[ { "id": "authors:4c1m4-76298", "collection": "authors", "collection_id": "4c1m4-76298", "cite_using_url": "https://authors.library.caltech.edu/records/4c1m4-76298", "type": "monograph", "title": "Hydrogel encapsulation of a designed fluorescent protein biosensor for continuous measurements of sub-100 nanomolar nicotine", "author": [ { "family_name": "Nichols", "given_name": "Aaron L.", "orcid": "0000-0001-9341-0049", "clpid": "Nichols-Aaron-L" }, { "family_name": "Marotta", "given_name": "Christopher B.", "orcid": "0000-0002-3110-0819", "clpid": "Marotta-Christopher-Bruno" }, { "family_name": "Wagenaar", "given_name": "Daniel A.", "orcid": "0000-0002-6222-761X", "clpid": "Wagenaar-D-A" }, { "family_name": "Mayo", "given_name": "Stephen L.", "orcid": "0000-0002-9785-5018", "clpid": "Mayo-S-L" }, { "family_name": "Dougherty", "given_name": "Dennis A.", "orcid": "0000-0003-1464-2461", "clpid": "Dougherty-D-A" }, { "family_name": "Lester", "given_name": "Henry A.", "orcid": "0000-0002-5470-5255", "clpid": "Lester-H-A" } ], "abstract": "

The reinforcing and addictive properties of nicotine result from concentration- and time-dependent activation, desensitization, and upregulation of nicotinic acetylcholine receptors. However, time-resolved [nicotine] measurement in people who consume nicotine is challenging, as current approaches are expensive, invasive, tedious, and discontinuous. To address the challenge of continuous nicotine monitoring in human biofluids, we report the encapsulation of a purified, previously developed fluorescent biosensor protein, iNicSnFR12, into acrylamide hydrogels and polyethylene glycol diacrylate (PEGDA) hydrogels. We optimized the hydrogels for optical clarity and straightforward slicing. With fluorescence photometry of the hydrogels in a microscope and an integrated miniscope, [nicotine] is detected within a few min at the smoking- and vaping-relevant level of 10 - 100 nM (1.62 – 16.2 ng/ml), even in a 250 µm thick hydrogel at the end of 400 µm dia multimode fiber optic. Concentration-response relations are consistent with previous measurements on isolated iNicSnFR12. Leaching of iN-icSnFR12 from the hydrogel and inactivation of iNicSnFR12 are minimal for several days, and nicotine can be detected for at least 10 months after casting. This work provides the molecular, photophysical, and mechanical bases for personal, wearable continuous [nicotine] monitoring, with straightforward extensions to existing, homologous “iDrugSnFR” proteins for other abused and prescribed drugs.

", "doi": "10.1101/2024.12.02.625538", "publisher": "bioRxiv", "publication_date": "2024-12-02" }, { "id": "authors:97swd-q7f69", "collection": "authors", "collection_id": "97swd-q7f69", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20230316-182493000.35", "type": "monograph", "title": "Interactive computational and experimental approaches improve the sensitivity of periplasmic binding protein-based nicotine biosensors for measurements in biofluids", "author": [ { "family_name": "Haloi", "given_name": "Nandan", "orcid": "0000-0003-3542-333X", "clpid": "Haloi-Nandan" }, { "family_name": "Huang", "given_name": "Shan", "orcid": "0000-0002-4436-3327", "clpid": "Huang-Shan" }, { "family_name": "Nichols", "given_name": "Aaron N.", "orcid": "0000-0001-9341-0049", "clpid": "Nichols-Aaron-N" }, { "family_name": "Fine", "given_name": "Eve J.", "orcid": "0000-0001-7404-897X", "clpid": "Fine-Eve-J" }, { "family_name": "Marotta", "given_name": "Christopher B.", "clpid": "Marotta-Christopher-B" }, { "family_name": "Dougherty", "given_name": "Dennis A.", "orcid": "0000-0003-1464-2461", "clpid": "Dougherty-D-A" }, { "family_name": "Lindahl", "given_name": "Erik", "orcid": "0000-0002-2734-2794", "clpid": "Lindahl-Erik" }, { "family_name": "Howard", "given_name": "Rebecca J.", "orcid": "0000-0003-2049-3378", "clpid": "Howard-Rebecca-J" }, { "family_name": "Mayo", "given_name": "Stephen L.", "orcid": "0000-0002-9785-5018", "clpid": "Mayo-S-L" }, { "family_name": "Lester", "given_name": "Henry A.", "orcid": "0000-0002-5470-5255", "clpid": "Lester-H-A" } ], "abstract": "To develop more sensitive fluorescent protein sensors for nicotine, we combined computational protein design, site-saturated, site-directed, and combinatorial mutagenesis with fluorescence assays, molecular dynamics simulations, and absorbance measurements. The data showed that the resulting molecules, iNicSnFR11 and iNicSnFR12, have higher sensitivity to nicotine than previously reported constructs. In the linear portion of the dose-response relation at sub-\u03bcM [nicotine] for iNicSnFR12, \u2206F/F\u2080 increased with a proportionality constant (S-slope) of 2.6 \u03bcM\u207b\u00b9, representing a 6.5-fold higher sensitivity than iNicSnFR3a. Molecular dynamics calculations