Serotonin Transporter Gene Expression Research Articles

Alternative splicing of the human serotonin transporter gene.

To explore the structural basis for regulation of human serotonin transporter (hSERT) gene expression, we used primer extension and 5' rapid amplification of cDNA ends (5'RACE) techniques to estimate levels of and identify 5'-noncoding elements of hSERT mRNAs and genomic cloning to place these elements within the overall map of the hSERT gene. Primer extension on JAR cell mRNA suggested the presence of significant hSERT mRNA sequence upstream of the 5' end of our cloned hSERT cDNA. Using 5'RACE and reverse transcription-PCR (RT-PCR) methodologies, we cloned these sequences from brain and placenta and found this material to be composed of alternatively spliced exons using a previously reported noncoding exon (1A) and a novel 97-bp noncoding exon (1B). RT-PCR of JAR cell mRNA blotted with exon-specific oligonucleotides revealed both exons 1A and 1B to be regulated in a cholera toxin-dependent manner. To clarify the structure of the hSERT gene including exon 1B, we isolated and characterized genomic hSERT clones from Lambda Fix II and P1 artificial chromosome libraries. In agreement with previous findings, a single hSERT gene was identified that accounts for hybridizing bands on genomic Southern blots and was found to utilize 13 exons to encode the transporter's coding sequences along with the two noncoding 5' exons. Exon 1B was identified approximately 14 kb downstream of exon 1A in the hSERT gene and 737 bp upstream of exon 2, where the initiation site for translation is located. Exon 1B is surrounded by several elements potentially suitable for regulating serotonin transporter gene expression in vivo, including consensus sites for transcription factors AP-1, AP-2, CREB/ATF, and NF-kappaB. These data reveal additional complexity in hSERT gene structure and expression that may be relevant to regulated and compromised transporter expression in vivo.

72-6 - Serotonin transporter gene expression: effects of mood-stabilizing drugs

72-6 - Serotonin transporter gene expression: effects of mood-stabilizing drugs

Estradiol-17 beta increases serotonin transporter (SERT) mRNA levels and the density of SERT-binding sites in female rat brain.

The aim of the present study was to determine the effect of estradiol-17 beta (E2), in its positive feedback mode for gonadotropin release, on the serotonin transporter (SERT) in female rat brain. Levels of SERT mRNA were determined by in situ hybridization and SERT-binding sites were measured by quantitative [3H]paroxetine receptor autoradiography. The injection of estradiol benzoate (EB) in acutely ovariectomized rats increased significantly (approximately 50%) the numbers of cells that expressed SERT mRNA in the dorsal raphe nucleus and the density of SERT-binding sites in lateral septum (90%), basolateral amygdala (20%), ventral nucleus of thalamus (250%) and ventromedial hypothalamic nucleus (250%). SERT-binding sites in EB-treated rats were significantly lower in periaqueductal central grey (15%). These findings indicate that effects on SERT gene expression may be involved in the E2-induction of the gonadotropin surge. Together with our previous findings, they also suggest that the sex differences in depression and the apparent psychotropic effect of E2 may be due to the action of E2 on the serotonin transporter as well as 5-HT2A receptors.

Functional link between tyrosine phosphorylation and human serotonin transporter gene expression

Treatment of the JAR human placental choriocarcinoma cells with herbimycin A, an inhibitor of tyrosine kinases, led to an increase in the activity of the serotonin transporter. This effect was accompanied by an increase in the serotonin transporter density and in the steady-state levels of the serotonin transporter mRNA. A treatment time of >4 h was necessary for herbimycin A to elicit its effect. Actinomycin D and cycloheximide blocked the effect. There was no increase in the steady-state levels of the serotonin transporter mRNA when cells were treated with herbimycin A in the presence of actinomycin D. The herbimycin A-induced increase in the transporter activity was abolished by genistein, another inhibitor of tyrosine kinases. But the increase in the transporter mRNA levels caused by herbimycin A was not affected by genistein. Treatment of cells with herbimycin A resulted in an increase in the tyrosine phosphorylation of specific cellular proteins, suggesting that herbimycin A directly or indirectly activates specific tyrosine kinases. It is concluded that tyrosine phosphorylation is an essential component in the signaling pathways participating in the regulation of the human serotonin transporter gene expression. © 1997 Elsevier Science B.V.

