Alzhemier's disease (AD) is a common form of dementia which is characterized by the deposition of amyloids in affected neurons and a cholinergic neurotransmission deficit in the brain. Current therapeutic intervention for AD is primarily based on inhibition of brain acetylcholinesterase (AChE) to restore the brain acetylcholine level. Cryptotanshinone (CT) is a diterprene which is extracted from the root of Salvia miltiorrhiza, an herb that is commonly prescribed in Chinese medicine to treat cardiovascular disease. The present study is aimed at verifying CT's property as an AChE inhibitor using different models. By AChE activity assay, CT was found to be a dual inhibitor which inhibits both human acetylcholinesterase (AChE) and butylcholinesterase (BuChE) with similar IC50. CT inhibited human AChE in a reversible manner, and the inhibition showed the characteristics of mixed-type. To human BuChE, CT is an uncompetitive inhibitor. CT can also inhibit AChE from rat cortical neurons. Apart from AChE inhibition, CT was demonstrated to have ameliorating effect on glutamate excitotoxicity, which is a cause of neuron death in AD. Further study showing that CT treatment can reduce cellular tau phosphorylation, which is the downstream effector of glutamate-induced excitotoxicity. In animal model, the effect of CT on learning impairment in scopolamine-treated rats was also evaluated by the acquisition protocol of Morris water maze. The task learning ability of scopolamine-treated rats was significantly reversed by CT, and the CT-fed rats were able to develop spatial searching strategy comparable to the control animals. Chronic administration of CT at effective doses did not cause significant hepatotoxicity. Cholinergic side effect of muscle weakness was not observed in CT treated rats. On the contrary CT was found to increase the locomotor activity of NIH mice in forced swimming test through reducing the lactic acid in the circulation. Data in this study gives further support on CT's potential as a therapeutic drug for treating AD. / by Wong, Kin Kwan Kelvin. / Source: Dissertation Abstracts International, Volume: 73-01, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 144-167). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Orientador: Hideko Yamanaka / Coorientador: Flávio Santos Damos / Banca: Cecilio Sadao Fugivara / Banca: Eder Tadeu Gomes Cavalheiro / Banca: Carla dos Santos Riccardi / Banca: Silvia Helena Pires Serrano / Resumo: Os pesticidas estão entre os poluentes mais preocupantes, devido à toxicidade e presença significativa no ambiente. A sua toxicidade é baseada na capacidade de inibir irreversivelmente a enzima acetilcolinesterase (AChE) que é chave na transmissão de impulsos nervosos. Este trabalho descreve o desenvolvimento de sensor contendo hexapeptídeo, bioinspirado em enzima acetilcolinesterase, para detecção de pesticidas organofosforados e carbamatos. A sequência peptídica (NH3+ - His - Glu - Trp - Arg - Pro - Ser - COO-) foi imobilizada sobre nanopartículas magnéticas (Fe3O4) previamente sintetizadas, modificadas com quitosana e posteriormente funcionalizadas com 1,12-diaminododecano. As condições experimentais de imobilização do peptídeo foram otimizadas, sendo estas: concentração 5,0 x 10-5 mol L-1 e tempo de incubação de 30 minutos a 25 ºC. O grupo carboxílico presente na sequência peptídica foi ativado com o uso de agentes de acoplamento 1-etil-3-(3-dimetilaminopropil) carbodiimida (EDC) e N-hidróxisuccinimida (NHS). A razão de concentração otimizada de EDC/NHS foi de 18,6/12,5 mmol L-1, respectivamente, e tempo de ativação de 60 minutos. O sinal eletroquímico do peptídeo foi monitorado pelo pico de oxidação da histidina, cujo valor é diminuído ao interagir com o pesticida. O perclorato de sódio (NaClO4) 0,1 mol L-1 pH 7,5 foi selecionado como eletrólito suporte. Os parâmetros da voltametria de onda quadrada foram otimizados (frequência de 100 Hz, amplitude de 90 mV e incremento ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Pesticides are among the most worrying pollutants due to toxicity and significant presence in the environment. Its toxicity is based on the ability to irreversibly inhibit the enzyme acetylcholinesterase (AChE) which is key in the transmission of nerve impulses. This work describes the development of a sensor containing hexapeptide, bioinspiring enzyme acetylcholinesterase, for the detection of organophosphorus pesticides and carbamates. The peptide sequence (NH3+ - His - Glu - Trp - Arg - Pro - Ser - COO-) was immobilized on previously synthesized magnetic nanoparticles (Fe3O4), modified with chitosan and subsequently functionalized with 1,12 - diaminododecane. The experimental conditions of immobilization of the peptide were optimized, being: 5,0 x 10-5 mol L-1 concentration and incubation time of 30 minutes at 25 ºC. The carboxyl group present in the peptide sequence was activated with the use of 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) coupling agents. The optimum concentration ratio of EDC / NHS was 18.6 / 12.5 mmol L-1, respectively, and activation time of 60 minutes. The electrochemical signal of the peptide was monitored by the histidine oxidation peak, whose value is decreased when interacting with the pesticide. Sodium perchlorate (NaClO4) 0.1 mol L-1 pH 7.5 was selected as supporting electrolyte. The parameters of the square wave voltammetry were optimized (frequency of 100 Hz, amplitude of 90 mV and increment of sweep of 6 mV) using a matrix of factorial planning. The preconcentration time of the peptide with the pesticide was fixed in 5 minutes. The sensor presented linear response in the studied concentration ranges, with detection limits of 6.0 x 10-11 mol L-1 and 4.0 x 10- 10 mol L- 1 for carbofuran and chlorpyrifos, respectively. The storage in the refrigerator at ± 4 °C allowed 85% stability of the immobilized peptide after a period of... / Doutor

