Graduation date: 2006 / Most vertebrates exhibit seasonality in many life history traits. Such seasonal rhythms are temporally organized via the transduction of environmental cues (e.g., photoperiod, temperature) into appropriate endocrine signals. However, among ectothermic vertebrates that undergo continuous winter dormancy, temperature is the only environmental cue available for synchronizing seasonal rhythms. Most intriguing is that in species where reproduction occurs immediately following spring emergence, the associated changes in neurophysiology and behavior that accompany reproduction likely occur during winter dormancy. The purpose of this dissertation research was to explore the mechanisms by which temperature cues affect the chronobiology and seasonal reproduction of red-sided garter snakes (Thamnophis sirtalis parietalis). Because of their roles in circadian organization and energy balance, melatonin and corticosterone are likely hormonal components of these time-keeping systems. I first characterized the interactions between melatonin and corticosterone to better understand the hormonal mechanisms facilitating temperature-induced reproduction. Melatonin and corticosterone additively inhibit reproductive behavior during the spring mating season. Experimental manipulations with a serotonin receptor antagonist suggest the mechanism underlying these effects involves a serotonin-regulated system. Although melatonin does not influence corticosterone responses to capture stress, capture stress significantly increases melatonin concentrations. To investigate the functional significance of these interactions in regulating temperature-induced reproduction, I measured body temperatures of snakes as well as circadian melatonin and corticosterone cycles during winter dormancy and spring emergence using a combination of field and laboratory experiments. Surprisingly, an increase in body temperature is not necessary for emergence from winter dormancy. Rather, critically low temperatures may serve as a zeitgeber entraining an endogenous circannual cycle that regulates emergence. Decreased environmental temperatures, in the absence of changing photoperiod cues, modulate circadian melatonin and corticosterone rhythms during hibernation. Such temperature-induced changes in hormone rhythms may facilitate seasonal reproductive behavior following spring emergence. Furthermore, a phase-shift in corticosterone rhythms during the mating season may regulate the seasonal transition between reproductive and non-reproductive states in red-sided garter snakes. Such studies investigating the environmental and hormonal mechanisms underlying time-keeping systems may provide valuable insight into the potential impact of environmental perturbations (e.g., climate change) on seasonal rhythms in physiology and behavior.

Objectives: To investigate the effects of serotonin and melatonin (MLT) on the regulation of malignant growth and the activity of serotonin receptors (5HTR1a/-1b) in prostate cancer (PCa) cell lines. Materials and Methods: In four PCa cell lines (LNCaP, 22RV1, PC3, DU145) and two reference cell lines 5HTR1a and -1b, relative mRNA expression levels were assessed. Different serotonin and MLT receptor agonists and antagonists were used in stimulation and inhibition experiments. Results: mRNA expression of 5HTR1b was higher than that of 5HTR1a in all PCa cell lines. Serotonin showed a significant growth stimulatory effect in all PCa lines. The 5HTR1a and -1b agonists/antagonists did not significantly affect viability. MLT inhibited viability only in PC3 cells. Similarly, the 5HTR1a antagonist induced apoptotic changes in PC3 cells only at 10–4M, while the 5HTR1b antagonist induced necrosis at 10–4M in all cell lines. Cell cycle alterations were seen in PC3 and DU145 cells under the influence of the 5HTR1a antagonist. Conclusions: Serotonin receptor antagonists and agonists as well as MLT influence viability and apoptosis of PCa cell lines at supraphysiologic concentrations. In contrast to other reports, our results do not support a regulatory role of serotonin or MLT receptor activation or inhibition in PCa growth. