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C. Claire Aickin^*, R. A. Deisz and H. D. Lux

Department of Neurophysiology, Max Planck Institute for Psychiatry, D-8000 Munich 40, Federal Republic of Germany

University Department of Pharmacology, South Parks Road, Oxford OX1 3QT

1. The reversal potential of the Cl^--dependent, inhibitory post-synaptic potential (E_i.p.s.p.) was measured in the isolated crayfish stretch receptor neurone using two intracellular micro-electrodes. The difference between E_i.p.s.p. and the resting membrane potential (E_m), the i.p.s.p. driving force, was reversibly decreased by addition of NH₃/NH₄⁺, and the mechanism of this decrease was investigated.

2. The NH₃/NH₄⁺-induced decrease in i.p.s.p. driving force was dose-dependent with an onset at about 0·2 mM. E_i.p.s.p. always remained more negative than E_m or, when the neurone was spontaneously firing, the threshold potential. E_m and resting membrane resistance (R_m) also decreased in a dose-dependent fashion. Synaptic conductance (g_s) increased with low doses, but decreased on application of 20 mM-NH₃/NH₄⁺. All the effects were fully reversible on return to normal Ringer solution.

3. Intracellular acidification (substitution of 50% Cl^- by acetate compared with isethionate) considerably reduced the i.p.s.p. driving force. Simultaneous application of NH₃/NH₄⁺ and acetate-substituted Ringer solution caused a similar decrease in the driving force to application of the same concentration of NH₃/NH₄⁺ under normal conditions. Increasing the extracellular pH at which a given concentration of NH₃/NH₄⁺ was applied caused a smaller decline in the i.p.s.p. driving force. These results suggest that intracellular acidification decreases the i.p.s.p. driving force and that the NH₃/NH₄⁺-induced decline is caused by an action of the ammonium ion.

4. Elevation of extracellular K⁺ (K⁺₀) decreased the i.p.s.p. driving force, E_m and R_m, and increased g_s. Reduction of K⁺₀ had the converse effects on all parameters.

5. Application of Rb⁺ or Cs⁺ mimicked the effects of NH₃/NH₄⁺. Substitution of K⁺₀ by Rb⁺, Cs⁺ or NH₃/NH₄⁺ opposed or even reversed the increase in i.p.s.p. driving force when Na⁺ was used as the substitute. The effectiveness of the various cations in decreasing the driving force was in the following order: Rb⁺ > NH₄⁺ > K⁺ > Cs⁺.

6. Inhibition of the Na pump by ouabain or K⁺-free Ringer solution caused a gradual reduction in the i.p.s.p. driving force. Since the driving force also decreased when the Na⁺ gradient probably was increased (elevated K⁺₀), this suggests a dependence on the K⁺ gradient rather than the Na⁺ gradient or the Na pump itself.

7. Frusemide (6 x 10^-4 M) reversibly decreased the i.p.s.p. driving force and E_m, and increased g_s. R_m was not significantly affected. Application of frusemide in the presence of 5 mM-Rb⁺ and vice versa, caused a further reduction in the driving force. The recovery of the driving force on removal of either agent was slowed by the presence of the other.

8. Application of 4,4-diisothiocyanostilbene-2,2-disulphonic acid (DIDS; 10^-4 M) caused spontaneous firing and reduced E_i.p.s.p. to the threshold potential. R_m and g_s increased. The effects were slowly reversible on removal of the drug.

9. It is proposed that the i.p.s.p. driving force is maintained by a K⁺—Cl^- co-transport mechanism, driven by the K⁺ gradient. The K⁺ site exhibits the binding selectivity: Rb⁺ > NH₄⁺ > K⁺ > Cs⁺ and the mechanism is inhibited partially by frusemide and completely by DIDS.

Laboratory in which the experiments were carried out.

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