Effect of Phospholipase Digestion and Lysophosphatidylcholine on Dopamine Receptor Binding

[ 3 H]Spiperone specific binding by microsomal membranes isolated from sheep caudate nucleus is decreased by trypsin and phospholipase A 2 ( Vipera russeli ), but is insensitive to neuraminidase. The inhibitory effect of phospholipase A 2 is correlated with phospholipid hydrolysis. After 15 min of phospholipase (5 μg/mg protein) treatment, a maximal effect is observed; the maximal lipid hydrolysis is about 56% and produces 82% reduction in [ 3 H]spiperone binding. Equilibrium binding studies in nontreated and treated membranes showed a reduction in B max from a value of 388 ± 9.2 fmol/mg protein before phospholipase treatment to a value of 52 ± 7.8 fmol/mg protein after treatment, but no change in affinity ( K D = 0.24 ± 0.042 n M ) was observed. Albumin washing of treated membranes removes 47% of lysophosphatidyl‐choline produced by phospholipid hydrolysis without recovering [ 3 H]spiperone binding activity. However, the presence of 2.5% albumin during phospholipase A 2 action (1.5 μg/mg protein) prevents the inhibitory effect of phospholipase on [ 3 H]spiperone binding to the membranes, although 28% of the total membrane phospholipid is hydrolysed. Lysophosphatidylcholine, a product of phospholipid hydrolysis, mimics the phospholipase A 2 effect on receptor activity, but the [ 3 H]spiperone binding inhibition can be reversed by washing with 2.5% defatted serum albumin. Addition of microsomal lipids to microsomal membranes pretreated with phospholipase does not restore [ 3 H]spiperone stereospecific binding. It is concluded that the phospholipase‐mediated inhibition of [ 3 H]spiperone binding activity results not only from hydrolysis of membrane phospholipids, but also from an alteration of the lipid environment by the end products of phospholipid hydrolysis.