Cl⁻ currents activated via purinergic receptors inXenopus follicles

Abstract

Ionic currents elicited via purinergic receptors located in the membrane of Xenopus follicles were studied using electrophysiological techniques. Follicles responded to ATP-activating inward currents with a fast time course ( F in). In Ringer solution, reversal potential ( E rev) of F in was −22 mV, which did not change with external substitutions of Na+ or K+, whereas solutions containing 50 or 5% of normal Cl−concentration shifted E rev to about +4 and +60 mV, respectively, and decreased F in amplitude, indicating that F in was carried by Cl−. F in had an onset delay of ∼400 ms, measured by application of a brief jet of ATP from a micropipette positioned near the follicle (50 μm). F in was inhibited by 50% in follicles pretreated with pertussis toxin. This suggests a G protein-mediated receptor channel pathway. F in was mimicked by 2-MeSATP and UTP, the potency order (half-maximal effective concentration) was 2-MeSATP (194 nM) > UTP (454 nM) > ATP (1,086 nM). All agonists generated Cl− currents and displayed cross-inhibition on the others. F in activation by acetylcholine also cross-inhibited F in-ATP responses, suggesting that all act on a common channel-activation pathway.

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

“…For example, it is known that responses to stimulation of Ado receptors conform closely to the pattern of the A2 subtype (5, 10), although a more precise molecular identification has not been made. However, F Cl currents elicited by ATP closely follow the pattern of UTP-sensitive P2Y receptors (8). To date, no molecule has been proposed in Xenopus or other species that has the characteristics shown for the putative ␤,␥-meATP sensitive P3-subtype receptor.…”

“…Electrodes (1-2 M⍀) were filled with 3 M KCl, unless otherwise stated. Follicles and oocytes were voltage clamped at Ϫ40 mV at room temperature, and brief (1-1.5 s) steps were applied periodically every 40 or 60 s to Ϫ20 mV, close to the Cl Ϫ equilibrium potential in follicles (2,8). All drugs were applied by superfusion.…”

“…preparations derived from them. UTP is a specific agonist of F Cl , and shows a weak or null capacity to generate I K,cAMP (8). In this series of experiments, I K,cAMP was elicited by Ado (10 M) or by follicle-stimulating hormone (0.5 g/mL) (20), and the amplitude of I K,cAMP was compared with that obtained in the presence of UTP (100 M), using a 40-s preincubation with UTP alone to ensure maximal effect (Fig.…”

“…Three main subtypes have been described: (i) A2 receptors responsive to Adenosine (Ado), coupled to cAMP synthesis and opening of K ϩ channels (cAMP-dependent K ϩ current, I K,cAMP ), that are sensitive to glibenclamide (3-7); (ii) P2Y receptors responsive to ATP and UTP that operate Cl Ϫ channels generating fast and slow Ca 2ϩ -independent currents (F Cl and S Cl ) (2,8,9); and (iii) a purinergic receptor, referred to as a P3 or novel P1 receptor, that is sensitive to both Ado and ATP and coupled to I K,cAMP generation (10,11). Also, there is a P2Y receptor that inhibits I K,cAMP and is probably the same as the receptor involved in F Cl and S Cl activation (2,12,13).…”

Ion Currents Induced by ATP and Angiotensin II in Cultured Follicular Cells of Xenopus laevis

“…However, only five FCs showed outward K + currents associated with a decrease in the membrane conductance (data not shown) (S out, see Arellano et al, 1996). This could be explained by the variety of ATP receptor subtypes that can potentially be expressed in the FC membranes (Arellano et al, 1998). It was of particular interest to find that ATP induced mainly chloride currents in the FCs, whereas in the epithelial cells, ATP generated cationic currents, even when chloride ions were replaced by aspartate (Wu and Mori, 1999).…”