Activation of protein kinase C by phorbol ester induces downregulation of the Na+/K+-ATPase in oocytes ofXenopus laevis

Abstract

Full-grown prophase-arrested oocytes of Xenopus laevis were treated with 50 nM phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, or with 50 nM 4 alpha-phorbol 12,13-didecanoate (4 alpha PDD) that does not activate protein kinase C. The effect on membrane currents and capacitance, inulin uptake and ouabain binding, and on membrane morphology were analyzed. (i) During application of PMA, current generated by the Na+/K+ pump decreases; in addition, Cl- and K+ channels become inhibited. This general decrease in membrane conductance reaches steady state after about 60 min. 4 alpha PDD was ineffective. (ii) Ouabain binding experiments demonstrate that PMA (K1/2 = 7 nM), but not 4 alpha PPD, induces a reduction of the number of pump molecules in the surface membrane. Permeabilization of oocytes by digitonin plus 0.02% SDS renders all binding sites present prior to PMA treatment again accessible for ouabain. The KD value for ouabain binding is not influenced. 4 alpha PDD was ineffective. (iii) Exposure of oocytes to PMA reduces membrane capacitance and stimulates uptake of inulin suggesting an increase in endocytosis. Electron micrographs show that PMA reduces the number and length of microvilli, leading finally to a smooth membrane surface with a reduced surface area. From these results we conclude that stimulation of protein kinase C leads to downregulation of the sodium pump. A major portion of this inhibition is brought about by reduction in area of surface membrane with a concomitant internalization of pump molecules. In addition to this mode of downregulation, a direct effect of stimulation of protein kinase C on the pump molecule cannot be excluded.

Store-operated Calcium Entry Inactivates at the Germinal Vesicle Breakdown Stage of Xenopus Meiosis

“…6A), which is responsible for the down-regulation of plasma membrane Na ϩ -K ϩ -ATPase (25,26). To determine whether SOCE channels are subject to the same regulation, we correlated I soce levels with the different phases of meiosis (see Figs.…”

“…Whereas membrane area decreases gradually after progesterone addition, SOCE inactivates acutely at GVBD. The Na ϩ -K ϩ -ATPase has also been shown to be down-regulated during meiotic maturation because of sequestration into intracellular vesicles (25,26). In this case Na ϩ -K ϩ -ATPase activity begins to decrease gradually after progesterone addition, even before GVBD, and continues after GVBD has occurred (28).…”

Modulation of cardiac Ca2+ channels in Xenopus oocytes by protein kinase C

“…2C. In order to verify that the decrease in lm by PMA was not a re.~ult of an accompanying reduction in the membrane surface area [46], the membrane capacitance was monitored along with the development of PMA effect on In,,, The capacitance was unchanged or slightly increased (after 10 rain in PMA, it was 105_+2% of control, n=56; after 30 rain in PMA, it still was 105__.2% of control, n=39; see Fig. 2), and only later returned to the control level or dropped below it (e.g, Fig.…”

Activation of the 5‐HT1c receptor expressed in Xenopus oocytes by the benzazepines SCH 23390 and SKF 38393

“…Application of SCH 23390 (1 pM) for 5min did elicit a response in every oocyte tested, but that application also caused a prolonged reduction in the sensitivity to SCH 23390 and to 5-HT that lasted for several hours in some oocytes. Such 'desensitization' of similar responses in oocytes has been commonly observed and may be related to depletion of internal Ca2 + or other aspects of the signal transduction process (Berridge, 1989;Vasilets et al, 1990 + +1 Figure 5 Benzazepine responses in oocytes injected with clonal 5-HT1c mRNA. In each oocyte, the responses to the benzazepines were determined and normalized to the 5-hydroxytryptamine response in Figure 3, (Hoyer et al, 1989); stereoselectivity was not tested for this choroid plexus action.…”