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R C de Sousa

Publications and source records attributed to R C de Sousa.

At least 19 recordsLinked to original sources
PubMed1993

Effects of anions and/or cell volume on the permeance of an apical water pathway induced by Hg in toad skin epithelium.

Hg compounds block membrane transport units behaving as water channels. Here we show that Hg induces an apical water pathway in toad skins pretreated with 10(-3) M CH3ClHg or HgCl2, added to the outer bathing medium. Washing with SO4-Ringer caused a several-fold increase in net water flow (Jw) and osmotic permeability coefficient (Pf) that was reversed by re-exposure to Cl- or NO3-Ringer and mimicked by gluconate-Ringer. These Pf changes could be elicited repeatedly and were present if, and only if, anion replacements took place in the inner bathing solution. Such inner polarity was related to the anion permeability of the epidermal basolateral membrane: impermeant anions (SO4, gluconate) increased Pf; permeant anions (Cl, NO3) did not change basal Pf but reversed the high Pf induced by impermeant anions. Hg induced the appearance of aggregates that persisted despite repeated washings of the skins during 4-5 h, and whether Pf was high (SO4-Ringer) or low (Cl-Ringer) before skin fixation. The Hg-induced apical water pathway in toad skin appears to be a unique model for studying the interplay between cell volume, cell ionic composition and water permeability.

Animals↗
PubMed1986

Effects of vasopressin on epithelial transport.

Vasopressin increases the permeability of responsive epithelia for a variety of chemical species, such as sodium, urea, water, and lipophilic molecules. The analysis of the mechanisms leading to the stimulation of water transport (hydrosmotic action) is the main topic of this review. A model implying the quantal insertion of water channels into the apical epithelial membrane by an exocytosislike process is discussed with reference to biophysical, biochemical, and ultrastructural results obtained in the past few years. It is also shown that, in nonexcitable cells, insertion of transport units packed in intracytoplasmic structures may be a rather general mechanism for controlling membrane transport rates.

Animals↗
PubMed1984

High [K+] alters the stimulus-hydrosmotic response coupling in toad bladder.

Substitution of K+ for Na+ in the Ringer solution bathing the inner surface of toad urinary bladders (Bufo marinus) had no effect on basal water permeability but significantly altered the stimulus-hydrosmotic response of this epithelium. In chloride-Ringer, high [K+] increased the hydrosmotic responses to submaximal stimulations induced by vasopressin or exogenous cAMP, while the responses to theophylline or serosal hypertonicity were decreased. In sulfate-Ringer, all these responses were enhanced but for that induced by serosal hypertonicity which was actually diminished. As a step towards determining if Ca2+ might mediate the K+-induced effects on water flow, experiments were conducted either in the presence of a Ca2+ "antagonist" (cobalt) or in nominally Ca2+-free Ringer. In both conditions the hydrosmotic effects of vasopressin and cAMP were markedly reduced. The results raise the possibility that a transient Ca2+ influx through voltage-sensitive, Co2+-blockade Ca2+ channels may play a role in the stimulus-hydrosmotic response of toad urinary bladder.

Animals↗
PubMed1982

Cellular and membrane events involved in the K-induced increase in water permeability of toad skin.

Exposure of the inner surface of toad skin (Bufo marinus) to high [K+] resulted in a marked (up to 7-fold) increase in water permeability (Pf) that was more marked in KC1-Ringer than in K2SO4-Ringer. Although high [K+] did not elicit a maximal increase in Pf, it blunted the hydrosmotic responses to vasopressin, isoproterenol and cAMP. Both "post-cAMP" inhibitors of stimulated water flow, such as diamide and vanadate, and "pre-cAMP" inhibitors, such as methohexital and propranolol, markedly reduced the K response, while exposure to Ca2+-free, KC1-Ringer did not inhibit water flow. Intramembrane particle aggregates, similar to those induced by cAMP-mediated hydrosmotic agents, were seen in the apical membrane of granular cells, just beneath the stratum corneum, in skins exposed to KC1. Available evidence indicates that cAMP might mediate, at least partially, the hydrosmotic effect of high [K+]. In contrast, a role of voltage-dependent Ca2+ channels, described in other cell systems depolarized with K, was not apparent in toad skin.

Animals↗
PubMed1982

Evidence for a role of calmodulin in the hydrosmotic action of vasopressin in toad bladder.

