Altered Circadian Rhythm of Melatonin Concentrations in Hypocretin-Deficient Men

Hypocretin deficiency causes narcolepsy. It is unknown whether melatonin secretion is affected in this sleep disorder. Therefore, in both narcolepsy patients and matched controls, the authors measured plasma melatonin levels hourly for 24?h before and after 5 days of sodium oxybate (SXB) administration. Although mean melatonin concentrations were similar between patients and controls, in narcoleptics the percentage of 24-h melatonin secreted during the daytime was significantly higher, and melatonin secretion exhibited a weaker coupling to sleep. SXB did not affect melatonin secretion. These findings suggest that hypocretin deficiency might disturb both the circadian control of melatonin release and its temporal association with sleep. (Author correspondence: c.e.h.m.donjacour@lumc.nl)     

Role of oxidants on calcium and sodium movement in healthy and diseased cardiac myocytes

In this review article we give an overview of current knowledge with respect to redox-sensitive alterations in Na⁺ and Ca²⁺ handling in the heart. In particular, we focus on redox-activated protein kinases including cAMP-dependent protein kinase A (PKA), protein kinase C (PKC), and Ca/calmodulin-dependent protein kinase II (CaMKII), as well as on redox-regulated downstream targets such as Na⁺ and Ca²⁺ transporters and channels. We highlight the pathological and physiological relevance of reactive oxygen species and some of its sources (such as NADPH oxidases, NOXes) for excitation—contraction coupling (ECC). A short outlook with respect to the clinical relevance of redox-dependent Na⁺ and Ca²⁺ imbalance will be given.     

Guanine-induced inhibition of renal Na(+)-ATPase activity: evidence for the involvement of the Gi protein-coupled receptor

There is some evidence to show a possible role of guanosine in the modulation of cellular function, in particular, in the neuronal system. However, nothing is known about the role of guanine in renal function. The aim of the present work was to investigate the role of guanine on modulation of Na⁺-ATPase activity in isolated basolateral membrane (BLM) of the renal cortex. Guanine inhibited the enzyme activity in a dose-dependent manner with maximal effect (56%) obtained at 10⁻⁶ M. This effect was reversed by DPCPX (8-cyclopentyl-1,3-dipropylxanthine), an antagonist of A₁ receptors, but it was not changed by 10⁻⁸ M DMPX (3,7-dimethyl-1-propargylxanthine) or 10⁻⁸ M MRS (2,3-diethyl-4,5-dipropyl-6-phenylpyridine-3-thiocarboxylate-5-carboxylate), antagonists of A₂ and A₃ receptors, respectively. Furthermore, it was observed that guanine increased [γ-³⁵S]GTP-specific binding with the maximal effect observed at 10⁻⁶ M and this effect was abolished by 10⁻⁶ M GDPβS. The inhibitory effect of 10⁻⁶ M guanine on Na⁺-ATPase activity was reversed by 10⁻⁶ M GDPβS, 10⁻⁶ M forskolin, 10⁻⁶ M pertussis toxin and 10⁻⁸ M cholera toxin. These results indicate that guanine binds to a DPCPX-sensitive receptor promoting the activation of Gi protein and leading to a decrease in cAMP level and, consequently, inhibition of BLM Na⁺-ATPase activity.     

On the surface properties of oleate micelles and oleic acid/oleate vesicles studied by spin labeling

Dilute aqueous systems composed of sodium oleate micelles and sodium oleate/oleic acid vesicles were investigated as a function of pH by electron spin resonance spectroscopy with TEMPO-stearate TEMPO-stearamide as well as with a positively charged water soluble spin label, TEMPO-choline. The dynamics of the three TEMPO-spin labels were found to be sensitive to changes in the interfacial region of the aggregates as a function of pH. The results obtained are consistent with the formation of a hydrogen bond network (RCOO⁻ ↔ HOOCR) at the surface of the sodium oleate/oleic acid system in the course of the transformation of micelles into the closed bilayers (vesicles). Vesicles formation below pH 10 was determined independently with a spin labeled glucose derivative.     

Immunohistological localization of serotonin in the CNS and feeding system of the stable fly Stomoxys calcitrans L. (Diptera: Muscidae)

Serotonin, or 5-hydroxytryptamine (5-HT), plays critical roles as a neurotransmitter and neuromodulator that control or modulate many behaviors in insects, such as feeding. Neurons immunoreactive (IR) to 5-HT were detected in the central nervous system (CNS) of the larval and adult stages of the stable fly, Stomoxys calcitrans, using an immunohistological technique. The location and pattern of the 5-HT IR neurons are described and compared for these two different developmental stages. Anatomical features of the fly feeding system were analyzed in third instar larvae and adult flies using a combination of histological and immunohistological techniques. In third instar larvae, the cibarial dilator muscles were observed within the cibarial pump skeleton and innervated by 5-HT IR neurons in nerves arising from the brain. There were four pairs of nerves arising from the frontal surface of the larval brain that innervate the cibarial pump muscles, pharynx, and muscles controlling the mouth hooks. A strong serotoninergic innervation of the anterior stomatogastric system was observed, which suggests 5-HT may play a role in the coordination of different phases of food ingestion by larvae. Similarly, many 5-HT IR neurons were found in both the brain and the thoracico-abdominal ganglia in the adult, some of which innervate the cibarial pump dilator muscles and the stomatogastric muscles. This is tnhe first report describing neuromuscular structures of the stable fly feeding system. The results reported here suggest 5-HT may play a critical role in feeding behaviors of stable fly larvae and adults.     

