A defective viral genome strategy elicits broad protective immunity against respiratory viruses.

RNA viruses generate defective viral genomes (DVGs) that can interfere with replication of the parental wild-type virus. To examine their therapeutic potential, we created a DVG by deleting the capsid-coding region of poliovirus. Strikingly, intraperitoneal or intranasal administration of this genome, which we termed eTIP1, elicits an antiviral response, inhibits replication, and protects mice from several RNA viruses, including enteroviruses, influenza, and SARS-CoV-2. While eTIP1 replication following intranasal administration is limited to the nasal cavity, its antiviral action extends non-cell-autonomously to the lungs. eTIP1 broad-spectrum antiviral effects are mediated by both local and distal type I interferon responses. Importantly, while a single eTIP1 dose protects animals from SARS-CoV-2 infection, it also stimulates production of SARS-CoV-2 neutralizing antibodies that afford long-lasting protection from SARS-CoV-2 reinfection. Thus, eTIP1 is a safe and effective broad-spectrum antiviral generating short- and long-term protection against SARS-CoV-2 and other respiratory infections in animal models.

Biomimetic Synthesis and Chemical Proteomics Reveal the Mechanism of Action and Functional Targets of Phloroglucinol Meroterpenoids.

Natural products perennially serve as prolific sources of drug leads and chemical probes, fueling the development of numerous therapeutics. Despite their scarcity, natural products that modulate protein function through covalent interactions with lysine residues hold immense potential to unlock new therapeutic interventions and advance our understanding of the biological processes governed by these modifications. Phloroglucinol meroterpenoids constitute one of the most expansive classes of natural products, displaying a plethora of biological activities. However, their mechanism of action and cellular targets have, until now, remained elusive. In this study, we detail the concise biomimetic synthesis, computational mechanistic insights, physicochemical attributes, kinetic parameters, molecular mechanism of action, and functional cellular targets of several phloroglucinol meroterpenoids. We harness synthetic clickable analogues of natural products to probe their disparate proteome-wide reactivity and subcellular localization through in-gel fluorescence scanning and cell imaging. By implementing sample multiplexing and a redesigned lysine-targeting probe, we streamline a quantitative activity-based protein profiling, enabling the direct mapping of global reactivity and ligandability of proteinaceous lysines in human cells. Leveraging this framework, we identify numerous lysine-meroterpenoid interactions in breast cancer cells at tractable protein sites across diverse structural and functional classes, including those historically deemed undruggable. We validate that phloroglucinol meroterpenoids perturb biochemical functions through stereoselective and site-specific modification of lysines in proteins vital for breast cancer metabolism, including lipid signaling, mitochondrial respiration, and glycolysis. These findings underscore the broad potential of phloroglucinol meroterpenoids for targeting functional lysines in the human proteome.

Strategies for assessing the limit of detection in voltammetric methods: comparison and evaluation of approaches.

The realm of analytical chemistry continues to struggle with defining and evaluating the limit of detection in analytical methods in the sense that a multitude of definitions, criteria, caveats, and methods have been proposed, developed, and adopted across disciplines. The last decade has seen a surge in the growth of electrochemical methods and studies in the field of forensic science and forensic chemistry. While many disciplines within forensic science have established method validation guidelines, the historical and current lack of electrochemical methods within forensic laboratories throughout the United States has left a major gap in knowledge, inhibiting the adoption and utilization of electrochemistry, which may serve as a powerful tool in many subdisciplines of forensics. As such, this work begins this discussion by focusing first on the limit of detection (LOD), with application toward both qualitative and quantitative methods. Both inorganic (ferrocyanide and lead) and organic (diphenylamine, naltrexone, and acetaminophen) target analytes were analyzed via two common voltammetry methods: cyclic voltammetry and square-wave voltammetry. The LOD for each analyte was estimated and/or calculated following a variety of literature-described methods and compared. The accuracy and reliability of these LOD characteristics based on the experimental data is described herein along with suggestions and recommendations. This manuscript is intended to compare the resulting LOD values from various methods and provide a starting point for the incorporation of electrochemistry into the forensic science laboratory, beginning a focused discussion on the development of validation guidelines and parameters needed for the adoption of this technology in forensic laboratories in order to meet the standards required by the criminal justice system.

Plasma-treated gold microelectrodes for subsecond detection of Zn(II) with fast-scan cyclic voltammetry.

