Mechanism of pHi regulation by locust neurones in isolated ganglia: a microelectrode study

Abstract

1. We have measured membrane potential (Em) and intracellular pH (pHi), and sodium and chloride activities (aNai and aCli) in exposed dorsal unpaired median neurones in isolated metathoracic ganglia from the desert locust, Schistocerca gregaria using eccentric double-barrelled ion-sensitive microelectrodes. 2. In the absence of added HCO3- the steady-state pHi was 7.21 +/- 0.13 (mean +/- S.D.) at a mean membrane potential of -37 +/- 7.0 mV (S.D.) (n = 44 cells). The pHi was always more alkaline than predicted for passive H+ distribution. 3. The pHi recovery from acid loads, induced by weak acid application or weak base removal, was pHi dependent and associated, in both the presence and absence of added CO2-HCO3-, with a transient increase in aNai. 4. In the absence of added HCO3-, application of the Na(+)-H+ exchange blocker amiloride or external Na+ removal caused intracellular acidification. Also in the absence of added HCO3- the inhibitor SITS (4-acetamido-4'-isothiocyanatostilbene-2,2'-disulphonic acid) caused an acidification of about 0.2 pH units which was not additive to the effects of the removal of external Na+. 5. We found that the application of a CO2-HCO3(-)-containing solution increased the rate of pHi recovery from acidification. 6. Intracellular chloride was decreased by intracellular acidification in the presence of added CO2-HCO3-. In the presence of amiloride, intracellular Cl- depletion inhibited pHi regulation. 7. Simultaneous application of SITS (160 microM) and removal of CO2-HCO3- revealed a continuous underlying acid load of 0.03-0.05 pH unit min-1. 8. We conclude that locust neurones possess at least two pHi-regulating mechanisms which operate against a continuous acid load. One is a Na(+)-H+ exchanger which can be blocked by amiloride, while the second is a Na(+)-dependent Cl(-)-HCO3- exchanger. The latter mechanism appears to be able to operate in the absence of added HCO3- and can recover pHi to around pH 7.4; it is probably the main pHi regulating mechanism. The Na(+)-H+ exchanger appears to activate at more acid pHi and being less energy efficient may serve a protective role.