Three distinct binding sites for [3H]‐prazosin in the rat cerebral cortex

Abstract

1 The putative α₁-adrenoceptor subtypes of rat cerebral cortex membranes were characterized in binding experiments with [³H]-prazosin. 2 Specific binding of [³H]-prazosin was saturable between 20–5000 pm. Scatchard plots of the binding data were non-linear, indicating the presence of two distinct affinity sites for prazosin (pK_{D, high} = 10.18, R_high = 308 fmol mg⁻¹ protein; pK_{D, low} = 8.96, R_low = 221 fmol mg⁻¹ protein). 3 In the membranes pretreated with chlorethylclonidine (CEC) two affinity sites for prazosin were also observed: the affinities were similar to those without CEC pretreatment, but the maximum numbers of binding sites were reduced by CEC pretreatment to 23 and 62% for prazosin-high (R_high) and low affinity sites (R_low), respectively. 4 The prazosin-high affinity sites were further subdivided into two subclasses by WB4101(2-(2,6-dimethoxyphenoxyethyl)aminomethyl-1,4-benzodioxane) and phentolamine; the low affinity sites for WB4101 and phentolamine were more potently inactivated by CEC as compared with the high affinity sites. On the other hand, prazosin, HV723 (α-ethyl-3,4,5-trimethoxy-α-(3-((2-(2-methoxyphenoxy)ethyl)-amino)-propyl)benzeneacetonitrile fumarate) and yohimbine inhibited [³H]-prazosin binding to prazosin-high affinity sites monophasically. 5 In addition to the high affinity sites, the prazosin-low affinity sites were labelled at high concentrations of [³H]-prazosin. Thus, prazosin and WB4101 showed shallow displacement curves. On the other hand, HV723 and yohimbine did not discriminate between prazosin-high and low affinity sites. 6 Two distinct α₁-adrenoceptor subclassifications have been recently proposed (α_1A, α_1B subtypes and α_1H, α_1L, α_1N subtypes). According to the criteria defined with competitive antagonists in both the subclassifications, the present results indicate that the α₁-adrenoceptors of rat cerebral cortex consist of three different subtypes, presumably α_1A, α_1B and α_1L, and suggest that the α_1A and α_1B subtypes are identified as a single affinity site for prazosin (α_1H). The results also indicate that care must be taken in the use of CEC for α₁-adrenoceptor subclassification because of its low selectivity.