a cell.

Passive electrical properties of toad urinary bladder epithelium. Intercellular electrical coupling and transepithelial cellular and shunt conductances.

The electrical resistances of the transcellular and paracellular pathways across the toad urinary bladder epithelium (a typical "tight" sodium-transporting epithelium) were determined by two independent sets of electrophysiological measurements: (a) the measurement of the total transepithelial resistance, the ratio of resistance of the apical to the basal cell membrane, and cable analysis of the voltage spread into the epithelium; (b) the measurement of the total transepithelial resistance and the ratio of resistances of both cell membranes before and after replacing all mucosal sodium with potassium (thus, increasing selectively the resistance of the apical membrane). The results obtained with both methods indicate the presence of a finite transepithelial shunt pathway, whose resistance is about 1.8 times the resistance of the transcellular pathway. Appropriate calculations show that the resistance of the shunt pathway is almost exclusively determined by the zonula occludens section of the limiting junctions. The mean resistance of the apical cell membrane is 1.7 times that of the basal cell membrane. The use of nonconducting materials on the mucosal side allowed us to demonstrate that apparently all epithelial cells are electrically coupled, with a mean space constant of 460 microm, and a voltage spread consistent with a thin sheet model.

Ultrastructural observations on the aldosterone-stimulated toad urinary bladder epithelium—ATPase activity

Ultrastructural evidence is presented indicating an increase in membrane-bound ATPase activity of the mucosal epithelium of the toad urinary bladder following exposure of its serosal surface to aldosterone. Although no conclusion could be made as to whether the observed increase in ATPase activity was a direct affect of AIP, or an event secondary to an effect of AIP on other components of the cells, various possibilities are discussed in terms of current physiological information regarding the effects of aldosterone on the epithelium of the isolated toad urinary bladder.

Cyclic 3',5'-AMP-dependent protein kinase activity in toad bladder epithelium.

SummaryA cyclic AMP-dependent protein kinase was partially purified from toad urinary bladder epithelium. Cyclic AMP-stimulated protein kinase activity as well as the unstimulated (control) activity of the enzyme was maximal at pH 6.5 using Tris-maleate, sodium cacodylate or sodium acetate buffers. The apparent Km value of cyclic AMP for the stimulation of the enzyme (in sodium acetate buffer and in the presence of 10 mM Mg2+) was 1.5 × 10-9M. The acyclic nucleotide 5'-AMP (in the concentration range 5 × 10-10 to 5 × 10-5M) failed to activate bladder protein kinase. Ca2+ was found to be inhibitory at concentrations higher than 5 × 10-4M. A role for cyclic AMP-dependent protein kinase in neurohypophyseal hormone-induced permeability changes is discussed.

Effects of aldosterone and vasopressin on adenyl cyclase activity of rat kidney.

ARTICLESEffects of aldosterone and vasopressin on adenyl cyclase activity of rat kidneyMA Lang, and IS EdelmanMA Lang, and IS EdelmanPublished Online:01 Jan 1972https://doi.org/10.1152/ajplegacy.1972.222.1.21MoreSectionsPDF (992 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInEmailWeChat Previous Back to Top Next Download PDF FiguresReferencesRelatedInformationCited ByThe nature of the increased sensitivity to injected GTP of the sodium efflux in barnacle muscle fibers pre-exposed to aldosteroneComparative Biochemistry and Physiology Part A: Physiology, Vol. 77, No. 1Membrane Recognition and Effector Sites in Steroid Hormone ActionThe Actions of AldosteroneMolecular Mechanisms of Steroid Hormone ActionThe Significance of the Hairpin Counter-current Principle in the Pathogenesis of Toxic Kidney Lesions An Investigation of the Influence of Antidiuretic Hormone and Papaverine on Hydroxyquinoline Nephropathy of the RatBeiträge zur Pathologie, Vol. 160, No. 2Cellular actions of vasopressin in the mammalian kidneyKidney International, Vol. 10, No. 1Drugs and Other Agents Affecting the Renal Adenylate Cyclase SystemThe mechanism of aldosterone functionPharmacology & Therapeutics. Part B: General and Systematic Pharmacology, Vol. 2, No. 2Renal adenylate cyclase — effects of diureticsEuropean Journal of Pharmacology, Vol. 34, No. 2The Relationship of Antidiuretic Hormone to the Control of Volume and Tonicity in The HumanContribution of long chain fatty acids to the energy supply of the rat kidney cortexPfl�gers Archiv European Journal of Physiology, Vol. 356, No. 1The Vasopressin-sensitive Adenylate Cyclase of the Rat KidneyJournal of Biological Chemistry, Vol. 249, No. 20Relationship of Binding to Biological ActivityUltrastructural observations on the aldosterone-stimulated toad urinary bladder epithelium—ATPase activityJournal of Ultrastructure Research, Vol. 44, No. 5-6 More from this issue > Volume 222Issue 1January 1972Pages 21-24 Copyright & PermissionsCopyright © 1972 by American Physiological Societyhttps://doi.org/10.1152/ajplegacy.1972.222.1.21PubMed4334058History Published online 1 January 1972 Published in print 1 January 1972 Metrics

Epinephrine antagonism of insulin-dependent glucose uptake in the toad bladder.

The serosal epithelium of the toad urinary bladder possesses an insulin-dependent glucose uptake system. Epinephrine in concentrations of 9 μg/ml antagonizes this action of insulin at a concentration of 227 mU/ml, significantly reducing glucose uptake. Similar results have been noted in mammalian muscle. The morphologic characteristics of this membrane may make it a suitable structure for the study of the mechanism of action and interrelationships of these hormones. (Endocrinology 80: 788, 1967)