Cyclic AMP and intracellular ionic activities innecturus gallbladder

Abstract

Open-tip and liquid ion-exchanger microelectrodes were used to study the effects of cAMP (6mM, added to the serosal medium) on apical membrane potential (Em) and intracellular sodium, potassium, and chloride activities (alphaiNa, alphaiK, alphaiCl) in Necturus gallbladder under open-circuit conditions. Transepithelial potential difference (ETr) was also measured. In the presence of cAMP, alphaiCl fell from about 1.5 times its equilibrium value to a level that corresponded to electrochemical equilibrium across the apical and basolateral cell membranes. Under these conditions alphaiNa decreased and alphaiK increased, Em was unchanged and ETr increased from virtually zero to a small but significant serosal positive value. The cAMP-induced increase in alphaiK was abolished when Cl- -free incubation media were used. Addition of the Ca++-ionophore A23187 (0.5 microgram/ml) to the serosal medium had no effect on Em, ETr, or alphaiCl. When A23187 was added to the mucosal medium, Em and the basolateral membrane potential hyperpolarized by about 20 mV and an increase in the outwardly directed electrochemical driving force for Cl- was observed. These results indicate that cAMP inhibits coupled transapical Na-Cl entry into epithelial cells of Necturus gallbladder and suggest that this inhibition may not be mediated by an increase in intracellular Ca++ concentration.

Cyclic AMP-induced changes in membrane conductance ofNecturus gallbladder epithelial cells

“…It seems reasonable to suppose that, had the tissues been exposed to IBMX for longer times than those corresponding to the data of Table 2, a~l would have reached a level that did not differ from the corresponding a~ values. Thus, the results shown in Table 2 are in essential agreement with those reported for the action of cAMP, 8-Br-cAMP and theophylline on abl in Necturus gallbladder (Diez de los Rios et al, 1981;Petersen & Reuss, 1983). …”

“…The average a~i value for Necturus gallbladder under control conditions (21.7 raM) is well within the range of values (about 12 to 31 mM) reported for this tissue by other investigators (Reuss & Grady, 1979;Garcia-Diaz & Armstrong, 1980;Diez de los Rios et al, 1981;Fisher, Persson & Spring, 1981;Zeuthen, 1982;Petersen & Reuss, 1983). Also, as found in these earlier investigations, a~ under control conditions was well above a~.…”

“…As expected on the basis of earlier observations with cAMP (Diez de los Rios et al, 1981), 8-BrcAMP, and theophylline (Petersen & Reuss, 1983), IBMX added to the mucosal medium caused a marked decline in all (25.7 to 19.6 mg). It is, however, noteworthy that this decline was much slower than the concomitant depolarization of Va.…”

“…Because of this we examined the effect of IBMX (added to the mucosal medium) on membrane conductances in Necturus gallbladder. Since cAMP and theophyiline have been reported to decrease the intracellular C1 activity (a~l) of gallbladder epithelial cells (Diez de los Rios et al, 1981;Petersen & Reuss, 1983) and since this decrease has been interpreted as a direct consequence of an increase in Pc~ (Petersen & Reuss, 1983), we also investigated the effects of IBMX on this parameter. Finally, the response of tissue cAMP levels in Necturus gallbladder to IBMX was measured directly.…”

Cyclic AMP inhibits Cl-/HCO3- exchange at the apical membrane of Necturus gallbladder epithelium.

“…Support for this view was later obtained in studies in which membrane voltages and intracellular ion activities were determined before and after exposure to CAMP. The expected falls of intracellular Na' and Cl-activities were observed, and there were no changes in membrane voltage (Diez de los Rios et al, 1981). In contrast with these observations, Duffey et al (1981), in a study primarily designed to assess the effect of CAMP on the junctional complexes of Necturus gallbladder epithelium, found a moderate depolarization of the cell membranes and a large fall in the ratio of cell membrane resistances (apical:basolateral).…”