Inwardly rectifying potassium (Kir) channels are critically important for regulating resting membrane potential in excitable cells, a job underscored by the severe pathophysiology associated with channel dysfunction. While all Kir channels require the activating lipid PIP2, many of these channels have diverse modulatory factors that couple to PIP2-dependent gating. Channels in the Kir3 (GIRK) family, in particular, have several co-activating elements, including G-protein betagamma subunits, ethanol, and sodium. During stimulation of Gq-coupled receptors, downstream activation of Protein Kinase C can phosphorylate and inhibit Kir3 channels, yet the mechanism of inhibition and phosphorylation sites are incompletely understood. We took a combined experimental and computational approach using neuronal Kir3.2 to investigate how phosphorylation at a putative PKC site identified in Kir3.1/3.4 could lead to channel inhibition. Kir3.2 inhibition was found to depend on the phosphorylation state of Ser-196, although mutagenesis data suggest it functions as an allosteric regulator of PKC inhibition. MD simulations identified a molecular switch whereby phosphorylation of Ser-196 recruits a critical gating residue, Arg-201, away from the sodium coordination site Asp-228. Neutralization of Ser-196 or Arg-201 resulted in less active channels which exhibited increased sensitivity to PKC inhibition. Additionally the interplay of PIP2 and PKC inhibition was examined in depth using homomeric Kir3.2, revealing that increases in channel-PIP2 interactions limit sensitivity to PKC inhibition, whereas low levels of PIP2 increase PKC sensitivity. Neutralization of Ser-196 uncoupled PKC inhibition from this PIP2 dependence. These studies suggest a model whereby PKC inhibition can occur along PIP2-dependent and PIP2-independent pathways, depending on the phosphorylation state of Ser-196.

No description available.

Master of Science / Department of Agronomy / Dorivar Ruiz Diaz / Corn (Zea mays) response to fertilization and placement methods has been studied extensively; however studies on soybean [Glycine max (L.) Merr.] response to placement have been limited. Three studies were completed to evaluate different aspects of crop response. The objective of the first study was to evaluate the effect of starter and broadcast fertilizer application on corn and soybean, in a typical corn-soybean rotation in Kansas. Treatments were unfertilized control, starter (N, P and K), broadcast P and K using mono ammonium phosphate (MAP) and potassium chloride (KCl) and the combination of starter and broadcast. Corn and soybean yield generally was not affected by starter and broadcast treatments. Thus fertilization may be recommended only under specific conditions. The objectives of the second study were (i) to evaluate the effect of residual and direct fertilization on soybeans after corn under a corn-soybean rotation system, and (ii) study the effect of fertilizer P and K application on soil test P (STP) and soil test K (STK) changes over time. Direct fertilization increased soybean yield while residual fertilizer did not. Therefore maintenance rates may be effective to improve soybean yield and likely maintain STP and STK levels. Application of P and K fertilizer generated significant increases in STP and STK after one year of application. The rate of P and K fertilizer required to increase 1 mg kg-1 yr-1 was between 2.8 - 5.1 kg ha-1 for P and between 1.0- 2.5 kg ha-1 for K, respectively. The objective of the third study evaluate both corn and soybean response to direct P fertilization including starter and broadcast. The treatments were a control, two starter fertilizers (with N-P and N only), five P rates (9.8, 19.6, 29.3, 39.1, 48.9 kg P ha-1) and one treatment with starter fertilizer in addition to the broadcast fertilizer application. Corn grain yield was not significantly affected by any broadcast or starter treatments. Broadcast application rates significantly increased soybean yield on low STP levels. Results of this study show that large corn or soybean yield response to starter and broadcast P application are likely with low STP levels.

No description available.

