Tau reduction with artificial microRNAs modulates neuronal physiology and improves tauopathy phenotypes in mice.

Abnormal tau accumulation is the hallmark of several neurodegenerative diseases, named tauopathies. Strategies aimed at reducing tau in the brain are promising therapeutic interventions, yet more precise therapies would require targeting specific nuclei and neuronal subpopulations affected by disease while avoiding global reduction of physiological tau. Here, we developed artificial microRNAs directed against the human MAPT mRNA to dwindle tau protein by engaging the endogenous RNA interference pathway. In human differentiated neurons in culture, microRNA-mediated tau reduction diminished neuronal firing without affecting neuronal morphology or impairing axonal transport. In the htau mouse model of tauopathy, we locally expressed artificial microRNAs in the prefrontal cortex (PFC), an area particularly vulnerable to initiating tau pathology in this model. Tau knockdown prevented the accumulation of insoluble and hyperphosphorylated tau, modulated firing activity of putative pyramidal neurons, and improved glucose uptake in the PFC. Moreover, such tau reduction prevented cognitive decline in aged htau mice. Our results suggest target engagement of designed tau-microRNAs to effectively reduce tau pathology, providing a proof of concept for a potential therapeutic approach based on local tau knockdown to rescue tauopathy-related phenotypes.

SMaRT modulation of tau isoforms rescues cognitive and motor impairments in a preclinical model of tauopathy.

Tau is a microtubule-associated protein predominantly expressed in neurons, which participates in microtubule polymerization and axonal transport. Abnormal tau metabolism leads to neurodegenerative diseases named tauopathies, such as Alzheimer's disease and frontotemporal dementia. The alternative splicing of exon 10 (E10) in the primary transcript produces tau protein isoforms with three (3R) or four (4R) microtubule binding repeats, which are found in equal amounts in the normal adult human brain. Several tauopathies are associated with abnormal E10 alternative splicing, leading to an imbalance between 3R and 4R isoforms, which underlies disease. Correction of such imbalance represents a potential disease-modifying therapy for those tauopathies. We have previously optimized a trans-splicing RNA reprogramming strategy to modulate the 3R:4R tau content in a mouse model of tauopathy related to tau mis-splicing (htau mice), and showed that local modulation of E10 inclusion in the prefrontal cortex prevents cognitive decline, neuronal firing impairments and hyperphosphorylated tau accumulation. Furthermore, local shifting of 3R-4R tau into the striatum of htau mice prevented motor coordination deficits. However, a major bottleneck of our previous work is that local splicing regulation was performed in young mice, before the onset of pathological phenotypes. Here we tested whether regulation of tau E10 splicing could rescue tau pathology phenotypes in htau mice, after the onset of cognitive and motor impairments, comparable to early stages of human tauopathies. To determine phenotypic time course and affected brain nuclei, we assessed htau mice using behavioural tests and microPET FDG imaging over time, similarly to diagnosis methods used in patients. Based on these analyses, we performed local delivery of pre-trans splicing molecules to regulate E10 inclusion either into the medial prefrontal cortex (mPFC) or the striatum at 6-month-old once behavioral phenotypes and metabolic changes were detected. Tau isoforms modulation into the mPFC restored cognitive performance in mice that previously showed mild to severe memory impairment while motor coordination deficit was rescued after striatal injection of trans-splicing molecules. Our data suggest that tau regulation could recover pathological phenotypes early after phenotypic onset, raising promising perspectives for the use of RNA based therapies in tauopathies related to MAPT abnormal splicing.

Tau mis-splicing correlates with motor impairments and striatal dysfunction in a model of tauopathy.

Tauopathies are neurodegenerative diseases caused by the abnormal metabolism of the microtubule associated protein tau (MAPT), which is highly expressed in neurons and critically involved in microtubule dynamics. In the adult human brain, the alternative splicing of exon 10 in MAPT pre-mRNA produces equal amounts of protein isoforms with either three (3R) or four (4R) microtubule binding domains. Imbalance in the 3R:4R tau ratio is associated with primary tauopathies that develop atypical parkinsonism, such as progressive supranuclear palsy and corticobasal degeneration. Yet, the development of effective therapies for those pathologies is an unmet goal. Here we report motor coordination impairments in the htau mouse model of tauopathy which harbour abnormal 3R:4R tau isoforms content, and in contrast to TauKO mice, are unresponsive to l-DOPA. Preclinical-PET imaging, array tomography and electrophysiological analyses indicated the dorsal striatum as the candidate structure mediating such phenotypes. Indeed, local modulation of tau isoforms by RNA trans-splicing in the striata of adult htau mice, prevented motor coordination deficits and restored basal neuronal firing. Together, these results suggest that abnormal striatal tau isoform content might lead to parkinsonian-like phenotypes and demonstrate a proof of concept that modulation of tau mis-splicing is a plausible disease-modifying therapy for some primary tauopathies.

Acute Regulation of the Arousal-Enhancing Drugs Caffeine and Modafinil on Class IIa HDACs In Vivo and In Vitro: Focus on HDAC7.