Made available in DSpace on 2016-08-10T10:39:07Z (GMT). No. of bitstreams: 1 Douglas Dantas Rodrigues.pdf: 1094258 bytes, checksum: be59e50a44653cbac4abda0ac7ca028f (MD5) Previous issue date: 2015-03-30 / The agent to combat epidemic (ACEs) are the more exposed classes to effect of pesticides, since their expositions is given to preparation to application of pesticides. The uses of PPE are essential to protection of these workers, yet they often dispense their uses. Factors such as lack of knowledge about the correct use of pesticides or no respect for the PPE uses may be related to the risk of intoxication. It is well known that pesticides may cause DNA damage in people who are occupationally exposed. Although there are many studies on the damage to farmers, few studies have been performed with ACEs. In this context, this study aimed to evaluate the genomic damage in ACEs of the city of Aparecida de Goiânia (GO) and to correlate the damage with intoxication reports and use of PPE and styles of life of the agents. We selected for research 39 ACEs and 39 people from the local community who were not occupationally exposed to pesticides. Based on a structured questionnaire was collected lifestyle information of the participants, such as age, gender, alcohol consumption, tobacco use, and data on the use of PPE and signs and symptoms of intoxication of ACEs. The evaluation of genomic damage was made by micronucleus technique, and the cholinesterase test was performed in ACEs to observe the presence of chronic intoxication. The exposed group consisted of 21 men (53.8%) and 18 women (46.2%). Of agents 39, 10 (25.6%) smoked and 29 (74.4%) did not smoke; 23 (58.9%) drank and 16 (41.1%) did not drink. The evaluation of cholinesterase was normal in all agents, but when comparing the frequency of micronuclei in the case group with control, a statistical difference (p> 0.05) was observed. By correlating the use of PPE with genomic damage, was not found a significant difference. Based on the findings of this research, it can be states that the use of pesticides by ACEs is a risk to it in genomic damage, which can lead to serious healths problems for the same. Worth pointing out that there are a need for education about the risk of using pesticides, particularly for occupationally exposed group. / Os agentes de combate a endemias (ACEs) são as categorias mais exposta aos efeitos de pesticidas, pois sua exposição se dá deste o preparo da calda dos pesticidas até a aplicação do mesmo. O uso de EPIs é essencial para a proteção desses trabalhadores, todavia eles muitas vezes os dispensam. Fatores como a falta de conhecimento acerca do uso correto dos pesticidas ou não respeito aos usos das EPIs podem estar relacionados com o risco de intoxicação. Sabe-se que os pesticidas podem causar danos no DNA em pessoas que são ocupacionalmente expostas. Apesar de existir muitos estudos sobre o dano em agricultores, poucos estudos foram realizados com ACEs. Neste contexto, o presente trabalho teve como objetivo avaliar o dano genômico em ACEs da cidade de Aparecida de Goiânia (GO) e correlacionar o dano com os quadros de intoxicação e com uso de EPIs e aos estilos de vidas dos agentes. Foram selecionados para pesquisa 39 ACEs e 39 pessoas da comunidade local que não foram ocupacionalmente expostos a pesticidas. Através de um questionário estruturado foram coletados dados sobre estilo de vida dos participantes, tais como idade, sexo, etilismo, tabagismo, além de dados acerca do uso de EPIs e sinais e sintomas de intoxicação dos ACEs. A avaliação do dano genômico foi realizada pela da técnica de micronúcleo, e o teste de colinesterase foi realizado nos ACEs para observar a presença de intoxicação crônica. O grupo exposto foi composto por 21 homens (53,8%) e 18 mulheres (46,2%). Dos 39 agentes, 10 (25,6%) fumavam e 29 (74,4%) não fumavam e 23 (58,9%) bebiam e 16 (41,1%) não bebiam. A avaliação da colinesterase foi normal em todos os agentes, porém, ao comparar a frequência dos micronúcleos do grupo caso com controle, foi observada uma diferença estatística (p<0,05). Ao correlacionar o uso de EPIs com dano genômico, não foi encontrada uma diferença significativa. Com base nos achados dessa pesquisa, pode-se afirmar que o uso de pesticidas por ACEs representa um risco de dano genômico, o que pode trazer sérios problemas de saúde para os mesmos. Vale salientar que há uma necessidade de educação sobre o risco do uso de pesticidas, principalmente para os grupos ocupacionalmente exposto.

