[Transculturation and validation of a Questionnaire Factors Influencing Organ Donation]. / Transculturación y validación del cuestionario Factores que influyen en la Donación de Órganos.

Background: There are validated questionnaires in Spanish that evaluate the factors that influence organ donation, but they are not designed for the open population or do not delve into various aspects such as the one proposed. Objective: Validate an instrument to evaluate the factors that influence organ donation in Mexico. Material and methods: Phase 1: Development of the instrument. Translation into Spanish of the questionnaire "Factors Inffluencing Organ Donation in Qatar", adapted by experts in donation and clinimetry. Simultaneously, the definitive version of the questionnaire "Factors that Influence Organ Donation" (FIDO) and the questionnaire "International Donor Collaborative Project" were applied to patients, relatives and staff of a tertiary hospital in Puebla, Mexico. Mind a week after 200 respondents. Cronbach's Alpha (AC) (internal consistency), Intraclass Correlation Coefficient (ICC) (external consistency), and Phi (Phi) and Chi square Coefficient (concurrent validity in intention to donate) were obtained. Results: AC and ICC by domain: Knowledge 0.625 and 0.372; Attitudes 0.776 and 0.761; Beliefs 0.649 and 0.633; Intentions 0.126 and 0.123; Phi: 0.976, Chi square: 2.358 (p = 0.125). Conclusions: The FIDO questionnaire is valid and consistent to assess knowledge, attitudes, beliefs and intentions in organ donation in the general Mexican population.

Introducción: existen cuestionarios validados en español que evalúan los factores que influyen en la donación de órganos, pero no están diseñados para población abierta o no exploran aspectos diversos como el propuesto. Objetivo: validar un instrumento para evaluar los factores que influyen en la donación de órganos en México. Material y métodos: fase 1: Elaboración del instrumento. Traducción al español del cuestionario Factors Influencing Organ Donation in Qatar, adaptado por expertos en donación y clinimetría. Se realizaron pruebas piloto hasta lograr acuerdo en dos rondas consecutivas. Fase 2: Validez y consistencia. Simultáneamente se aplicó la versión definitiva del cuestionario Factores que Influyen en la Donación de Órganos (FIDO) y el cuestionario Proyecto Colaborativo Internacional Donante a pacientes, familiares y personal de un hospital de tercer nivel de Puebla, México. Se reaplicó telefónicamente una semana después a 200 respondientes. Se utilizó alfa de Cronbach (AC) (consistencia interna), coeficiente de correlación intraclase (CCI) (consistencia externa), y coeficiente de Phi (Phi) y Chi cuadrada (validez concurrente en intención de donar). Resultados: AC y CCI por dominio: Conocimiento 0.625 y 0.372; Actitudes 0.776 y 0.761; Creencias 0.649 y 0.633; Intenciones 0.126 y 0.123; Global 0.774 y 0.675, respectivamente (p = 0.000); Phi: 0.976, Chi cuadrada: 2.358 (p = 0.125). Conclusiones: el cuestionario FIDO es válido y consistente para explorar: conocimiento, actitudes, creencias e intenciones en donación de órganos en población general mexicana.

Optimized design and in vivo application of optogenetically functionalized Drosophila dopamine receptors.

Neuromodulatory signaling via G protein-coupled receptors (GPCRs) plays a pivotal role in regulating neural network function and animal behavior. The recent development of optogenetic tools to induce G protein-mediated signaling provides the promise of acute and cell type-specific manipulation of neuromodulatory signals. However, designing and deploying optogenetically functionalized GPCRs (optoXRs) with accurate specificity and activity to mimic endogenous signaling in vivo remains challenging. Here we optimize the design of optoXRs by considering evolutionary conserved GPCR-G protein interactions and demonstrate the feasibility of this approach using two Drosophila Dopamine receptors (optoDopRs). These optoDopRs exhibit high signaling specificity and light sensitivity in vitro. In vivo, we show receptor and cell type-specific effects of dopaminergic signaling in various behaviors, including the ability of optoDopRs to rescue the loss of the endogenous receptors. This work demonstrates that optoXRs can enable optical control of neuromodulatory receptor-specific signaling in functional and behavioral studies.