Psychostimulant drugs, such as modafinil and caffeine, induce transcriptional alterations through the dysregulation of epigenetic mechanisms. We have previously demonstrated that acute modafinil administration is accompanied by multiple changes in the expression of histone deacetylases (HDACs) within the mouse medial prefrontal cortex (mPFC). Herein, we compared alterations in class IIa HDACs in the mouse mPFC and dorsal striatum (DS) after a single exposure to each psychostimulant. We treated male C57BL/6 mice with modafinil (90 mg/kg, i.p.), caffeine (10 mg/kg, i.p.), or vehicle and evaluated locomotor activity. Following, we examined hdac4, hdac5, and hdac7 mRNA expression using qRT-PCR and HDAC7, pHDAC7, and pHDACs4/5/7 using Western blot. Last, we explored generalized effects in N2a cell line using modafinil (100 µM and 1 mM) or caffeine (80 µM and 800 µM). Our results indicate that modafinil had greater effects on locomotor activity compared with caffeine. qRT-PCR experiments revealed that modafinil decreased hdac5 and hdac7 mRNA expression in the DS, while caffeine had no effects. In the mPFC, modafinil increased hdac7 mRNA expression, with no effects observed for caffeine. Western blot revealed that within the DS, modafinil induced increases in HDAC7, pHDAC7, and pHDACs4/5/7 protein expression, while, in the mPFC, caffeine induced decreases in HDAC7, pHDAC7, and pHDACs4/5/7 protein levels. In vitro studies revealed that modafinil increased hdac4, hdac5, and hdac7 mRNA levels in N2a, while caffeine only increased hdac5 at a higher dose. These findings support the notion that modafinil and caffeine exert distinct regulation of class IIa HDAC family members and that these transcriptional and translational consequences are region-specific.

Molecular changes in the nucleus accumbens and prefrontal cortex associated with the locomotor sensitization induced by coca paste seized samples.

RATIONALE: In previous studies, we have demonstrated that seized samples of a smokable form of cocaine, also known as coca paste (CP), induced behavioral sensitization in rats. Interestingly, this effect was accelerated and enhanced when the samples were adulterated with caffeine. While the cocaine phenomenon is associated with persistent functional and structural alterations in the prefrontal cortex (PFC) and nucleus accumbens (NAc), the molecular mechanisms underlying the CP sensitization and the influence of caffeine remains still unknown. OBJECTIVE: We examined the gene expression in NAc and mPFC after the expression caffeine-adulterated and non-adulterated CP locomotor sensitization. METHODS: The locomotor sensitization was established in C57BL/6 mice, repeatedly treated with a CP-seized sample adulterated with caffeine (CP-2) and a non-adulterated one (CP-1). We then assessed the mRNA expression of receptor subunits of the dopaminergic and glutamatergic systems in the medial PFC (mPFC) and NAc. Other molecular markers (e.g., adenosinergic, endocannabinoid receptor subunits, and synaptic plasticity-associated genes) were also analyzed. RESULTS: Only CP-2-treated mice expressed locomotor sensitization. This phenomenon was associated with increased Drd1a, Gria1, Cnr1, and Syn mRNA expression levels in the NAc. Drd3 mRNA expression levels were only significantly increased in mPFC of CP-2-treated group. CONCLUSIONS: Our results demonstrated that caffeine actively collaborates in the induction of the molecular changes underlying CP sensitization. The present study provides new knowledge on the impact of active adulterants to understand the early dependence induced by CP consumption.

Social inequalities in care for the elderly with diabetes in Brazil

[ABSTRACT]. Objective. To measure the prevalence of various care services offered to the elderly with diabetes mellitus in Brazil, and to assess the social inequalities in these services. Methods. This cross-sectional, population-based study was carried out in 2013. The care services offered were evaluated in terms of the following eight indicators: recommendations to lower carbohydrates, to measure blood glucose, and to examine the feet; requests made for blood tests, for glycated hemoglobin tests, and for glycemic curve tests; and whether service users had had their eyes or feet examined in the previous year. We used the slope index of inequality and the concentration index to assess the inequalities among wealth quintiles. Results. A total of 1 685 elderly persons with diabetes were evaluated. Overall, 41.7% of them had had their eyes examined in the preceding year, 35.4% had had their feet examined in the preceding year, and 10.9% had been offered all eight of the care services. The largest absolute differences (in percentage points) between the first (poorest) and fifth (richest) wealth quintiles in terms of the care services that were offered to the users were for: a recommendation to measure blood glucose (25.8), a glycated hemoglobin test request (27.4), a glycemic curve test request (31.9), having the eyes examined in the preceding year (29.3), and having the feet examined in the preceding year (27.0). Conclusion. There were notable inequalities in the prevalences of the care services. In the future, measurement of blood glucose and examination of the feet should be emphasized, especially for elderly persons in a lower socioeconomic level.

