Serotonergic Actions and Interactions on the SCN Circadian Pacemaker: In Vitro Investigations

The phase of the mammalian circadian pacemaker located in the suprachiasmatic nuclei (SCN) is controlled by a multitude of stimuli. While phase control is undoubtedly dominated by photic input, the serotonergic input from the raphe nuclei also influences SCN clock phase. In this article I review the evidence for serotonergic modulation of the SCN pacemaker, and the cellular mechanisms underlying these effects, obtained from in vitro experiments performed during the past decade. Serotonin can advance the SCN pacemaker when applied during the subjective day, and delay the pacemaker when applied during the subjective night. The daytime advances appear due to stimulation of 5HT₇ receptors, activation of adenylate cyclase and protein kinase A, and opening of K⁺ channels. The synthesis of new proteins may also be critical for these phase shifts. Serotonergic phase advances can be inhibited by a variety of other modulatory inputs to the SCN, including neuropeptide Y, melatonin, and glutamate. Together, these data demonstrate that SCN circadian pacemaker phase is controlled by a complex interplay between multiple afferent stimuli, and that serotonin plays a critical role in this process.     

Allelic variation in the serotonin transporter (5HTT) gene contributes to idiopathic pulmonary hypertension in children

Pulmonary hypertension is a potentially lethal condition, which affects adults and children alike. Genetic factors are implicated in the causation of primary pulmonary hypertension. We investigate the role of polymorphism in the 5HTT gene in the etiology of pulmonary hypertension in children aged 1-18.8 years. We have tested the hypothesis that the 5HTT gene does contribute to the pathogenesis of this disease in children by comparing the allelic frequencies of both the long and short variants between children with idiopathic pulmonary hypertension and pulmonary hypertension secondary to underlying pulmonary disease. We found that homozygosity for the long variant of 5HTT was highly associated with idiopathic pulmonary hypertension in children, suggesting perhaps a more important role for 5HTT gene function in the pathogenesis of early onset disease.     

Intrastriatal Injection of 5,7-Dihydroxytryptamine Decreased 5-HT Levels in the Striatum and Suppressed Locomotor Activity in C57BL/6 Mice

Effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 5,7-dihydroxytryptamine (5,7-DHT) on striatal levels of dopamine (DA), 5-hydroxytryptamine (5-HT), and their metabolites, as well as on locomotor activity were investigated in C57BL/6 mice. The results showed that MPTP significantly increased locomotor activity and decreased striatal DA levels. However, injection of the serotonergic neurotoxin 5,7-DHT in the striatum, either alone or following high doses of MPTP, significantly decreased locomotor activity, and concomitantly decreased striatal levels of 5-HT and 5-HIAA. This study suggests that the increased locomotor activity may be due to increased striatal serotonergic activity which overcompensates for the DA deficiency. The locomotor hypoactivity, induced by 5,7-DHT, might be due to the decreased striatal levels of 5-HT and 5-HIAA.     

Discovery of new anti-depressants from structurally novel 5-HT3 receptor antagonists: design, synthesis and pharmacological evaluation of 3-ethoxyquinoxalin-2-carboxamides

A novel series of 3-ethoxyquinoxalin-2-carboxamides were designed as per the pharmacophoric requirements of 5-HT₃ receptor antagonist using ligand-based approach. The desired carboxamides were synthesized from the key intermediate, 3-ethoxyquinoxalin-2-carboxylic acid by coupling with appropriate amines in the presence of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl) and 1-hydroxybenzotriazole (HOBt). The 5-HT₃ receptor antagonism was evaluated in longitudinal muscle myenteric plexus preparation from guinea pig ileum against 5-HT₃ agonist, 2-methy-5-HT, which was expressed in the form of pA₂ values. Compound 6h (3-ethoxyquinoxalin-2-yl)(4-methylpiperazin-1-yl)methanone was found to be the most active compound, which expressed a pA₂ value of 7.7. In forced swim test, the compounds with higher pA₂ value exhibited good anti-depressant-like activity and compounds with lower pA₂ value failed to show activity as compared to the vehicle-treated group.     

Locomotor activity is regulated by D2-like receptors in Drosophila: an anatomic and functional analysis

In mammals, dopamine 2-like receptors are expressed in distinct pathways within the central nervous system, as well as in peripheral tissues. Selected neuronal D2-like receptors play a critical role in modulating locomotor activity and, as such, represent an important therapeutic target (e.g. in Parkinson's disease). Previous studies have established that proteins required for dopamine (DA) neurotransmission are highly conserved between mammals and the fruit fly Drosophila melanogaster. These include a fly dopamine 2-like receptor (DD2R; Hearn et al. PNAS 2002 99(22):14554) that has structural and pharmacologic similarity to the human D2-like (D2R). In the current study, we define the spatial expression pattern of DD2R, and functionally characterize flies with reduced DD2 receptor levels. We show that DD2R is expressed in the larval and adult nervous systems, in cell groups that include the Ap-let cohort of peptidergic neurons, as well as in peripheral tissues including the gut and Malpighian tubules. To examine DD2R function in vivo, we generated RNA-interference (RNAi) flies with reduced DD2R expression. Behavioral analysis revealed that these flies show significantly decreased locomotor activity, similar to the phenotype observed in mammals with reduced D2R expression. The fly RNAi phenotype can be rescued by administration of the DD2R synthetic agonist bromocriptine, indicating specificity for the RNAi effect. These results suggest Drosophila as a useful system for future studies aimed at identifying modifiers of dopaminergic signaling/locomotor function.     