The sensitivity of zinc (Zn(II)) detection using fast-scan cyclic voltammetry (FSCV) with carbon fiber microelectrodes (CFMEs) is low compared to other neurochemicals. We have shown previously that Zn(II) plates to the surface of CFME's and we speculate that it is because of the abundance of oxide functionality on the surface. Plating reduces sensitivity over time and causes significant disruption to detection stability. This limited sensitivity and stability hinders Zn(II) detection, especially in complex matrices like the brain. To address this, we developed plasma-treated gold fiber microelectrodes (AuMEs) which enable sensitive and stable Zn(II) detection with FSCV. Typically, gold fibers are treated using corrosive acids to clean the surface and this step is important for preparing the surface for electrochemistry. Likewise, because FSCV is an adsorption-based technique, it is also important for Zn(II) to adsorb and desorb to prevent irreversible plating. Because of these requirements, careful optimization of the electrode surface was necessary to render the surface for Zn(II) adsorption yet strike a balance between attraction to the surface vs. irreversible interactions. In this study, we employed oxygen plasma treatment to activate the gold fiber surface without inducing significant morphological changes. This treatment effectively removes the organic layer while functionalizing the surface with oxygen, enabling Zn(II) detection that is not possible on untreated gold surfaces. Our results demonstrate significantly improved Zn(II) detection sensitivity and stability on AuME compared to CFME's. Overall, this work provides an advance in our understanding of Zn(II) electrochemistry and a new tool for improved metallotransmitter detection in the brain.

Measuring neuron-regulated immune cell physiology via the alpha-2 adrenergic receptor in an ex vivo murine spleen model.

The communication between the nervous and immune systems plays a crucial role in regulating immune cell function and inflammatory responses. Sympathetic neurons, which innervate the spleen, have been implicated in modulating immune cell activity. The neurotransmitter norepinephrine (NE), released by sympathetic neurons, influences immune cell responses by binding to adrenergic receptors on their surface. The alpha-2 adrenergic receptor (α2AR), expressed predominantly on sympathetic neurons, has received attention due to its autoreceptor function and ability to modulate NE release. In this study, we used fast-scan cyclic voltammetry (FSCV) to provide the first subsecond measurements of NE released in the white pulp region of the spleen and validated it with yohimbine, a known antagonist of α2AR. For further application of FSCV in neuroimmunology, we investigated the extent to which subsecond NE from sympathetic neurons is important for immune cell physiology and cytokine production, focusing on tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), and interleukin-6 (IL-6). Our findings provide insights into the regulatory mechanisms underlying sympathetic-immune interactions and show the significance of using FSCV, a traditional neurochemistry technique, to study these neuroimmune mechanisms.

Use of the Vineland-3, a measure of adaptive functioning, in CLN3.

Progressive vision loss and neurocognitive impairment are early and frequent presentations in CLN3 disease. This highlights neurodevelopmental functioning as critical to the disease, but limits the neuropsychological test repertoire. We evaluated the convergent validity of the Vineland Adaptive Behavior Scales as a potential outcome measure. In a prospective observational study of 22 individuals (female:male 11:11; 6-20 years-old) with a molecular diagnosis of CLN3, we used generalized linear models and Spearman correlations to quantify the relationship of the adaptive behavior composite (ABC) standard score with established outcomes of verbal IQ (VIQ) and disease severity (Unified Batten Disease Rating Scale, UBDRS) scores. We analyzed ABC changes in 1-year follow-up data in a subset of the same cohort (n = 17). The ABC and VIQ, both standard scores, exhibited a strong positive correlation in cross-sectional data (r = 0.81). ABC and UBDRS scores were strongly and positively correlated in cross-sectional data (rrange  = 0.87-0.93). Participants' ABC scores decreased slightly over the 1-year follow-up period (mean change, 95% CI: -5.23, -2.16). The convergent validity of the Vineland-3 for use in CLN3 is supported by its relationships with the established outcomes of VIQ and UBDRS. Future longitudinal research, including replication in other cohorts and evaluation of sensitivity to change, will be important to establish utility of the Vineland-3 for monitoring change in CLN3.

Glutamate Electropolymerization on Carbon Increases Analytical Sensitivity to Dopamine and Serotonin: An Auspicious In Vivo Phenomenon in Mice?

