Modulation of serotonin 2C receptor editing by sustained changes in serotonergic neurotransmission

Abstract

Serotonin 2C (5-HT2C) receptor pre-mRNA is a substrate for RNA editing enzymes that convert five adenosines (named A, B, C', C, and D editing sites) to inosines. Editing of two of these sites (C' and C) is crucial for decreasing the efficiency of the receptor to activate G-protein. Nucleotide sequence analysis of mouse forebrain neocortical 5-HT2C mRNA isoforms revealed that editing at these two sites is regulated in a serotonin-dependent manner. In serotonin-depleted mice, C'- and C-site editing is significantly decreased. This results in an increased expression of 5-HT2C mRNA isoforms encoding receptors with higher sensitivity to serotonin. In contrast, a 4 d treatment with the 5-HT2A/2C agonist (+/-)-1-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane significantly increases the editing frequency at the C' site and leads to increased expression of 5-HT2C mRNA isoforms encoding receptors that activate G-protein least efficiently. None of the drug treatments led to alterations in cytoplasmic 5-HT2C mRNA levels. These data indicate that editing of 5-HT2C pre-mRNA is a mechanism that retains basic response properties of 5-HT2C receptors in the face of changing synaptic input to keep receptor activation within an optimal range for information processing. Key words: serotonin; 5.

Fig. 1.

Tissue levels of 5-HT/5-HIAA in the forebrain neocortex (CTX) and hindbrain of pCPA-treated mice. pCPA treatment regimens are indicated on theabscissa. Tissues levels of 5-HT and 5-HIAA levels were determined by C₁₈ reverse-phase HPLC and are expressed as nanomoles per gram of tissue. Data represent the means ± SEM of measurements obtained from four animals per treatment group. A Student's t test was used to test for the significance of differences between groups. *p < 0.05; **p < 0.001.

Fig. 2.

Expression of 5-HT_2C mRNA isoforms in the forebrain neocortex of drug-naive mice and mice treated with either pCPA or DOI. Comparison of the percentages of the major edited isoforms (top) and percentages of editing at the five editing sites (bottom). The data represent the means ± SEM of percentages determined for controls (n = 4; 212 sequences), pCPA-treated animals (n = 4 for each of the two treatment regimens; 218 and 240 sequences), and DOI-treated animals (n = 4; 228 sequences). Data were compared by a one-way ANOVA, followed by Tukey–Kramer multiple comparisons test. Top, *p < 0.05; **p < 0.01. Bottom, *p < 0.04; **p < 0.02. Partially edited transcripts involving the C′ site are mRNA isoforms resulting from the editing combinations ABC′D, AC′D, C′D, and C′. Transcripts edited at both C′ and C sites included mRNA isoforms resulting from the editing combinations ABC′CD, AC′CD, AC′C, ABC′C, C′CD, and C′C. The latter two were found only in DOI-treated animals.

Fig. 3.

Expression levels of 5-HT_2C mRNA in the forebrain neocortex of drug-naive mice and pCPA- and DOI-treated mice. Southern blot of exponential RT-PCR amplification of cDNA encoding a 270-nucleotide-long sequence of 5-HT_2CmRNA sequence containing the edited region. Aliquots of the PCR reactions were removed after 10, 15, 20, and 25 cycles of amplification. For each treatment group, representative results obtained from two mice are shown, and, for comparison, PCR amplification of 0.01 ng of plasmid DNA encoding the full-length 5-HT_2C receptor is shown on top. Blots were exposed to film for 3 hr. CMV, Cytomegalovirus;pRC, plasmid vector RC (Invitrogen).