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / Levetiracetam (LEV), nova droga antiepilÃptica, apresenta eficÃcia na terapia adicional das convulsÃes e em modelos experimentais. Clonazepam (CNZP) à um benzodiazepÃnico utilizado no tratamento de convulsÃes mioclÃnicas e crises generalizadas. Melatonina (MEL), hormÃnio pineal, demonstra atividade anticonvulsivante em diversos modelos animais. Objetivando investigar novos mecanismos relacionados aos efeitos dessas drogas, foi realizado estudo comparativo, a partir da anÃlise da influÃncia do prÃ-tratamento com LEV, CNZP ou MEL sobre o estresse oxidativo neuronal e sobre a modulaÃÃo de sistemas de neurotransmissÃo (colinÃrgico e dopaminÃrgico) durante as convulsÃes induzidas por pilocarpina (400mg/Kg; P400). Camundongos Swiss, machos, 25-30g receberam LEV, 200 mg/Kg, CNZP, 0,5mg/Kg ou MEL, 25mg/Kg, i.p., (doses escolhidas à partir de curvas dose-resposta) 30min antes de P400. Hipocampo e corpo estriado foram removidos para as anÃlises neuroquÃmicas. Experimentos in vitro, onde homogenatos cerebrais foram incubados com as drogas em estudo (50, 100 ou 200g/mL), tambÃm permitiram analisar alteraÃÃes no estresse oxidativo apÃs a induÃÃo de choque tÃrmico e, ainda, a densidade de receptores muscarÃnicos no hipocampo. Os estudos sobre os efeitos sobre o sistema de neurotransmissores colinÃrgicos demonstraram que o prÃ-tratamento com LEV, CNZP ou MEL causou reduÃÃo nos tremores induzidos por oxotremorina e elevou a atividade da acetilcolinesterase no hipocampo. LEV e CNZP alteraram o binding dos receptores muscarÃnicos hipocampais in vivo, revertendo a downregulation induzida por P400, e ensaios in vitro demostraram alteraÃÃo no binding muscarÃnico hipocampal pela prÃvia incubaÃÃo com LEV, CNZP ou MEL. Os ensaios de binding demonstraram, ainda, a downregulation dos receptores D2 hipocampais nos animais tratados LEV, CNZP ou prÃ-tratados com MEL antes de P400. As anÃlises para investigaÃÃo da atividade antioxidante de LEV e CNZP e do papel da aÃÃo antioxidativa da MEL na proteÃÃo contra as convulsÃes permitiram observar que a associaÃÃo com vitamina E potencializou os efeitos anticonvulsivantes de todas as drogas estudas. A administraÃÃo prÃvia de LEV, CNZP ou MEL, antes de P400, reverteu o aumento nos nÃveis de peroxidaÃÃo lipÃdica e nitrito-nitrato e normalizou a atividade da catalase e os nÃveis fisiolÃgicos do antioxidante glutationa em hipocampo e/ou corpo estriado. Nos experimentos de stress oxidativo in vitro, o aumento da peroxidaÃÃo lÃpÃdica, dos nÃveis de nitrito-nitrato e da atividade da catalase nos homogenatos cerebrais submetidos ao choque tÃrmico, foram alterados de forma significativa pela incubaÃÃo prÃvia com LEV, CNZP ou MEL, onde estas drogas foram capazes de reduzir os nÃveis de MDA, de nitrito-nitrato e, ainda, estabilizar a atividade da catalase, potencializando, assim, a atividade enzimÃtica antioxidante endÃgena e a capacidade de inativaÃÃo de radicais livres. Dessa forma, o estudo sugere uma aÃÃo moduladora, exercida por LEV, CNZP e MEL sobre o funcionamento dos sistemas muscarÃnico e dopaminÃrgico, em nÃvel central, como mecanismo alternativo para a proteÃÃo contra as convulsÃes no modelo de P400, bem como a participaÃÃo de propriedades antioxidantes diretas ou indiretas dessas drogas, atravÃs da capacidade de modificar a resposta ao estresse oxidativo neuronal. / Levetiracetam (LEV), nova droga antiepilÃptica, apresenta eficÃcia na terapia adicional das convulsÃes e em modelos experimentais. Clonazepam (CNZP) à um benzodiazepÃnico utilizado no tratamento de convulsÃes mioclÃnicas e crises generalizadas. Melatonina (MEL), hormÃnio pineal, demonstra atividade anticonvulsivante em diversos modelos animais. Objetivando investigar novos mecanismos relacionados aos efeitos dessas drogas, foi realizado estudo comparativo, a partir da anÃlise da influÃncia do prÃ-tratamento com LEV, CNZP ou MEL sobre o estresse oxidativo neuronal e sobre a modulaÃÃo de sistemas de neurotransmissÃo (colinÃrgico e dopaminÃrgico) durante as convulsÃes induzidas por pilocarpina (400mg/Kg; P400). Camundongos Swiss, machos, 25-30g receberam LEV, 200 mg/Kg, CNZP, 0,5mg/Kg ou MEL, 25mg/Kg, i.p., (doses escolhidas à partir de curvas dose-resposta) 30min antes de P400. Hipocampo e corpo estriado foram removidos para as