Abstract
Regulation of 5-hydroxytryptamine (5-HT2) receptor expression by SR 46349B, a potent and selective 5-HT2 receptor antagonist, was investigated in cultured rat aortic smooth muscle cells. Binding of [3H]SR 46349B to rat vascular smooth muscle cells was time-dependent, reversible, and saturable. [3H]SR 46349B bound to one class of specific binding sites with high affinity (KD = 1.3 +/- 0.3 nM; Bmax = 176 +/- 42 fmol/10(5) cells). Exposure of cells to a 1 microM concentration of the 5-HT2 agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane ((+/-)-DOI) or the antagonist ketanserin led to a significant decrease in 5-HT2 receptor density as measured by [3H]SR 46349B binding. In contrast, exposure of cells to 1 microM SR 46349B caused a marked increase in the maximal binding capacity of [3H]SR 46349B, with a maximal effect at 24 h (73% increase). The affinity constant was not affected by prior exposure to (+/-)-DOI, ketanserin, or SR 46349B. Furthermore, exposure of cells to 1 microM (+/-)-DOI or ketanserin produced, 48 h later, a decrease in the ability of (+/-)-DOI to stimulate phosphoinositide turnover in the cells, whereas treatment with SR 46349B induced a significant stimulation of the 5-HT2 receptor-linked signal transduction. This effect occurred with no changes in the amount of 5-HT2 receptor mRNAs as measured by quantitative polymerase chain reaction. These results indicate that SR 46349B increases 5-HT2 receptor binding and functions without altering steady-state 5-HT2 mRNA levels in cultured rat aortic smooth muscle cells.
Highlights
Recent work from our laboratory showed that repeated administration of SR 46349B, a novel and selective 5-HT2 receptorantagonist (Fig.1)(RinaldiCarmona et al, 19921, produces a parallelenhancementin
(+)-DO1to stimulate phosphoinositide turnover in the mRNA levels were investigated in cultured rat aortic smooth cells, whereas treatmentwith SR 46349B induced asig- muscle cells
The mechanisms of regulation of receptor expression have been extensively studied for 5-HT2 receptors, but atypical desensitization phenomena have been reported following treatment with receptor antagonists such as ritanserin or ketanserin (Leysen et al, 1986; Twist et al, 1990)
Summary
The results reported hereshow that, as observed nificant stimulationof the 5-HT2receptor-linkedsignal in vivo, exposure of aortic smooth muscle cells to SR 46349B transduction. This effect occurred with no changes in leads to up-regulation of the 5-HT2 receptor protein and functhe amount of 5-HT2receptor mRNAs as measured by tions without altering thegene transcription. It has been verse transcriptase enzyme and buffer werefromInvitrogen (San widely reported that the densityof 5-HT2 receptors in rodent brain is down-regulated in response tochronic treatment with antidepressant or neuroleptic drugs (Goodwin et al, 1984; Peroutka and Snyder, 1980; Andree et al, 19861, with 5-HT2 receptor agonists (Buckholtz et al, 1988; McKenna et al, 1989; Diego, CA).
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