[RESUMEN]. Objetivo. Medir la prevalencia de los diversos servicios de atención que se ofrecen a las personas mayores con diabetes mellitus en Brasil y evaluar las desigualdades sociales en dichos servicios. Métodos. Este estudio transversal y basado en la población se llevó a cabo en el 2013. Los servicios de atención que se ofrecen se evaluaron en cuanto a los siguientes ocho indicadores: recomendaciones para reducir los carbohidratos, medir la glucemia y revisarse los pies; solicitudes de análisis de sangre, pruebas de glucohemoglobina y pruebas de la curva de glucemia; y si a los usuarios de los servicios les habían examinado los ojos o los pies en el último año. Usamos el índice de pendiente de la desigualdad y el índice de concentración para evaluar las desigualdades entre los quintiles de riqueza. Resultados. Se evaluó en total a 1.685 personas mayores con diabetes. En términos generales, en el último año les habían examinado los ojos a 41,7% de ellos, en el último año les habían examinado los pies a 35,4% y les habían ofrecido los ocho servicios de atención a 10,9%. Las mayores diferencias absolutas en puntos porcentuales entre el primer quintil de riqueza (los más pobres) y el quinto (los más ricos) en cuanto a los servicios de atención ofrecidos a los usuarios se encontraron en la recomendación de medir la glucemia (25,8), la solicitud de pruebas de glucohemoglobina (27,4), la solicitud de pruebas de curvas de glucemia (31,9), el examen de los ojos en el último año (29,3) y el examen de los pies en el último año (27,0). Conclusiones. Hubo notables desigualdades en la prevalencia de los servicios de atención. En el futuro, debe hacerse énfasis en la medición de la glucemia y el examen de los pies, especialmente en las personas mayores de un nivel socioeconómico bajo.

[RESUMO]. Objetivo. Medir a prevalência de vários serviços de atenção oferecidos a idosos portadores de diabetes mellitus no Brasil e avaliar desigualdades sociais nestes serviços. Métodos. Estudo transversal, de caráter populacional, realizado em 2013. Os serviços de atenção oferecidos foram avaliados quanto aos oito indicadores a seguir: recomendação de diminuir o consumo de carboidratos, medir a glicemia e examinar os pés; pedido de exames de sangue, hemoglobina glicada e curva glicêmica; e realização de exame dos olhos ou pés nos usuários do serviço durante o ano anterior. Foram utilizados o índice absoluto de desigualdade e o índice de concentração para avaliar as desigualdades entre quintis de renda. Resultados. Foram avaliados 1.685 idosos com diabetes. Destes, 41,7% haviam sido submetidos a exame dos olhos no ano anterior, 35,4% a exame dos pés, e a 10,9% haviam sido oferecidos todos os oito serviços de atenção. As maiores diferenças absolutas (em pontos percentuais) entre o primeiro (mais pobre) e quinto (mais rico) quintis de renda foram constatadas para os seguintes serviços de atenção: recomendação de medir a glicemia (25,8), pedido de exame de hemoglobina glicada (27,4), pedido de exame de curva glicêmica (31,9), exame de olhos no ano anterior (29,3) e exame dos pés no ano anterior (27,0). Conclusão. Foram constatadas desigualdades notáveis na prevalência dos serviços de atenção. Futuramente, deve-se dar ênfase à medição da glicemia e do exame dos pés, principalmente em idosos de nível socioeconômico mais baixo.

[Structure of primary healthcare units for treating persons with diabetes: Cycles I and II of the Brazilian National Program to Improve Access and Quality]. / Estrutura das unidades básicas de saúde para atenção às pessoas com diabetes: Ciclos I e II do Programa Nacional de Melhoria do Acesso e da Qualidade.

The objective was to describe the necessary structure for treating diabetes patients in the primary healthcare system, as evaluated in Cycles I and II of the Brazilian National Program for the Improvement of Access and Quality (PMAQ) in 2012 and 2014, according to the municipalities' characteristics. A descriptive study was used to assess primary care units whose teams participated in Cycles I and II of the PMAQ in 2012 and 2014. The study used variables from Module I of the external evaluation of the PMAQ that addresses the primary care units' structure. Materials (150kg scale, sphygmomanometer, adult stethoscope, tape measure, blood glucose monitor, monofilament packs, ophthalmoscope, and capillary blood glucose strips); medicines (NPH and regular insulin, glyburide, and metformin); and physical space (clinical consultation room, pharmacy, reception/waiting room, and meeting room). All the medicines and the reception/waiting room increased by more than 10p.p. from 2012 to 2014. The prevalence rates for adequate structure of materials, medicines, and physical space in the primary care units were higher in 2014. Adequate structure increased as follows: for materials, from 3.9% to 7.8%, for medicines, from 31.3% to 49.9%, for physical space, from 15.3% to 23.3%. Municipalities with more than 300,000 inhabitants, higher Human Development Index (HDI), and lower coverage of the Family Health Strategy (FHS) showed higher prevalence rates for adequate primary care units. Units that adhered to Cycles I and II of the PMAQ showed improvement in their structures. However, there was a low prevalence of primary care units with adequate structures, besides differences in the services' structure according to population size, HDI, and FHS coverage.

