HYPEROSMOLALITY STIMULATES PHOSPHOLIPASE-A2 ACTIVITY IN RABBIT RENAL MEDULLA AND IN MADIN-DARBY CANINE KIDNEY (MDCK) CELLS

Renal medullary cells are able to accumulate glycerophosphocholine dur ing adaptation to the high extracellular osmolality, The aim of this s tudy was to investigate the effect of hyperosmolality on both phosphol ipase A2 activity and the rate of choline incorporation into glyceroph osphocholine in rabbit renal medulla and Madin-Darby Canine Kidney cel ls, Phospholipase A2 activity was assayed in cellular subfractions iso lated from both rabbit kidney medulla and Madin-Darby Canine Kidney ce lls in the presence of either 1-palmitoyl-2-[1-C-14]palmitoyl phosphat idylcholine or 1-stearoyl-2-[1-C-14]arachidonyl phosphatidylcholine as substrate, The rate of choline incorporation into glycerolphosphochol ine was measured in Madin-Darby Canine Kidney cells growing in the pre sence of [methyl-H-3]choline in the growth medium, Water deprivation o f rabbits resulted in an increase of phospholipase A2 activity from 2. 7 +/- 0.4 (n = 5) and 5.7 +/- 0.7 (n = 5) to 5.0 +/- 0.8 (n = 5) and 1 0.8 +/- 1.3 (n = 5) pmol of fatty acid released/min per mg protein in mitochondrial and microsomal fractions, respectively, using dipalmitoy l phosphatidilcholine as substrate while the activity of cytosolic enz yme remained unchanged, Similarly, the addition of sodium chloride in order to increase growth medium osmolality (from 320 mOsm/kg to 520 mO sm/kg) resulted in an elevation of both mitochondrial (from 1.8 +/- 0. 1 to 4.9 +/- 0.8 pmol of fatty acid released/min per mg protein, (n = 4) and microsomal (from 8.7 +/- 0.5 to 15.9 +/- 1.7 pmol of fatty acid released/min per mg protein, n = 4) phospholipase A2 activities, An a cceleration of phospholipase A2 activity in Madin-Darby Canine Kidney cells was accompanied by 1.5-fold stimulation of choline incorporation into intracellular glycerophosphocholine. In the presence of raffinos e to increase the medium osmolality, phospholipase A2 was mainly activ ated in microsomal fraction of MDCK cells (up to 380% of the control v alue), The cytosolic PLA2 exhibited still low activity (1.3 +/- 0.1 pm ol of fatty acids released/min per mg protein) despite relatively high stimulation of this enzyme in MDCK cells growing in the raffinose con taining, hyperosmotic medium, The data indicate that both animal antid iuresis in vivo as well as hyporosmolality of extracellular fluids in vitro result in an enhancement of phospholipase A2 activity in kidney medulla and renal cells in culture, It is concluded that activation of this enzyme is involved in the adaptation process of kidney to increa sed extracellular osmolality.