Full-grown immature oocytes of Xenopus laevis express in their membrane a great number and variety of ion channels, including Ca²⁺-dependent Cl^– channels (CaCCs). The aim of this work was to determine the properties of CaCCs in immature and mature oocytes, since heterogeneity of CaCCs has been proposed in immature oocytes (Boton et al. 1989; Kuruma & Hartzell, 1999).

To evoke Ca²⁺-dependent Cl^– currents (I_Cl(Ca)) in immature oocytes, cells were permeabilized to Ca²⁺ with the ionophore A-23187 (2 µM, 15 min incubation in a Ca²⁺-free Ringer solution) and challenged with 32 s pulses of Ringer solution containing 10 mM Ca²⁺. At a holding potential of -60 mV, the elicited I_Cl(Ca) showed two components: (i) a fast peak, which was almost fully blocked after adding to the Ringer solution either 100 µM niflumic acid (NFA), flufenamic acid (FFA), 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) or 1 mM 9-anthracene carboxylic acid (9-AC), and (ii) a slower one, much less sensitive to these blockers (the percentages of blockage were 29 ± 16 % (mean ± S.E.M.), n = 5; 77 ± 2 %, n = 3; and 50 ± 10 %, n = 3, for 100 µM NFA, FFA or NPPB, respectively; and less than 10 % for 1 mM 9-AC, n = 3). Eggs were obtained by incubating oocytes with progesterone (10 µM) for 12 h. The egg activation potential (AP) was evoked by either mechanical stimulation or by incubation with A-23187 (2 µM). The I-V relationship of the AP current showed a reversal potential of +30 ± 1 mV (n = 20), close to the E_rev for Cl^– (eggs were recorded in 10 % Ringer solution), indicating that it is due to CaCCs. Furthermore, replacement of Cl^– by other anions markedly affected the I-V curve. Neither 100 µM NFA, FFA or NPPB, nor 1 mM 9-AC blocked the AP, although it could be fully blocked by 1 mM DIDS. In fact, in eggs, I_Cl(Ca) was potentiated by NFA, FFA and 9-AC at potentials more negative than -60 mV and was only moderately reduced at positive potentials. The mean open time of oocyte or egg CaCCs was assessed by voltage jumps. In immature oocytes, current relaxation curves of the slow component of the I_Cl(Ca) could be fitted to a single exponential function, but two exponentials were required for egg I_Cl(Ca) relaxation. In eggs, the values of the first component were similar to those obtained for immature oocytes (at -60 mV, τ = 21 ± 1 ms, n = 15, in oocytes, versus τ₁ = 23 ± 2 ms, n = 10, in eggs), but the second component was much slower (τ₂ = 125 ± 9 ms, n = 10).

These results suggest that either there are different subtypes of CaCCs in oocytes and eggs or, alternatively, they are differently regulated at these stages.

This work was supported by grant BFI2001-0756 from MCYT.