O objetivo foi descrever a estrutura necessária à atenção às pessoas com diabetes, usuárias da rede de atenção primária à saúde, avaliada nos Ciclos I e II do Programa Nacional de Melhoria do Acesso e da Qualidade (PMAQ) no Brasil, 2012 e 2014, considerando-se as características dos municípios. Utilizando-se um estudo descritivo foram avaliadas as unidades básicas de saúde (UBS) cujas equipes participaram dos Ciclos I e II do PMAQ, em 2012 e 2014. Utilizaram-se variáveis do Módulo I da avaliação externa do PMAQ que aborda a estrutura das UBS. Materiais (balança de 150kg, esfigmomanômetro, estetoscópio adulto, fita métrica, glicosímetro, kit de monofilamentos, oftalmoscópio e tiras de glicemia capilar); medicamentos (insulina NPH e regular, glibenclamida e metformina) e espaço físico (consultório clínico, farmácia, recepção, sala de acolhimento e de reunião). Todos os medicamentos avaliados e a sala de acolhimento apresentaram um aumento de mais de 10p.p. de 2012 para 2014. As prevalências de estrutura adequada de materiais, medicamentos e física nas UBS foram maiores em 2014. A estrutura adequada de materiais passou de 3,9% para 7,8%, de medicamentos de 31,3% para 49,9% e física de 15,3% para 23,3%. Os municípios com mais de 300 mil habitantes, melhor Índice de Desenvolvimento Humano (IDH) e menor cobertura de Estratégia Saúde da Família (ESF) registraram maiores prevalências de UBS adequadas. As unidades que aderiram aos Ciclos I e II do PMAQ obtiveram melhoria em suas estruturas. Entretanto, foi encontrada baixa prevalência de UBS com estrutura adequada, além de diferenças na estrutura dos serviços, de acordo com o porte populacional, IDH e cobertura de ESF.

El objetivo fue describir la estructura necesaria para la atención a las personas con diabetes, usuarias de la red de atención primaria en salud, evaluada en los Ciclos I y II del Programa Nacional de Mejoría del Acceso y de la Calidad (PMAQ) en Brasil, 2012 y 2014, considerándose las características de los municipios. Utilizándose un estudio descriptivo se evaluaron las unidades básicas de salud (UBS), cuyos equipos participaron en los Ciclos I y II del PMAQ, en 2012 y 2014. Se utilizaron variables del Módulo I de evaluación externa del PMAQ que aborda la estructura de las UBS. Materiales (peso de 150kg, esfigmomanómetro, estetoscopio adulto, cinta métrica, glucómetro, kit de monofilamentos, oftalmoscopio y tiras de glucemia capilar); medicamentos (insulina NPH y regular, glibenclamida y metformina) y espacio físico (consultorio clínico, farmacia, recepción, sala de acogida y reunión). Todos los medicamentos evaluados y la sala de acogida presentaron un aumento de más de 10p.p. de 2012 a 2014. Las prevalencias sobre estructura adecuada de materiales, medicamentos y física en las UBS fueron mayores en 2014. La estructura adecuada de materiales pasó de 3,9% a 7,8%, de medicamentos de 31,3% a 49,9% y física de 15,3% a 23,3%. Los municipios con más de 300.000 habitantes, mejor Índice de Desarrollo Humano (IDH) y menor cobertura de Estrategia Salud de la Familia (ESF), registraron mayores prevalencias de UBS adecuadas. Las unidades que se adhirieron a los Ciclos I y II del PMAQ obtuvieron una mejoría en sus estructuras. No obstante, se encontró una baja prevalencia de UBS con estructura adecuada, además de las diferencias en la estructura de los servicios, de acuerdo con la dimensión poblacional, IDH y cobertura de ESF.

Social inequalities in care for the elderly with diabetes in Brazil.

OBJECTIVE: To measure the prevalence of various care services offered to the elderly with diabetes mellitus in Brazil, and to assess the social inequalities in these services. METHODS: This cross-sectional, population-based study was carried out in 2013. The care services offered were evaluated in terms of the following eight indicators: recommendations to lower carbohydrates, to measure blood glucose, and to examine the feet; requests made for blood tests, for glycated hemoglobin tests, and for glycemic curve tests; and whether service users had had their eyes or feet examined in the previous year. We used the slope index of inequality and the concentration index to assess the inequalities among wealth quintiles. RESULTS: A total of 1 685 elderly persons with diabetes were evaluated. Overall, 41.7% of them had had their eyes examined in the preceding year, 35.4% had had their feet examined in the preceding year, and 10.9% had been offered all eight of the care services. The largest absolute differences (in percentage points) between the first (poorest) and fifth (richest) wealth quintiles in terms of the care services that were offered to the users were for: a recommendation to measure blood glucose (25.8), a glycated hemoglobin test request (27.4), a glycemic curve test request (31.9), having the eyes examined in the preceding year (29.3), and having the feet examined in the preceding year (27.0). CONCLUSION: There were notable inequalities in the prevalences of the care services. In the future, measurement of blood glucose and examination of the feet should be emphasized, especially for elderly persons in a lower socioeconomic level.