anÃlises neuroquÃmicas. Experimentos in vitro, onde homogenatos cerebrais foram incubados com as drogas em estudo (50, 100 ou 200g/mL), tambÃm permitiram analisar alteraÃÃes no estresse oxidativo apÃs a induÃÃo de choque tÃrmico e, ainda, a densidade de receptores muscarÃnicos no hipocampo. Os estudos sobre os efeitos sobre o sistema de neurotransmissores colinÃrgicos demonstraram que o prÃ-tratamento com LEV, CNZP ou MEL causou reduÃÃo nos tremores induzidos por oxotremorina e elevou a atividade da acetilcolinesterase no hipocampo. LEV e CNZP alteraram o binding dos receptores muscarÃnicos hipocampais in vivo, revertendo a downregulation induzida por P400, e ensaios in vitro demostraram alteraÃÃo no binding muscarÃnico hipocampal pela prÃvia incubaÃÃo com LEV, CNZP ou MEL. Os ensaios de binding demonstraram, ainda, a downregulation dos receptores D2 hipocampais nos animais tratados LEV, CNZP ou prÃ-tratados com MEL antes de P400. As anÃlises para investigaÃÃo da atividade antioxidante de LEV e CNZP e do papel da aÃÃo antioxidativa da MEL na proteÃÃo contra as convulsÃes permitiram observar que a associaÃÃo com vitamina E potencializou os efeitos anticonvulsivantes de todas as drogas estudas. A administraÃÃo prÃvia de LEV, CNZP ou MEL, antes de P400, reverteu o aumento nos nÃveis de peroxidaÃÃo lipÃdica e nitrito-nitrato e normalizou a atividade da catalase e os nÃveis fisiolÃgicos do antioxidante glutationa em hipocampo e/ou corpo estriado. Nos experimentos de stress oxidativo in vitro, o aumento da peroxidaÃÃo lÃpÃdica, dos nÃveis de nitrito-nitrato e da atividade da catalase nos homogenatos cerebrais submetidos ao choque tÃrmico, foram alterados de forma significativa pela incubaÃÃo prÃvia com LEV, CNZP ou MEL, onde estas drogas foram capazes de reduzir os nÃveis de MDA, de nitrito-nitrato e, ainda, estabilizar a atividade da catalase, potencializando, assim, a atividade enzimÃtica antioxidante endÃgena e a capacidade de inativaÃÃo de radicais livres. Dessa forma, o estudo sugere uma aÃÃo moduladora, exercida por LEV, CNZP e MEL sobre o funcionamento dos sistemas muscarÃnico e dopaminÃrgico, em nÃvel central, como mecanismo alternativo para a proteÃÃo contra as convulsÃes no modelo de P400, bem como a participaÃÃo de propriedades antioxidantes diretas ou indiretas dessas drogas, atravÃs da capacidade de modificar a resposta ao estresse oxidativo neuronal. / Levetiracetam (LEV), a new antiepileptic drug, shows efficacy in the treatment of additional seizures and in experimental models. Clonazepam (CNZP) is a benzodiazepine used to treat myoclonic seizures and generalized seizures. Melatonin (MEL), the pineal hormone, shows anticonvulsant activity in several animal models. To investigate new mechanisms related to the effects of these drugs, comparative study was conducted, from the analysis of the influence of pretreatment with LEV, CNZP or MEL on the oxidative stress and neuronal modulation of neurotransmitter systems (cholinergic and dopaminergic) during seizures induced by pilocarpine (400mg/Kg; P400). Male Swiss mice, 25-30g received LEV, 200 mg / kg, CNZP, 0.5 mg / kg or MEL, 25mg/kg, ip (doses chosen from the dose-response curves) 30min before P400. Hippocampus and striatum were removed for neurochemical analysis. In vitro experiments, where brain homogenates were incubated with drugs under study (50, 100 ou 200g/mL) also allowed us to analyze changes in oxidative stress after induction of heat shock and also the density of muscarinic receptors in the hippocampus. Studies on the muscarinic modulation demonstrated that pretreatment with LEV, CNZP or MEL resulted in lower oxotremorina induced tremors and increased acetylcholinesterase activity in the hippocampus. LEV and CNZP altered the binding of hippocampal muscarinic receptors in vivo, reversing the P400-induced downregulation and in vitro tests showed changes in hippocampal muscarinic binding by previous incubation with LEV, CNZP or MEL. The binding assays also showed a downregulation of hippocampal D2 receptors in treated animals LEV, CNZP or pretreated with MEL before P400. Analyses to investigate the antioxidant activity of LEV and CNZP and role of antioxidative action of MEL in the protection against seizures propose that