Carbon is the material of choice for electroanalysis of biological systems, being particularly applicable to neurotransmitter analysis as carbon fiber microelectrodes (CFMs). CFMs are most often applied to dopamine detection; however, the scope of CFM analysis has rapidly expanded over the last decade with our laboratory's focus being on improving serotonin detection at CFMs, which we achieved in the past via Nafion modification. We began this present work by seeking to optimize this modification to gain increased analytical sensitivity toward serotonin under the assumption that exposure of bare carbon to the in vivo environment rapidly deteriorates analytical performance. However, we were unable to experimentally verify this assumption and found that electrodes that had been exposed to the in vivo environment were more sensitive to evoked and ambient dopamine. We hypothesized that high in vivo concentrations of ambient extracellular glutamate could polymerize with a negative charge onto CFMs and facilitate response to dopamine. We verified this polymerization electrochemically and characterized the mechanisms of deposition with micro- and nano-imaging. Importantly, we identified that the application of 1.3 V as a positive upper waveform limit is a crucial factor for facilitating glutamate polymerization, thus improving analytical performance. Critically, information gained from these dopamine studies were extended to an in vivo environment where a 2-fold increase in sensitivity to evoked serotonin was achieved. Thus, we present here the novel finding that innate aspects of the in vivo environment are auspicious for detection of dopamine and serotonin at carbon fibers, offering a solution to our goal of an improved fast-scan cyclic voltammetry serotonin detection paradigm.

Ultra-High-Resolution Ion Mobility Separations Over Extended Path Lengths and Mobility Ranges Achieved using a Multilevel Structures for Lossless Ion Manipulations Module.

Over the past few years, structures for lossless ion manipulations (SLIM) have used traveling waves (TWs) to move ions over long serpentine paths that can be further lengthened by routing the ions through multiple passages of the same path. Such SLIM "multipass" separations provide unprecedentedly high ion mobility resolving powers but are ultimately limited in their ion mobility range because of the range of mobilities spanned in a single pass; that is, higher mobility ions ultimately "overtake" and "lap" lower mobility ions that have experienced fewer passes, convoluting their arrival time distribution at the detector. To achieve ultrahigh resolution separations over broader mobility ranges, we have developed a new multilevel SLIM possessing multiple stacked serpentine paths. Ions are transferred between SLIM levels through apertures (or ion escalators) in the SLIM surfaces. The initial multilevel SLIM module incorporates four levels and three interlevel ion escalator passages, providing a total path length of 43.2 m. Using the full path length and helium buffer gas, high resolution separations were achieved for Agilent tuning mixture phosphazene ions over a broad mobility range (K0 ≈ 3.0 to 1.2 cm2/(V*s)). High sensitivity was achieved using "in-SLIM" ion accumulation over an extended trapping region of the first SLIM level. High transmission efficiency of ions over a broad mobility range (e.g., K0 ≈ 3.0 to 1.67 cm2/(V*s)) was achieved, with transmission efficiency rolling off for the lower mobility ions (e.g., K0 ≈ 1.2 cm2/(V*s)). Resolving powers of up to ∼560 were achieved using all four ion levels to separate reverse peptides (SDGRG1+ and GRGDS1+). A complex mixture of phosphopeptides showed similar coverage could be achieved using one or all four SLIM levels, and doubly charged phosphosite isomers not significantly separated using one SLIM level were well resolved when four levels were used. The new multilevel SLIM technology thus enables wider mobility range ultrahigh-resolution ion mobility separations and expands on the ability of SLIM to obtain improved separations of complex mixtures with high sensitivity.

Pubertal Development: What's Normal/What's Not.

Onset of puberty, as defined by breast stage 2, appears to be starting at younger ages since the 1940s. There is an ongoing controversy regarding what is normative, as well as what is normal, and the evaluation that is deemed necessary for girls maturing before 8 years of age. There are potential implications of earlier pubertal timing, including psychosocial consequences during adolescence, as well as longer term risks, such as breast cancer and cardiometabolic risks. There are additional consequences derived from slower pubertal tempo, for age of menarche has not decreased as much as age of breast development; these include longer interval between sexual initiation and intentional childbearing, as well as a broadened window of susceptibility to endocrine-related cancers.

Demonstration of electron acceleration in a laser-driven dielectric microstructure.