Social inequalities in care for the elderly with diabetes in Brazil / Desigualdades sociales en la atención de las personas mayores con diabetes en Brasil / Desigualdades sociais na atenção a idosos com diabetes no Brasil

ABSTRACT Objective. To measure the prevalence of various care services offered to the elderly with diabetes mellitus in Brazil, and to assess the social inequalities in these services. Methods. This cross-sectional, population-based study was carried out in 2013. The care services offered were evaluated in terms of the following eight indicators: recommendations to lower carbohydrates, to measure blood glucose, and to examine the feet; requests made for blood tests, for glycated hemoglobin tests, and for glycemic curve tests; and whether service users had had their eyes or feet examined in the previous year. We used the slope index of inequality and the concentration index to assess the inequalities among wealth quintiles. Results. A total of 1 685 elderly persons with diabetes were evaluated. Overall, 41.7% of them had had their eyes examined in the preceding year, 35.4% had had their feet examined in the preceding year, and 10.9% had been offered all eight of the care services. The largest absolute differences (in percentage points) between the first (poorest) and fifth (richest) wealth quintiles in terms of the care services that were offered to the users were for: a recommendation to measure blood glucose (25.8), a glycated hemoglobin test request (27.4), a glycemic curve test request (31.9), having the eyes examined in the preceding year (29.3), and having the feet examined in the preceding year (27.0). Conclusion. There were notable inequalities in the prevalences of the care services. In the future, measurement of blood glucose and examination of the feet should be emphasized, especially for elderly persons in a lower socioeconomic level.

RESUMEN Objetivo. Medir la prevalencia de los diversos servicios de atención que se ofrecen a las personas mayores con diabetes mellitus en Brasil y evaluar las desigualdades sociales en dichos servicios. Métodos. Este estudio transversal y basado en la población se llevó a cabo en el 2013. Los servicios de atención que se ofrecen se evaluaron en cuanto a los siguientes ocho indicadores: recomendaciones para reducir los carbohidratos, medir la glucemia y revisarse los pies; solicitudes de análisis de sangre, pruebas de glucohemoglobina y pruebas de la curva de glucemia; y si a los usuarios de los servicios les habían examinado los ojos o los pies en el último año. Usamos el índice de pendiente de la desigualdad y el índice de concentración para evaluar las desigualdades entre los quintiles de riqueza. Resultados. Se evaluó en total a 1.685 personas mayores con diabetes. En términos generales, en el último año les habían examinado los ojos a 41,7% de ellos, en el último año les habían examinado los pies a 35,4% y les habían ofrecido los ocho servicios de atención a 10,9%. Las mayores diferencias absolutas en puntos porcentuales entre el primer quintil de riqueza (los más pobres) y el quinto (los más ricos) en cuanto a los servicios de atención ofrecidos a los usuarios se encontraron en la recomendación de medir la glucemia (25,8), la solicitud de pruebas de glucohemoglobina (27,4), la solicitud de pruebas de curvas de glucemia (31,9), el examen de los ojos en el último año (29,3) y el examen de los pies en el último año (27,0). Conclusiones. Hubo notables desigualdades en la prevalencia de los servicios de atención. En el futuro, debe hacerse énfasis en la medición de la glucemia y el examen de los pies, especialmente en las personas mayores de un nivel socioeconómico bajo.

RESUMO Objetivo. Medir a prevalência de vários serviços de atenção oferecidos a idosos portadores de diabetes mellitus no Brasil e avaliar desigualdades sociais nestes serviços. Métodos. Estudo transversal, de caráter populacional, realizado em 2013. Os serviços de atenção oferecidos foram avaliados quanto aos oito indicadores a seguir: recomendação de diminuir o consumo de carboidratos, medir a glicemia e examinar os pés; pedido de exames de sangue, hemoglobina glicada e curva glicêmica; e realização de exame dos olhos ou pés nos usuários do serviço durante o ano anterior. Foram utilizados o índice absoluto de desigualdade e o índice de concentração para avaliar as desigualdades entre quintis de renda. Resultados. Foram avaliados 1.685 idosos com diabetes. Destes, 41,7% haviam sido submetidos a exame dos olhos no ano anterior, 35,4% a exame dos pés, e a 10,9% haviam sido oferecidos todos os oito serviços de atenção. As maiores diferenças absolutas (em pontos percentuais) entre o primeiro (mais pobre) e quinto (mais rico) quintis de renda foram constatadas para os seguintes serviços de atenção: recomendação de medir a glicemia (25,8), pedido de exame de hemoglobina glicada (27,4), pedido de exame de curva glicêmica (31,9), exame de olhos no ano anterior (29,3) e exame dos pés no ano anterior (27,0). Conclusão. Foram constatadas desigualdades notáveis na prevalência dos serviços de atenção. Futuramente, deve-se dar ênfase à medição da glicemia e do exame dos pés, principalmente em idosos de nível socioeconômico mais baixo.

Estrutura das unidades básicas de saúde para atenção às pessoas com diabetes: Ciclos I e II do Programa Nacional de Melhoria do Acesso e da Qualidade / Structure of primary healthcare units for treating persons with diabetes: Cycles I and II of the Brazilian National Program to Improve Access and Quality / Estructura de las unidades básicas de salud para la atención a personas con diabetes: Ciclos I y II del Programa Nacional de Mejora al Acceso y Calidad