Reproductive science plays an important role in conservation biology. Quantitative studies of basic reproductive biology in wildlife are critical for the development of successful assisted reproductive technologies. Investigation of the volume regulatory mechanism of spermatozoa could produce options to improve the cryopreservation of spermatozoa and provide a non-hormonal contraceptive option for men, both of which could have significant impacts on global biodiversity preservation. Volume regulation of somatic cells involves the movement of osmolytes through various channels, including potassium channels. The potassium channels involved in volume regulation of human, monkey, and murine spermatozoa were investigated. Flow cytometry was used to gauge the sensitivity of the volume regulatory process of spermatozoa to various potassium channel inhibitors and a simultaneous hypotonic challenge. Channels potentially involved in regulatory volume decrease of spermatozoa varied with species but included voltage-gated (Kv) channels 1.4, 1.5, 1.7, 4.1, 4.2 and 4.3 as well as TWIK1, TWIK2, TASK1, TASK2, TASK3, TREK2 , and minK. The presence of some of these channels was confirmed by western blotting and immunocytochemistry. Changes in the motility patterns of human and monkey spermatozoa in the presence of potassium channel inhibitors during hypotonic stress were also observed, suggesting a relationship between volume regulation and motility. To evaluate potential organic osmolytes involved in, and compare effects of CPAs on, volume regulation, the isotonicity of murine epididymal spermatozoa was measured using a null point method. Spermatozoa were then exposed to high concentrations of various osmolytes and cryoprotective agents in isotonic medium to evaluate which compounds were able to penetrate the sperm plasma membrane. The osmotic responses of spermatozoa from strains of mice known to have spermatozoa of high (B6D2F1) and low (C57BL6) post-thaw fertility were compared during various osmotic challenges in various media. These experiments indicated that spermatozoa from B6D2F1 mice may have better volume regulation capabilities than spermatozoa from C57BL6 mice, suggesting that better post-thaw fertility of murine spermatozoa could be influenced by the volume regulatory process. The knowledge gained from these experiments could contribute to improved sperm handling and preservation techniques and be used to develop non-hormonal male contraceptives based on inhibiting volume regulation.

Orientador: Antonio Ismael Inácio Cardoso / Banca: Dirceu Maximino Fernandes / Banca: Luis Felipe Villani Purqueiro / Resumo: Embora existam recomendações de adubação para o cultivo comercial de abobrinha-de-moita, são escassos os trabalhos de pesquisa que relacionam o efeito da adubação potássica em cobertura sobre a produtividade e qualidade desta cultura. Assim, faz-se a necessidade de determinar doses deste nutriente em cobertura que auxiliem estas recomendações. Dessa forma, foram conduzidos ensaios no outono e na primavera na Fazenda Experimental São Manuel, da FCA-UNESP, em São Manuel-SP, Brasil. O delineamento experimental utilizado foi o de blocos ao acaso com cinco tratamentos e quatro repetições. Os tratamentos utilizados foram cinco doses de potássio em cobertura (0, 50, 100, 200 e 400 kg K2O ha-1). Nos dois experimentos foram avaliadas as características químicas do solo ao final do cultivo, características vegetativas das plantas ao final do cultivo (altura, número de folhas, massa fresca e seca da parte vegetativa), características químicas da parte aérea (teor de macronutrientes na fase de diagnose foliar, teor de macronutrientes na parte vegetativa e no fruto ao fim do cultivo e a extração de macronutrientes na parte aérea ao fim do cultivo), características de produção de frutos (número de frutos totais e comerciais, produção total e comercial). No experimento de primavera, além destas características, também foram avaliadas o teor de macronutrientes nos frutos das colheitas semanais e as características de qualidade de frutos (pH, acidez total titulável, sólidos solúveis, relação sólidos solúveis e acidez total titulável, açúcares redutores e textura). Para a comparação das características comuns das duas épocas de experimentação foi realizada a análise estatística conjunta. Ocorreu aumento linear para as características teor de potássio no solo no outono e primavera, nas folhas diagnose... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Although there are fertilizer recommendations for commercial cultivation of summer squash, there are few research papers that relate the effect of potassium fertilizer in topdressing on productivity and quality of the crop. Thus, is necessary to determine doses of this nutrient to assist in covering these recommendations. Thus, experiments were conducted in the fall and spring on the Experimental Farm Manuel, the FCA-UNESP, São Manuel-SP, Brazil. The experimental design was randomized blocks with five treatments and four replications. The treatments were five levels of potassium fertilization (0, 50, 100, 200 and 400 kg K2O ha-1). In both experiments were evaluated soil chemical characteristics at the end of the cycle, vegetative characteristics of plants at the end of the cycle (plant height, leaf number, fresh and dry mass of shoots) at the end of the cycle, the chemical characteristics of the plant ( macronutrients in the phase of leaf analysis, macronutrients in the vegetative plant parts and fruit at the end of the cycle and the extraction of nutrients in the plant at the end of the cycle), characteristics of fruit yield (fruit number and total trade, total output and commercial). In the spring experiment, and these characteristics were also assessed the macronutrient composition of the fruits of the weekly harvest and fruit quality characteristics (pH, total acidity, soluble solids, soluble solids and total acidity, reducing sugars and texture). For comparison of common features of two seasons of experimentation was performed statistical joint analysis. There was a linear increase of the characteristics of the potassium content in soil in autumn and spring, diagnosis leaves in autumn, the vegetative plant parts in spring and fruit in late season in the fall to the extent that increased doses... (Complete abstract click electronic access below) / Mestre

O uso agrícola de resíduos de efluentes orgânicos é uma alternativa interessante de disposição, pois permite o aproveitamento potencial das águas e dos nutrientes para o crescimento das plantas. A utilização destes efluentes na agricultura constitui uma importante contribuição para a minimização da contaminação devido à redução de seu lançamento em corpos hídricos, além de se tornar uma alternativa econômica para a propriedade rural, que diminui os custos para aquisição de fertilizantes. O aumento no resíduo de efluente produzido pelas usinas de cana-de-açúcar, a vinhaça, é reflexo do aumento da produção do álcool no Brasil. A vinhaça é um subproduto rico em nutrientes, principalmente em matéria orgânica, característica essa que a torna um grande poluidor quando lançado em corpos hídricos. Dentre as soluções para destinação final da vinhaça, a que se destaca é sua aplicação no solo, por meio da técnica de fertirrigação. Dentro desse contexto, o presente trabalho consiste em avaliar o efeito da aplicação de diferentes doses de vinhaça no crescimento, produção e qualidade dos frutos do cultivar de abacaxi vitória, visando à disposição final deste efluente de forma a garantir a reciclagem de água e nutrientes, sem comprometimento das características químicas e físicas do solo. O experimento foi conduzido na área experimental do Departamento de Engenharia de Biossistemas da Escola Superior de Agricultura \"Luiz de Queiroz\" (ESALQ/USP) em ambiente protegido (casa de vegetação). Adotou-se delineamento inteiramente aleatorizado com 5 tratamentos e 4 repetições, totalizando 20 parcelas experimentais, sendo a unidade experimental representada por trincheiras com dimensões 3,20 m de comprimento por 0,80 m de largura por 0,40 m de profundidade. Os tratamentos foram formados pela aplicação de doses de vinhaça referentes a 0, 50, 75, 100 e 125% da demanda de 462 kg ha-1 de potássio pelo abacaxizeiro. O potássio foi tomado como elemento de referência para obtenção dos volumes de vinhaça, sendo que para os cálculos de volume, utilizou-se a norma CETESB P4.231. As fertirrigações foram ministradas, via gotejamento, de acordo com a demanda nutricional do abacaxizeiro, respeitando suas fases de desenvolvimento. Verificou-se que a aplicação de vinhaça no solo aumentou a CTC e a matéria orgânica, porém não promoveu alterações nas características físicas. O aumento das doses de vinhaça afetaram diretamente as variáveis biométricas: altura de plantas, número de folhas, comprimento e largura da folha \"D\". A área foliar, matéria seca da parte aérea e raiz sofreram influência positiva do aumento das doses de vinhaça aplicadas. Verificou-se que as maiores concentrações de nitrato e potássio na solução do solo foram encontradas nos tratamentos com maiores doses de vinhaça aplicadas. Os tratamentos fertirrigados com vinhaça apresentaram menor produtividade quando comparados ao tratamento