the association with vitamin E increased the anticonvulsant effects of all studied drugs. The prior administration of LEV, MEL or CNZP before P400, reversed the increased levels of lipid peroxidation and nitrite-nitrate and normalized the activity of catalase and the physiological levels of the antioxidant glutathione in the hippocampus and / or striatum. According to in vitro experiments, increased lipid peroxidation, levels of nitrite-nitrate and catalase activity in brain homogenates subjected to thermal shock were significantly altered by incubation with LEV, CNZP or MEL, where these drugs were able to reduce the levels of MDA, nitrite-nitrate, and also stabilize the activity of catalase, enhancing thus the endogenous antioxidant enzyme activity and the ability to inactivate free radicals.Thus, the study suggests a modulatory action exerted by LEV, CNZP and MEL on the functioning of muscarinic and dopaminergic systems in the central nervous system as an alternative mechanism to protect against seizures in the model P400, and the participation of direct and indirect antioxidant properties of these drugs, through the ability to modify the neuronal response to oxidative stress. / Levetiracetam (LEV), a new antiepileptic drug, shows efficacy in the treatment of additional seizures and in experimental models. Clonazepam (CNZP) is a benzodiazepine used to treat myoclonic seizures and generalized seizures. Melatonin (MEL), the pineal hormone, shows anticonvulsant activity in several animal models. To investigate new mechanisms related to the effects of these drugs, comparative study was conducted, from the analysis of the influence of pretreatment with LEV, CNZP or MEL on the oxidative stress and neuronal modulation of neurotransmitter systems (cholinergic and dopaminergic) during seizures induced by pilocarpine (400mg/Kg; P400). Male Swiss mice, 25-30g received LEV, 200 mg / kg, CNZP, 0.5 mg / kg or MEL, 25mg/kg, ip (doses chosen from the dose-response curves) 30min before P400. Hippocampus and striatum were removed for neurochemical analysis. In vitro experiments, where brain homogenates were incubated with drugs under study (50, 100 ou 200g/mL) also allowed us to analyze changes in oxidative stress after induction of heat shock and also the density of muscarinic receptors in the hippocampus. Studies on the muscarinic modulation demonstrated that pretreatment with LEV, CNZP or MEL resulted in lower oxotremorina induced tremors and increased acetylcholinesterase activity in the hippocampus. LEV and CNZP altered the binding of hippocampal muscarinic receptors in vivo, reversing the P400-induced downregulation and in vitro tests showed changes in hippocampal muscarinic binding by previous incubation with LEV, CNZP or MEL. The binding assays also showed a downregulation of hippocampal D2 receptors in treated animals LEV, CNZP or pretreated with MEL before P400. Analyses to investigate the antioxidant activity of LEV and CNZP and role of antioxidative action of MEL in the protection against seizures propose that the association with vitamin E increased the anticonvulsant effects of all studied drugs. The prior administration of LEV, MEL or CNZP before P400, reversed the increased levels of lipid peroxidation and nitrite-nitrate and normalized the activity of catalase and the physiological levels of the antioxidant glutathione in the hippocampus and / or striatum. According to in vitro experiments, increased lipid peroxidation, levels of nitrite-nitrate and catalase activity in brain homogenates subjected to thermal shock were significantly altered by incubation with LEV, CNZP or MEL, where these drugs were able to reduce the levels of MDA, nitrite-nitrate, and also stabilize the activity of catalase, enhancing thus the endogenous antioxidant enzyme activity and the ability to inactivate free radicals.Thus, the study suggests a modulatory action exerted by LEV, CNZP and MEL on the functioning of muscarinic and dopaminergic systems in the central nervous system as an alternative mechanism to protect against seizures in the model P400, and the participation of direct and indirect antioxidant properties of these drugs, through the ability to modify the neuronal response to oxidative stress.