by Zhang Xiang. / "June 2003." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (p. 165-174). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2018-04-24T20:39:20Z No. of bitstreams: 1 2014 - Maria Clara da Silva Negreiros Botelho.pdf: 2378053 bytes, checksum: bfc9cb9e12f70927d727da1fba44c017 (MD5) / Made available in DSpace on 2018-04-24T20:39:21Z (GMT). No. of bitstreams: 1 2014 - Maria Clara da Silva Negreiros Botelho.pdf: 2378053 bytes, checksum: bfc9cb9e12f70927d727da1fba44c017 (MD5) Previous issue date: 2014-02-26 / CNPq / FAPUR (UFRRJ) / The aim of this study was to evaluate the efficacy of a collar impregnated with deltamethrin and propoxur to control infestations R. sanguineus and Ctenocephalides felis felis in beagle dogs and to evaluate the safety of treatment by clinical, haematological and biochemical tests, and to assess the dosage of serum cholinesterase in dogs after the treatment with a carbamate. Twenty beagles dogs were selected, divided into two groups: control and treated with a collar containing a combination of deltamethrin 4g and propoxur 12g. Each animal was infested with 50 R. sanguineus ticks (25 males and 25 females) and 100 C. felis felis fleas adults (50 males and 50 females). For the assessment of efficacy, 48 hours after each infestation, the mechanical removal of the parasites was performed. Each animal was infested with fleas and ticks on days -7 (before treatment), -2, +5, +12, +19 and +26. After day 26, every 14 days infestations were performed until the day +208. After this experimental day, the animals were infested with fleas only every 7 days until the termination of the study on day +271. For haematological and biochemical evaluations, blood samples were collected from animals in both groups every seven days until day +14. This experimental day up to +266, samples were collected every 14 days. After removal of the collars (day 274), three collections were made on days: +275, +281 and +288 , to assess the possible occurrence of changes in serum levels of serum enzyme butyrylcholinesterase (BChE). The acaricides effectiveness of the test product was 100 % on days +7, +21 and +28. The efficacy values were above 90% until day 126. As from day +182, the efficacy reached values lower than 80%. Only after day +168, levels declined below 80%. Pulicide efficiencies of the test product was 100 % on days +21, +28, +42, +56, +98 and +112. The efficacy values were above 90% until day +266, reducing 89.48% on day 273. The animals showed no changes in clinical and laboratory examinations that could be associated with adverse effects from the use of the product tested. It is concluded that a collar containing a combination of deltamethrin 4g and propoxur 12g was effective in controlling R. sanguineus ticks for up to 126 days and control C. felis felis fleas for up to 266 days in addition to proving to be safe for use in dogs. The animals showed no clinical and laboratory changes that could be associated with adverse effects from the use of the product tested, thus demonstrating that the association deltamethrin and propoxur in an impregnated collar is safe for use in dogs. / O objetivo do presente estudo foi avaliar a efic?cia de uma coleira impregnada com deltametrina e propoxur no controle de infesta??es por Rhipicephalus sanguineus e Ctenocephalides felis felis em c?es da ra?a beagle e avaliar a seguran?a cl?nica do tratamento atrav?s de exames cl?nicos, hematol?gicos e bioqu?micos, al?m de avaliar a dosagem da colinesterase s?rica em c?es submetidos ao tratamento com um carbamato. Para isso, foram selecionados 20 animais da ra?a beagle em bom estado sanit?rio, distribu?dos em dois grupos: controle e tratado com uma coleira impregnada com 4g de deltametrina e 12g de propoxur. Cada animal foi infestado com 50 carrapatos R. sanguineus (25 machos e 25 f?meas) e 100 pulgas C. felis felis adultas (50 machos e 50 f?meas). Para a avalia??o da efic?cia, 48 horas ap?s cada infesta??o, foi realizada a remo??o mec?nica dos parasitos. Cada