Elsevier

Neuropharmacology

Volume 39, Issue 11, October 2000, Pages 1996-2005
Neuropharmacology

Regulation of 5-HT2A receptor mRNA levels and binding sites in rat frontal cortex by the agonist DOI and the antagonist mianserin

https://doi.org/10.1016/S0028-3908(00)00026-5Get rights and content

Abstract

In the present study we have characterized the time course of effect of administration of the serotonin2 (5-HT2) receptor antagonist mianserin, or the 5-HT2 receptor agonist (±)-2,5-dimethoxy-4-iodophenyl-2-aminopropane (DOI), on 5-HT2A receptor binding sites and mRNA levels in rat frontal cortex. Radioligand binding and ribonuclease protection assays were performed with separate hemispheres of frontal cortex from each animal to examine concomitant changes in 5-HT2A receptor sites and mRNA levels. The decrease in cortical 5-HT2A receptor sites in response to chronic DOI administration was not accompanied by changes in 5-HT2A receptor mRNA. A single injection of DOI produced a transient decrease in 5-HT2A receptor mRNA levels detected 1 h post-injection. The density of 5-HT2A receptor sites, however, was not significantly reduced following a single injection of DOI. The down-regulation of cortical 5-HT2A receptor sites in response to a single injection of mianserin was accompanied by reductions in 5-HT2A receptor mRNA levels. Following 4 days of mianserin administration, however, we did not observe a change in 5-HT2A receptor mRNA levels, although 5-HT2A receptor density was decreased. Thus, changes in receptor mRNA may initially contribute to the down-regulation of 5-HT2A receptors in response to acute mianserin administration. Sustained changes in 5-HT2A receptor mRNA, however, appear not to be involved in maintaining the down-regulation of 5-HT2A receptor number with chronic mianserin administration. Mechanisms other than the regulation of receptor mRNA levels appear to underlie the down-regulation of 5-HT2A receptor sites in response to chronic administration of the agonist DOI.

Introduction

Numerous subtypes of receptor for the neurotransmitter serotonin (5-hydroxytryptamine, 5-HT) have been identified. Serotonin2A (5-HT2A) receptors in the central nervous system have been implicated in a variety of behaviors and have high affinity for psychoactive drugs such as hallucinogens (see Glennon, 1990), atypical antipsychotics (Meltzer et al., 1989; Leysen et al., 1993) and many antidepressants (see Frazer, 1997). The 5-HT2A receptor is enriched in many brain areas including the frontal cortex, claustrum, nucleus accumbens, and striatum (Pazos et al., 1985, Pazos et al., 1987). In the cortex, 5-HT2A receptors are thought to be located postsynaptically on intrinsic interneurons and pyramidal cells (Morilak et al., 1993; Willins et al., 1997). 5-HT2A receptors are coupled to the stimulation of phospholipase C and the hydrolysis of membrane phosphoinositides (see Sanders-Bush et al., 1990).

The regulation of the 5-HT2A receptor does not appear to follow the pattern established for many other G-protein-coupled receptors. Although repeated administration of 5-HT2 receptor agonists results in the expected desensitization and down-regulation of central 5-HT2A receptors (Buckholtz et al., 1988; McKenna et al., 1989), chronic administration of 5-HT2 receptor antagonists, including atypical neuroleptics, results in a paradoxical decrease in the density of 5-HT2A receptors (see Frazer et al., 1988; Kuoppamaki et al., 1995). Furthermore, the down-regulation of 5-HT2A receptors following chronic administration of many antidepressant drugs appears to be related to the potent 5-HT2A receptor antagonist properties of these drugs and not to the blockade of serotonin receptor reuptake or changes in serotonin neurotransmission (see Frazer et al., 1988; Eison et al., 1991; Frazer, 1997). The desensitization and the down-regulation of 5-HT2A receptors occur following chronic administration of a wide variety of antidepressants, as well as atypical neuroleptics, and may be important to the therapeutic action of these drugs. Interestingly, 5-HT2A receptors are down-regulated after a single systemic injection of the atypical antidepressant and 5-HT2 receptor antagonist mianserin (Sanders-Bush et al., 1987).