Resumo: O objetivo foi descrever a estrutura necessária à atenção às pessoas com diabetes, usuárias da rede de atenção primária à saúde, avaliada nos Ciclos I e II do Programa Nacional de Melhoria do Acesso e da Qualidade (PMAQ) no Brasil, 2012 e 2014, considerando-se as características dos municípios. Utilizando-se um estudo descritivo foram avaliadas as unidades básicas de saúde (UBS) cujas equipes participaram dos Ciclos I e II do PMAQ, em 2012 e 2014. Utilizaram-se variáveis do Módulo I da avaliação externa do PMAQ que aborda a estrutura das UBS. Materiais (balança de 150kg, esfigmomanômetro, estetoscópio adulto, fita métrica, glicosímetro, kit de monofilamentos, oftalmoscópio e tiras de glicemia capilar); medicamentos (insulina NPH e regular, glibenclamida e metformina) e espaço físico (consultório clínico, farmácia, recepção, sala de acolhimento e de reunião). Todos os medicamentos avaliados e a sala de acolhimento apresentaram um aumento de mais de 10p.p. de 2012 para 2014. As prevalências de estrutura adequada de materiais, medicamentos e física nas UBS foram maiores em 2014. A estrutura adequada de materiais passou de 3,9% para 7,8%, de medicamentos de 31,3% para 49,9% e física de 15,3% para 23,3%. Os municípios com mais de 300 mil habitantes, melhor Índice de Desenvolvimento Humano (IDH) e menor cobertura de Estratégia Saúde da Família (ESF) registraram maiores prevalências de UBS adequadas. As unidades que aderiram aos Ciclos I e II do PMAQ obtiveram melhoria em suas estruturas. Entretanto, foi encontrada baixa prevalência de UBS com estrutura adequada, além de diferenças na estrutura dos serviços, de acordo com o porte populacional, IDH e cobertura de ESF.

Abtstract: The objective was to describe the necessary structure for treating diabetes patients in the primary healthcare system, as evaluated in Cycles I and II of the Brazilian National Program for the Improvement of Access and Quality (PMAQ) in 2012 and 2014, according to the municipalities' characteristics. A descriptive study was used to assess primary care units whose teams participated in Cycles I and II of the PMAQ in 2012 and 2014. The study used variables from Module I of the external evaluation of the PMAQ that addresses the primary care units' structure. Materials (150kg scale, sphygmomanometer, adult stethoscope, tape measure, blood glucose monitor, monofilament packs, ophthalmoscope, and capillary blood glucose strips); medicines (NPH and regular insulin, glyburide, and metformin); and physical space (clinical consultation room, pharmacy, reception/waiting room, and meeting room). All the medicines and the reception/waiting room increased by more than 10p.p. from 2012 to 2014. The prevalence rates for adequate structure of materials, medicines, and physical space in the primary care units were higher in 2014. Adequate structure increased as follows: for materials, from 3.9% to 7.8%, for medicines, from 31.3% to 49.9%, for physical space, from 15.3% to 23.3%. Municipalities with more than 300,000 inhabitants, higher Human Development Index (HDI), and lower coverage of the Family Health Strategy (FHS) showed higher prevalence rates for adequate primary care units. Units that adhered to Cycles I and II of the PMAQ showed improvement in their structures. However, there was a low prevalence of primary care units with adequate structures, besides differences in the services' structure according to population size, HDI, and FHS coverage.

El objetivo fue describir la estructura necesaria para la atención a las personas con diabetes, usuarias de la red de atención primaria en salud, evaluada en los Ciclos I y II del Programa Nacional de Mejoría del Acceso y de la Calidad (PMAQ) en Brasil, 2012 y 2014, considerándose las características de los municipios. Utilizándose un estudio descriptivo se evaluaron las unidades básicas de salud (UBS), cuyos equipos participaron en los Ciclos I y II del PMAQ, en 2012 y 2014. Se utilizaron variables del Módulo I de evaluación externa del PMAQ que aborda la estructura de las UBS. Materiales (peso de 150kg, esfigmomanómetro, estetoscopio adulto, cinta métrica, glucómetro, kit de monofilamentos, oftalmoscopio y tiras de glucemia capilar); medicamentos (insulina NPH y regular, glibenclamida y metformina) y espacio físico (consultorio clínico, farmacia, recepción, sala de acogida y reunión). Todos los medicamentos evaluados y la sala de acogida presentaron un aumento de más de 10p.p. de 2012 a 2014. Las prevalencias sobre estructura adecuada de materiales, medicamentos y física en las UBS fueron mayores en 2014. La estructura adecuada de materiales pasó de 3,9% a 7,8%, de medicamentos de 31,3% a 49,9% y física de 15,3% a 23,3%. Los municipios con más de 300.000 habitantes, mejor Índice de Desarrollo Humano (IDH) y menor cobertura de Estrategia Salud de la Familia (ESF), registraron mayores prevalencias de UBS adecuadas. Las unidades que se adhirieron a los Ciclos I y II del PMAQ obtuvieron una mejoría en sus estructuras. No obstante, se encontró una baja prevalencia de UBS con estructura adecuada, además de las diferencias en la estructura de los servicios, de acuerdo con la dimensión poblacional, IDH y cobertura de ESF.

Cocaine and Caffeine Effects on the Conditioned Place Preference Test: Concomitant Changes on Early Genes within the Mouse Prefrontal Cortex and Nucleus Accumbens.