animal foi infestado com pulgas e carrapatos nos dias -7 (antes do tratamento), -2, +5, +12, +19 e +26. Ap?s o dia +26 foram realizadas infesta??es a cada 14 dias at? o dia +208. Ap?s este dia experimental, os animais foram infestados somente com pulgas a cada sete dias, at? o encerramento do estudo, no dia +271. Para avalia??o hematol?gica e bioqu?mica, amostras de sangue foram coletadas dos animais de ambos os grupos a cada sete dias at? o dia +14. Deste dia experimental at? o dia + 266, as coletas foram realizadas a cada 14 dias. Ap?s a retirada das coleiras (dia +274), foram realizadas mais tr?s coletas nos dias +275, +281 e +288, para avaliar a poss?vel ocorr?ncia de altera??es nos n?veis s?ricos da enzima butirilcolinesterase s?rica (BChE). A efic?cia carrapaticida do produto em teste foi de 100% nos dias +7, +21 e +28. Os valores de efic?cia foram superiores a 90% at? o dia+126. Apenas a partir do dia +182, a efic?cia atingiu valores inferiores a 80%. Apenas ap?s o dia +168 os n?veis declinaram abaixo dos 80%. A efic?cia pulicida do produto em teste foi de 100% nos dias +21, +28, +42, +56, +98 e +112. Os valores de efic?cia foram superiores a 90% at? o dia+266, reduzindo a 89,48% no dia +273. Os animais n?o apresentaram altera??es em exame cl?nico e laboratoriais que pudessem ser associados a efeitos adversos provenientes do uso do produto testado. Conclui-se que a coleira impregnada com 4g de deltametrina e 12g de propoxur foi eficaz no controle de carrapatos R. sanguineus por at? 126 dias e no controle de pulgas C. felis felis por at? 266 dias. Os animais n?o apresentaram altera??es cl?nicas e laboratoriais que pudessem ser associadas a efeitos adversos provenientes do uso do produto testado, demonstrando assim que a associa??o deltametrina e o propoxur em uma coleira impregnada ? segura para uso em c?es.

While life expectancy increases worldwide, treatment of neurodegenerative diseases such as AD becomes a major task for industrial and academic research. Currently, a treatment of AD is only symptomatical and limited to an early stage of the disease by inhibiting AChE. A cure for AD might even seem far away. A rethinking of other possible targets is therefore necessary. Addressing targets that can influence AD even at later stages might be the key. Even if it is not possible to find a cure for AD, it is of great value for AD patients by providing an effective medication. The suffering of patients and their families might be relieved and remaining years may be spent with less symptoms and restrictions. It was shown that a combination of hCB2R agonist and BChE inhibitor might exactly be a promising approach to combat AD. In the previous chapters, a first investigation of dual-acting compounds that address both hCB2R and BChE was illustrated (figure 6.1). A set of over 30 compounds was obtained by applying SARs from BChE inhibitors to a hCB2R selective agonist developed by AstraZeneca. In a first in vitro evaluation compounds showed selectivity over hCB1R and AChE. Further investigations could also prove agonism and showed that unwanted off-target affinity to hMOP receptor could be designed out. The development of a homology model for hCB2R (based on a novel hCB1R crystal) could further elucidate the mode of action of the ligand binding. Lastly, first in vivo studies showed a beneficial effect of selected dual-acting compounds regarding memory and cognition. Since these first in vivo studies mainly aim for an inhibition of the BChE, it should be the aim of upcoming projects to proof the relevance of hCB2R agonism in vivo as well. In addition, pharmacokinetic as well as solubility studies may help to complete the overall picture. Currently, hybrid-based dual-acting hCB2R agonists and selective BChE inhibitors are under investigation in our lab. First in vitro evaluations showed improved BChE inhibition and selectivity over AChE compared to tacrine.78 Future in vitro and in vivo studies will clarify their usage as drug molecules with regard to hepatotoxicity and