SELECTIVE ENRICHMENT WITH ALPHA(1A)-ADRENOCEPTOR AND BETA(1B)-ADRENOCEPTOR SUBTYPES IN RAT-BRAIN CORTICAL MEMBRANES

Recent evidences from molecular biology, radioreceptor binding and fun ctional studies indicate that the alpha(1)-adrenoceptor population is heterogeneous and can at least be divided into two subclasses named al pha(1A) and alpha(1B). The present study was designed to obtain, a sel ective enrichment of rat brain cortical membranes with each subtype of alpha(1)-adrenoceptor using alkylating agents. [H-3]prazosin binding to rat cortical membranes was saturable and of high affinity (K-D = 0. 11 +/- 0.02 nM; B-max = 132.5 +/- 7.2 fmol/mg protein). All ligands co mpeted for specific [H-3]prazosin binding in a statistically significa nt biphasic manner (%R(high) = 30-40%; %R(low) = 60-70%). These sites meet generally accepted and recently described pharmacologic criteria for their identification as the alpha(1A)- and alpha(1B)-adrenoceptors . After pretreatment of membranes with benextramine (1 mu M) in the pr esence of clonidine (1 mu M), the antagonists, WB4101, (+)-niguldipine and phentolamine, displaced the radioligand with an inhibition curve steeper than in control membranes and with K-i values that agree with those obtained for the low affinity site present in control membranes. On the other hand, after pretreatment with chloroethylclonidine (10 m u M) in the presence of WB4101 (1 nM), Hill coefficients for the displ acement of the radioligand by WB4101, (+)-niguldipine, and phentolamin e, were also increased, but in contrast to the situation described abo ve, the K-i values agree with those obtained for the high affinity sit e present in control membranes. In conclusion, this method of partial alkylation of receptors could be a valuable tool for separately studyi ng the pharmacological characteristics of the alpha(1)-adrenoceptor su btypes in native membranes of cerebral tissue.