Sodium influx induced by external calcium chelation decreases human sperm motility.

BACKGROUND: Calcium removal from the medium promptly reduces human sperm motility and induces a Na(+)-dependent depolarization that is accompanied by an increase in intracellular sodium concentration ([Na(+)](i)) and a decrease in intracellular calcium concentration ([Ca(2+)](i)). Sodium loading activates a Na(+)/K(+)-ATPase. METHODS: Membrane potential (Vm) and [Ca(2+)](i) were simultaneously detected in human sperm populations with the fluorescent probes diSC(3)(5) and fura 2. [Na(+)](i) and was measured independently in a similar fashion using sodium-binding benzofuran isophthalate. Motility was determined in a CASA system, ATP was measured using the luciferin-luciferase assay, and cAMP was measured by radioimmunoassay. RESULTS: Human sperm motility reduction after calcium removal is related to either Na(+)-loading or Na(+)-dependent depolarization, because, under conditions that inhibit the calcium removal-induced Na(+)-dependent depolarization and [Na(+)](i) increase, sperm motility was unaffected. By clamping sperm Vm with valinomycin, we found that the motility reduction associated with the calcium removal was related to sodium loading, and not to membrane potential depolarization. Mibefradil, a calcium channel blocker, markedly inhibited the Na(+)-dependent depolarization and sodium loading, and also preserved sperm motility. In the absence of calcium, both ATP and cAMP concentrations were decreased by 40%. However ATP levels were unchanged when calcium removal was performed under conditions that inhibit the calcium removal-induced Na(+)-dependent depolarization and [Na(+)](i) increase. CONCLUSIONS: Human sperm motility arrest induced by external calcium removal is mediated principally by sodium loading, which would stimulate the Na(+)/K(+)-ATPase and in turn deplete the ATP content.

Embarazo pretérmino intrabdominal exitoso: Reporte de caso / Successful intrabdominal preterm pregnancy: A case report

RESUMEN Introducción: El embarazo ectópico es toda gestación, en la que el sitio de implantación del óvulo fecundado se localiza fuera de la cavidad endometria y representa el 1.4 % de estos. Caso clínico: Paciente de 28 años con dos cesáreas previas, sometida a salpingoclasia durante la última intervención. La paciente ingresó al hospital con presión arterial de 180/130 mm Hg, no respondió a tratamiento médico y presentó falla en la progresión de trabajo de parto, por lo que se decidió interrupción por operación cesárea. Durante la cirugía, se observó un pequeño útero con embarazo extrauterino adherido a la serosa del ciego, el colon ascendente y el apéndice. Se obtuvo un producto femenino pretérmino vivo; la madre cursó con evolución favorable y sin complicaciones posoperatorias. Conclusión: La presentación del embarazo abdominal ectópico es rara, por lo que un control prenatal adecuado por personal capacitado puede orientar a la sospecha diagnóstica. La madre y la recién nacida no presentaron ninguna complicación, a pesar de ser un embarazo abdominal avanzado y la inserción multifocal de la placenta. Se resalta la importancia del manejo oportuno y multidisciplinario cuando se enfrentan embarazos con curso anormal para la mejor evolución de la madre y del producto.

ABSTRACT Introduction: Ectopic pregnancy is any gestation in which the implantation site of the fertilized egg is located outside the endometrial cavity. Abdominal ectopic pregnancy represents 1.4% of these. Case report: 28-year-old patient with two previous cesarean sections; bilateral tubal obstruction during the last operation. The patient was admitted to the hospital with blood pressure of 180/130 mm Hg that did not respond to medical treatment and not progression to labor so it was decided to interrupt the pregnancy by cesarean section. During surgery, a small uterus with extrauterine pregnancy was observed adhered to the serosa of the cecum, ascending colon, and appendix. A live preterm female product was obtained; the mother had a favorable evolution and no postoperative complications. Conclusion: Ectopic pregnancies are rera. An adequate prenatal control by well trained personnel is essential for an accurate diagnosis. The mother and the newborn did not present any complication. It is very important to have and accurate an opportune diagnosis so trained personnel can offer an adequate management.