1. The informational role of cytosolic Ca2+ appears to be mediated by a ubiquitous protein--calmodulin--in most cell systems. 2. Evidence has been accumulating that not only cAMP, but also Ca2+, behaves as an intracellular messenger in the stimulation of water transport by vasopressin (hydrosmotic effect). 3. To examine whether calmodulin plays a role in the hydrosmotic effect of vasopressin, we used a specific antagonist of calmodulin--trifluoperazine--and looked at its effects on water transport in the urinary bladder of toads Bufo marinus. 4. The results showed that trifluoperazine, at micromolar concentrations, blocked the hydrosmotic effects of vasopressin or cAMP, thus indicating a post-cAMP site of action. 5. Two other psychotropic drugs--amitriptyline and harmaline--had similar effects, but higher concentrations were required to induce the same degree of inhibition of water flow. 6. Calmodulin was detected in the membrane and in the cytosolic fractions of isolated epithelial cells of toad bladder by means of the phosphodiesterase test. The content of both fractions was similar to that found in bovine brain. 7. The results suggest that calmodulin plays a regulatory role in the hydrosmotic action of vasopressin by possibly interacting with proteins associated with microfilaments and/or microtubules.

Amitriptyline↗
PubMed1981

Quercetin enhances water transport in toad bladder.

A highly significant enhancement of the hydrosmotic actions both of vasopressin and of exogenous cAMP was seen in the presence of quercetin. The hypothesis is advanced that quercetin affects the intracellular coupling between Ca++ in cAMP.

Animals↗
PubMed1981

The mode of action of vasopressin: membrane microstructure and biological transport.

Vasopressin affects a variety of cell systems. This review is focused on permeability changes induced by vasopressin in tight epithelia such as the collecting duct of the mammalian kidney and the skin and the bladder of anurans. These vasopressin effects are discussed with reference to current concepts and models of the microstructure of the plasma membrane. The transport of three major chemical species--Na, urea and water--is analyzed. In each instance, the hormone appears to activate selective membrane pathways situated at the rat-limiting barrier of the epithelium, i.e., the apical membrane. Available data suggest that two intra-cellular messengers -- cAMP and calcium -- plan a key role in the coupling between stimulus (receptor occupancy) and biological effect (permeability change). The enhancement of Na transport (natriferic effect) depends on the opening and/or the insertion of Na channels, the biophysical and biochemical characteristics of which have been investigated by fluctuation analysis and by means of several chemical blockers of Na transport, particularly the amiloride molecule and its congeners. Likewise, the finding of inhibitors and activators of urea transport, which do not cause any appreciable change in Na or water permeability, led to the notion of selective urea channels or pores. Finally, the enhancement of water transport (hydrosmotic effect) possibly results from the insertion in the apical membrane of water channels already present in vesicular cytoplasmic structures. The restructuring of the apical membrane underlying the transition from a low to a higher state of water permeability is very likely related to the appearance of intramembrane particle aggregates detectable with the freeze-fracture technique in epithelia exposed to vasopressin. The putative water channels (or pores) appear to be so narrow that trans-apical water movement is constrained to single-file diffusion. Recent data also suggest that, in addition to cAMP, microtubules and microfilaments, the calmodulin-Ca complex is a major element in the hydrosmotic effect of vasopressin.

Amphibians↗
PubMed1980

Isoproterenol-induced intramembrane particle aggregation and water flux in toad epidermis.

Stimulation of toad skin with isproterenol resulted in a dramatic increase in water flow, and in the appearance of aggregates of intramembrane particles in the apical membrane of granular cells of the replacement layer, just beneath the stratum corneum. This membrane structural modification appears to be a general prerequisite for the change in water permeability of vasopressin-sensitive epithelia.

Animals↗
PubMed1979

Inhibitory and stimulatory effects of amiloride analogues on sodium transport in frog skin.

Effects of amiloride analogues on Na transport were studied in isolated skins of the frog Rana ridibunda. The pattern of structure-activity relationship of these compounds showed that both the -NH2 group at position 5 and Cl at position 6 of the pyrazine ring of the amiloride molecule were important for their biological activity. The paramount role of the groups at position 5 was further demonstrated by the striking properties of an analogue resulting from dimethylation of that -NH2 group. A stimulation of Na transport, opposite to the effect of amiloride itself, was observed in this instance. The increase in Na transport could already be seen at 10(-6) M and was equivalent to the measured increase in Na influx, reversible, dose-dependent, and additive to the natriferic action of oxytocin. Such characteristics resemble those reported with "external" agents like propranolol and La3+. Furthermore, mutual inhibition was observed between the stimulatory effects of this analogue and those of propranolol or La3+. These results suggest that the analogue may be considered as another "external" agent acting at sites of the external membrane distinct from those activated by cAMP but similar to the Ca sites described by Herrera and Curran (Herrera, F.C., Curran, P.F. 1963. J. Gen. Physiol. 46:999).

Amiloride↗
PubMed1978

Cytochalasin B and water transport. A scanning electron microscope study of the toad urinary bladder.