Caffeine is the world's most popular psychostimulant and is frequently used as an active adulterant in many illicit drugs including cocaine. Previous studies have shown that caffeine can potentiate the stimulant effects of cocaine and cocaine-induced drug seeking behavior. However, little is known about the effects of this drug combination on reward-related learning, a key process in the maintenance of addiction and vulnerability to relapse. The goal of the present study was thus to determine caffeine and cocaine combined effects on the Conditioned Place Preference (CPP) test and to determine potential differential mRNA expression in the Nucleus Accumbens (NAc) and medial prefrontal cortex (mPFC) of immediate-early genes (IEGs) as well as dopamine and adenosine receptor subunits. Mice were treated with caffeine (5 mg/kg, CAF), cocaine (10 mg/kg, COC), or their combination (caffeine 5 mg/kg + cocaine 10 mg/kg, CAF-COC) and trained in the CPP test or treated with repeated injections inside the home cage. NAc and mPFC tissues were dissected immediately after the CPP test, after a single conditioning session or following psychostimulant injection in the home cage for mRNA expression analysis. CAF-COC induced a marked change of preference to the drug conditioned side of the CPP and a significant increase in locomotion compared to COC. Gene expression analysis after CPP test revealed specific up-regulation in the CAF-COC group of Drd1a, cFos, and FosB in the NAc, and cFos, Egr1, and Npas4 in the mPFC. Importantly, none of these changes were observed when animals received same treatments in their home cage. With a single conditioning session, we found similar effects in both CAF and CAF-COC groups: increased Drd1a and decreased cFos in the NAc, and increased expression of Drd1a and Drd2, in the mPFC. Interestingly, we found that cFos and Npas4 gene expression were increased only in the mPFC of the CAF-COC. Our study provides evidence that caffeine acting as an adulterant could potentiate reward-associated memories elicited by cocaine. This is associated with specific changes in IEGs expression that were observed almost exclusively in mice that received the combination of both psychostimulants in the context of CPP memory encoding and retrieval. Our results highlight the potential relevance of caffeine in the maintenance of cocaine addiction which might be mediated by modifying neural plasticity mechanisms that strengthen learning of the association between drug and environment.

Combined Effects of Simultaneous Exposure to Caffeine and Cocaine in the Mouse Striatum.

Caffeine is the world's most popular psychoactive drug and is also an active adulterant found in many drugs of abuse, including seized cocaine samples. Despite several studies which examine the effects of caffeine or cocaine administered as single agents, little data are available for these agents when given in combination. The purpose of the present study was to determine if combined intake of both psychostimulants can lead to maladaptive changes in striatal function. Mice were injected with a binge regimen (intermittent treatment for 13 days) of caffeine (3 × 5 mg/kg), cocaine (3 × 10 mg/kg), or combined administration. We found that chronic caffeine potentiated locomotion induced by cocaine and that both caffeine-treated groups showed sensitization. Striatal tissue was obtained 24 h and 7 days after last injection (withdrawal) for immunohistochemistry and mRNA expression. Our results show that combined intake of both psychostimulants can increase GFAP immunoreactivity in the striatum at both times post treatment. Gene expression analysis, targeted at dopamine, adenosine, and glutamate receptor subunit genes, revealed significant transcript down-regulation in the dorsal striatum of AMPA, NMDA, D1 and D2 receptor subunit mRNA expression in the group that received combined treatment, but not after individual administration. At withdrawal, we found increased D1 receptor mRNA expression along with increased A1, AMPA, NMDA, and metabotropic subunit expression. A2A mRNA showed decreased expression after both times in all experimental groups. Our study provides evidence that there are striatal alterations mediated by combined caffeine and cocaine administration, and highlights negative outcomes of chronic intake of both psychostimulants.

Methamphetamine blunts Ca(2+) currents and excitatory synaptic transmission through D1/5 receptor-mediated mechanisms in the mouse medial prefrontal cortex.

Psychostimulant addiction is associated with dysfunctions in frontal cortex. Previous data demonstrated that repeated exposure to methamphetamine (METH) can alter prefrontal cortex (PFC)-dependent functions. Here, we show that withdrawal from repetitive non-contingent METH administration (7 days, 1 mg/kg) depressed voltage-dependent calcium currents (ICa ) and increased hyperpolarization-activated cation current (IH ) amplitude and the paired-pulse ratio of evoked excitatory postsynaptic currents (EPSCs) in deep-layer pyramidal mPFC neurons. Most of these effects were blocked by systemic co-administration of the D1/D5 receptor antagonist SCH23390 (0.5 and 0.05 mg/kg). In vitro METH (i.e. bath-applied to slices from naïve-treated animals) was able to emulate its systemic effects on ICa and evoked EPSCs paired-pulse ratio. We also provide evidence of altered mRNA expression of (1) voltage-gated calcium channels P/Q-type Cacna1a (Cav 2.1), N-type Cacna1b (Cav 2.2), T-type Cav 3.1 Cacna1g, Cav 3.2 Cacna1h, Cav 3.3 Cacna1i and the auxiliary subunit Cacna2d1 (α2δ1); (2) hyperpolarization-activated cyclic nucleotide-gated channels Hcn1 and Hcn2; and (3) glutamate receptors subunits AMPA-type Gria1, NMDA-type Grin1 and metabotropic Grm1 in the mouse mPFC after repeated METH treatment. Moreover, we show that some of these changes in mRNA expression were sensitive D1/5 receptor blockade. Altogether, these altered mechanisms affecting synaptic physiology and transcriptional regulation may underlie PFC functional alterations that could lead to PFC impairments observed in METH-addicted individuals.