Calidad de vida y características clínicas de pacientes con trasplante renal con y sin rinosinusitis crónica / Quality of life and clinical characteristics of renal transplant patients with and without chronic rhinosinusitis

RESUMEN Introducción: El trasplante es el tratamiento de elección en pacientes con enfermedad renal crónica (ERC). Requiere inmunosupresión, que predispone al desarrollo de complicaciones; la rinosinusitis crónica (RSC) es una de las más importantes. Objetivo: Comparar las características de pacientes con ERC con y sin RSC en protocolo de trasplante en un hospital de concentración en Puebla, en México. Métodos: Estudio comparativo, transversal, retrospectivo, en pacientes de un hospital de tercer nivel de atención, con ERC y en protocolo de trasplante renal. Se aplicaron las escalas SNOT-22 y Lund-Mackay. Se utilizó estadística descriptiva y pruebas U de Mann-Whitney, exacta de Fisher y coeficiente de Phi; p<0.05, por lo que se consideró significativa. Resultados: Se reclutaron 360 pacientes: 49 presentaron RSC; prevalencia, 13.61 %; medias edad, 39.22 ±12.09 años y tiempo de evolución, 17.73 ±5.91 semanas. Presentaron poliposis nasosinusal 14.3 %; obstrucción nasal, 95.9 %; algia facial, 67.3%; rinorrea, 49 % e hiposmia/anosmia, 40.8 %. Factores de riesgo asociados: alergia a ácido acetilsalicílico (p=0.014) y atopia (p=0.000). Variantes anatómicas en pacientes con y sin RSC, respectivamente: Celdilla Agger-Nasi 95 % y 15.4 %; desviación septal, 50 % y 6.4 %; hipertrofia de cornetes, 50 % y 1.3 %; concha bullosa, 30 % y 4.2 % y cornete paradójico, 10 % y 1.6 %. La afectación de calidad de vida predominante en pacientes con y sin RSC fue moderada con un 53.1 % y leve, 97.1 % (p=0.000). Conclusión: La prevalencia de RSC fue 13.61 %; los factores de riesgo asociados, alergia al ácido acetilsalicílico y atopia y la variante anatómica predominante, Agger-nasal. La severidad tomográfica fue leve y la afectación de la calidad de vida, moderada.

ABSTRACT Introduction: Transplantation is the treatment of choice for patients with chronic kidney disease (CKD). It requires immunosuppression, which predisposes to the development of complications; chronic rhinosinusitis (CRS) is one of the most significant. Objetive: To compare the characteristics of patients with CKD and without CRS in a renal transplant protocol at a specialized hospital in Puebla, Mexico. Methods: A comparative, cross-sectional, retrospective study in patients with CKD and in renal transplant protocol at a third-level care hospital. The SNOT-22 and Lund-Mackay scales were applied. Descriptive statistics, Mann-Whitney U tests, Fisher's exact test, and Phi coefficient were used; p<0.05 was considered significant. Results: 360 patients were recruited: 49 presented with CRS; prevalence: 13.61%; mean age: 39.22 ± 12.09 years and duration of progression: 17.73 ± 5.91 weeks. Naso-sinusal polyposis was present in 14.3%; nasal obstruction in 95.9%; facial pain in 67.3%; rhinorrhea in 49% and hyposmia/anosmia in 40.8%. Associated risk factors: allergy to acetylsalicylic acid (p=0.014) and atopy (p=0.000). Anatomical variants in patients with and without CRS, respectively: Agger nasi cell 95% and 15.4%; septal deviation, 50% and 6.4%; turbinate hypertrophy, 50% and 1.3%; concha bullosa, 30% and 4.2%; and paradoxical turbinate, 10% and 1.6%. The predominant quality of life impact in patients with and without CRS was moderate at 53.1% and mild at 97.1% (p=0.000). Conclusion: The prevalence of CRS was 13.61%; the associated risk factors were allergy to acetylsalicylic acid and atopy, and the predominant anatomical variant was Agger nasi. The tomographic severity was mild, and the impact on quality of life was moderate.

Intracellular sodium increase induced by external calcium removal in human sperm.