A morpho-functional study of the effects of cytochalasin B (CB) on Na and water transport was made in amphibian epithelia. The functional studies confirmed the dissociation of the natriferic and hydrosmotic effects of vasopressin in toad urinary bladders exposed to CB and showed in addition that the block of the hydrosmotic effect was reversible and could still be induced in epithelia maximally stimulated with the hormone. Scanning electron microscopy revealed that CB, per se, did not alter the apical surface of the bladders. An almost total loss of microvilli of granular cells was seen, however, if CB was associated with vasopressin and an osmotic gradient. The results suggest two points: a) the block of the hydrosmotic flow induced by CB is due to factors beyond the apical membrane; b) microfilaments may be important mechanochemical transducers in the chain of events leading to the hydrosmotic effect of vasopressin.

Animals↗
PubMed1978

Vasopressin-like effects of a hallucinogenic drug--harmaline--on sodium and water transport.

To determine if harmala alkaloids affect transport systems other than (Na +K)-ATPase, effects of harmaline on Na and water fluxes were studied in amphibian skins. Net Na flux was evaluated from short-circuit current, and water flux monitored with automatic, volumetric methods. At 2 to 5 mM, harmaline consistently inhibited SCC and prevented the natriferic effects of oxytocin and norepinephrine. However, at 0.1 to 0.5 mM, harmaline produced an increase in SCC inhibitable with amiloride. The stimulatory effects of harmaline and oxytocin were either nonadditive or additive depending on whether the hallucinogen was present in the inner solution or in the outer solution bathing the skin, respectively. Water flow was not modified by harmaline on the outer medium. In contrast, addition of the drug to the inner medium elicited a conspicuous, sustained, vasopressin-like, hydrosmotic effect, comparable to and competive with those of vasopressin and norepinephrine. The ensemble of these results suggests that harmaline may affect three distinct transport systems: (i) the Na pump; (ii) the cyclic nucleotide system; (iii) the Na entry pathway at the outer membrane of the skin that is also activated by agents such as diphenylhydantoin, lanthanides and propranolol.

Alkaloids↗
PubMed1978

Vasopressin-like effects of psychotropic drugs in amphibian epithelia.

Amphibian epithelia have been used as models for studying the effects of psychotropic drugs on membrane transport. Several of these agents added to the internal or to the external media, at concentrations greater than 10(-3) M, had inhibitory, "ouabain-like" effects on Na transport. In contrast, stimulatory, "vasopressin-like" effects were seen at lower concentrations. The stimulation was additive to that of oxytocin if the drug was present in the external solution but nonadditive if in the internal solution. On water transport, harmala alkaloids had a vasopressinomimetic action in toad skin, while inhibition was seen with Li and amitriptyline. To account for these multiple effects, it is hypothesized that psychotropic drugs act on the following cell targets: the Na pump, the cyclic nucleotide system, microtubules, and membrane calcium sites at the outer barrier of the epithelium. Direct, biochemical evidence is needed to substantiate this hypothesis.

Amitriptyline↗
PubMed1978

Cyclic AMP levels in isolated frog skin epithelium: effects of phosphodiesterase inhibitors, oxytocin and catecholamines.

Direct measurements of cyclic AMP were performed in the isolated epithelium of frog skin. Phosphodiesterase inhibitors (methylxanthines, papaverine) and activators of adenylyl cyclase (oxytocin, catecholamines) significantly increased the cyclic AMP content. Propranolol completely blocked the generation of cAMP induced by beta-adrenergic agonists but had little or no effect on that induced by oxytocin. Phentolamine enhanced the cAMP production by adrenalin and noradrenalin. At supramaximal concentrations, oxytocin and isoproterenol produced similar increments in cAMP, while exposure to both agents roughly doubled the increase in cAMP. The results suggest the presence of independent receptors for oxytocin and catecholamines in frog skin, with additive effects on cAMP generation.

Animals↗
PubMed1977

[The cell membrane: a frontier between 2 worlds].

The structure and function of the plasma membrane is reviewed. Three of its main functional roles are discussed, i.e. of 1. selective, physico-chemical barrier; 2. key regulator of cell tolerance and cell recognition; 3. effector organelle for long range and short range cell communication. From the structural viewpoint, the fluid mosaic model is presented as a particularly dynamic picture of the membrane machine and its components (pathways for active and passive transport, enzymes, hormonal receptors and other active sites). Some recent aspects fo the modulation of membrane permeability by hormonal or pharmacological agents are discussed by considering two examples of the author's own research: the hydrosmotic action of vasopressin and the vasopressin-like effects of a hallucinogenic drug, harmaline. Finally, the growing list of anomalies of membrane transport and membrane receptors found both in rare and in common clinical entities emphasizes the importance of membrane research in the understanding of human disease.

Animals↗