com adubação química, no entanto as plantas que foram fertirrigadas apresentaram frutos com qualidades físico-químicas superiores, apesar de na análise sensorial não ter sido verificado diferença significativa entre os tratamentos. / The using of effluent residues in agriculture it is a good alternative because allows the use of the water and the nutrients to cultivate some crops. Also it is an important contribution with the mitigation of the contamination promoted by these effluents when disposed in a hydric system and could be an economic alternative to reduce costs with fertilizers. The increase in the production of effluent in the sugarcane mill, the vinasse, it is the result of the increase in the production of ethanol. The vinasse it is a residue rich in nutrients, mainly organic material that can be a problem when disposed in the hydric systems. One alternative to solve this problem it is to utilize the vinasse in the soil by fertigation. This study aim to evaluate the growing, yield and fruit quality in pineapple variety vitória with different levels of vinasse applied by fertigation, and the recycling of water and nutrients by vinasse without compromise the chemical and physical soil characteristics. The experiment was conduct in a greenhouse located in the Biossystem Engineering department, college of agriculture \"Luiz de Queiroz\" ESALQ/USP. The experimental design adopted was completely randomly with 5 treatments and 4 replications, being the experimental unit one trench with 3.2 of length, 0.8 m of width and 0.4 m depth. The treatments was composed by different levels of vinasse: 0, 50, 75, 100 and 125% of the total pineapple potash necessity (462 kg ha-1). The potassium was used as reference element to obtain the total volume of vinasse and was used the standard CETESB P4.261. The fertigation was applied by drip irrigation and respected the pineapple nutrient requirement in the different development phases. The vinasse application promoted the increase in the soil cation-exchange capacity (CEC) and organic material, but not changed their physical characteristics. The increment in the vinasse levels effected directly the biometrics variables: plant high, number of leaves, length and width of leaf \"D\". The leaf area, dry material of aerial part and root showing positive correlation with the vinasse levels applied. The higher level of nitrate and potash in the soil solution was observed in the treatments that received the higher levels of vinasse. Treatments that receive vinasse by fertigation presented lesser yield when compare to treatments that receive just chemical fertilization, but fruits with better physic-chemicals quality, although sensory analysis have not been verified significant difference between treatments.

No presente trabalho apresentamos um outro método para criação de centros de cor, em matrizes de KI puro, através da absorção de dois fótons. Este processo possui uma característica útil: a coloração é realizada a baixa temperatura, ao contrário de outros métodos, que necessitam aquecer o cristal até próximo a ponto de fusão, fazendo com que as impurezas adicionadas durante o crescimento migrem para a superfície do cristal. Com este método, colorimos alguns cristais de KI puro, a fim de demonstrar a eficiência do processo, e algumas aplicações deste como em cristais de KI: \'Cu POT.+\', onde foi realizada a tentativa de criação de centros de cor do tipo \'F IND.A\', e cristais de KI:\'Yb POT.2+\', nos quais foram identificados centros de cor do tipo Z, com a possibilidade de formação de novos centros. Outras propriedades interessantes, como a suposta formação de \'Yb POT.3+\' nessas matrizes a partir da criação de centros de cor, abrem espaço para mais um amplo estudo desse processo, que ainda deverá ser investigado. / In this work we present an other method to produce color centers in pure KI lattice by two photon absorption. This process has a useful feature: coloring is achieved at low temperature as opposed to other methods that require heating of crystal up to near fusion temperature in such a way that impurities added during growing migrate to the surface of crystal. By this method we colored some pure KI crystals to demonstrate the efficiency and some applications of the process to some crystals such as KI: \'Cu POT.+\' crystals trying to create FA centers and KI:\'Yb POT.2+\' crystals where we identified Z centers with possibilities of formation of other centers. Other interesting properties as the supposed formation of \'Yb POT.3+\' in these lattices through color center creation widen the study of the process to be yet investigated.

