Rapid hypertonic cell volume regulation in the perfused inner medullary collecting duct

Abstract

Differential interference contrast microscopic images were used to assess the cell volume regulatory increase (VRI) response of rat IMCD segments isolated from the mid-inner medullary region of pathogen-free Sprague-Dawley rats and perfused in vitro at 37 degrees C. In the absence of ADH. IMCD cells behaved in an osmometric fashion over the range of extracellular osmolalities 290 to 386 mOsm/kg H2O and had an osmotic space equal to 54.2% of total geometric volume. After initial shrinkage in hypertonic perfusing and bathing solutions (340 mOsm/kg H2O using sucrose), cell volume increased rapidly to the isotonic value only in tubules preincubated in ADH (100 microU/ml). The rates of VIR were: (-ADH) 0.0142 +/- 0.0046 nl.min-1.cm-1 or 0.30 +/- 0.10%/min and (+ADH) 0.7225 +/- 0.1278 nl.min-1.cm-1 or 15.42 +/- 2.31%/min (N = 4; P less than 0.01). An overshoot in cell volume was observed on return to isotonic media only in the ADH exposed tubules showing a hypertonic VRI response, indicating that IMCD cells accumulated solute during hypertonic VRI. In the absence of ADH, one mM dibutyryl cyclic AMP mimicked the effect of hormone on hypertonic VRI. This ADH-dependent VRI process required Na+ and (CO2 + HCO3-) in external media and was reduced or abolished by 0.1 mM amiloride, 0.1 mM 4,4'-diisothiocyanatostilbene-2,2-'-disulfonic acid (DIDS) in peritubular solutions. These data suggest that ADH-dependent, rapid hypertonic cell volume regulation in rat inner medullary collecting duct depends on NA+ uptake, which may be mediated by parallel Na+-H+ and an HCO3(-)-dependent. DIDS-sensitive pathway (such as, Cl+-HCO3- exchanger) in basolateral cell membrane. In addition, a luminal amiloride-sensitive pathway (most likely the cation-selective channel) may contribute to cell volume regulation in the rat IMCD.