scholarly journals Fluorescence Activation Mechanism and Imaging of Drug Permeation with New Sensors for Smoking-Cessation Ligands

2021 ◽  
Author(s):  
Aaron L. Nichols ◽  
Zack Blumenfeld ◽  
Chengcheng Fan ◽  
Laura Luebbert ◽  
Annet E. M. Blom ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Hippocampal Neurons ◽  
Decay Rates ◽  
Atomic Scale ◽  
Activation Mechanism ◽  
Fluorescent Reporters ◽  
Partial Agonists ◽  
Pharmacokinetic Properties ◽  
Almost All ◽  
New Sensors

ABSTRACTNicotinic partial agonists provide an accepted aid for smoking cessation and thus contribute to decreasing tobacco-related disease. Improved drugs constitute a continued area of study. However, there remains no reductionist method to examine the cellular and subcellular pharmacokinetic properties of these compounds in living cells. Here, we developed new intensity-based drug sensing fluorescent reporters (“iDrugSnFRs”) for the nicotinic partial agonists dianicline, cytisine, and two cytisine derivatives – 10-fluorocytisine and 9-bromo-10-ethylcytisine. We report the first atomic-scale structures of liganded periplasmic binding protein-based biosensors, accelerating development of iDrugSnFRs and also explaining the activation mechanism. The nicotinic iDrugSnFRs detect their drug partners in solution, as well as at the plasma membrane (PM) and in the endoplasmic reticulum (ER) of cell lines and mouse hippocampal neurons. At the PM, the speed of solution changes limits the growth and decay rates of the fluorescence response in almost all cases. In contrast, we found that rates of membrane crossing differ among these nicotinic drugs by > 30 fold. The new nicotinic iDrugSnFRs, in combination with previously described nicotine and varenicline sensors, provide insight into the real-time pharmacokinetic properties of nicotinic agonists and provide a methodology whereby iDrugSnFRs can inform both pharmaceutical neuroscience and addiction neuroscience.

scholarly journals Fluorescence activation mechanism and imaging of drug permeation with new sensors for smoking-cessation ligands

eLife ◽  
2022 ◽  
Vol 11 ◽  
Author(s):  
Aaron L Nichols ◽  
Zack Blumenfeld ◽  
Chengcheng Fan ◽  
Laura Luebbert ◽  
Annet EM Blom ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Hippocampal Neurons ◽  
Decay Rates ◽  
Atomic Scale ◽  
Activation Mechanism ◽  
Fluorescent Reporters ◽  
Partial Agonists ◽  
Pharmacokinetic Properties ◽  
Almost All ◽  
New Sensors

Nicotinic partial agonists provide an accepted aid for smoking cessation and thus contribute to decreasing tobacco-related disease. Improved drugs constitute a continued area of study. However, there remains no reductionist method to examine the cellular and subcellular pharmacokinetic properties of these compounds in living cells. Here, we developed new intensity-based drug sensing fluorescent reporters ('iDrugSnFRs') for the nicotinic partial agonists dianicline, cytisine, and two cytisine derivatives - 10-fluorocytisine and 9-bromo-10-ethylcytisine. We report the first atomic-scale structures of liganded periplasmic binding protein-based biosensors, accelerating development of iDrugSnFRs and also explaining the activation mechanism. The nicotinic iDrugSnFRs detect their drug partners in solution, as well as at the plasma membrane (PM) and in the endoplasmic reticulum (ER) of cell lines and mouse hippocampal neurons. At the PM, the speed of solution changes limits the growth and decay rates of the fluorescence response in almost all cases. In contrast, we found that rates of membrane crossing differ among these nicotinic drugs by > 30 fold. The new nicotinic iDrugSnFRs provide insight into the real-time pharmacokinetic properties of nicotinic agonists and provide a methodology whereby iDrugSnFRs can inform both pharmaceutical neuroscience and addiction neuroscience.

scholarly journals A role for BDNF- and NMDAR-induced lysosomal recruitment of mTORC1 in the regulation of neuronal mTORC1 activity