enabled identification of a binding pose for nicotine previously indeterminate from experimental data. Further comparative simulations based on this model revealed a tilt in helix 4 in the optimized sensor, likely altering allosteric networks involving the ligand binding site. The absorbance data showed that the fluorescence activation results from increased absorption rather than increased quantum yield for fluorescence. iNicSnFR12 resolved nicotine in diluted mouse and human serum at the peak concentration (100-200 nM) that occurs during smoking or vaping, but also at the decaying concentrations (< 100 nM) during the intervals between smoking or vaping sessions. NicSnFR12 was roughly as sensitive to varenicline or acetylcholine as to nicotine; the sensitivity to choline was at least one order of magnitude less. None of these drugs would markedly distort measurements in human biofluids such as sweat and interstitial fluid. Therefore, iNicSnFR12 is a promising candidate as the molecular sensor that could underlie a continuous nicotine monitor for human biofluids.", "doi": "10.1101/2023.01.16.524298", "publication_date": "2023-01-19" }, { "id": "authors:zcnhw-fxm65", "collection": "authors", "collection_id": "zcnhw-fxm65", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20220812-829310000", "type": "monograph", "title": "Selective Serotonin Reuptake Inhibitors Within Cells: Temporal Resolution in Cytoplasm, Endoplasmic Reticulum, and Membrane", "author": [ { "family_name": "Nichols", "given_name": "Aaron L.", "orcid": "0000-0001-9341-0049", "clpid": "Nichols-Aaron-L" }, { "family_name": "Blumenfeld", "given_name": "Zack", "orcid": "0000-0002-4627-5582", "clpid": "Blumenfeld-Zachary" }, { "family_name": "Luebbert", "given_name": "Laura", "orcid": "0000-0003-1379-2927", "clpid": "Luebbert-Laura" }, { "family_name": "Knox", "given_name": "Hailey J.", "orcid": "0000-0003-0608-2855", "clpid": "Knox-Hailey-J" }, { "family_name": "Muthusamy", "given_name": "Anand K.", "orcid": "0000-0003-1041-914X", "clpid": "Muthusamy-Anand-K" }, { "family_name": "Marvin", "given_name": "Jonathan S.", "orcid": "0000-0003-2294-4515", "clpid": "Marvin-Jonathan-S" }, { "family_name": "Kim", "given_name": "Charlene H.", "orcid": "0000-0003-4048-5244", "clpid": "Kim-Charlene-M" }, { "family_name": "Grant", "given_name": "Stephen N.", "orcid": "0000-0003-0923-8886", "clpid": "Grant-Stephen-N" }, { "family_name": "Walton", "given_name": "David P.", "orcid": "0000-0002-9557-6461", "clpid": "Walton-David-P" }, { "family_name": "Cohen", "given_name": "Bruce N.", "orcid": "0000-0003-2913-6238", "clpid": "Cohen-Bruce-N" }, { "family_name": "Hammar", "given_name": "Rebekkah", "orcid": "0000-0001-5740-0353", "clpid": "Hammar-Rebekkah" }, { "family_name": "Looger", "given_name": "Loren L.", "orcid": "0000-0002-7531-1757", "clpid": "Looger-Loren-L" }, { "family_name": "Artursson", "given_name": "Per", "orcid": "0000-0002-3708-7395", "clpid": "Artursson-Per" }, { "family_name": "Dougherty", "given_name": "Dennis A.", "orcid": "0000-0003-1464-2461", "clpid": "Dougherty-D-A" }, { "family_name": "Lester", "given_name": "Henry A.", "orcid": "0000-0002-5470-5255", "clpid": "Lester-H-A" } ], "abstract": "Selective serotonin reuptake inhibitors (SSRIs) are the most prescribed treatment for individuals experiencing major depressive disorder (MDD). The therapeutic mechanisms that take place before, during, or after SSRIs bind the serotonin transporter (SERT) are poorly understood, partially because no studies exist of the cellular and subcellular pharmacokinetic properties of SSRIs in living cells. We studied escitalopram and fluoxetine using new intensity-based drug-sensing fluorescent reporters (\u2033iDrugSnFRs\u2033) targeted to the plasma membrane (PM), cytoplasm, or endoplasmic reticulum (ER) of cultured neurons and mammalian cell lines. We also employed chemical detection of drug within cells and phospholipid membranes. The drugs attain equilibrium in neuronal cytoplasm and ER, at approximately the same concentration as the externally applied solution, with time constants of a few s (escitalopram) or 200-300 s (fluoxetine). Simultaneously, the drugs accumulate within lipid membranes by \u2265 18-fold (escitalopram) or 180-fold (fluoxetine), and possibly by much larger factors. Both drugs leave cytoplasm, lumen, and membranes just as quickly during washout. We synthesized membrane-impermeant quaternary amine derivatives of the two SSRIs. The quaternary derivatives are substantially excluded from membrane, cytoplasm, and ER for > 2.4 h. They inhibit SERT transport-associated currents 6- or 11-fold less potently than the SSRIs (escitalopram or fluoxetine derivative, respectively), providing useful probes for distinguishing compartmentalized SSRI effects. Although our