Determination of adrenoceptors of the alpha₂-subtype on isolated human fat cells

Abstract

Binding state for the selective alpha₂-adrenergic antagonist (³H)-yohimbine on isolated intact human subcutaneous abdominal fat cells is described in the present study. Specific binding was rapid, reversible, saturable and stereospecific. Adrenergic antagonists competed with (³H)-yohimbine binding in the order indicative of alpha-receptors. Competition studies with adrenergic antagonists revealed that (³H)-yohimbine binding sites displayed properties consistent with those of alpha₂-subtype. Saturation experiments demonstrated that (³H)-yohimbine specific binding was of high affinity with a dissociation constant (K_d) of 3.7 ± 0.4 nmol/l and a maximum occupancy (B₀) of 10.5 ± 2.1 pmol/10⁷ cells. The latter corresponded to approximately 600,000 alpha₂-receptors/fat cell. A significant (P < 0.002) linear relationship was found between the fat cell volume and B₀. A complete saturation experiment for the determination of K_d and B₀ could be performed using 0.5 g adipose tissue (0.5 ml isolated fat cells), which is 30 times less required when adipocyte membranes are used for these studies. It is concluded that the method can be used for the study of alpha₂-receptor properties and regulation in subcutaneous adipose tissue in a given individual.

Key Words:

• alpha agonist
• alpha blocker
• radioligand