blood-brain barrier penetration. Since the role of hCB2R is not yet completely elucidated, the use of photochromic toolcompounds becomes an area of interest. These tool-compounds (and their biological effect) can be triggered upon irradiation with light and thus help to investigate time scales and ligand binding. A set of 5-azobenzene benzimidazoles was developed and synthesized. In radioligand binding studies, affinity towards hCB2R could be increased upon irradiation with UV-light (figure 6.2). This makes the investigated compounds the first GPCR ligands that can be activated upon irradiation (not vice versa). The aim of upcoming research will be the triggering of a certain intrinsic activity by an “efficacy-switch”. For this purpose, several attempts are currently under investigation: an introduction of an azobenzene moiety at the 2-position of the benzimidazole core already led to a slight difference in efficacy upon irradiation with UV light. Another approach going on in our lab is the development of hCB1R switches based on the selective hCB1R inverse agonist rimonabant. First in vitro results are not yet available (figure 6.3). / Durch die weltweit steigende Lebenserwartung rückt die Behandlung von neurodegenerativen Krankheiten, wie der Alzheimer’schen Krankheit, immer mehr in den Fokus der industriellen und akademischen Forschung. Momentan erfolgt die Behandlung der Alzheimer’schen Krankheit durch die Blockade der AChE nur symptomatisch und in einem Frühstadium. Eine Heilung scheint dabei in weiter Ferne zu liegen, weshalb ein Umdenken nach neuen Ansätzen stattfinden sollte. Der Schlüssel könnte darin liegen, dass man biologischen Funktionen adressiert, die den Verlauf der Alzheimer’schen Krankheit auch in einem späteren Stadium beeinflussen. Selbst wenn eine Heilung in absehbarer Zeit unmöglich bleibt, ist es für die betroffenen Patienten eine erhebliche Erleichterung auf eine effektive Medikation zurückgreifen zu können. Das Leid der Patienten und ihrer Familien könnte dadurch gelindert und die verbleibenden Lebensjahre ohne Symptome und Einschränkungen genossen werden. In den vorangegangenen Kapiteln wurde bereits gezeigt, dass die Kombination aus einem hCB2R Agonisten und einem BChE Hemmer genau diesen vielversprechenden Ansatz verfolgt. Ein erster Entwicklungsansatz von dual-aktiven hCB2R Agonisten / BChE Hemmern wurde in den Kapiteln 3 und 4 ausführlich dargestellt (Abb. 7.1). Ein Set von 30 verschiedenen Verbindungen wurde synthetisiert, indem die Erkenntnisse der Struktur-Wirkungsbeziehungen von anderen BChE Hemmern auf einen von AstraZeneca entwickelten selektiven hCB2R Agonisten angewendet wurden. Erste in vitro Untersuchungen zeigten eine hohe Selektivität gegenüber hCB1R und AChE auf. Desweiteren verhielten sich alle getesteten Substanzen wie Agonisten. Nachdem ausgewählte Substanzen auf ihre „off-target“ Wechselwirkung mit dem hMOP Rezeptor untersucht wurden, konnten diese strukturellen Merkmale in nachfolgenden Entwicklungsbemühungen berücksichtigt werden. Die Entwicklung eines hCB2R Homologiemodells (basierend auf einer erst kürzlich veröffentlichten hCB1R Kristallstruktur) lieferte wertvolle Informationen zum Bindemodus und der Wirkweise der Liganden am Rezeptor. Schlussendlich konnte in einer ersten in vivo Studie bewiesen werden, dass ausgewählte dual-aktive Substanzen eine positive Auswirkung auf das Gedächtnis und die kognitiven Eigenschaften haben. Da diese in vivo Untersuchungen hauptsächlich die Hemmung der BChE berücksichtigen, wäre es sinnvoll, in zukünftigen Studien den Einfluss der hCB2R Agonisten zu untersuchen. Pharmakokinetik- und Löslichkeitsstudien könnten zudem helfen, das Gesamtbild zu komplettieren. Im Moment befinden sich auch dual-aktive hCB2R Agonisten / BChE Hemmer in der Entwicklung, die den Hybrid-Ansatz verfolgen. Erste in vitro Untersuchungen dazu ergaben vielversprechende Ergebnisse mit einer guten Selektivität gegenüber AChE und einer erhöhten Hemmung der BChE verglichen mit Tacrin.78 Es wird Gegenstand zukünftiger