measurements are orders of magnitude faster than the \u2033therapeutic lag\u2033 of SSRIs, these data suggest that SSRI-SERT interactions within organelles or membranes may play roles during either the therapeutic effects or the \u2033antidepressant discontinuation syndrome\u2033.", "doi": "10.1101/2022.08.09.502705", "publication_date": "2022-08-12" }, { "id": "authors:vxg86-ydj89", "collection": "authors", "collection_id": "vxg86-ydj89", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200526-135356434", "type": "monograph", "title": "Successful Cessation Programs that Reduce Comorbidity May Explain Surprisingly Low Smoking Rates Among Hospitalized COVID-19 Patients", "author": [ { "family_name": "Cohen", "given_name": "Bruce N.", "clpid": "Cohen-Bruce-N" }, { "family_name": "Nichols", "given_name": "Aaron L.", "orcid": "0000-0001-9341-0049", "clpid": "Nichols-Aaron-L" }, { "family_name": "Grant", "given_name": "Stephen", "orcid": "0000-0003-0923-8886", "clpid": "Grant-Stephen" }, { "family_name": "Blumenfeld", "given_name": "Zach", "orcid": "0000-0002-4627-5582", "clpid": "Blumenfeld-Zachary" }, { "family_name": "Dougherty", "given_name": "Dennis A.", "orcid": "0000-0003-1464-2461", "clpid": "Dougherty-D-A" }, { "family_name": "Alvarez", "given_name": "R. Michael", "orcid": "0000-0002-8113-4451", "clpid": "Alvarez-R-M" }, { "family_name": "Ritz", "given_name": "Beate", "orcid": "0000-0001-6976-7339", "clpid": "Ritz-Beate" }, { "family_name": "Lester", "given_name": "Henry A.", "orcid": "0000-0002-5470-5255", "clpid": "Lester-H-A" } ], "abstract": "A recent, non-peer-reviewed meta-analysis suggests that smoking may reduce the risk of hospitalization with COVID-19 because the prevalence of smoking among hospitalized COVID-19 is less than that of the general population. However, there are alternative explanations for this phenomena based on (1) the failure to report, or accurately record, smoking history during emergency hospital admissions and (2) a pre-disposition to avoid smoking among COVID-19 patients with tobacco-related comorbidities (a type of \"reverse\" causation). For example, urine testing of hospitalized patients in Australia for cotinine showed that smokers were under-counted by 37% because incoming patients failed to inform staff about their smoking behavior. Face-to-face interviews can introduce bias into the responses to attitudinal and behavioral questions not present in the self-completion interviews typically used to measure smoking prevalence in the general population. Subjects in face-to-face interviews may be unwilling to admit socially undesirable behavior and attitudes under direct questioning. Reverse causation may also contribute to the difference between smoking prevalence in the COVID-19 and general population. Patients hospitalized with COVID-19 may be simply less prone to use tobacco than the general population. A potentially robust \"reverse causation\" hypothesis for reduced prevalence of smokers in the COVID-19 population is the enrichment of patients in that population with serious comorbidities that motivates them to quit smoking. We judge that this \"smoking cessation\" mechanism may account for a significant fraction of the reduced prevalence of smokers in the COVID-19 population. Testing this hypothesis will require a focused research program.", "doi": "10.32388/WURFH0", "publication_date": "2020-05-26" }, { "id": "authors:vezvm-wdv47", "collection": "authors", "collection_id": "vezvm-wdv47", "cite_using_url": "https://resolver.caltech.edu/CaltechAUTHORS:20200210-132609996", "type": "monograph", "title": "A fast genetically encoded fluorescent sensor for faithful in vivo acetylcholine detection in mice, fish, worms and flies", "author": [ { "family_name": "Borden", "given_name": "Philip M.", "clpid": "Borden-P-M" }, { "family_name": "Shivange", "given_name": "Amol V.", "clpid": "Shivange-A-V" }, { "family_name": "Lester", "given_name": "Henry A.", "orcid": "0000-0002-5470-5255", "clpid": "Lester-H-A" }, { "family_name": "Fan", "given_name": "Chengcheng", "orcid": "0000-0003-4213-5758", "clpid": "Fan-Chengcheng" }, { "family_name": "Rees", "given_name": "Douglas C.", "orcid": "0000-0003-4073-1185", "clpid": "Rees-D-C" } ], "abstract": "Here we design and optimize a genetically encoded fluorescent indicator, iAChSnFR, for the ubiquitous neurotransmitter acetylcholine, based on a bacterial periplasmic binding protein. iAChSnFR shows large fluorescence changes, rapid rise and decay kinetics, and insensitivity to most cholinergic drugs. iAChSnFR revealed large transients in a variety of slice and in vivo preparations in mouse, fish, fly and worm. iAChSnFR will be useful for the study of acetylcholine in all organisms.", "doi": "10.1101/2020.02.07.939504", "publication_date": "2020-02-08" } ]