Postsynaptic fall in intracellular pH and increase in surface ph caused by efflux of formate and acetate anions through GABA-gated channels in crayfish muscle fibres

Abstract

H +-selective microelectrodes and a two- or three-microelectrode voltage clamp were used to examine the influence of weak-acid, car☐ylate anions on the actions of GABA on postsynaptic intracellular pH, surface pH and on membrane potential in fibres of the crayfish leg opener muscle. Substitution of 30 mM Cl − by formate or acetate promoted a GABA-induced decrease in intracellular pH, which was coupled to an increase in surface pH and to a depolarization. Such effects were not seen in the presence of an equivalent amount of lactate, methanesulphonate or glucuronate. Both the GABA-induced depolarization and the fall in internal pH promoted by formate and acetate were blocked by picrotoxin, and the fall in pH was reversibly inhibited by a K +-induced depolarization. The rate of the fall in intracellular pH produced by GABA (0.2 mM) was about 0.02 pH units/min in the presence of formate and 0.03 pH units/min in the presence of acetate. Under steady-state conditions, both 30 mM formate and acetate (but not lactate) induced a positive shift in the reversal potential of GABA-activated current, which was accounted for by a relative permeability vs Cl − of formate and acetate of 0.5 and 0.15, respectively. The conductance sequence of the anions was identical to the permeability sequence, i.e. Cl − > formate > acetate > lactate ≅ 0. This sequence is strictly correlated to the Stokes diameter of the anions. The relative permeabilities of the anions indicate that the effective diameter of the GABA-gated channel is about 0.5 nm. The fact that the GABA-induced acidosis was slower in the presence of formate than in the presence of acetate suggests that, in the former case, the rate-limiting step in the fall in internal pH is the entry of non-dissociated formic acid. All the above results are consistent with a scheme where GABA induces a channel-mediated efflux of permeant weak-acid anions, which gives rise to an inward (depolarizing) current and to an intracellular acidosis. A comparison of the permeability properties of crayfish and vertebrate GABA-gated channels suggests that effects similar to those seen in this work are likely to occur in mammalian and other vertebrate neurons in the presence of permeant weak-acid anions.