RNA editing is a post-transcriptional modification resulting in an alteration of the primary nucleotide sequence of RNA transcripts by mechanisms other than splicing. The enzymatic conversion of adenosine to inosine by RNA editing has been identified within an increasing number of RNA transcripts, indicating that this modification represents an important mechanism for the generation of molecular diversity. Several of these editing events have been shown to have significant consequences for cellular function. Transcripts encoding the Bsubunit (GluR-B) of the ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid subtype of glutamate receptor undergo RNA editing events that modulate both the ion permeation and electrophysiological characteristics of this glutamate-gated ion channel (1-3). Mice that are deficient in their ability to edit GluR-B transcripts die at 3 weeks of age due to epileptic seizures, suggesting that editing of GluR-B RNA is important in the modulation of normal glutamatergic neurotransmission (4). These results suggest that the consequences of editing events within other, diverse RNA molecules might also have important ramifications for cellular function.The monoamine 5-hydroxytryptamine (serotonin; 5-HT) 1 interacts with a large family of receptors to induce signal transduction events important in the modulation of neurotransmission (5). The 2C subtype of serotonin receptor (5-HT 2C R) is a member of the G protein-coupled receptor superfamily and stimulates phospholipase C, resulting in the production of inositol phosphates and diacylglycerol (6). We have recently shown that RNA transcripts encoding the rat, mouse, and human 5-HT 2C R undergo adenosine-to-inosine RNA editing events at five positions, termed A, B, C, D, and E (previously termed CЈ) (7,8), resulting in an alteration of amino acid coding potential within the putative second intracellular loop of the encoded protein. Editing at the A site, or at the A and B sites concurrently, converts an isoleucine to a valine at amino acid 156 of the human receptor, while editing at the B position alone generates a methionine codon at this site (Fig. 1A). Editing at C converts asparagine 158 to a serine; editing at E generates an aspartate at this site, and conversion at both C and E generates a glycine triplet. Editing at D results in the substitution of a valine for an isoleucine at position 160.We have previously demonstrated a decrease in 5-HT potency when interacting with the rat 5-HT 2C R isoform 5-HT 2C-VSV , which is simultaneously edited at the A, B, C, and D positions encoding valine, serine, and valine at positions 157, 159, 161, respectively. This decrease in potency was reflected as a rightward shift in the dose-response curve for [ 3 H]inositol monophosphate accumulation (7). We proposed that the decreased potency resulted from a reduced G protein coupling efficiency induced by the introduction of novel amino acids into the second intracellular loop, a region known to be important for G protein coupling (9 -16). In the present study,...

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Evidence from studies with phenylisopropylamine hallucinogens indicates that the 5HT2A receptor is the likely target for the initiation of events leading to hallucinogenic activity associated with LSD and related drugs. Recently, lisuride (a purported non-hallucinogenic congener of LSD) was reported to be a potent antagonist at the 5HT2C receptor and an agonist at the 5HT2A receptor. LSD exhibited agonist activity at both receptors. These data were interpreted as indicating that the 5HT2C receptor might be the initiating site of action for hallucinogens. To test this hypothesis, recombinant cells expressing 5HT2A and 5HT2C receptors were used to determine the actions of LSD and lisuride. LSD and lisuride were potent partial agonists at 5HT2A receptors with EC50 values of 7.2 nM and 17 nM, respectively. Also, LSD and lisuride were partial agonists at 5HT2C receptors with EC50 values of 27 nM and 94 nM, respectively. We conclude that lisuride and LSD have similar actions at 5HT2A and 5HT2C receptors in recombinant cells. As agonist activity at brain 5HT2A receptors has been associated with hallucinogenic activity, these results indicate that lisuride may possess hallucinogenic activity, although the psychopharmacological effects of lisuride appear to be different from the hallucinogenic effects of LSD.

The issue of G protein-coupled receptor (GPCR) oligomer status has not been resolved. Although many studies have provided evidence in favor of receptor-receptor interactions, there is no consensus as to the exact oligomer size of class A GPCRs. Previous studies have reported monomers, dimers, tetramers, and higher-order oligomers. In the present study, this issue was examined using fluorescence correlation spectroscopy (FCS) with photon counting histogram (PCH) analysis, a sensitive method for monitoring diffusion and oligomer size of plasma membrane proteins. Six different class A GPCRs were selected from the serotonin (5-HT 2A ), adrenergic (a 1b -AR and b 2 -AR), muscarinic (M 1 and M 2 ), and dopamine (D 1 ) receptor families. Each GPCR was C-terminally labeled with green fluorescent protein (GFP) or yellow fluorescent protein (YFP) and expressed in human embryonic kidney 293 cells. FCS provided plasma membrane diffusion coefficients on the order of 7.5 Â 10 29 cm 2 /s. PCH molecular brightness analysis was used to determine the GPCR oligomer size. Known monomeric (CD-86) and dimeric (CD-28) receptors with GFP and YFP tags were used as controls to determine the molecular brightness of monomers and dimers. PCH analysis of fluorescence-tagged GPCRs revealed molecular brightness values that were twice the monomeric controls and similar to the dimeric controls. Reduced x 2 analyses of the PCH data best fit a model for a homogeneous population of homodimers, without tetramers or higher-order oligomers. The homodimer configuration was unaltered by agonist treatment and was stable over a 10-fold range of receptor expression level. The results of this study demonstrate that biogenic amine receptors freely diffusing within the plasma membrane are predominantly homodimers.