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Dany Khamsing ◽  
Solène Lebrun ◽  
Isabelle Fanget ◽  
Nathanaël Larochette ◽  
Christophe Tourain ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nmda Receptors ◽  
Hippocampal Neurons ◽  
De Novo ◽  
Long Term Potentiation ◽  
Activation Mechanism ◽  
Functional Link ◽  
Term Potentiation ◽  
Endocytic Compartments ◽  
Mtorc1 Activation

AbstractMemory and long term potentiation require de novo protein synthesis. A key regulator of this process is mTORC1, a complex comprising the mTOR kinase. Growth factors activate mTORC1 via a pathway involving PI3-kinase, Akt, the TSC complex and the GTPase Rheb. In non-neuronal cells, translocation of mTORC1 to late endocytic compartments (LEs), where Rheb is enriched, is triggered by amino acids. However, the regulation of mTORC1 in neurons remains unclear. In mouse hippocampal neurons, we observed that BDNF and treatments activating NMDA receptors trigger a robust increase in mTORC1 activity. NMDA receptors activation induced a significant recruitment of mTOR onto lysosomes even in the absence of external amino acids, whereas mTORC1 was evenly distributed in neurons under resting conditions. NMDA receptor-induced mTOR translocation to LEs was partly dependent on the BDNF receptor TrkB, suggesting that BDNF contributes to the effect of NMDA receptors on mTORC1 translocation. In addition, the combination of Rheb overexpression and artificial mTORC1 targeting to LEs by means of a modified component of mTORC1 fused with a LE-targeting motif strongly activated mTOR. To gain spatial and temporal control over mTOR localization, we designed an optogenetic module based on light-sensitive dimerizers able to recruit mTOR on LEs. In cells expressing this optogenetic tool, mTOR was translocated to LEs upon photoactivation. In the absence of growth factor, this was not sufficient to activate mTORC1. In contrast, mTORC1 was potently activated by a combination of BDNF and photoactivation. The data demonstrate that two important triggers of synaptic plasticity, BDNF and NMDA receptors, synergistically power the two arms of the mTORC1 activation mechanism, i.e., mTORC1 translocation to LEs and Rheb activation. Moreover, they unmask a functional link between NMDA receptors and mTORC1 that could underlie the changes in the synaptic proteome associated with long-lasting changes in synaptic strength.

scholarly journals Concurrent Daily and Non-Daily Use of Heated Tobacco Products with Combustible Cigarettes: Findings from the 2018 ITC Japan Survey

2020 ◽  
Vol 17 (6) ◽  
pp. 2098 ◽  
Author(s):  
Edward Sutanto ◽  
Connor Miller ◽  
Danielle M. Smith ◽  
Ron Borland ◽  
Andrew Hyland ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Tobacco Use ◽  
Lower Number ◽  
Web Survey ◽  
Tobacco Products ◽  
Concurrent Use ◽  
Frequency Of Use ◽  
Concurrent User ◽  
Nationally Representative ◽  
Almost All

Use of heated tobacco products (HTPs) among current smokers is becoming increasingly popular in Japan. This study aims to compare characteristics and tobacco-related behaviors among concurrent users of HTPs and combustible cigarettes (n = 644) with exclusive smokers (n = 3194) or exclusive HTP users (n = 164). The secondary aim was to explore heterogeneity within concurrent use subgroups. Data were from Wave 1 of the ITC Japan Survey, a nationally representative web survey conducted from February to March 2018. Concurrent cigarette-HTP users were younger and wealthier than exclusive smokers. However, there were no difference in the frequency of smoking, number of cigarettes per day, and smoking cessation behaviors between the two groups, suggesting that HTPs reinforce nicotine dependence. Compared to exclusive HTP users, concurrent cigarette-HTP users reported higher frequency of non-daily HTP use, and lower number of tobacco-containing inserts per day. Almost all concurrent cigarette-HTP users smoked every day (93.9%); 48.4% both smoked and used HTPs daily (dual daily users, n = 396), while 45.5% were daily smokers and non-daily HTP users (predominant smokers, n = 213). Concurrent user subgroups differed from each other on age, tobacco use behaviors, and quit intention. Alongside heterogeneity between concurrent and exclusive product users, differences across concurrent use subgroups highlight the importance of considering frequency of use in characterizing poly-tobacco users.

