Extracellular Ca2+ and its effect on acid extrusion in the crayfish stretch receptor neurone

1. In the stretch receptor neurones of the crayfish Astacus astacus, the intracellular pH (pHi), the intracellular Na+ concentration ([Na+]i) and the membrane potential (Em) were measured simultaneously using ion-selective and conventional microelectrodes. Normal Astacus saline (NAS), and salines containing varying amounts of Ca2+ (Ca2+-NAS) but of constant ionic strength, with Na+, Mg2+ or Ba2+ as substituting ions, were used to investigate the effects of extracellular Ca2+ concentration ([Ca2+]o) on pHi and pHi regulation, on [Na+]i and on Em. The maximum rate of pHi recovery was used as a measure of pHi regulation. Acid loads were imposed using the NH4+/NH3 rebound technique. 2. [Ca2+]o affected pHi, pHi regulation, [Na+]i and Em. The magnitudes of the effects were inversely related to [Ca2+]o and were specific to the ion used for [Ca2+]o substitution. 3. Compared with controls, increasing [Ca2+]o threefold (in exchange for Na+) elicited some alkalization, a 7 % faster maximum rate of pHi recovery and generally lower values of [Na+]i. 4. In low-Ca2+ or Ca2+-free NAS (substitutions by Na+ or Mg2+), pHi became more acid, the maximum rate of pHi recovery was reduced by up to 50 % and [Na+]i was generally higher. The effects were faster and larger at lower [Ca2+]o, and stronger with Na+ than with Mg2+ as the substituting ion. 5. In Ca2+-free NAS (Ca2+ substituted for by Ba2+), the effects on pHi, on the maximum rate of pHi recovery and on [Na+]i were generally small. In this respect, Ba2+ had similar physiological properties to Ca2+ and was almost equally effective. 6. Changes in Em, including rapid depolarizations and occasional burst activity in Ca2+-free NAS, indicate that alterations in the properties of the membrane, such as a change in its permeability or selectivity, are occurring. Measurements of [Na+]i support this view. In addition, Ba2+ per se induced a (small) depolarization, as shown when Ba2+ was present in NAS or in low-Ca2+ NAS. 7. Changes in [Ca2+]o affected [Na+]i. *[Na+]i is defined as [Na+]i determined at the onset of the maximum rate of pHi recovery, and the ratio *[Na+]i/[Na+]o at that instant was calculated. A linear relationship between the maximum rate of pHi recovery and the *[Na+]i/[Na+]o ratio was found, irrespective of the amount and of the ion species used for [Ca2+]o substitution. This is strong evidence that pHi and pHi regulation were indirectly affected by [Ca2+]o, which altered membrane properties and thus caused a change in [Na+]i. We could find no evidence for a direct contribution of [Ca2+]o to acid extrusion or to a direct modulatory action on the transport protein of the Na+/H+ antiporter.