Regulation of the Human Serotonin Transporter by Interleukin-1β

The serotonin transporter expressed in brain, platelets, and placenta is a primary target for antidepressants, cocaine, and amphetamines. Here we report that interleukin-1β is a potent regulator of the expression of this transporter. The activity of the serotonin transporter in human JAR choriocarcinoma cells is stimulated by this cytokine and the stimulation is accompanied by an increase in the steady state levels of the transporter mRNAs and in the transporter density. The increase in mRNA levels is effectively blocked by actinomycin D. The stimulatory effect of interleukin-1β is not associated with any change in the cellular levels of cAMP, indicating involvement of a cAMP-independent pathway. Even though cholera toxin, which is known to increase cAMP levels, also upregulates the serotonin transporter in JAR cells, the effects of interleukin-1β and cholera toxin on the transporter activity are additive. This constitutes the first report on the identification of a physiologically occurring hormone/cytokine which upregulates the expression of the human serotonin transporter gene.

Increased expression of serotonin transporter messenger RNA in raphe neurons of the aged rat

The action of serotonin (5-hydroxytryptamine; 5HT) in the nervous system is terminated by reuptake of the neurotransmitter into the presynaptic nerve terminal via a plasma membrane-bound transporter protein. Recently, cDNAs encoding serotonin transporter protein were cloned. Using in situ hybridization we have studied the expression of serotonin transporter mRNA in the bulbospinal 5HT system of adult (2-3 months) and aged (30 months) rats. In adult rats, serotonin transporter mRNA labelled neurons were detected in the nucleus raphe obscurus, pallidus and magnus. In all aged rats, the optical density of the serotonin transporter mRNA labelling of individual cell profiles was increased by 20-30% (P < 0.001) when compared with adult rats. Five of the six aged rats studied had various degrees of hindlimb motor dysfunctions. In rats with the most pronounced signs of hindlimb motor dysfunction the number of labelled neurons was increased by 25-75% (P < 0.01) when compared with young adult rats. It is concluded that an increased serotonin transporter gene expression is present in the bulbospinal 5HT system of aged animals.

Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of monoamine transporters in neuroblastoma cell lines: Correlations to meta-iodobenzylguanidine (MIBG) uptake and tyrosine hydroxylase gene expression

Radiolabelled meta-iodobenzylguanidine (MIBG) has been widely used in scintigraphy and targeted radiotherapy in patients with neuroblastoma. Recently, it has been demonstrated that MIBG is incorporated into neuroblastoma cells by the noradrenaline transporter. In vitro experiments on SK-N-SH human neuroblastoma cells performed in the present study showed that uptake of MIBG is inhibited by noradrenaline, more so by dopamine and to a lesser extent, by serotonin, indicating that the respective transporters may also contribute to MIBG uptake. However, neither dopamine nor serotonin transporter gene expression was detected. Noradrenaline transporter gene expression was found in 4 of 6 investigated cell lines, which correlated with specific MIBG uptake. Furthermore, an inverse correlation of noradrenaline transporter and tyrosine hydroxylase gene expression, the key regulatory enzyme of catecholamine synthesis, was observed. These data show that MIBG is specifically incorporated only in neuroblastoma cells in which there is noradrenaline transporter gene expression. Furthermore, the catecholamine status in neuroblastoma cells is regulated by a coordinate expression of the key elements of catecholamine synthesis and reuptake systems.

Expression of serotonin transporter messenger RNA in the human brain.

The characterization and cellular localization of human brain mRNA encoding the serotonin transporter were investigated using northern blot analysis and in situ hybridization histochemistry. In contrast to results from rodent studies, in which single hybridizing mRNAs are detected in brain and periphery, northern analysis of human midbrain raphe tissue revealed the presence of three mRNA species absent from samples prepared from substantia nigra/ventral tegmental area. In situ hybridization studies revealed dense hybridization signal corresponding to serotonin transporter mRNA highly localized to neurons of the dorsal and median raphe nuclei and the caudal linear nucleus. No hybridization signal was observed in neurons of the substantia nigra or locus ceruleus. These findings demonstrate the first anatomical visualization of human brain serotonin transporter gene expression and reveal heterogeneity associated with serotonin transporter transcripts, similar to, but distinct from the pattern of expression visualized in human peripheral tissues.