In human sperm, removal of external calcium produces a fast Na(+)-dependent depolarization that is presumably due to sodium permeation through calcium channels. Calcium restoration produces a ouabain-sensitive hyperpolarization that brings the membrane potential to values frequently more negative than resting. In this work, we show evidence indicating that external calcium removal induces an increase in the intracellular sodium ([Na(+)](i)) and that this phenomenon is related to the Na(+)-dependent depolarization. Calcium restoration blocked the [Na(+)](i) increase and then produced a slow decrease that was inhibited by ouabain. The [Na(+)](i) increase was inhibited by nanomolar-micromolar calcium or by millimolar magnesium, which has been previously shown to inhibit the Na(+)-dependent depolarization. This evidence supports the hypothesis that, in zero-calcium medium, a calcium channel that would contribute to resting intracellular calcium levels allows sodium permeation, producing depolarization and a significant [Na(+)](i) increase. Sodium loading would stimulate the Na(+),K(+)-ATPase, the activity of which contributes to the sperm hyperpolarization observed upon calcium restoration.

Evidence for simultaneous 1Na+:1Mg2+ and ping pong 2Na+:1Mg2+ exchangers in rat thymocyte.

Rat thymocytes showed two Na+/Mg2+ exchangers with high- and low- affinities for external Na+ (Na+o) at physiological internal Mg2+content. The total internal Mg2+ content (Mg2+it) was enhanced by loading with MgCl2 and the ionophore A-23187. Under these conditions, Na+/Mg2+ exchangers were dramatically stimulated by the Mg2+it increase. Na+-induced Mg2+ effluxes were independent of Cl-o or H+. The Na+/Mg2+ exchangers, which we named HANao (high affinity for Na+o) and LANao (low affinity for Na+o), were dissected in Mg2+-loaded thymocytes according to their kinetics and stoichiometries. HANao, which showed an apparent dissociation constant for Na+o (KNa H) = 9.2 +/- 1.6 mmol l(-1) Na+o and a maximal Na+ influx rate (VNa(Na H)max) = 30.5 +/- 6.1 mmol (l cells)(-1) h(-1), was a 1Na+:1Mg2+ simultaneous antiporter insensitive to external magnesium (Mg2+o) whereas that LANao, with KNa L = 65.1 +/- 8.6 mmol l(-1) Na+ and a VNa(Na L)max = 79.5 +/- 14.3 mmol (l cells)(-1) Na+ h(-1), was a 2Na+:1Mg2+ "ping-pong" antiporter which was strongly inhibited by Mg2+o. At physiological concentration of Mg2+o (1 mM), the Na+/Mg2+ exchange through the LANao was inhibited by approximately 50%. Amiloride (10(-4) M) inhibited at similar extent both Na+ and Mg2+ fluxes at high and at low Na+o.

Peroxynitrite activates glucose uptake in 3T3-L1 adipocytes through a PI3-K-dependent mechanism.

Peroxynitrite, the product of the reaction between *NO and O2*-, is a strong oxidant and nitrating molecule, and it has been recently consideredas a component of some important signaling pathways. Herein, we report the effect of peroxynitrite on glucose uptake in 3T3-L1 adipocytes. Peroxynitrite stimulated glucose uptake and this effect was inhibited by citochalasin B, indicating the participation of facilitated GLUT transporters. Peroxynitrite-induced glucose uptake was not related to intracellular ATP, nor to external or internal calcium, but it was inhibited by the phosphatidylinositol 3-kinase (PI3-K) inhibitor, wortmannin. Additionally, we also found that peroxynitrite did not activate the insulin receptor nor the PI3-K downstream signaling protein kinase B (PKB/Akt). The dose-dependent inhibitory action of wortmannin suggests that peroxynitrite activates glucose transport without affecting GLUT transporters translocation.

Fluorometric evidence for different stoichiometries for the Na+/Mg2+ exchange in Mg-loaded rat thymocytes.

