Feedback control of intracellular pH by means of iontophoretic H+/OH- injection.
A new method for controlling intracellular pH (pH(i)) within a single cell has been developed that is based on the iontophoretic injection of the pH-affecting ions H+ and OH-. The H+/OH- injection currents are governed by an electronic feedback system, which compares the actual pH(i), monitored by pH-sensitive microelectrodes, with a command value. To avoid membrane potential (Em) alterations caused by the H+/OH- currents, the pH(i) control is performed under voltage-clamp conditions, using a two-electrode voltage-clamp design. This feedback-controlled iontophoretic injection system was capable of adjusting pH(i) between pH 6.8 and 8.0 with a precision of 0.031+/-0.026 pH units (n=51) and without affecting Em. We illustrate the application of this system by investigating the pH(i) dependence of glutamate receptors in identified leech Retzius neurons. Stimulation of these receptors with the glutamatergic agonist kainate (50 microM) under control conditions induced an intracellular acidification of 0.14+/-0.07 pH units (n=13). The feedback control system for pH(i) reduced this intracellular acidification to 0.02+/-0.01 pH units (n=15). The kainate-induced inward current was not affected by pH(i) alterations. This result demonstrates that the feedback system for pH(i) control was feasible for investigating the pH(i) dependence of physiological processes and suggests that the glutamate receptors are not modulated by pH(i).
Kilb
W
W
Institut für Neurobiologie, Heinrich-Heine-Universität Düsseldorf, 40255 Düsseldorf, Germany.
Schlue
W R
WR
D000818:Animals; D000838:Anions; D004553:Electric Conductivity; D005246:Feedback; D006863:Hydrogen-Ion Concentration; D006878:Hydroxides / Q000008:administration & dosage*; D007478:Iontophoresis; D007608:Kainic Acid / Q000494:pharmacology; D007865:Leeches; D008839:Microelectrodes; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D011522:Protons; D017470:Receptors, Glutamate / Q000502:physiology
D000838:Anions; D006878:Hydroxides; D011522:Protons; D017470:Receptors, Glutamate; D007608:Kainic Acid