Phenylphosphonate: a 31P-NMR indicator of extracellular pH and volume in the isolated perfused rabbit bladder.

Abstract

31P-NMR has been used extensively to estimate intracellular pH. It also can be used to measure extracellular pH and volume when an NMR-detectable extracellular phosphorous probe is used. Phosphonic acids have been suggested as useful 31P-NMR extracellular markers. The present study was designed to assess the utility of phenylphosphonic acid (PPA) as a 31P-NMR extracellular marker in perfused smooth muscle. Rabbit bladder strips were exposed to PPA concentrations of 1-20 mM. Tension development in response to maximal carbachol challenges (10 microM) was independent of PPA concentration. Addition of PPA (6 mM) to the perfusate supplying the isolated resting rabbit bladder had no effect on 31P-NMR-detectable phosphatic compounds. PPA's resonance frequency was distinctly downfield from endogenous phosphates and demonstrated a pH-dependent chemical shift of +/- 1.12 ppm/pH unit over the range of 6.4 to 7.6 with a pK' of 7.09 at 23 degrees C. The time courses for washing PPA in and out of the resting bladder were best described by monotonic exponential growth (r = 0.972; n = 3) and decay (r = 0.972; n = 3) equations, respectively. Rate and time constants for PPA wash-in (0.039 +/- 0.004 min-1 and 25.7 +/- 2.3 minutes) and washout (0.038 +/- 0.000 min-1 and 26.3 +/- 0.0 minutes) were not significantly different. Using steady state PPA and ATP peak intensities and concentrations, an extracellular-to-intracellular ratio was calculated to be 0.31 +/- 0.03 (n = 3). These data indicate that PPA remains distributed exclusively in the extracellular spaces.(ABSTRACT TRUNCATED AT 250 WORDS)