Activity-related transient changes in extracellular K+ concentration ([K+]c), extracellular pH (pHc), and extracellular volume (EC volume) were studied by means of ion-selective microelectrodes in the adult rat spinal cord in vivo and in neonatal rat spinal cords isolated from pups 3-14 days of age. Repetitive electrical nerve stimulation (10-100 Hz) in adults elicited increases in [K+]c by about 2.0-3.5 mM, followed by a poststimulation K+ undershoot and triphasic alkaline-acid-alkaline changes in pHc. In 3- to 6-day-old pups, the [K+]c increased by as much as 6.5 mM at a stimulation frequency of 10 Hz, and this was accompanied by an alkaline shift. Increases in [K+]c as large as 1.3-2.5 mM accompanied by an alkaline shift were evoked by a single electrical stimulus. Stimulation in 10- to 13-day-old pups produced smaller [K+]c change and an acid shift, which was preceded by a small initial alkaline shift, as in adult rats. We conclude that glial cells buffer the activity-related [K+]c increase and alkaline pHc shifts. Mg2+ blocked the alkaline but not the acid shift. Acetazolamide had no effect on the alkaline shift but blocked the acid shift. The alkaline shift was enhanced and the acid shift blocked by Ba2+, amiloride, 4-acetamido-4'-isothiocyanotostilbene-2,2'-disulfonic acid (SITS), and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS). Activity-related acid shifts therefore have a complex mechanism, which includes Na+H+ exchange, Cl-/HCO3- exchange, or Na+/Cl-/H+/HCO3- antiport, Na(+)-HCO3- cotransport, and H+ efflux through voltage-sensitive H+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)