STIMULUS-EVOKED CHANGES OF EXTRACELLULAR AND INTRACELLULAR PH IN THE LEECH CENTRAL-NERVOUS-SYSTEM .1. BICARBONATE DEPENDENCE

1. We have measured the effect of repetitive electrical nerve root sti mulation on the extracellular potassium activity (aK(e)) and the extra cellular pH (pH(e)) and intracellular pH (pH(i)) in segmental ganglia of the leech Hirudo medicinalis with double-barreled K+- and pH-sensit ive microelectrodes. To investigate the influence of CO2/HCO3-, we com pared the stimulus-evoked changes in aK(e), pH(e), and pH(i) in the pr esence and absence of 5% CO2-24 mM HCO3- in the saline. 2. An electric al nerve root stimulation at 20-30 Hz for 1 min caused a rapid increas e of 1.11 +/- 0.79 (SD) mM in aK(e), followed by an aK(e) undershoot o f 0.17 +/- 0.15 mM when the stimulation was discontinued (n = 6). aK(e ) transients were not significantly affected by CO2/HCO3-. 3. In 5 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered, nominally CO2/HCO3--free saline, low stimulus intensities or stimulus durations up to a few seconds resulted in a fast alkaline pH(e) transi ent. This alkalinization was followed by a larger and longer-lasting e xtracellular acidification when the stimulation was intensified and pr olonged. A stimulation at 20 Hz, 5 V for 1 min caused an average alkal ine shift of 0.083 +/- 0.055 pH units, followed by an acidosis of 0.07 9 +/- 0.038 pH units (n = 63). A change from 5 mM HEPES-buffered salin e to 20 mM HEPES-buffered saline attenuated the stimulus-evoked pH(e) transients by 50-60%. 4. pH(e) transients were strongly dependent on t he presence of CO2/HCO3-. In a saline containing 5% CO2-24 mM HCO3-, a n extracellular alkaline shift could only be resolved in about half of the experiments. The subsequent acidification amounted to 0.035 +/- 0 .012 pH units (n = 21), which is a reduction by 60% in comparison with that in 5 mM HEPES-buffered, CO2/HCO3--free solution. 5. In sensory p ressure neurons, the electrical stimulation induced an intracellular a cidification of 0.071 +/- 0.025 pH units in 5 mM HEPES-buffered, CO2/H CO3--free saline (n = 21). This intraneuronal acidification was attenu ated by 45% in CO2/HCO3-, buffered saline (n = 12). 6. Neuropile glial cells were depolarized by 8.4 +/- 2.3 mV, on average, during the stim ulation (n = 49). In 5 mM HEPES-buffered, CO2/HCO3--free saline this d epolarization was accompanied by an acidification of 0.025 +/- 0.009 p H units, which was preceded by a small and brief alkalinization in 10 experiments (n = 25). In contrast, in the presence of CO2/HCO3- all gl ial cells showed a distinct alkalinization of 0.025 +/- 0.009 pH units (n = 24), which lasted during the entire stimulation. This alkaliniza tion was followed by a weak acidification in nine experiments. 7. Both the reduction of the stimulus-evoked extracellular alkalinization and the appearance of a distinct intraglial stimulus evoked alkalinizatio n in CO2/HCO3--buffered saline compared with CO2/HCO3--free saline wer e a result of the increase in physicochemical buffering power due to C O2/HCO3-, as well as of active transport of acid/base equivalents acro ss the,glial membrane, which may contribute to the H+-muffling of the stimulus-induced pH(e) changes.