The study of drug-induced changes in 5-HT2A receptor mRNA, for which we have employed the ribonuclease protection assay, allows assessment of the potential contribution that changes in gene expression may make to drug-induced changes in receptor number. It has been previously demonstrated that chronic administration of the 5-HT2 receptor agonist (±)-2,5-dimethoxy-4-iodophenyl-2-aminopropane (DOI) results in down-regulation of 5-HT2A receptors (Buckholtz et al., 1988; Hensler and Truett, 1998). However, whether this agonist-induced decrease in 5-HT2A receptor number is accompanied by changes in 5-HT2A receptor mRNA levels has not been described previously. It has been demonstrated that a single injection of the 5-HT2 receptor antagonist mianserin results in down-regulation of cortical 5-HT2A receptors (Sanders-Bush et al., 1987; Hensler and Truett, 1998). However, whether changes in 5-HT2A receptor mRNA levels accompany this decrease in receptor number after acute mianserin administration has not been determined.

One of the goals of the present study was to characterize the time course of effect of chronic DOI administration on 5-HT2A receptor binding sites and mRNA levels in frontal cortex at 2, 4 and 8 days of treatment. Although acute treatment of rats with DOI does not alter the 5-HT2A receptor number (Buckholtz et al., 1988; McKenna et al., 1989), we also examined the effect of a single injection of DOI on 5-HT2A receptor mRNA levels at several time points post-injection. A second goal of the present study was to characterize the time course of effect of a single injection of mianserin on 5-HT2A receptor binding sites and mRNA in rat frontal cortex 1, 3, 6 and 24 h post-injection. Experiments were also performed to examine the effect of repeated mianserin administration for 4 days on cortical 5-HT2A receptors and mRNA levels. The combined use of radioligand binding and ribonuclease protection assay, performed with separate hemispheres of frontal cortex from each animal, allowed us to examine concomitant changes in 5-HT2A receptor binding sites and receptor mRNA levels in this brain region. Our data indicate that the decrease in cortical 5-HT2A receptor sites in response to chronic DOI administration is not accompanied by changes in 5-HT2A receptor mRNA. A reduction in cortical 5-HT2A receptor mRNA levels accompanies the down-regulation of 5-HT2A receptors following a single injection of mianserin. Sustained changes in 5-HT2A receptor mRNA, however, appear not to be involved in maintaining the down-regulation of 5-HT2A receptor number with chronic mianserin administration.

Section snippets

Animals

All experiments used male Sprague-Dawley rats (250–300 g). Animals were housed three per cage, with lighting on a 14:10 h day:night cycle and with constant access to food and water. These studies were carried out in accordance with the Guide for the Care and Use of Laboratory Animals as adopted and promulgated by the National Institutes of Health.

Drug treatments

In one experimental group, rats received a single injection of mianserin (5 mg/kg, s.c.) and were sacrificed 1, 3, 6 or 24 h after mianserin

Effect of DOI administration on cortical 5-HT2A receptor mRNA levels and 5-HT2A receptor binding sites

To characterize the time course of regulation of 5-HT2A receptor binding sites and mRNA levels in frontal cortex following repeated administration of the 5-HT2 receptor agonist DOI, rats were treated with DOI (1 mg/kg, s.c., once daily) for 2, 4 or 8 days. Animals were sacrificed 6 h after the last injection. Radioligand binding and ribonuclease protection assays were performed with separate hemispheres of frontal cortex from each animal to examine concomitant changes in 5-HT2A receptors and

Discussion

In the present study we have characterized the time course of effect of administration of the 5-HT2 receptor antagonist mianserin, or the 5-HT2 receptor agonist DOI, on 5-HT2A receptor binding sites and mRNA levels in rat frontal cortex. Radioligand binding and ribonuclease protection assays were performed with separate hemispheres of frontal cortex from each animal to examine concomitant changes in 5-HT2A receptor sites and mRNA levels. We found that although repeated administration of DOI for

Acknowledgements

The authors thank Hymavathi Durgam for excellent technical assistance. This work was supported by US PHS grant MH 52369.

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