1. The performance of skeletal muscle during repetitive stimulation may be limited by the development of an intracellular
acidosis due to
lactic acid accumulation. To study this, we have measured the intracellular pH (pHi) with the fluorescent
indicator BCECF (2',7'-bis(carboxyethyl)-5(6)- carboxyfluorescein) during
fatigue produced by repeated, short tetani in intact, single fibres isolated from the mouse flexor brevis muscle. 2. The pHi at rest was 7.33 +/- 0.02 (mean +/- S.E.M., n = 29, 22 degrees C). During fatiguing stimulation pHi initially went alkaline by about 0.03 units (maximum alkalinization after about ten tetani). Thereafter pHi declined slowly and at the end of fatiguing stimulation (tetanic tension reduced to 30% of the original; 0.3Po), pHi was only 0.063 +/- 0.011 units (n = 14) more
acid than in control. 3. We considered three possible causes of
acidosis being so small in
fatigue: (i) a high oxidative capacity so that
fatigue occurs without marked production of
lactic acid; (ii) an effective transport of H+ or H+ equivalents out of the fibres; a high intracellular
buffer power. 4. The oxidative metabolism was inhibited by 2 mM-
cyanide in three fibres. After being exposed to
cyanide for 5 min without stimulation, the tetanic tension was reduced to about 0.9 Po and pHi was alkaline by about 0.1 units. The fibres fatigued faster in
cyanide and the pHi decline in
fatigue was more than twice as large as that under control conditions. 5. Inhibition of Na(+)-H+ exchange with
amiloride resulted in a slow acidification of rested fibres; resting pHi was not affected by either inhibition of HCO3(-)-Cl- exchange with
DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid) or inhibition of the
lactate transporter with
cinnamate. 6. Fibres fatigued in
cinnamate displayed a markedly larger acidification (approximately 0.4 pH units) and tension fell more rapidly than under control conditions; inhibition of Na(+)-H+ and HCO3(-)-Cl- exchange did not have any significant effect on
fatigue. 7. The intracellular
buffer power, assessed by exposing fibres to the weak base
trimethylamine, was about 15 mM (pH unit)-1 in a
HEPES-buffered
solution (non-CO2 or intrinsic
buffer power) and about 33 mM (pH unit)-1 in a
bicarbonate-buffered
solution. Somewhat higher values of the intrinsic
buffer power was obtained from changes of the partial pressure of CO2 (PCO2) of the bath
solution. Application of
lactate or
butyrate frequently gave an infinite
buffer power, which indicates that powerful pH-regulating mechanisms operate in these cases.(ABSTRACT TRUNCATED AT 400 WORDS)