The enormous size and cost of current state-of-the-art accelerators based on conventional radio-frequency technology has spawned great interest in the development of new acceleration concepts that are more compact and economical. Micro-fabricated dielectric laser accelerators (DLAs) are an attractive approach, because such dielectric microstructures can support accelerating fields one to two orders of magnitude higher than can radio-frequency cavity-based accelerators. DLAs use commercial lasers as a power source, which are smaller and less expensive than the radio-frequency klystrons that power today's accelerators. In addition, DLAs are fabricated via low-cost, lithographic techniques that can be used for mass production. However, despite several DLA structures having been proposed recently, no successful demonstration of acceleration in these structures has so far been shown. Here we report high-gradient (beyond 250 MeV m(-1)) acceleration of electrons in a DLA. Relativistic (60-MeV) electrons are energy-modulated over 563 ± 104 optical periods of a fused silica grating structure, powered by a 800-nm-wavelength mode-locked Ti:sapphire laser. The observed results are in agreement with analytical models and electrodynamic simulations. By comparison, conventional modern linear accelerators operate at gradients of 10-30 MeV m(-1), and the first linear radio-frequency cavity accelerator was ten radio-frequency periods (one metre) long with a gradient of approximately 1.6 MeV m(-1) (ref. 5). Our results set the stage for the development of future multi-staged DLA devices composed of integrated on-chip systems. This would enable compact table-top accelerators on the MeV-GeV (10(6)-10(9) eV) scale for security scanners and medical therapy, university-scale X-ray light sources for biological and materials research, and portable medical imaging devices, and would substantially reduce the size and cost of a future collider on the multi-TeV (10(12) eV) scale.

Preoperative Testing for Urethral Sling Surgery for Stress Urinary Incontinence: Overuse, Underuse and Cost Implications.

PURPOSE: We identify areas of overuse and underuse in the preoperative evaluation of patients undergoing mid urethral sling surgery. We also estimate the effect of overuse of preoperative testing on health care costs. MATERIALS AND METHODS: We conducted a retrospective review of women who underwent sling surgery with or without concomitant prolapse repair between 2012 and 2013. Physician orders for preoperative electrocardiogram, chest x-ray, basic metabolic panel, complete blood count, coagulation studies and urinalysis were classified as appropriate or inappropriate based on summary guidelines from the American Academy of Family Physicians. The additional costs of inappropriate tests were estimated using the 2014 Medicare clinical laboratory and physician fee schedules. RESULTS: A total of 101 women who underwent mid urethral sling surgery were identified and 346 preoperative tests were ordered. Overall 76% of coagulation profiles, 73% of complete blood counts, 47% of basic metabolic panels, 39% of chest x-rays and 21% of electrocardiograms ordered did not have an appropriate clinical indication. In addition, 6% of electrocardiograms, 22% of chest x-rays and 10% of urinalyses were not ordered despite an appropriate indication. The estimated charges of overused tests were $1,844.15 for the cohort, or $18 per patient. CONCLUSIONS: Preoperative testing is overused as well as underused in patients undergoing sling surgery. The greatest variation occurred with the use of electrocardiograms, chest x-rays and urinalysis. Poor adherence to national guidelines leads to increased health care costs and warrants increased awareness in following evidence-based guidelines.

A microfluidic chip for sustained oxygen gradient formation in the intestine ex vivo.

The oxygen gradient across the intestine influences intestinal physiology and the microbial environment of the microbiome. The microbiome releases metabolites that communicate with enterochromaffin cells, neuronal cells, and resident immune cells to facilitate the bidirectional communication across the gut-brain axis. Measuring communication between various cell types within the intestine could provide essential information about key regulators of gut and brain health; however, the microbial environment of the intestine is heavily dependent on the physiological oxygen gradient that exists across the intestinal wall. Likewise, there exist a need for methods which enable real-time monitoring of intestinal signaling ex vivo yet this remains challenging due to the inability to adequately culture intestinal tissue ex vivo while also exposing the appropriate locations of the intestine for probe insertion and monitoring. Here, we designed and fabricated a 3D printed microfluidic device to maintain the oxygen gradient across precision cut murine intestinal slices with the capability to couple to external neurochemical recording techniques. The gradient is maintained from outlets below while allowing access to the slice from above for detection with fast scan cyclic voltammetry (FSCV) and carbon-fiber microelectrodes. A series of 11 outlet ports were designed to lay underneath the slice which were connected to channels to deliver oxygenated vs. deoxygenated media. Outlet ports were designed in an oval shape where deoxygenated media was delivered to the center of the slice and oxygenated media is delivered to the outer portion of the slice to mimic the location of oxygen across the intestine. An oxygen sensitive fluorescent dye, tris(2,2'-bipyridyl)dichlororuthenium(II), was used to characterize the tunability of the gradient. Viability of the tissue was confirmed by both fluorescence microscopy and FSCV. Additionally, we measured simultaneous serotonin and melatonin signaling with FSCV in the intestine for the first time. Overall, this chip provides a significant advance in our ability to culture intestinal slices ex vivo with the added benefit of direct access for measurements and imaging.