Objectives: To investigate the effects of serotonin and melatonin (MLT) on the regulation of malignant growth and the activity of serotonin receptors (5HTR1a/-1b) in prostate cancer (PCa) cell lines. Materials and Methods: In four PCa cell lines (LNCaP, 22RV1, PC3, DU145) and two reference cell lines 5HTR1a and -1b, relative mRNA expression levels were assessed. Different serotonin and MLT receptor agonists and antagonists were used in stimulation and inhibition experiments. Results: mRNA expression of 5HTR1b was higher than that of 5HTR1a in all PCa cell lines. Serotonin showed a significant growth stimulatory effect in all PCa lines. The 5HTR1a and -1b agonists/antagonists did not significantly affect viability. MLT inhibited viability only in PC3 cells. Similarly, the 5HTR1a antagonist induced apoptotic changes in PC3 cells only at 10–4M, while the 5HTR1b antagonist induced necrosis at 10–4M in all cell lines. Cell cycle alterations were seen in PC3 and DU145 cells under the influence of the 5HTR1a antagonist. Conclusions: Serotonin receptor antagonists and agonists as well as MLT influence viability and apoptosis of PCa cell lines at supraphysiologic concentrations. In contrast to other reports, our results do not support a regulatory role of serotonin or MLT receptor activation or inhibition in PCa growth. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Estrogen therapy (ET) is most effective to reduce menopausal symptoms and prevent other disorders associated with estrogen deficiency. However, Women's Health Initiative studies found that hormone therapy (HT) containing estrogen plus progestogen, but not estrogen-alone increases breast cancer (BC) risk. To prevent the increase in BC risk and yet relieve menopausal symptoms, a novel HT with 17β-estradiol (E2) for symptom relief, progesterone (P4) for uterine protection and melatonin (Mel) for both BC and uterine protection was designed. Inclusion of Mel was postulated to offer uterine protection with lower P4 dose and protect against BC. The goal of this study was to assess the efficacy of E2, P4 and Mel Therapy (EPMT) on mammary cancer (MC) and uterine protection in MMTV-Neu mouse model that mimics HER2 BC. Starting at 2 months age, female mice received Mel in drinking water at night to supplement endogenous Mel surge; while E2 and P4 Therapy (EPT) was provided continuously in diet until 14 months with weekly MC onset and growth monitoring. Normal mammary, uterus and mammary tumors harvested by month 14 were analyzed for potential mechanisms. The results from this study revealed that EPMT delayed tumor onset leading to a decrease in MC incidence. In addition, mice in the EPMT group had no increase in relative uterine weight as opposite to an increase of this parameter in EPT group versus control. The percent tumor-bearing mice with gross metastatic lung lesions were reduced in Mel, EPT and EPMT groups. Mel receptor, estrogen receptor (ER) and progesterone receptor (PR) expression revealed that all tissues examined have Mel receptors. However, ER and PR expression varied. In normal mammary tissue, both ERα and PR were detected by immunohistochemistry. However, no ERα and PR were detected in mammary tumors of same mice. In uterus, mice given Mel or EPMT had significant decreases in PR expression but no change in ERα expression compared to control suggesting that Mel-mediated inhibition of ER binding to estrogen response elements may be involved in the down regulation of uterine PRs. Overall, this study reveal that EPMT prevents mammary cancer and may protect against uterotrophy. / Mylan School of Pharmacy and the Graduate School of Pharmaceutical Sciences; / Pharmacology / PhD; / Dissertation;

Circadian rhythm is a biological rhythm with period of about 24 hours. Circadian rhythm is universal in phyla from bacteria to mammals and exists in different level from gene expression to behavior. Circadian system consists of three components: 1) a self-sustained oscillator; 2) an input pathway which can alter the phase of the oscillator; and 3) an output such as gene expression, enzyme activity, hormone production, heart rate, body temperature or locomotor activities. The way the oscillator regulates its outputs is complicated, in that on one hand usually the oscillator is not the only factor affecting the outputs, and on the other, the oscillator itself is incorporated in intricate pathways. Chicken pineal cell culture is a well-established in vitro model to study circadian rhythm. It contains a self-sustained oscillator which can be phase-shifted by light as input and rhythmically releases melatonin as an output. Here I have characterized the role of norepinephrine (NE), the sympathetic regulatory input of pineal gland, and the microenvironment of pineal cells in melatonin rhythmicity of cultured chicken pineal cells. Chapter 1 of this dissertation provides a review of circadian rhythm with a focus on melatonin regulation in pineal gland. Chapter 2 describes the methods to build up a fraction collector which offers high time resolution of sampling for a superfusion system. Chapter 3 is a technical report of a melatonin enzyme-linked immunosorbent assay suitable for high throughput measurement of melatonin. Chapter 4 presents