in vitro und in vivo Untersuchungen sein, herauszufinden, ob sich diese Hybride mit Hinblick auf Hepatotoxizität und Blut-Hirnschrankengängigkeit als Wirkstoffe eignen. Da die Rolle des hCB2R noch nicht komplett erforscht ist, erfreut sich die Entwicklung von sog. „tool-compounds“ großen wissenschaftlichen Interesses. Durch die Bestrahlung mit Licht können diese „tool-compounds“ (und ihr nachgeschalteter biologischer Effekt) gesteuert werden. Eine genauere Untersuchung von Zeitskalen und Ligandbindung an den Rezeptor wird dadurch ermöglicht. Ein Set von 5-Azobenzolbenzimidazolen wurde zu diesem Zwecke entwickelt und synthetisiert. In Radioligandbindungsstudien konnte gezeigt werden, dass sich die Affinität gegenüber dem hCB2R durch die Bestrahlung mit UV-Licht erhöhen lässt (Abb. 7.2). Diese Eigenschaft macht die entwickelten Substanzen zu den ersten GPCR-Liganden, die durch Licht aktiviert werden können (nicht umgekehrt wie bei den bisher beschriebenen photochromen GPCR-Liganden). Ziel zukünftiger Forschungsbemühungen wird die Steuerung einer bestimmten intrinsischen Aktivität / Effekts durch die Bestrahlung mit Licht sein. Zu diesem Zwecke werden aktuell mehrere Herangehensweisen untersucht: die Einführung eines Azobenzol-Strukturelements an Position 2 des Benzimidazol-Grundgerüsts zeigte in ersten in vitro Untersuchungen bereits Unterschiede bei Bestrahlung mit UV-Licht. Eine weitere Herangehensweise ist die Entwicklung von „Photo-Schaltern“ auf Basis von Rimonabant, einem selektiven hCB1R inversen Agonisten. Hier stehen erste in vitro Ergebnisse jedoch noch aus (Abb. 7.3).

myotubesProper muscle function depends upon the fine tuning of the different molecular components of the neuromuscular junction (NMJ). Synaptic acetylcholinesterase (AChE) is responsible for rapidly terminating neurotransmission. Neuroscientists in the field have elucidated many aspects of synaptic AChE structure, function, and localization during the last 75 years. Nevertheless, how the enzyme is regulated and targeted to the NMJ is not completely understood. In skeletal muscle the synaptic AChE form derives from two separate genes encoding the catalytic and the collagenic tail (ColQ) subunits respectively. ColQ-AChE expression is regulated by muscle activity; however, how this regulation takes place remains poorly understood. We found that over or down-regulation of ColQ is sufficient to change the levels of AChE activity by promoting assembly of higher order oligomeric forms including the collagen-tailed forms. Furthermore, when peptides containing the Proline Rich Attachment Domain (PRAD), the region of ColQ that interacts with the AChE, are fed to muscle cells or cell lines expressing AChE, they are taken up by the cells and retrogradely transported to the endoplasmic reticulum (ER)/Golgi network where they induce assembly of newly synthesize AChE into tetramers. This results in an increase, as a consequence, in total cell associated AChE activity and active tetramer secretion, making synthetic PRAD peptides potential candidates for the treatment of organophosphate pesticides and nerve gas poisoning. To study the developmental regulation of ColQ-AChE we determined the levels of ColQ and ColQ mRNA in primary quail muscle cells in culture and as a function of muscle activity. Surprisingly, we found dissociation between transcription and translation of ColQ from its assembly into ColQ-AChE indicating the importance of posttranslational controls in the regulation of AChE folding and assembly. Furthermore, we found that the vast majority of the ColQ molecules in QMCs are not assembled into ColQ-AChE, suggesting that they can have alternative function(s). Finally, we found that the levels of ER molecular chaperones calnexin, calreticulin, and particularly protein disulfide isomerase are regulated by muscle activity and they correlate with the levels of ColQ-AChE. More importantly, our results suggest that newly synthesized proteins compete for chaperone assistance during the folding process.

