COMPARISON OF THE EXPRESSION, TRANSCRIPTION AND GENOMIC ORGANIZATION OF D(2) DOPAMINE-RECEPTORS IN OUTBRED AND INBRED STRAINS OF RAT

Three outbred (Sprague-Dawley, Wistar and Long-Evans) and five inbred (Brown-Norway, Buffalo, DA, Fisher and Lewis) strains of rat were used to investigate the extent of genetic variation in the expression and organization of the rat D2 receptor locus. Radioligand binding studies were performed using I-125-iodobenzamide ([I-125]IBZM), a high-affinity antagonist for D2 dopamine receptors, to determine the extent of variation in the expression of D2 receptors in these strains of rat. A comparison of the affinities (K(d) = 0.26-0.38 nM) and densities (450-580 fmol/mg of protein) of binding sites for [I-125]IBZM in the striatum of the eight strains of rat did not reveal statistically significant differences. Solution hybridization using P-32-labeled riboprobes complementary to the coding region of the third intracellular loop of the D2 receptor was used to investigate the extent of variation in transcription of the long (D2L) and short (D2S) isoforms of D2 receptor mRNA in rat striatal tissue. The level of expression of these two mRNA isoforms was found to be invariant in the strains of rats that were examined. The genomic organization of the D2 receptor locus for each strain of rat was compared using Southern blot hybridization. Southern blots were hybridized with a DNA probe that codes for the D2L receptor isoform. Restriction fragment lengths were conserved between each rat strain for genomic DNA digested with BamHI, EcoRI, HindIII, PstI and TaqI. Restriction fragment length polymorphisms (RFLPs) were identified when genomic DNA was digested with XbaI or MspI. The XbaI polymorphism was mapped to within 2 kb of the exon coding for the third intracellular loop of the D2 receptor. Both RFLPs differentiated Sprague-Dawley and Brown-Norway rats from Wistar, Long-Evans, Buffalo, DA, Lewis and Fisher rats' The RFLPs for the rat D2 receptor locus provide genetic markers that can be used with classic genetic studies to determine whether strain differences in behavior and/or in the response to pharmacologic intervention are determined by genetic elements linked to the D2 receptor locus.