D2 Receptor Isoforms Research Articles

Differential plasticity of the dopaminergic D2 receptor mRNA isoforms under haloperidol treatment, as evidenced by in situ hybridization in rat anterior pituitary

Quantitative in situ hybridization with 35S-radiolabelled oligonucleotidic probes complementary either to the common sequence of both isoforms (D2(415) and D2(444)) of the D2 receptor mRNA, or to the specific sequence of the long isoform (D2(444)), and also to prolactin mRNA, was performed on pituitaries from control male rats and from rats injected daily with haloperidol (5 mg/kg, i.m.) for 3 or 30 days. It was shown that the D2(444)-mRNA is expressed in both anterior and intermediate lobes. In the anterior lobe, haloperidol treatment did not modify the hybridization signal specific for the D2(444)-mRNA, whereas long term (30 days)--but not short term (3 days)--treatment significantly increased the hybridization signal for the common sequence (D2(415) + D2(444)) by about 20%, suggesting an increased expression of the short isoform (D2(415)). Prolactin mRNA content was increased by the third day of haloperidol treatment. This neuroleptic-induced alteration in the ratio of both D2 receptor isoforms suggests changes in mRNA splicing regulatory processes, which could represent an alternative way to modulate receptor sensitivity, through differential plasticity of each isoform's expression.

Transcription mediated by a cAMP-responsive promoter element is reduced upon activation of dopamine D2 receptors.

Dopaminergic D2 receptors mediate the effect of dopamine on cellular effector systems by means of guanine nucleotide-binding proteins (G proteins). The major biochemical effect evoked by these receptors is the inhibition of adenylyl cyclase. As a consequence, the activation of D2 receptors lowers the intracellular cAMP level. Two cDNAs, originated by alternative splicing of the same gene, have been isolated: D2A and D2B. They code for two proteins of 444 and 415 amino acids. These proteins display high affinity for selective D2 dopamine ligands. D2A differs from D2B by an insertion of 87 nucleotides in its cDNA, which is located in a region of the protein considered important for the coupling to G proteins. To investigate functional differences between the two dopamine D2 receptor isoforms, we transiently expressed them in cultured cells. To do so we developed an assay to study membrane receptors that are coupled to the adenylyl cyclase. Using this assay, we were able to show that the stimulation of the adenylyl cyclase induced by the activation of the beta 2-adrenergic receptor is inhibited more efficiently by D2B than D2A. The effects elicited by the D2 receptors are mediated by pertussis toxin-sensitive G proteins. Treatment of transfected cells with pertussis toxin abolishes the inhibitory effects in a dose- and receptor isoform-dependent manner. Our results suggest that the two dopamine receptor isoforms are differentially coupled to G proteins.

Alternative splicing directs the expression of two D2 dopamine receptor isoforms

Dopamine receptors are classified into D1 and D2 subtypes on the basis of their pharmacological properties and the intracellular responses they mediate. The cerebral D2 dopamine receptor is the target of drugs used to alleviate the main symptoms of schizophrenia. Although it is considered to be a single molecular entity, there is evidence that multiple D2-receptor subtypes exist. A complementary DNA encoding a D2 receptor has recently been cloned and the deduced 415-amino-acid sequence indicates that it belongs to the large superfamily of receptors coupled to G proteins, and that its topology consists of seven transmembrane domains. In this family, the genes are frequently without introns and each is believed to encode a unique polypeptide product. Here we show that the gene for the D2 receptor produces two receptor isoforms by alternative messenger RNA splicing, providing a route to receptor diversity in this family. One isoform corresponds to the D2(415) receptor, but the second contains an additional sequence encoding a 29-amino-acid fragment, defining a novel D2(444) receptor isoform. Expression of the two isoforms is tissue-specific, and both are regulated by guanyl nucleotides. As the extra sequence is located within a putative cytoplasmic loop that binds to G proteins, the two isoforms might interact with different G proteins and thereby initiate distinct intracellular signals.