The opioid antagonist naltrexone inhibits activity and alters expression of α7 and α4β2 nicotinic receptors in hippocampal neurons: implications for smoking cessation programs

2000 ◽  
Vol 39 (13) ◽  
pp. 2740-2755 ◽  
Author(s):  
Luis E.F Almeida ◽  
Edna F.R Pereira ◽  
Manickavasagom Alkondon ◽  
William P Fawcett ◽  
William R. Randall ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Hippocampal Neurons ◽  
Nicotinic Receptors ◽  
Opioid Antagonist

Rationale, pharmacology and clinical efficacy of partial agonists of α4β2 nACh receptors for smoking cessation

2007 ◽  
Vol 28 (7) ◽  
pp. 316-325 ◽  
Author(s):  
Hans Rollema ◽  
Jotham W. Coe ◽  
Leslie K. Chambers ◽  
Raymond S. Hurst ◽  
Stephen M. Stahl ◽  
...  
Keyword(s):  
Smoking Cessation ◽  
Clinical Efficacy ◽  
Partial Agonists

scholarly journals Flutamide and Cyproterone Acetate Exert Agonist Effects: Induction of Androgen Receptor-Dependent Neuroprotection

Endocrinology ◽  
2007 ◽  
Vol 148 (6) ◽  
pp. 2936-2943 ◽  
Author(s):  
Thuy-Vi V. Nguyen ◽  
Mingzhong Yao ◽  
Christian J. Pike
Keyword(s):  
Gene Expression ◽  
Nervous System ◽  
Hippocampal Neurons ◽  
Cyproterone Acetate ◽  
Androgen Receptors ◽  
Partial Agonists ◽  
Protective Actions ◽  
And Function ◽  
Insight Into ◽  
Cultured Hippocampal Neurons

Androgens can exert profound effects on the organization, development, and function of the nervous system through activation of androgen receptors (ARs). Nonsteroidal and steroidal antiandrogens antagonize AR-mediated, classic genomic actions of androgens. However, emerging studies in nonneuronal cells indicate that antiandrogens can act as partial agonists for the AR. Here we investigated the effects of the antiandrogens flutamide and cyproterone acetate on neuroprotection induced by dihydrotestosterone (DHT). We observed that, although flutamide and cyproterone acetate blocked androgen-induced gene expression, they failed to inhibit DHT protection against apoptotic insults in cultured hippocampal neurons. Interestingly, flutamide and cyproterone acetate alone, like DHT, significantly reduced apoptosis. Furthermore, the protective actions of flutamide and cyproterone acetate were observed specifically in AR-expressing cell lines, suggesting a role for AR in the agonist effects of antiandrogens. Our results indicate that, in contrast to the classic antiandrogen properties of flutamide and cyproterone acetate, these AR modulators display agonist activities at the level of neuroprotection. These findings provide new insight into the agonist vs. antagonist properties of antiandrogens, information that will be crucial to understanding the neural implications of clinically used AR-modulating drugs.

scholarly journals Ekspresi Caspase-3 pada Sel Epitel Rongga Mulut (Kb Cell Line) setelah Paparan Ekstrak Kopi

2014 ◽  
Vol 21 (2) ◽  
pp. 122
Author(s):  
Suryani Hutomo ◽  
Yanti Ivana Suryanto ◽  
Heni Susilowati ◽  
Agustinus Rudolf Phym ◽  
Devi Chretella Maheswara
Keyword(s):  
Cell Death ◽  
Cell Line ◽  
Caspase 3 ◽  
Positive Control ◽  
Activation Mechanism ◽  
Kb Cells ◽  
Direct Exposure ◽  
Almost All ◽  
Oral Epithelial ◽  
Active Caspase

