Enhancement of alcohol drinking in mice depends on alterations in RNA editing of serotonin 2C receptors

Abstract

Serotonin 2C receptors (5-HT2CR) are G-protein-coupled receptors with various actions, including involvement in drug addiction. 5-HT2CR undergoes mRNA editing, converting genomically encoded adenosine residues to inosines via adenosine deaminases acting on RNA (ADARs). Here we show that enhanced alcohol drinking behaviour in mice is associated with the degree of 5-HT2CR mRNA editing in the nucleus accumbens and dorsal raphe nuceus, brain regions important for reward and addiction. Following chronic alcohol vapour exposure, voluntary alcohol intake increased in C57BL/6J mice, but remained unchanged in C3H/HeJ and DBA/2J mice. 5-HT2CR mRNA editing frequency in both regions increased significantly in C57BL/6J mice, as did expressions of 5-HT2CR, ADAR1 and ADAR2, but not in other strains. Moreover, mice that exclusively express the unedited isoform (INI) of 5-HT2CR mRNA on a C57BL/6J background did not exhibit increased alcohol intake compared with wild-type mice. Our results indicate that alterations in 5-HT2CR mRNA editing underlie alcohol preference in mice.

Development of the 5-HT2CR-Tango System Combined with an EGFP Reporter Gene

“…Two-way ANOVA analysis showed a significant difference in receptor activity (isoform: F 1,40 =80.26, p<0.01; 5-HT treatment: F 3,40 =21.66 p<0.01; isoform×5-HT: F 3,40 =3.01, p<0.05). This isoform difference has previously been reported in various studies (Burns et al 1997;Fitzgerald et al 1999;Wang et al 2000;Gurevich et al 2002a;Watanabe et al 2014); therefore, our results demonstrate correct functioning of the 5-HT 2C R-Tango system.…”

“…editing of 5-HT 2C R mRNA may have causative relevance to neuropsychiatric disorders (Maas et al 2006;Iwamoto et al 2009). Moreover, we recently demonstrated that after long-term alcohol exposure, alterations in 5-HT 2C R RNA editing are involved in alcohol preference (Yoshimoto et al 2012;Watanabe et al 2014). Overall, these findings indicate that modulation of 5-HT 2C R activity is involved in various brain functions.…”

“…Detection of an activity difference between the isoforms by the 5-HT 2C R-Tango assay is essential, as alterations in 5-HT 2C R isoform expression pattern by RNA editing is implicated in psychiatric disorders and alcohol dependence (Gurevich et al 2002b;Watanabe et al 2014). It was previously reported using the IP production assay, that basal activity of the VGV isoform was 2-to 8-fold lower than the INI isoform (Niswender et al 1999;Wang et al 2000;Watanabe et al 2014). However, we found an approximately 1.2-fold reduction with the Tango system.…”

“…The 5-HT 2C R INI and VGV forms were amplified by PCR using 5′-GGTACCCGCCACCATGGTGAACCTGGGCAC-3′ and 5′-GCGGCCGCCCACACTACTAATCCTCTCG from pEF1-5HT 2C R-INI and pEF1-5HT 2C R-VGV, respectively (Watanabe et al 2014), followed by TA cloning into pGEM-T easy (Promega,, Madison, WI, USA). The KpnI-NotI fragment containing 5-HT 2C R was inserted into partially KpnINotI digested pEF1-Tango plasmid (Fig.…”

A short history of the 5-HT2C receptor: from the choroid plexus to depression, obesity and addiction treatment

“…Stimulation of the 5-HT2CR is also suppressing ethanol self-administration (Maurel et al 1999;Tomkins et al 2002;Kasper et al 2013;Rezvani et al 2014) and reinstatement in rodents (Kasper et al 2013). Further, ethanol vapor exposure leads to an increased corticostriatal and hypothalamic 5-HT 2C R mRNA, increased 5-HT 2C R protein in the NAc and 5-HT2CR pre-mRNA editing (Yoshimoto et al 2012;Watanabe et al 2014). The behavioral effects of d-amphetamine (O'Neill et al 1999;Rippberger et al 2015;Wohr et al 2015), 3,4-methylenedioxymethamphetamine (MDMA) (Bankson and Cunningham 2002;Fletcher et al 2002b), methamphetamine (Steed et al 2011;Graves and Napier 2012), and the marijuana alkaloid 9 -THC (Ji et al 2006) can be modulated by the administration of 5-HT2CR agonists, thus extending the potential therapeutic value of selective 5-HT2CR agonists in the treatment of substance abuse.…”