data demonstrating that daily administration of NE recovers damped melatonin rhythm in constant darkness. In addition, NE does not change the expression of clock genes but the recovery effect of NE depends on the internal clock. Furthermore, the data indicates that NE administration stimulates the gene expression of phosphodiesterase 4D (PDE4D) and adenylate cyclase 1 (AC1) in a time order, potentially corresponding to the trough and peak of recovered melatonin rhythm. Chapter 5 presents data showing that the amplitude of melatonin rhythm in cultured pineal cells is affected by microenvironments of the cell culture and connexin plays a role in this effect. Finally, in Chapter 6 I discuss how the results of each chapter demonstrate multiple regulatory mechanism of the melatonin rhythm of chicken pineal cells. Furthermore, I discuss the implications of this work in the field of developmental biology and how the current data will shape future investigations. My dissertation incorporates engineering, immunocytochemistry, chicken genetics, and biochemical analyses, and will help in better understanding the regulation mechanism of output in a circadian system.

The purpose of these studies was to determine whether the inhibitory effects of short photoperiod exposure on prolactin cell activity in male Syrian hamsters and/or the inhibitory effects of melatonin treatment on the growth and activity of diethylstilbestrol- (DES) induced prolactinomas in Fisher 344 (F344) rats were possibly mediated through alterations in dopaminergic regulatory mechanisms. In both the hamster and the rat, changes in hypothalamic dopamine neuronal activity and changes in pituitary responsiveness to dopamine have been suggested as possible mechanisms in the prolactin-inhibitory effects of light deprivation or melatonin administration. The present studies in the male Syrian hamster addressed two issues. First, it was of interest to determine if anterior pituitaries of long photoperiod-exposed male hamsters possess dopamine receptors, which are presumably necessary for responsiveness to dopamine. This was accomplished by analysis of ³H-spiperone binding to anterior pituitary membranes. Second, possible changes in pituitary sensitivity to dopamine were assessed by comparison of dose response curves for the inhibition by dopamine of prolactin release from hemipituitaries incubated in vitro from both long and short photoperiod-exposed animals over a series of time points from three to fifteen weeks. In the second series of experiments, adult female F344 rats received daily injection of melatonin or saline vehicle. After two weeks, half of the animals were sacrificed for analysis of ³H-spiperone binding to anterior pituitary membranes, measurement of hypothalamic dopamine turnover and analysis of in vitro pituitary sensitivity to dopamine. The remaining animals received subcutaneous implants containing DES and injections were continued on the same schedule until sacrifice four weeks later for measurement of the same parameters. In both the hamster and rat models, treatments exerted profound inhibitory effects on indices of prolactin cell activity. However, these studies provide no evidence for the involvement of altered dopaminergic regulation in the production of such effects. Neither pituitary sensitivity to dopamine in vitro nor hypothalamic dopamine neuronal activity was enhanced by short photoperiod exposure or melatonin treatment. Prolactin-inhibitory effects of these treatments appear to be mediated through as yet unidentified dopamine-independent mechanisms.

It is estimated that 25 % of children suffer from sleeping disorders. This is worrying as sleep is essential for children's health and is required for normal development of both body and brain. When changes in sleep hygiene are not sufficient to improve sleep, there is a need for pharmacological options. One of those is to use the endogenous substance melatonin, which for instance controls the body's sleep/wake phases. In Sweden the use of exogenous melatonin as a hypnotic for children has increased significantly in recent years. The aim of this study was to examine the existing evidence for the use of melatonin in children with sleeping disorders.  In this literature review clinical studies, in which melatonin's efficacy in children with sleeping disorders is evaluated, have been analyzed. The studies were found by searching the data base PubMed.  Earlier sleep onset, shortened sleep onset latency, extended total sleep time and an earlier DLMO (dim light melatonin onset, the time when plasma melatonin levels reaches 10 pg/ml) were endpoints used in the majority of the studies. Most of the results for these sleep-specific parameters show a statistically significant improvement in melatonin treated subjects compared with placebo treatment. This is also confirmed by results from questionnaires. No dose-response relationship was found from analysis of the studies. Overall, the results show that melatonin has a good effect on sleeping disorders and that adverse effects that occur are mild.  The conclusion is that there is strong evidence for the use of melatonin for sleepingdisorders in children and that the increased use of melatonin can be motivated by this.

