pH recovery from intracellular alkalinization in Retzius neurones of the leech central nervous system.
1. Neutral-carrier pH-sensitive microelectrodes were used to investigate intracellular pH (pHi) recovery from alkalinization in leech Retzius neurones in Hepes- and in CO2-HCO3(-)-buffered solution. The Retzius neurones were alkaline loaded by the addition and subsequent removal of 16 mM acetate, by changing from 5% CO2-27 mM HCO3- to 2% CO2-11 mM HCO3- or by changing from CO2-HCO3(-)- to Hepes-buffered solution. 2. In Hepes-buffered solution (pH 7.4) the mean pHi was 7.29 +/- 0.11 and the mean membrane potential -44.7 +/- 5.9 mV (mean +/- S.D.; n = 83). 3. The rate of pHi recovery from alkalinization increased with decreasing pH of the bathing medium (pHb). pHi changed about 0.30 pH units for a pHb unit change. 4. A decrease of extracellular buffer concentration (Hepes concentration lowered from 20 to 5 mM) caused an acidification of extracellular and intracellular pH and an acceleration of pHi recovery from alkalinization. 5. A depolarization of the Retzius cell membrane-induced by increasing the K+ concentration of the bathing medium from 4 to 20 mM (delta Em = 16.5 +/- 5.5 mV) or from 4 to 40 mM (delta Em = 24.8 +/- 3.5 mV)--evoked a decrease of pHi and an acceleration of pHi recovery from alkalinization. 6. The H+ current blocker Zn2+ (0.5 mM) inhibited pHi recovery from alkalinization at resting membrane potential as well as during depolarization. The inhibition was more pronounced during depolarization. 7. In Cl(-)-free, CO2-HCO3(-)-buffered solution pHi recovery from an alkaline load by changing from 5% CO2-27 mM HCO3- to 2% CO2-11 mM HCO3- was slowed by 48-71%. The rate of pHi recovery from an alkaline load induced by changing from CO2-HCO3- to Hepes buffer was reduced by 33-56% in Cl(-)-free solution. The removal of external Cl- did not affect pHi recovery in Hepes-buffered solution. 8. The pHi recovery from alkalinization was DIDS-insensitive in CO2-HCO3(-)- as in Hepes-buffered solutions and was not slowed in the absence of external Na+. 9. It is concluded that in Retzius neurones pHi recovery from alkalinization is mediated by a passive voltage-dependent H+ influx along the electrochemical proton gradient. In the presence of CO2-HCO3- buffer a DIDS-insensitive Cl(-)-HCO3- exchanger additionally regulates pHi after an intracellular alkaline load. It cannot be excluded that intracellular processes (e.g. H+ release from organelles, metabolic H+ production) are also involved in pHi recovery from alkalinization.
Frey
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Institut für Zoologie/Lehrstuhl für Neurobiologie, Heinrich-Heine-Universität, Düsseldorf, Germany.
Schlue
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D000468:Alkalies / Q000378:metabolism*; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D002021:Buffers; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002490:Central Nervous System / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007424:Intracellular Fluid / Q000378:metabolism*; D007865:Leeches; D008564:Membrane Potentials / Q000502:physiology; D008565:Membrane Proteins / Q000378:metabolism; D008839:Microelectrodes; D009474:Neurons / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D015032:Zinc / Q000378:metabolism
D000468:Alkalies; D002021:Buffers; D002352:Carrier Proteins; D008565:Membrane Proteins; D000584:Amiloride; D012964:Sodium; D015032:Zinc; D011188:Potassium; D002118:Calcium