The regulation of the cytosolic free magnesium concentration ([Mg2+]i) is a fundamental cellular process that requires magnesium extruding mechanisms. Here, we present evidence indicating that rat thymocytes are endowed with different Na/Mg exchange systems. Fluxes of magnesium were measured using the fluorescent magnesium indicator magfura-2. Cells were loaded with magnesium using the calcium ionophore A-23187 to 0.6-8.0 mM [Mg2+]i (resting [Mg2+]i = 0.38 +/- 0.06 mM, n = 5). The presence of extracellular sodium was required for magnesium exit. The initial rate of [Mg2+]i was stimulated by extracellular sodium with Michaelis-Menten kinetics. The Vmax of the sodium-dependent magnesium exit was markedly increased by [Mg2+]i. Holding the membrane potential at either -84 mV or at -10 mV had different effects on the sodium-stimulated magnesium efflux, depending on the extracellular sodium concentration ([Na+]o). At 10-30 mM [Na+]o, the magnesium efflux was faster at -10 mV than at -84 mV. Conversely, at 50-200 mM [Na+]o, the efflux of magnesium was faster at -84 mV that at -10 mV. At 75 mM [Na+]o, the activities where nearly the same at both membrane potential values. These observations suggest that the stochiometry of the Na+/Mg2+ exchange changes with [Na+]o.

Lack of voltage-dependent calcium channel opening during the calcium influx induced by progesterone in human sperm. Effect of calcium channel deactivation and inactivation.

Progesterone induces calcium influx and acrosomal exocytosis in human sperm. Pharmacologic evidence suggests that voltage-dependent calcium channels (VDCCs) are involved. In this study, membrane potential (Vm) and intracellular calcium concentration ([Ca(2+)](i)) were monitored simultaneously to assess the effect of VDCC gating on the calcium influx triggered by progesterone. Holding the Vm to values that maintained VDCCs in a deactivated (-71 mV) closed state inhibited the calcium influx induced by progesterone by approximately 40%. At this Vm, the acrosomal reaction induced by progesterone, but not by A23187, was inhibited. However, when the Vm was held at -15 mV (which maintains VDCCs in an inactivated closed state), the progesterone-induced calcium influx was stimulated. Furthermore, the progesterone and voltage-dependent calcium influxes were additive. These findings indicate that progesterone does not produce VDCC gating in human sperm.

Activation of protein kinase A stimulates the progesterone-induced calcium influx in human sperm exposed to the phosphodiesterase inhibitor papaverine.

Progesterone induces a fast transient calcium influx in human sperm though the activation of nongenomic receptors. During sperm capacitation, a complex process required for sperm to be able to fertilize the egg, the calcium influx induced by progesterone is enhanced. Sperm capacitation is mediated by an increase in cAMP content and subsequent protein kinase A (PKA) activation. In this work, we examined the effect of increasing intracellular cAMP on the calcium influx induced by progesterone in noncapacitated human sperm. To do this, sperm were exposed to the phosphodiesterase inhibitor papaverine for 5 minutes, a treatment that increased both the cAMP content and the PKA activity several-fold. The calcium influx induced by progesterone was increased by papaverine to levels close to those found in capacitated sperm. This effect was partially inhibited by H89 (48%) and by genistein (41%), and the sum of both inhibitors reduced the stimulating effect of papaverine by 89%. The inhibitory effect of genistein on the progesterone-induced calcium influx could be related to its capability to inhibit the papaverine-stimulated increase in cAMP content and PKA activity. The results presented here suggest that the calcium influx induced by progesterone is up-regulated by the PKA activity.

A remarkable increase in the pHi sensitivity of voltage-dependent calcium channels occurs in human sperm incubated in capacitating conditions.

Human sperm are endowed with voltage-dependent calcium channels (VDCC) that produce increases in [Ca2+]i in response to depolarization with KCl. These channels are stimulated during "capacitation", a complex biochemical process, accompanied by a slight pHi alkalization, that sperm must accomplish to acquire the ability to fertilize the egg. The stimulation can be explained in part by the fact that in non-capacitated sperm, calcium influx through VDCC is stimulated by pHi alkalization in the range of pHi observed during capacitation. In this work, we explored the effect of pHi on VDCC in capacitated sperm loaded with fura ff. Strikingly, the pHi sensitivity of VDCC increased approximately 7-fold when sperm was capacitated, as compared with non-capacitated sperm. This finding suggests that the pHi sensitivity of VDCC can be modulated during capacitation so that a combined effect of pHi alkalization and biochemical regulation enhances calcium influx through these channels.

