Native ion current coupled to purinergic activation via basal and mechanically induced ATP release in xenopus follicles

Abstract

Xenopus follicle-enclosed oocytes are endowed with purinergic receptors located in the follicular cell membrane; their stimulation by ATP elicits an electrical response that includes generation of a fast inward current (F(Cl)) carried by Cl(-). Here, it was found that mechanical stimulation of the follicle provoked a native electrical response named I(mec). This was dependent on coupling between oocyte and follicular cells, because I(mec) was eliminated by enzymatic defolliculation or application of uncoupling drugs, such as heptanol or carbenoxolone. Moreover, the characteristics of I(mec) suggested that it corresponded with opening of the Cl(-) channel involved in F(Cl). For example, I(mec) showed cross-talk with the membrane mechanism that activates the F(Cl) response and anionic selectivity similar to that displayed by F(Cl). Also like F(Cl), I(mec) was independent of extracellular or intracellular Ca(2+). Furthermore, I(mec) was inhibited by superfusion with a purinergic antagonist, suramin, or by an enzyme that rapidly hydrolyzes ATP, apyrase. The response to mechanical stimulation was reconstituted in defolliculated oocytes expressing P2X channels as an ATP sensor. Recently, it has been shown that ATP release from the Xenopus oocyte is triggered by mechanical stimulation. Together, these observations seemed to indicate that I(mec) is activated through a mechanism that involves oocyte release of ATP that diffuses and activates purinergic receptors in follicular cells, with subsequent opening of F(Cl) channels. Thus, I(mec) generation disclosed a paracrine communication system via ATP between the oocyte and its companion follicular cells that might be of physiological importance during the growth and development of the gamete.

Functional expression and intracellular signaling of UTP-sensitive P2Y receptors in theca-interstitial cells

“…It has been shown in other tissues that ATP is co-released with noradrenaline by sympathetic terminals and that it participates in several physiological events such as the induction and regulation of smooth muscle contraction [5] and the modulation of cardiac muscle excitation [6]. In addition, several cell types are able to release ATP in a basal manner and/or in response to different stimuli, such as mechanical stimulation, changes in pH, or hypotonic stress [7-9]. …”

Functional interaction between native G protein-coupled purinergic receptors in Xenopus follicles

“…First, the results presented strongly suggest that the formation of a receptor-receptor complex in this native system is dynamic and reversible, and this interaction seems to exist without a prior stimulus, although basal ATP-eff lux from the oocyte (13,25) might have a modulatory role (26), and thereby, have physiological significance in follicular development. Second, to our knowledge, this putative interaction between purinergic receptors in the follicle is the first reported between native receptors coupled positively to cAMP synthesis; given the advantages that the follicleenclosed oocyte represents for studies using diverse methodologies, it will be an important tool to probe the physiological consequences of these protein-protein interactions and their modulation.…”

Differential role of STIM1 and STIM2 during transient inward (Tin) current generation and the maturation process in the Xenopus oocyte

“…During the maturation process, molecular elements that control the Ca 2+ dynamics in the Xenopus oocyte undergo an important reconfiguration; this observation has been extended to different species, and similar changes are known to occur during mitosis [23,31,41]. Given the importance of these events for cell cycle control, we asked whether or not the knockdown of STIM proteins affected the maturation process.…”

“…In general, after injection, oocytes were incubated at 18°C in Barth’s solution, and the effects of these procedures on protein expression and current responses were examined by biochemical and electrophysiological methods. Unless otherwise stated, groups of injected oocytes that were induced to express purinergic receptors were incubated in Barth’s solution containing 5 U/ml apyrase to hydrolyze the ATP that is released from the oocyte into the medium, thus avoiding stimulation of purinergic receptors during the incubation period [31]. …”