Psychostimulant-Induced Testicular Toxicity in Mice: Evidence of Cocaine and Caffeine Effects on the Local Dopaminergic System.

Several organ systems can be affected by psychostimulant toxicity. However, there is not sufficient evidence about the impact of psychostimulant intake on testicular physiology and catecholaminergic systems. The aim of the present study was to further explore potential toxic consequences of chronic exposure to cocaine, caffeine, and their combination on testicular physiology. Mice were injected with a 13-day chronic binge regimen of caffeine (3x5mg/kg), cocaine (3×10mg/kg), or combined administration. Mice treated with cocaine alone or combined with caffeine showed reduced volume of the seminiferous tubule associated to a reduction in the number of spermatogonia. Cocaine-only and combined treatments induced increased lipid peroxidation evaluated by TBARS assay and decreased glutathione peroxidase mRNA expression. Importantly, caffeine-cocaine combination potentiated the cocaine-induced germ cell loss, and induced pro-apoptotic BAX protein expression and diminished adenosine receptor A1 mRNA levels. We analyzed markers of dopaminergic function in the testis and detected the presence of tyrosine hydroxylase (TH) in the cytoplasm of androgen-producing Leydig cells, but also in meiotic germs cells within seminiferous tubules. Moreover, using transgenic BAC-Drd1a-tdTomato and D2R-eGFP mice, we report for the first time the presence of dopamine receptors (DRs) D1 and D2 in testicular mouse Leydig cells. Interestingly, the presence of DRD1 was also detected in the spermatogonia nearest the basal lamina of the seminiferous tubules, which did not show TH staining. We observed that psychostimulants induced downregulation of DRs mRNA expression and upregulation of TH protein expression in the testis. These findings suggest a potential role of the local dopaminergic system in psychostimulant-induced testicular pathology.

Differential effects of environment-induced changes in body temperature on modafinil's actions against methamphetamine-induced striatal toxicity in mice.

Methamphetamine (METH) exposure can produce hyperthermia that might lead to toxicity and death. Modafinil is a wake-promoting compound that is also been prescribed off-label to treat METH dependence. Modafinil has shown neuroprotective properties against METH harmful effects in animal models. The goal of the present study was to test if the prevention of hyperthermia might play a role on the neuroprotective actions of modafinil against METH toxicity using various ambient temperatures. METH was administered to female C57BL/6 mice in a binge regimen: 4 × 5 mg/kg, 2 h apart; modafinil (90 mg/kg) was injected twice, 1 h before first and fourth METH injections. Drugs were given at cold ambient temperature (14 °C) or hot ambient temperature (29 °C). Body temperature was measured during treatments. Brains were dissected out 6 days after treatments and processed for tyrosine hydroxylase (TH), dopamine transporter (DAT), GFAP and c-Fos immunohistochemistry. Exposure to hot ambient temperature exacerbated METH toxicity evidenced by striatal reductions in TH and DAT and increased GFAP immmunoreactivity. Modafinil counteracted reductions in TH and DAT, but failed to block astroglial activation. At both ambient temperatures tested modafinil did induce increments in GFAP, but the magnitude was significantly lower than the one induced by METH. Both drugs induced increases in c-Fos positive nuclei; modafinil did not block this effect. Our results suggest that protective effects of modafinil against METH-induced neurotoxicity may be dependent, in part, to its hypothermic effects. Nevertheless, modafinil maintained some protective properties on METH-induced alterations in the striatum at different ambient temperatures.

Neuroprotective effects of hypothermia on synaptic actin cytoskeletal changes induced by perinatal asphyxia.

Cerebral hypoxia-ischemia damages synaptic proteins, resulting in cytoskeletal alterations, protein aggregation and neuronal death. In the previous works, we have shown neuronal and synaptic changes in rat neostriatum subjected to hypoxia that leads to ubi-protein accumulation. Recently, we also showed that, changes in F-actin organization could be related to early alterations induced by hypoxia in the Central Nervous System. However, little is known about effective treatment to diminish the damage. The main aim of this work is to study the effects of birth hypothermia on the actin cytoskeleton of neostriatal post-synaptic densities (PSD) in 60 days olds rats by immunohistochemistry, photooxidation and western blot. We used 2 different protocols of hypothermia: (a) intrahypoxic hypothermia at 15°C and (b) post-hypoxia hypothermia at 32°C. Consistent with previous data at 30 days, staining with phalloidin-Alexa(488) followed by confocal microscopy analysis showed an increase of F-actin fluorescent staining in the neostriatum of hypoxic animals. Correlative photooxidation electron microscopy confirmed these observations showing an increment in the number of mushroom-shaped F-actin staining spines in neostriatal excitatory synapses in rats subjected to hypoxia. In addition, western blot revealed ß-actin increase in PSDs in hypoxic animals. The optic relative density measurement showed a significant difference between controls and hypoxic animals. When hypoxia was induced under hypothermic conditions, the changes observed in actin cytoskeleton were blocked. Post-hypoxic hypothermia showed similar answer but actin cytoskeleton modifications were not totally reverted as we observed at 15°C. These data suggest that the decrease of the body temperature decreases the actin modifications in dendritic spines preventing the neuronal death.