Cb. Farrell et Km. Oboyle, THE KINETICS OF [H-3] SCH-23390 DISSOCIATION FROM RAT STRIATAL DOPAMINE D-1 RECEPTORS - EFFECT OF DOPAMINE, European journal of pharmacology. Molecular pharmacology section, 268(1), 1994, pp. 79-88
The present study investigated possible allosteric interactions betwee
n dopamine and [H-3]SCH 23390 o-3-methyl-5-phenyl-1H-3-benzazepin-7-ol
)-labelled dopamine D-1 receptors in rat striatum. As previously descr
ibed, dopamine prevented [H-3]SCH 23390 binding in a mixed competitive
/non-competitive manner, causing both a loss of ligand affinity and a
decrease in B-max. The effect of dopamine was largely reversed followi
ng pretreatment of the membranes with 100 mu M Gpp(NH)p (5'-guanylylim
idodiphosphate) and was significantly enhanced by omission of Na+ from
the incubation buffer. In dissociation kinetic studies, two methods o
f initiating ligand dissociation were used: dilution into 100-fold vol
ume excess of buffer or addition of a molar excess of drug. Both metho
ds yielded similar rates of [H-3]SCH 23390 dissociation. Inclusion of
dopamine in the volume excess of buffer did not alter the k(-1) for [H
-3]SCH 23390 dissociation. However, when 100 mu M dopamine was used in
stead of 1 mu M piflutixol to initiate dissociation, a significant slo
wing of the rate of dissociation of [H-3]SCH 23390 occurred. This effe
ct of dopamine on k(-1) was Na+-dependent since in the absence of Nathe dopamine-induced rate of dissociation was only slightly slower tha
n control values. Under neither condition did dopamine accelerate the
rate of ligand dissociation, indicating that dopamine does not interac
t allosterically with [H-3]SCH 23390 binding sites. These data, theref
ore, preclude an allosteric mechanism to explain the dopamine-induced
decrease in dopamine D-1 receptor density and provide direct evidence
that dopamine masks ligand binding by binding to a high affinity site
which can be modulated by Gpp(NH)p and Na+.