Neste trabalho buscou-se avaliar a suscetibilidade dos diferentes modelos de tubos gotejadores ao processo de entupimento quando expostos ao uso de cloreto de potássio (branco e vermelho), aplicados via diferentes qualidades de água. O experimento foi realizado em três fases, por um período de doze meses, analisando o desempenho de 22 modelos de gotejadores (autocompensante e convencionais), com quatro tratamentos e dez repetições, sendo cada repetição representada por um gotejador. Na primeira fase, foram aplicados os seguintes tratamentos: (T1) água com fitoplâncton (lago) e cloreto de potássio branco; (T2) água com fitoplâncton (lago) e cloreto de potássio vermelho; (T3) água potável (laboratório) e cloreto de potássio branco; (T4) água potável (laboratório) e cloreto de potássio vermelho. Na segunda fase foram adicionadas partículas sólidas (solo) às soluções dos tratamentos 1 (T1-Lg/B) e 2 (T2-Lg/V), mantendo a mesma solução do tratamento 3 (T3-Lb/B) e adicionado sulfato de ferro à solução do tratamento 4 (T4-Lb/V). Na terceira fase acrescentou-se hidróxido de ferro às soluções referentes a T1 (T1+So) e T2 (T2+So), mais uma aplicação de uma solução concentrada diretamente nas linhas (tubos gotejadores) sem passar pelo sistema de filtragem, sendo os tratamentos: (T1) Água do lago, cloreto de potássio branco, partículas sólidas e hidróxido de ferro, com o orifício do gotejador posicionado para baixo. (T2) água do lago, cloreto de potássio vermelho, partículas sólidas e hidróxido de ferro, com o orifício do gotejador posicionado para cima. (T3) água do lago , cloreto de potássio branco, partículas sólidas e hidróxido de ferro, com o orifício do gotejador posicionado para cima. (T4) água do lago, cloreto de potássio vermelho, partículas sólidas e hidróxido de ferro, com o orifício do gotejador posicionado para baixo. Os modelos de tubos gotejadores analisados apresentaram desempenhos variáveis, tanto na suscetibilidade ao entupimento quanto no coeficiente de variação de vazão, sugerindo que a arquitetura interna dos gotejadores, foi o fator determinante na caracterização do processo de entupimento. Não foi observada diferença significativa da dinâmica de entupimento com relação à aplicação dos cloretos de potássio branco e vermelho na ausência ou presença de fitoplâncton, mostrando que é possível fazer uso do cloreto de potássio vermelho em fertirrigação, tomando o cuidado de verificar a ausência do elemento ferro no adubo (análise de laboratório). A adição de partículas sólidas e do sulfato de ferro aos tratamentos da primeira fase, passando pelo sistema de filtragem, não potencializou mudanças significativas do cenário de suscetibilidade ao entupimento, nas condições de irrigação da segunda fase. A entrada de partículas sólidas e hidróxido de ferro no sistema sem passar pelo sistema de filtragem (fase 3) e o posicionamento do orifício dos gotejadores para baixo intensificaram o processo de entupimento, onde os modelos convencionais foram os mais sensíveis e alguns modelos autocompensantes foram mais tolerantes a presença destes elementos na água de irrigação. O modelo C3 foi o que apresentou melhor desempenho entre os modelos convencionais nas três fases estudadas, sendo que os modelos A2 e A3 destacaram-se entre os modelos autocompensantes, com uma boa recuperação da vazão ao final dos ensaios. / This work aims to evaluate the susceptibility of several dripline models against clogging process when exposed to potassium chloride (white and red), applied through different water qualities (fertirrigation). The experiment was accomplished in three phases, during a period of twelve months, analyzing the performance of 22 drip models (compensating and conventional), with four treatments and ten repetitions, being each repetition a dripper. In the first phase, it was applied the following treatments: (T1) water with fitoplancton (lake) and white potassium chloride; (T2) water with fitoplancton (lake) and red potassium chloride; (T3) potable water (laboratory) and white potassium chloride; (T4) potable water (laboratory) and red potassium chloride. In the second phase, solid particles were added to the solutions of previous treatments 1 (T1-Lg/B) and 2 (T2-Lg/V), maintaining the same solution for treatment 3 (T3-Lb/B) and added iron sulfate to the solution of treatment 4 (T4-Lb/V). In the third phase iron hydroxide was added to the solutions of T1 (T1+So) and T2 (T2+So) treatments, one