Inhibition of c-Kit, VEGFR-2 (KDR), and ABCG2 by analogues of OSI-930

The quinoline domain of OSI-930, a dual inhibitor of receptor tyrosine kinases (RTKs) c-Kit and KDR, was modified in an effort to further understand the SAR of OSI-930, and the binding site characteristics of c-Kit and KDR. A series of 16 compounds with heteroatom substituted pyridyl and phenyl ring systems was synthesized and evaluated against a panel of kinases including c-Kit and KDR. Aminopyridyl derivative 6 was found to be the most active member of the series with 91% and 57% inhibition of c-Kit at 10μM and 1μM, respectively and 88% and 50% inhibition of KDR at 10μM and 1μM, respectively. The target compounds were also tested for their ability to inhibit efflux of mitoxantrone through inhibition of ATP dependent ABCG2 pump. Nitropyridyl derivative 5 and o-nitrophenyl derivative 7 exhibited complete inhibition of the ABCG2 pump with IC(50) values of 13.67μM and 16.67μM, respectively.     

Synthesis and DNA photocleavage by a pyridine-linked bis-acridine chromophore in the presence of copper(II): ionic strength effects

We report the synthesis of photonuclease 3 consisting of two acridine rings joined by a 2,6-bis(aminomethyl)pyridine copper-binding linker. In reactions containing micromolar concentrations of 3, irradiation at 419 nm produces efficient, copper(II)-dependent cleavage of plasmid DNA in the presence of the high concentrations of salt that exist in the cell nucleus (150 mM NaCl and 260 mM KCl). The DNA interactions of 3 are compared to an analogous bis-acridine (4) containing a more flexible 2,6-bis{[(methoxycarbonylamino)-ethyl]methylaminomethyl}pyridine unit.     

Rings of charge within the extracellular vestibule influence ion permeation of the 5-HT3A receptor

The determinants of single channel conductance (γ) and ion selectivity within eukaryotic pentameric ligand-gated ion channels have traditionally been ascribed to amino acid residues within the second transmembrane domain and flanking sequences of their component subunits. However, recent evidence suggests that γ is additionally controlled by residues within the intracellular and extracellular domains. We examined the influence of two anionic residues (Asp(113) and Asp(127)) within the extracellular vestibule of a high conductance human mutant 5-hydroxytryptamine type-3A (5-HT(3)A) receptor (5-HT(3)A(QDA)) upon γ, modulation of the latter by extracellular Ca(2+), and the permeability of Ca(2+) with respect to Cs(+) (P(Ca)/P(Cs)). Mutations neutralizing (Asp → Asn), or reversing (Asp → Lys), charge at the 113 locus decreased inward γ by 46 and 58%, respectively, but outward currents were unaffected. The D127N mutation decreased inward γ by 82% and also suppressed outward currents, whereas the D127K mutation caused loss of observable single channel currents. The forgoing mutations, except for D127K, which could not be evaluated, ameliorated suppression of inwardly directed single channel currents by extracellular Ca(2+). The P(Ca)/P(Cs) of 3.8 previously reported for the 5-HT(3)A(QDA) construct was reduced to 0.13 and 0.06 by the D127N and D127K mutations, respectively, with lesser, but clearly significant, effects caused by the D113N (1.04) and D113K (0.60) substitutions. Charge selectivity between monovalent cations and anions (P(Na)/P(Cl)) was unaffected by any of the mutations examined. The data identify two key residues in the extracellular vestibule of the 5-HT(3)A receptor that markedly influence γ, P(Ca)/P(Cs), and additionally the suppression of γ by Ca(2+).     

A conserved asparagine residue in transmembrane segment 1 (TM1) of serotonin transporter dictates chloride-coupled neurotransmitter transport

Na(+)- and Cl(-)-dependent uptake of neurotransmitters via transporters of the SLC6 family, including the human serotonin transporter (SLC6A4), is critical for efficient synaptic transmission. Although residues in the human serotonin transporter involved in direct Cl(-) coordination of human serotonin transport have been identified, the role of Cl(-) in the transport mechanism remains unclear. Through a combination of mutagenesis, chemical modification, substrate and charge flux measurements, and molecular modeling studies, we reveal an unexpected role for the highly conserved transmembrane segment 1 residue Asn-101 in coupling Cl(-) binding to concentrative neurotransmitter uptake.