Usabamycins A-C: new anthramycin-type analogues from a marine-derived actinomycete

New anthramycin-type analogues, designated usabamycin A-C (1, 2 and 3), have been isolated from cultures of Streptomyces sp. NPS853, a bacterium found in marine sediments. The structures of the new compounds were established on the basis of extensive spectroscopic analyses including 1D- and 2D-NMR ((1)H-(1)H COSY, HSQC, and HMBC) experiments. The usabamycins show weak inhibition of HeLa cell growth and selective inhibition of serotonin (5-hydroxytrypamine) 5-HT(2B) uptake.     

Antibodies raised against different extracellular loops of the melanocortin-3 receptor affect energy balance and autonomic function in rats

Melanocortin receptors (MCR) play an important role in the regulation of energy balance and autonomic function. In the present studies, we used active immunization against peptide sequences from the first and the third extracellular loop (EL1 and EL3) of the MC3R to generate selective antibodies (Abs) against this MCR subtype in rats. Immunization with the EL1 peptide resulted in Abs that enhanced the effects of the endogenous ligand α-melanocyte-stimulating hormone (α-MSH), whereas immunization with the EL3 peptide resulted in Abs acting as non-competitive antagonists. The phenotype of immunized rats chronically instrumented with telemetry transducers was studied under four different conditions: a high-fat diet was followed by standard lab chow, by fasting, and finally by an intraperitoneal injection of lipopolysaccharide (LPS). Under high-fat diet, food intake and body weight were higher in the EL3 than in the EL1 or the control group. Blood pressure was increased in EL3 rats and locomotor activity was reduced. Plasma concentrations of triglycerides, insulin, and leptin tended to rise in the EL3 group. After switching to standard lab chow, the EL1 group showed a small significant increase in blood pressure that was more pronounced and associated with an increase in heart rate during food restriction. No differences between the EL1 or the EL3 group were observed after LPS injection. These results show that immunization against the MC3R resulted in the production of Abs with positive or negative allosteric properties. The presence of such Abs induced small changes in metabolic and cardiovascular parameters.     

A Role for Testosterone in the Maintenance of Seasonally Appropriate Body Mass but not in Lipectomy-Induced Body Fat Compensation in Siberian Hamsters

The purpose of this study was to test whether serum testosterone (T) concentrations characteristic of reproductively active, long-day-housed Siberian hamsters are necessary for compensatory increases in nonexcised fat pads following removal of epididy-mal white adipose tissue (EWAT) and/or for the maintenance of seasonally appropriate body weights in these hamsters. Long-day-housed hamsters were castrated or left intact, sham or EWAT lipectomized, and given T or cholesterol (C) implants. All groups had ad libitum food access except for two castrated T-treated groups that were pair-fed to their C-treat-ed counterparts to control for effects of T on food intake. C-treated castrates had decreased body weights compared with all other groups, suggesting a role of T in the maintenance of seasonally appropriate body mass. Since the T-treated hamsters pair-fed to these T-deficient animals exhibited seasonally appropriate body weights and fat pad masses, T does not appear to affect these parameters through the modulation of food intake. All fat pads of C-treated animals were smaller than those of ad libitum- or pair-fed, T-treated castrates; however, EWAT was the only fat pad that was smaller in the C-treated sham-lipectomized group than in gonad-intact sham-lipectomized hamsters. This result may indicate an enhanced sensitivity of EWAT to T. The effects of T on fat pad mass were not associated with proportionate changes in lipoprotein lipase activity, suggesting that the major effect of T on fat accumulation occurs through other mechanisms in this species. C-treated lipectomized hamsters compensated for the body fat deficit 8 weeks after lipectomy via statistically nonsignificant increases in retroperitoneal and inguinal WAT mass. This finding suggests that, whereas T is necessary for maintenance of seasonally-appropriate body weight, it is not necessary for fat pad compensation after EWAT lipectomy.     

Effects of dark-light step transitions on circadian locomotor activity rhythms of the hagfish,Eptatretus burgeri

Captive hagfish,Eptatretus burgeri, were subjected to step transitions from continuous dark (DD) to continuous light (LL) and their locomotor activity patterns recorded. Free-running activity rhythms occurred in both the DD and LL regimes. The timing of the transition influenced the circadian period (τ), the relationship between the period response (δτ, or τ{inLL}–τ{inDD}, change in the circadian period) and the former being represented by a cosine curve δτ became most positive and most negative when hagfish underwent a DD-LL transition shortly before the beginning of the duration of activity (α) and shortly after the end of such, respectively. The phase response (δ-phase, change in the relative timing of the activity phase) was characterized by a delay in the activity phase after the DD-LL transition, although its magnitude bore no relation to the timing of the transition.