Induction of a sodium-dependent depolarization by external calcium removal in human sperm.

Removal of external calcium with EGTA (from 2.5 mm to nanomolar levels) caused a remarkable depolarization in human sperm. This depolarization was initially fast. It was followed by a slow phase that brought the Vm to values of over 0 mV in 1-2 min. The slow and sustained phase correlated with a sustained decrease in intracellular calcium. However, calcium removal still induced depolarization in sperm with enhanced intracellular calcium (induced by progesterone), indicating that the sustained depolarization was not caused by a sustained intracellular calcium decrease. The depolarization was reduced as the external sodium content was substituted with choline, indicating that it was due to a sodium current, and was observed in lithium but not in tetramethylammonium-containing medium. In low sodium medium, the addition of sodium after calcium removal induced depolarization to the extent of which slightly increased in 2 min. The depolarization was completely inhibited by external magnesium (Ki = 1.16 mm). The addition of calcium or magnesium to calcium removal-induced depolarized sperm induced hyperpolarization that was inhibited by ouabain and was also prevented in medium without potassium, suggesting that the activity of the electrogenic Na+,K+-ATPase was involved. The conductance activated by calcium removal might unveil the presence of a calcium channel that in the absence of external calcium allows sodium permeation and that in normal conditions might contribute to the resting intracellular calcium concentration.

Effect of intracellular pH on depolarization-evoked calcium influx in human sperm.

Human sperm are endowed with putative voltage-dependent calcium channels (VDCC) that produce measurable increases in intracellular calcium concentration ([Ca(2+)](i)) in response to membrane depolarization with potassium. These channels are blocked by nickel, inactivate in 1-2 min in calcium-deprived medium, and are remarkably stimulated by NH(4)Cl, suggesting a role for intracellular pH (pH(i)). In a previous work, we showed that calcium permeability through these channels increases approximately onefold during in vitro "capacitation," a calcium-dependent process that sperm require to fertilize eggs. In this work, we have determined the pH(i) dependence of sperm VDCC. Simultaneous depolarization and pH(i) alkalinization with NH(4)Cl induced an [Ca(2+)](i) increase that depended on the amount of NH(4)Cl added. VDCC stimulation as a function of pH(i) showed a sigmoid curve in the 6.6-7.2 pH(i) range, with a half-maximum stimulation at pH approximately 7.00. At higher pH(i) (> or =7.3), a further stimulation occurred. Calcium release from internal stores did not contribute to the stimulating effect of pH(i) because the [Ca(2+)](i) increase induced by progesterone, which opens a calcium permeability pathway that does not involve gating of VDCC, was unaffected by ammonium. The ratio of pH(i)-stimulated-to-nonstimulated calcium influx was nearly constant at different test depolarization values. Likewise, depolarization-induced calcium influx in pH(i)-stimulated and nonstimulated cells was equally blocked by nickel. In our capacitating conditions pH(i) increased 0.11 pH units, suggesting that the calcium influx stimulation observed during sperm capacitation might be partially caused by pH(i) alkalinization. Additionally, a calcium permeability pathway triggered exclusively by pH(i) alkalinization was detected.

Stimulation of voltage-dependent calcium channels during capacitation and by progesterone in human sperm.

To fertilize, mammalian sperm must undergo two sequential steps that require activation of calcium entry mechanisms, capacitation and acrosomal exocytosis, induced in the latter case by the egg zona pellucida glycoprotein ZP3 or by progesterone. Voltage-dependent calcium channels (VDCC) could participate in these processes. Since patch clamp recordings are extremely difficult in mature sperm, the activity of VDCC has been alternatively analyzed with optical detectors of membrane potential and intracellular calcium in sperm populations. Using this approach, we previously reported that in human sperm there is a voltage-dependent calcium influx system that strongly indicates that human sperm are endowed with functional VDCC. In this study we developed evidence indicating that calcium influx through VDCC is significantly stimulated during sperm in vitro capacitation and by progesterone action, which is present in the follicular fluid that surrounds the egg. The observed effects of capacitation and progesterone on VDCC may be physiologically significant for sperm-egg interaction.