more application of a concentrated solution directly in drip lines without passing through the filtration system, resulting the following treatments: (T1) water with fitoplancton, white potassium chloride, solid particles and iron hydroxide, with dripper facing down. (T2) water with fitoplancton, red potassium chloride, solid particles and iron hydroxide, with drippers facing up, (T3) water with fitoplancton , white potassium chloride, solid particles and iron hydroxide, with drippers facing up, (T4) water with fitoplancton, red potassium chloride, solid particles and hydroxide of iron, with drippers facing down. Driplines presented a variable performance, regarding the original flow rate levels and variation coefficient, suggesting that internal architecture of emitters, it is a major factor related to clogging resistance to treatments imposed. Statistical differences were not observed for clogging dynamics under the application of white or red potassium chloride, under different water quality conditions, showing that it is possible to use the red potassium chloride for fertirrigation without problems. It is recommended to certify the absence of iron element in the fertilizer based on laboratory analysis. The addition of solid particles and iron sulfate to first phase treatments, going through the filtration system, did not increase the clogging rate or emitters (second phase). The application of solid particles and iron hydroxide in the system without passing through the filtration system (phase 3) and the positioning of drippers facing down, intensified the clogging process. Conventional models were more sensitive and compensating models were more tolerant of this water quality conditions. Models C3 performance better among the conventional models in all phases studied. Models A2 and A3 stood out among compensating models, presenting a good recovery flow rate at the end of the experiment.

Sickle cell disease (SCD) is one of the most common genetic diseases worldwide. HbS polymerisation causes altered red blood cell (RBC) rheology and fragility, increase in blood viscosity with blockage of small blood vessels, and RBC membrane permeability changes. Excessive levels of cell-free Hb, high autoxidation of Hb, contribute to the production of reactive oxygen species (ROS) in SCD patients. In this work, oxidants showed direct and indirect effects on the main cation permeability pathways involved in dehydration of HbS/S RBCs - Psickle, the Gardos channel and the KCl cotransporter (KCC) - and thus on RBC volume causing polymersation. Psickle and Gardos channel activities showed significant correlation, consistent with the hypothesis that Ca2+ entry via Psickle causes activation of the Ca2+-dependent K+ channel. Treatment of SCD remains inadequate relying on the blood transfusion and supportive therapy depending on the organ affected. In the present study antioxidants and aromatic aldehydes showed some promising results towards future alternative treatments for SCD. Antioxidants showed inhibitory effects on the cation permeability pathways leading to inhibition of polymerisation and haemolysis and thus maintained RBC volume. Aromatic aldehydes interact with HbS and are usually given to increase oxygen affinity, thereby reducing its tendency to polymerise. GBT1118 had a marked inhibitory effect on all three cation permeability pathways. It reduced sickling, Psickle and Gardos activity. It inhibited KCC by affecting the regulatory protein phosphorylation cascade. It maintained RBC hydration, and stabilised RBCs. Historically Oman was the principal trading port of the Persian Gulf region, resulting in the complex mix of social and ethnic backgrounds. In 1989 a second mutation in the β chain of Hb, at position β121 was found in an Omani patient in addition to the usual HbS mutation at the β6 position, and termed HbS-Oman. At low percentage of HbS-Oman patients show severe SCD symptoms. Despite RBCs containing at most 25% HbS-Oman, there was high sickling percentage and K+ permeability showed many features similar to those seen in homozygous HbS/S patients. The presence of α thalassaemia was protective and represents an obvious potential prognostic marker for this rare SCD genotype. Overall, the present work contributes to elucidation of the pathogenesis of SCD, suggests approaches to the development of novel therapies and increases our understanding of a rare SCD genotype, HbS-Oman.