Kopi adalah minuman yang biasa dikonsumsi oleh masyarakat sehari-hari. Telah diketahui bahwa kopi mengandung kafein seperti yang terdapat juga pada teh dan coklat. Kandungan terbanyak kafein terdapat pada kopi. Kafein mempunyai struktur kimia 1, 3, 7- trimethylxanthine dan merupakan derivat xanthine. Senyawa ini dapat menginduksi kematian sel yang mengarah pada apoptosis, namun mekanisme yang terlibat belum diketahui dengan jelas. Tingginyakonsumsi kopi di dunia yang selalu meningkat mengindikasikan perlunya dilakukan penelitian untuk mengetahui efek kafein pada epitel rongga mulut yang berkontak langsung dengan kafein. Penelitian terdahulu melaporkan bahwaekstrak kopi menyebabkan kerusakan sel yang sebagian besar mengarah pada apoptosis, tetapi mekanismenya belum jelas. Tujuan penelitian ini adalah untuk menganalisis mekanisme kematian sel KB yang diinduksi oleh kafein melaluiaktivasi caspase-3. Sel KB sebagai model epitel oral (5x10⁴ sel) dikultur dalam DMEM menggunakan 24 wells microplate selama 24 jam sebelum perlakuan. Sel selanjutnya dipapar dengan kafein dengan konsentrasi 100 μg/ml, 200 μg/ml, 400 μg/ml dan diinkubasi selama 24 dan 48 jam dalam DMEM. Doxorubicin (0,5625 μg/ml) digunakan sebagai kontrol positif induksi apoptosis. Teknik imunositokimia terhadap caspase-3 dilakukan pada sel setelah dipapar kafeinuntuk mengamati adanya ekspresi caspase-3 sebagai ciri apoptosis. Identifikasi caspase-3 dilakukan menggunakan mikroskop fase kontras. Ekspresi protein caspase-3 terdeteksi pada sitoplasma sel KB. Hasil penelitian ini menunjukkanadanya ekspresi caspase-3 aktif yang ditandai dengan warna cokelat dengan intensitas kuat pada sitoplasma sebagian besar sel setelah dipapar kafein dengan konsentrasi 100 μg/ml dan 200 μg/ml selama 24 jam. Disimpulkan bahwa ekstrak kopi menyebabkan apoptosis sel KB melalui jalur aktivasi caspase-3. ABSTRACT: The Expression of Caspase-3 in Oral Cavity (Kb Cell Line) after Exposure to Coffee Extract. People widely consume coffee in daily meals. It is known there is caffeine found in coffee like it is found in tea and chocolate.Caffeine is found in the greatest amount of coffee. This 1, 3, 7- trimethyl xanthine substance is a derivate of xanthine that is consumed by almost all people in the world. This substance could induce cell death that mainly is apoptosis, but how the mechanism has not been clearly understood. Considering that coffee is widely consumed in the whole world, it is necessary to conduct an experiment to find any possible effect of caffeine to oral epitel that make direct exposure to caffeine. This experiment is targeted to analyze the mechanism of cell death which caused by caffeine through activation of caspase-3. KB cells as oral epithelial model (5x10⁴ sel) were cultured in DMEM using 24 well microplate for 24 hours before treatment. Then caffeine was given with concentration of 100 μg/ml, 200 μg/ml and 400 μg/ml. Cells were then incubated for 24 and 48 hours period in DMEM. Doxorubicin (0,5625 μg/ml) was used as a positive control of apoptosis induction. Immunocytochemistry technique was then done to observe any caspase three expression as amarker for apoptosis. Identification of active caspase-3 was then done using contrast phase microscope. The results showed expression of caspase-3 in KB cells cytoplasm which observed as high intensity of brown colored molecules incell cytoplasm after 100 μg/ml and 200 μg/ml caffeine exposure in 24 hours. It was concluded that coffee extract induce KB cells apoptosis through caspase-3 activation mechanism.

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