Region‐specific regulation of 5‐HT1A receptor expression by Pet‐1‐dependent mechanisms in vivo

Abstract

J. Neurochem. (2011) 116, 1066–1076. Abstract Serotonin (5‐hydroxytryptamine, 5‐HT) neurotransmission is negatively regulated by 5‐HT1A autoreceptors on raphe neurons, and is implicated in mood disorders. Pet‐1/FEV is an ETS transcription factor expressed exclusively in serotonergic neurons and is essential for serotonergic differentiation, although its regulation of 5‐HT receptors has not yet been studied. Here, we show by electrophoretic mobility shift assay that recombinant human Pet‐1/FEV binds directly to multiple Pet‐1 elements of the human 5‐HT1A receptor promoter to enhance its transcriptional activity. In luciferase reporter assays, mutational analysis indicated that while several sites contribute, the Pet‐1 site at ‐1406 bp had the greatest effect on 5‐HT1A promoter activity. To address the effect of Pet‐1 on 5‐HT1A receptor regulation in vivo, we compared the expression of 5‐HT1A receptor RNA and protein in Pet‐1 null and wild‐type littermate mice. In the raphe nuclei of Pet‐1−/− mice tryptophan hydroxylase 2 (TPH2) RNA, and 5‐HT and TPH immunostaining were greatly reduced, indicating a deficit in 5‐HT production. Raphe 5‐HT1A RNA and protein levels were also reduced in Pet‐1‐deficient mice, consistent with an absence of Pet‐1‐mediated transcriptional enhancement of 5‐HT1A autoreceptors in serotonergic neurons. Interestingly, 5‐HT1A receptor expression was up‐regulated in the hippocampus, but down‐regulated in the striatum and cortex. These data indicate that, in addition to transcriptional regulation by Pet‐1 in raphe neurons, 5‐HT1A receptor expression is regulated indirectly by alterations in 5‐HT neurotransmission in a region‐specific manner that together may contribute to the aggressive/anxiety phenotype observed in Pet‐1 null mice.

Increased Serotonin-1A (5-HT1A) Autoreceptor Expression and Reduced Raphe Serotonin Levels in Deformed Epidermal Autoregulatory Factor-1 (Deaf-1) Gene Knock-out Mice

“…TPH2) in 5-HT neurons was not affected in Deaf-1 knock-outs, arguing against a nonspecific global effect, which would be expected to produce a general inhibition or induction of 5-HT1A and other genes. For example, a developmental abnormality that leads to impaired serotonergic differentiation such as Pet-1 knock-out, globally reduces 5-HT1A and other serotonin markers like TPH2 (37,38), which we did not observe in the Deaf-1 Ϫ/Ϫ mice. Thus, the effects of Deaf-1 deficiency that we observed are both cell-and gene-specific and indicate a direct effect.…”

The Native Serotonin 5-HT5AReceptor: Electrophysiological Characterization in Rodent Cortex and 5-HT1A-Mediated Compensatory Plasticity in the Knock-Out Mouse

“…Equal amounts of denatured protein extracts (20 g) were separated by SDS-PAGE on 12% gels and transferred to nitrocellulose membranes. Membranes were incubated overnight at 4°C with an anti-5-HT 1A receptor polyclonal primary antibody (1:4000, AB15350; Millipore) (Jacobsen et al, 2011), incubated for 1 h with a peroxidaseconjugated secondary antibody (1:7000; Jackson Immunoresearch), and visualized using chemiluminescence. Band intensities were quantified using ImageJ and normalized to ␤-actin.…”

Transcriptional Dys-regulation in Anxiety and Major Depression: 5-HT1A Gene Promoter Architecture as a Therapeutic Opportunity

“…Pet-1 is a transcription factor that is expressed exclusively in raphe nuclei and is required for differentiation of serotonin neurons and expression of serotonergic genes [154][155][156]. Several conserved Pet-1 sites are present in the 5-HT1A promoter (Fig.…”

“…4), but the upstream site (−1400 bp) shows the greatest conservation ( Fig. 5B) and is the most critical for raphespecific expression of the 5-HT1A autoreceptor [156]. Due to its exclusive expression in 5-HT neurons in the brain, inhibiting Pet-1 may be useful to reduce 5-HT1A autoreceptor levels (Box 4), but since Pet-1 also affects TPH and 5-HTT levels, inhibiting Pet-1 could undesirably reduce 5-HT levels [155].…”