Molecular Cloning, Stable Expression and Desensitization of the Human Dopamine D1B / D5 Receptor

Abstract

The sub-family of dopamine D1-like receptors is now known to be comprised of at least two members: the originally cloned D1 receptor (herein referred to as the D1a receptor) and a related receptor referred to as the D1b, D1 beta or D5 dopamine receptor (herein referred to as the D1b/D5 receptor). Here, we characterize the D1b/D5 receptor expressed transiently in COS-7 cells and permanently in Ltk- cells. Transiently expressed human D1b/D5 receptors bind the D1 specific ligand [125I]SCH 23982 saturably and with high affinity (KD = 500 pM). Competition for [125I]SCH 23982 binding to rat D1b/D5 and human D1a and D1b/D5 receptors supports the contention that the two D1b/D5 receptors are species homologues. Furthermore, in COS-7 cells, as previously observed, dopamine competes for the binding of [125I]SCH 23982 to human D1b/D5 receptors with a higher affinity than that seen at the human D1a receptor. These results are similar to those seen in Ltk- cells permanently transfected with the human D1b/D5 receptor. In these cells, dopamine competition for [125I]SCH 23982 binding is complex, sensitive to guanine nucleotides and of a higher affinity than that observed for dopamine binding to the human D1a receptor expressed in these same cells. In both D1a and D1b/D5 expressing Ltk- cells, dopamine stimulates adenylyl cyclase with an EC50 of approximately 200 nM. Furthermore, preincubation of Ltk- cells expressing the D1a and D1b/D5 receptors with dopamine results in desensitization of the response of adenylyl cyclase to subsequent agonist stimulation.

Developing a Neuronal Model for the Pathophysiology of Schizophrenia Based on the Nature of Electrophysiological Actions of Dopamine in the Prefrontal Cortex

“…2 Prefrontal Dopamine Electrophysiology and Schizophrenia 167 cellular recordings at 34 Њ C). The short duration application time 30-90 s) implemented in these studies served to minimize rapid D1 receptor desensitization (Jarvie et al 1993;Ng et al 1994Ng et al , 1995 Dumartin et al 1998).…”

“…2 Prefrontal Dopamine Electrophysiology and Schizophrenia 167 cellular recordings at 34 Њ C). The short duration application time 30-90 s) implemented in these studies served to minimize rapid D1 receptor desensitization (Jarvie et al 1993;Ng et al 1994Ng et al , 1995 Dumartin et al 1998).More recently, other intracellular and patch-clamp studies also detected an early transient suppression of input resistance that results in a late onset of first spike latency (responses start at ‫ف‬ 2 min after a 5-min long dopamine or D2/3/4 agonist application with the responses lasting for ‫ف‬ 5 min) (Geijo-Barrientos and Pastore 1995; Gulledge and Jaffe 1998). Responses from some of the same cells also show a delayed increase in neuronal excitability, as shown by a rebound earlier onset of first spike latency evoked by the same depolarizing pulse in PFC pyramidal neurons (Gulledge and Jaffe 1998).…”

Classification of Dopamine Receptor Genes in Vertebrates: Nine Subtypes in Osteichthyes

“…Later studies revealed that the human D 5 and the rat D 1B receptor sequences are in fact orthologous [Jarvie et al, 1993]. Genes are referred to as 'orthologous' when homologous sequences present in different species originate from a gene already present in their common ancestor.…”

Early Emergence of Three Dopamine D1 Receptor Subtypes in Vertebrates

“…It suggests that one functional differentiating characteristic of the D 1B receptor is its constitutive activity, a property inherent in the mammalian D 5 /D 1B receptor (34,35) and which appears to have been absolutely conserved throughout the evolutionary course of the D 1B receptor subfamily. As such, constitutive activation of adenylate cyclase by the D 1B receptor system appears fundamental to this receptor subtype and functionally relevant to the physiology of dopamine in the vertebrate nervous system.…”