Le chlorpyrifos (CPF) est un insecticide organophosphoré qui est capable de produire des effets développementaux chez les rongeurs, tels que des perturbations de la mémoire spatiale, des altérations du niveau de lactivité, des modifications du niveau danxiété et de la dépression. Cependant, ces troubles comportementaux persistants ont été décrits à partir de différentes périodes dexposition courtes (4 jours), et à des doses juste sous le seuil de toxicité de ce produit, et ne peuvent donc pas renseigner directement sur le type de perturbations comportementales que pourrait produire une exposition à des doses plus basses de CPF administrées durant une plus grande partie du développement du cerveau. Pour répondre à cette question, nous avons exposé des souris CD1 au CPF (in utero et via le lait ; par lintermédiaire du gavage de la mère ; 5 mg/kg) durant la fin de la gestation et pendant les deux semaines postnatales. Ensuite, des tests comportementaux ont été réalisés avec les animaux adultes. Nous avons également étudié dans les mêmes conditions la phosalone (PHO), un insecticide organophosphoré dont la toxicité est similaire à celle du CPF, mais qui, contrairement à celui-ci, na pas encore donné lieu à des études de toxicité développementale. Nous avons déterminé que le CPF et la PHO sont capables de produire une altération de la mémoire spatiale à long terme (dans un labyrinthe de Lashley), avec un effet plus marqué chez les souris femelles que chez les mâles et pour la PHO. Par contre, seul le CPF altère la mémoire spatiale à court terme des souris mâles (test dalternance spontanée dans un labyrinthe en T). Le CPF et la PHO produisent également une augmentation du niveau danxiété (« elevated plus maze » et « light and dark box ») et donnent des signes de dépression (« forced swimming test ») chez les souris femelles uniquement. Par contre, le CPF et la PHO diminuent lactivité locomotrice spontanée (test dactométrie et vidéotracking) des souris mâles et non des souris femelles. Nous avons ensuite approfondi létude de lanxiété développementale produite dans les mêmes conditions expérimentales mais par différentes doses subtoxiques. Cela nous a permis de préciser quune dose de CPF plus faible, soit 1 mg/kg, produisait une augmentation du niveau danxiété plus importante que 5 mg/kg chez la souris femelle. Nous nous sommes intéressés ensuite à une exposition au CPF directement chez le jeune, afin de déterminer les troubles de lanxiété conséquents à quatre périodes néonatales différentes de 4 jours. Dans ce contexte, 1 mg/kg de CPF durant 4 jours dexposition par voie sous-cutanée suffirait à produire une augmentation du niveau danxiété chez les souris femelles, sauf pour une période dexposition située en fin de la seconde semaine après la naissance. La dose et la période dexposition jouent donc un rôle sur le type deffet produit. Ces expériences confirment également que plusieurs mécanismes daltération du développement sont mis en route par le CPF. Toujours dans le cadre de lanxiété développée chez les souris femelles adultes et liée à une exposition périnatale au CPF en doses subtoxiques (1 mg/kg, via gavage de la mère), lutilisation danxiolytiques spécifiques a révélé que les troubles de lanxiété seraient en relation avec une modification du fonctionnement des récepteurs sérotoninergique 5HT1A. Une des implications de ces résultats est que lexposition au CPF durant le développement du cerveau serait susceptible de produire des troubles comportementaux persistants apparaissant chez ladulte, à partir de doses bien inférieures à celles décrites jusquici dans la littérature. Il en va de même pour la PHO qui pourtant nétait pas encore renseignée comme représentant un danger pour le développement du cerveau des mammifères./Perinatal exposure to apparently subtoxic doses of the organophosphorus insecticide chlorpyrifos (CPF) elicits some form of developmental effects in adult rodents, like alteration of spatial memory, locomotor activity, anxiety and depression disorders. However, these persistent neurobehavioral effects are described in adults exposed during 4 days of perinatal treatment with doses close to the toxicity threshold, and do not indicate the consequences of a prolonged perinatal exposure to lower doses of CPF. Therefore, we orally treated CD1 pregnant and lactating mice with subtoxic doses of CPF (5 mg/kg, daily) during the 4 last days of gestation and the following 2 weeks after delivery and assessed the behaviour of adult offspring. We also studied the phosalone (PHO), another organophosphorus insecticide, which has the same toxicity as CPF but whose developmental toxicity in mammals has not been examined yet. We have found that CPF and PHO produced an alteration of long-term spatial memory (in the Lashley maze). This effect was more pronounced in female mice and in PHO-treated mice. On the other hand, only CPF (and not PHO) produced a decrease in short-term spatial memory of male mice (spontaneous alternation in a transparent T-maze). Both insecticides increased the anxiety level (in the elevated plus maze and in the light and dark box) and induced signs of depression disorder (forced swimming test) in female mice. We showed that CPF and PHO also decreased the level of locomotor activity only in male mice. Then we further studied the anxiety disorder developed by CPF using several different subtoxic doses of CPF. 1 mg/kg CPF daily treated female mice showed greater increase of anxiety level compared to 5 mg/kg. To determine the developmental effects of short exposure to low subtoxic CPF doses on anxiety levels, we directly (subcutaneously) administrated 1 mg/kg (daily) during 4 different postnatal periods of 4 days to CD1 offspring and evaluated their anxiety level once adults. Exposed female mice presented a general increase of anxiety level except when administration occurred during the end of the second postnatal week. The level and the time of the subtoxic exposure to CPF would influence anxiety. We concluded that CPF induced several mechanisms of developmental toxicity. We used specific anxiolytic drugs to show that 5HT1A serotoninergic receptors are one of the mechanisms altered by a long perinatal period of subtoxic CPF exposure that produces an increase of anxiety level in adult female mice. These results imply that exposure to CPF during brain development may represent a risk to produce persistent behavioural troubles, even at doses lower than previously described in the literature. Exposure to PHO for which no developmental effect has been described in mammals, may produce similar behavioural troubles.

