The effect of extracellular weak acids and bases on the intracellular buffering power of snail neurones.
1. Intracellular pH (pHi) was measured in snail neurones using pH-sensitive glass microelectrodes. The influence of externally applied weak acids and bases on the total intracellular buffering power (beta T) was investigated by monitoring the pHi changes caused by the intracellular ionophoretic injection of HCl. 2. In the absence of weak acids or bases a reduction in the extracellular HEPES concentration had no effect on pHi or on beta T. It did, however, reduce slightly the rate of pHi recovery following HCl injection. 3. The presence of CO2 greatly increased beta T. However, as predicted for an open buffer system, the contributions to intracellular buffering by CO2 (beta CO2) decreased as pHi decreased. 4. When added to the superfusate, procaine, 4-aminopyridine, trimethylamine and NH4Cl (1-10 mM) all increased steady-state pHi. Procaine was fastest at increasing pHi and 4-aminopyridine the slowest. All four of these weak bases increased beta T. 5. The intracellular buffering action by these weak bases varied. HCl injection in the presence of procaine usually resulted in steady-state pHi changes with no pHi transients. In the presence of the other three weak bases HCl injections resulted in intracellular acidifications which were followed by pHi recovery-like transients. However, these were not blocked by SITS (4-acetamido-4'-isothiocyanatostilbene-2,2'-disulphonic acid) or by CaCl2 and I thus conclude that these transients were as a result of slow or incomplete intracellular buffering by the weak bases. 6. In many cells there was a good correlation between the measured contributions to intracellular buffering by the weak bases (beta base) and those predicted assuming a simple two-compartment open system. In all cases, as predicted, beta base increased as pHi decreased. 7. I found a clear relationship between the concentration of external buffer (HEPES) and the rate at which weak bases, applied to the superfusate, were able to increase pHi. The greater the extracellular buffer concentration the greater was the speed of intracellular alkalinization. 8. Lowering the extracellular buffer concentration reduced the efficiency of intracellular buffering by weak bases in response to an intracellular acid load. HCl injection in the presence of weak base caused a larger initial intracellular acidification if the extracellular HEPES concentration was reduced. 9. In conclusion, both weak acids and weak bases can make very large, pHi-dependent contributions to intracellular buffering by way of open buffer systems.(ABSTRACT TRUNCATED AT 400 WORDS)
Szatkowski
M S
MS
Department of Physiology, School of Medical Sciences, University of Bristol.
D015761:4-Aminopyridine / Q000494:pharmacology; D000143:Acids / Q000494:pharmacology*; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002021:Buffers; D005724:Ganglia / Q000502:physiology*; D006531:HEPES / Q000494:pharmacology; D006372:Helix, Snails; D006851:Hydrochloric Acid / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects; D008744:Methylamines / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects*; D011343:Procaine / Q000494:pharmacology
D000143:Acids; D002021:Buffers; D008744:Methylamines; D000643:Ammonium Chloride; D011343:Procaine; D015761:4-Aminopyridine; C023336:trimethylamine; D006851:Hydrochloric Acid; D006531:HEPES