Alzheimer’s disease is a devastating neurodegenerative disorder and the leading cause of dementia. The disease manifests via several pathologies including neuroinflammation, oxidative stress, metal ion dyshomeostasis, and cell death. To address the multifaceted nature of this disorder, the design of several diverse compounds, targeting many pathological effects, was generated. First, a series of compounds based on curcumin and diosgenin were synthesized following the bivalent design strategy. Two compounds were discovered to have neuroprotective ability, anti-oxidative function, and anti-Aß oligomerization (AßO) properties. A second set of molecules was also designed, wherein a hybrid compound strategy was utilized. Three hybrids were to shown to protect MC65 cells from Aß-induced toxicity and to have significant anti-oxidative activity. Mechanistic studies propose that protection is through disruption of interactions between AßOs and partner proteins. Furthermore, one hybrid was also shown to be able to pass the BBB. Lastly, studies of glyburide, an anti-diabetic medication, have shown an off-target anti-inflammatory effect specific for the NLRP3 inflammasome, which has been implicated in AD development. Therefore, a series of glyburide analogs were synthesized and characterized. One analog was able to successfully inhibit the NLRP3 inflammasome and reduce IL-1ß expression without affecting blood glucose. In vivo studies demonstrated an ability to prevent or ameliorate adverse inflammation-related outcomes in murine inflammatory models. Altogether, these investigations have yielded three novel series of compounds, all capable of modifying Alzheimer’s disease pathology. These results warrant future investigations into the development, optimization, and characterization of these analogs as potential treatments for Alzheimer’s disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder and the leading cause of dementia, affecting ~5.2 million Americans. Current FDA approved medications provide mainly symptomatic relief and there are no agents available to delay or cure this disease. Multiple factors such as amyloid-β aggregates, dyshomeostasis of biometals, oxidative stress, and neuroinflammation have been implicated in the development of AD. Even though significant advances have been made in understanding the mechanisms leading to AD, the exact etiology still remains elusive. Given AD’s multifactorial nature, a multifunctional strategy of small molecule design would help to identify novel chemical templates. Recently our lab has developed hybrid molecules of curcumin and melatonin that exhibited potent neuroprotective ability in various AD models. Further modifications identified a lead compound with potent neuroprotective and antioxidative activity in MC65 cells, while also establishing the hybrid strategy as a viable approach in providing unique chemotypes with novel pharmacology. Further preliminary biological studies suggest that the lead is orally available and exhibits multifunctional properties both in vitro and in vivo on AD pathologies, thus strongly encouraging further structural examination. Herein, we report the structural exploration of this chemical template through structure-activity relationship studies at three domains: the phenyl domain, α,β-unsaturated β-ketone amide domain, and the indole domain. Collectively, the results show that the chemical space around the curcumin portion doesn’t favor electronic or steric/hydrophobic interactions, but might favor pi-pi (π-π) and hydrogen-bond interactions. Additionally, the α,β-unsaturated β-ketone amide domain is not as important as the linearity of the β-ketone acetamide. Moreover, the results indicate that a free rotatable β-OH might be the actual moiety that is important for the observed biological activity through favorable hydrogen bonds. Finally, steric interactions are not favored in the chemical space surrounding the indole nitrogen, suggesting that hydrogen bond interactions are required for the observed neuroprotective activity. Conversely, a hydrogen bond acceptor is necessary at the 5-position of the indole ring and bulky substitutions can be accommodated, with restrictions, suggesting steric tolerance and hydrophobic interactions at this position. These modifications have yielded a series of novel compounds that are capable of modifying AD pathology while shedding further light onto the chemical scaffold thus warranting future investigations into the development, optimization, and characterization of these curcumin/melatonin hybrids as potential treatments for AD.