• 1
    Hypoxia (achieved by bubbling with N2 instead of O2) reduces the force of a KCl (40 mm)-induced contracture to ?10 % of the control value in guinea-pig taenia caeci. The underlying mechanism of this relaxation in response to hypoxia was investigated by measuring the major cell signalling parameters, intracellular Ca2+ concentration ([Ca2+]i) and myosin regulatory light chain (LC20) phosphorylation (MLC-Pi), as well as intracellular pH (pHi), a factor often suggested to mediate hypoxic relaxation of muscle.
  • 2
    [Ca2+]i, measured using the ratiometric fluorescent dye fura-2, increased when 40 mm KCl was added to physiological saline solution (PSS) (peak value assigned 100%), and the steady state after 15 min was 92.8 %. There were no detectable decreases in [Ca2+]i during hypoxia.
  • 3
    MLC-Pi, measured using isoelectric focusing-polyacrylamide gel electrophoresis and identified using Western blotting, increased from 9 % of the total LC20 in Ca2+-free PSS to a peak value of 51 % in 40 mM KCl-PSS. The steady-state value in hypoxia of 43 % was not significiantly different from that in control oxygenated conditions at the same point in time.
  • 4
    pHi, measured using the ratiometric fluorescent dye 2′,7′-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF), under quiescent conditions (Ca2+-free PSS) was 7.23 and increased to 7.36 with 40 mM KCl. After imposition of hypoxia pHi remained unchanged despite the known increase in both lactate content and production.
  • 5
    As [Ca2+]i and MLC-Pi, key factors in activation, were not decreased by hypoxia and changes in pHi were minor, hypoxic relaxation in guinea-pig taenia caeci appears to be directly related to energy limitation rather than any oxygen-sensing mechanism.