Fluorescent nucleoside derivatives as a tool for the detection of concentrative nucleoside transporter activity using confocal microscopy and flow cytometry.

The abundance and function of transporter proteins at the plasma membrane are likely to be crucial in drug responsiveness. Functional detection of human concentrative nucleoside transporters (hCNTs) is of interest for predicting drug sensitivity because of their ability to transport most nucleoside-derived drugs. In the present study, two fluorescent nucleoside analogues, uridine-furan and etheno-cytidine, were evaluated as tools to study in vivo nucleoside transporter-related functions. These two molecules showed high affinity interactions with hCNT1 and hCNT3 and were shown to be substrates of both transporters. Both fluorescence microscopy and flow cytometry experiments showed that uridine-furan uptake was better suited for distinguishing cells that express hCNT1 or hCNT3. These data highlight the usefulness of fluorescent nucleoside derivatives, as long as they fulfill the requirements of confocal microscopy and flow cytometry, for in vivo analysis of hCNT-related function.

Further characterization of the electrogenicity and pH sensitivity of the human organic anion-transporting polypeptides OATP1B1 and OATP1B3.

Organic anion-transporting polypeptides (OATPs) are involved in the liver uptake of many endogenous and xenobiotic compounds, such as bile acids and drugs, respectively. Using Xenopus laevis oocytes and Chinese hamster ovary (CHO) cells expressing rat Oatp1a1, human OATP1B1, or OATP1B3, the sensitivity of these transporters to extracellular/intracellular pH (pHo/pHi) and changes in plasma membrane potential (ΔΨ) was investigated. In X. laevis oocytes, nonspecific plasma membrane permeability increased only at pHo below 4.5. Above this value, both using oocytes and CHO cells, extracellular acidification affected differently the specific transport of taurocholic acid (TCA) and estradiol 17ß-d-glucuronide (E(2)17ßG) by Oatp1a1 (stimulation), OATP1B1 (inhibition), and OATP1B3 (stimulation). Changes in substrate uptake in the presence of valinomycin (K(+)-ionophore), carbonyl cyanide 3-chlorophenylhydrazone and nigericin (protonophores), and amiloride (Na(+)/H(+)-inhibitor) and cation replacement in the medium were studied with fluorescent probes for measuring substrate uptake (cholylglycyl amidofluorescein) and changes in pHi (SNARF-4F) and ΔΨ [DilC(1)(5)]. The results suggest that activity of these three carriers is sodium/potassium-independent and affected differently by changes in pHo and ΔΨ: Oatp1a1 was confirmed to be an electroneutral anion exchanger, whereas the function of both OATP1B1 and OATP1B3 was markedly affected by the magnitude of ΔΨ. Moreover, electrophysiological measurements revealed the existence of a net anion influx associated to OATP1B1/OATP1B3-mediated transport of TCA, E(2)17ßG, and estrone-3-sulfate. Furthermore, a leakage of Na(+) through OATP1B1 and OATP1B3, which is not coupled to substrate transport, was found. In conclusion, these results suggest that OATP1B1 and OATP1B3 are electrogenic transporters whose activity may be strongly affected under circumstances of displacement of local pH.

[Assessment of comorbidity and social anxiety in adolescents with attention deficit hyperactivity disorder: the SELFIE study]. / Evaluación de la comorbilidad y la ansiedad social en adolescentes con trastorno por déficit de atención con hiperactividad: Estudio SELFIE.

INTRODUCTION: Attention-deficit/hyperactivity disorder (ADHD) and its comorbidities have an impact on the social anxiety of children and adolescents, but there are practically no studies addressing this topic in adolescence. Our objective was to assess the degree of social anxiety and to analyse the presence of psychiatric comorbidities (PSCs) in adolescents with ADHD. METHODOLOGY: We conducted a cross-sectional observational study in patients aged 12 to 18 years with a confirmed diagnosis of ADHD (DSM-5). We collected data on the presence and type of PSCs and assessed social anxiety by means of the Social Anxiety Scale for Adolescents (SAS-A). RESULTS: Forty-six child and adolescent psychiatrists and paediatric neurologists participated in the study and recruited 234 patients. Of the total patients, 68.8% (159) were male and 31.2% (72) female, with a mean age in the sample of 14.9 years (95% CI, 14.6-15.1). The type of ADHD was combined type (C) in 51.7% (121), predominantly inattentive (PI) in 37.2% (87), and predominantly hyperactive-impulsive (PH) in 9% (21). Of all patients, 97.9% (229) received pharmacological therapy: 78.6% (184) methylphenidate, 15% (35) lisdexamfetamine and 4.3% (10) atomoxetine.We found PSCs in 50.4% of the patients (118), of which the most frequent were learning and communication disorders (20.1%, n=47) and anxiety disorders (19.2%, n=45). The patients scored significantly higher in the SAS-A compared to reference values in the healthy population. The scores in the SAS-A were less favourable in adolescents with the PI type compared to those with the PH type (P=.015). The presence of a comorbid anxiety disorder was associated with worst scores in SAS-A (P<.001) showing an increased social anxiety. CONCLUSION: Adolescents with ADHD classified as PI and those with comorbid anxiety had a higher degree of social anxiety as measured by the SAS-A. This psychological aspect must be identified and controlled in adolescents with ADHD to promote their social adaptation.