Abstract Background: Home care is an important care setting for those with Alzheimer’s disease (AD). It provides support that allows individuals with AD to remain at home and may delay the transition to long-term care homes. Many clients with AD receive medications that are used for managing the symptoms of AD: cholinesterase inhibitors (ChEIs) and memantine. Ontario’s provincial drug benefit plan (ODB) provides subsidies for some of these medications based on specific clinical criteria. These AD medications are costly and can have significant side effects, so it is important to understand how they are being used in practice. Objectives: The objectives of this study were to report the proportion taking AD medications and which types were taken, show the change in receipt of AD medications over time, and show the covariates that were independently associated with receiving AD medications. Methods: Analysis of secondary data was performed on the provincial home care dataset. All home care clients receiving long-term home care services were assessed using the RAI-Home Care (RAI-HC), which is a comprehensive and standardized assessment. One assessment from each individual over the age of 65 who was assessed between January 2004 and September 2008 was used, for a final sample size of 321,013. Results: Overall, 65% of clients with a diagnosis of AD were receiving an AD medication. Logistic regression analysis among those diagnosed with AD showed that increased physical impairment and clinical complexity were associated with decreased odds of receiving AD medication. Contraindicating diagnoses such as congestive heart failure, lack of medical oversight and needing to make economic tradeoffs were also associated with decreased odds of receiving AD medication. Conclusions: The multivariate model showed trends of rational prescribing, such as clients with contraindicating diagnoses or very high clinical complexity having decreased odds of receiving AD medications. At the same time, evidence of structural barriers to receiving the medications was shown. There is debate about the cost-effectiveness of these medications. The provincial government could consider expanding ODB guidelines to include all AD medications for those with all levels of cognitive impairment, but further analyses involving longitudinal outcomes available in this dataset should be performed to ensure it would be in the public interest.

Abstract Background: Home care is an important care setting for those with Alzheimer’s disease (AD). It provides support that allows individuals with AD to remain at home and may delay the transition to long-term care homes. Many clients with AD receive medications that are used for managing the symptoms of AD: cholinesterase inhibitors (ChEIs) and memantine. Ontario’s provincial drug benefit plan (ODB) provides subsidies for some of these medications based on specific clinical criteria. These AD medications are costly and can have significant side effects, so it is important to understand how they are being used in practice. Objectives: The objectives of this study were to report the proportion taking AD medications and which types were taken, show the change in receipt of AD medications over time, and show the covariates that were independently associated with receiving AD medications. Methods: Analysis of secondary data was performed on the provincial home care dataset. All home care clients receiving long-term home care services were assessed using the RAI-Home Care (RAI-HC), which is a comprehensive and standardized assessment. One assessment from each individual over the age of 65 who was assessed between January 2004 and September 2008 was used, for a final sample size of 321,013. Results: Overall, 65% of clients with a diagnosis of AD were receiving an AD medication. Logistic regression analysis among those diagnosed with AD showed that increased physical impairment and clinical complexity were associated with decreased odds of receiving AD medication. Contraindicating diagnoses such as congestive heart failure, lack of medical oversight and needing to make economic tradeoffs were also associated with decreased odds of receiving AD medication. Conclusions: The multivariate model showed trends of rational prescribing, such as clients with contraindicating diagnoses or very high clinical complexity having decreased odds of receiving AD medications. At the same time, evidence of structural barriers to receiving the medications was shown. There is debate about the cost-effectiveness of these medications. The provincial government could consider expanding ODB guidelines to include all AD medications for those with all levels of cognitive impairment, but further analyses involving longitudinal outcomes available in this dataset should be performed to ensure it would be in the public interest.