Evaluation and classification of fentanyl-related compounds using EC-SERS and machine learning.

Multiple analytical techniques for the screening of fentanyl-related compounds exist. High discriminatory methods such as GC-MS and LC-MS are expensive, time-consuming, and less amenable to onsite analysis. Raman spectroscopy provides a rapid, inexpensive alternative. Raman variants such as electrochemical surface-enhanced Raman scattering (EC-SERS) can provide signal enhancements with 1010 magnitudes, allowing for the detection of low-concentration analytes, otherwise undetected using conventional Raman. Library search algorithms embedded in instruments utilizing SERS may suffer from accuracy when multicomponent mixtures involving fentanyl derivatives are analyzed. The complexing of machine learning techniques to Raman spectra demonstrates an improvement in the discrimination of drugs even when present in multicomponent mixtures of various ratios. Additionally, these algorithms are capable of identifying spectral features difficult to detect by manual comparisons. Therefore, the goal of this study was to evaluate fentanyl-related compounds and other drugs of abuse using EC-SERS and to process the acquired data using machine learning-convolutional neural networks (CNN). The CNN was created using Keras v 2.4.0 with Tensorflow v 2.9.1 backend. In-house binary mixtures and authentic adjudicated case samples were used to evaluate the created machine-learning models. The overall accuracy of the model was 98.4 ± 0.1% after 10-fold cross-validation. The correct identification for the in-house binary mixtures was 92%, while the authentic case samples were 85%. The high accuracies achieved in this study demonstrate the advantage of using machine learning to process spectral data when screening seized drug materials comprised of multiple components.

Dynamic and Rapid Detection of Guanosine during Ischemia.

Guanosine acts in both neuroprotective and neurosignaling pathways in the central nervous system; in this paper, we present the first fast voltammetric measurements of endogenous guanosine release during pre- and post-ischemic conditions. We discuss the metric of our measurements via analysis of event concentration, duration, and interevent time of rapid guanosine release. We observe changes across all three metrics from our normoxic to ischemic conditions. Pharmacological studies were performed to confirm that guanosine release is a calcium-dependent process and that the signaling observed is purinergic. Finally, we show the validity of our ischemic model via staining and fluorescent imaging. Overall, this paper sets the tone for rapid monitoring of guanosine and provides a platform to investigate the extent to which guanosine accumulates at the site of brain injury, i.e., ischemia.

Serotonin is a common thread linking different classes of antidepressants.

Depression pathology remains elusive. The monoamine hypothesis has placed much focus on serotonin, but due to the variable clinical efficacy of monoamine reuptake inhibitors, the community is looking for alternative therapies such as ketamine (neurogenesis theory of antidepressant action). There is evidence that different classes of antidepressants may affect serotonin levels; a notion we test here. We measure hippocampal serotonin in mice with voltammetry and study the effects of acute challenges of escitalopram, fluoxetine, reboxetine, and ketamine. We find that pseudo-equivalent doses of these drugs similarly raise ambient serotonin levels, despite their differing pharmacodynamics because of differences in Uptake 1 and 2, rapid SERT trafficking, and modulation of serotonin by histamine. These antidepressants have different pharmacodynamics but have strikingly similar effects on extracellular serotonin. Our findings suggest that serotonin is a common thread that links clinically effective antidepressants, synergizing different theories of depression (synaptic plasticity, neurogenesis, and the monoamine hypothesis).

Serotonin is a Common Thread Linking Different Classes of Antidepressants.

Depression pathology remains elusive. The monoamine hypothesis has placed much focus on serotonin, but due to the variable clinical efficacy of monoamine reuptake inhibitors, the community is looking for alternative therapies such as ketamine (synaptic plasticity and neurogenesis theory of antidepressant action). There is evidence that different classes of antidepressants may affect serotonin levels; a notion we test here. We measure hippocampal serotonin in mice with voltammetry and study the effects of acute challenges of antidepressants. We find that pseudo-equivalent doses of these drugs similarly raise ambient serotonin levels, despite their differing pharmacodynamics because of differences in Uptake 1 and 2, rapid SERT trafficking and modulation of serotonin by histamine. These antidepressants have different pharmacodynamics but have strikingly similar effects on extracellular serotonin. Our findings suggest that serotonin is a common thread that links clinically effective antidepressants, synergizing different theories of depression (synaptic plasticity, neurogenesis and the monoamine hypothesis).

Evidence of high harmonics from echo-enabled harmonic generation for seeding x-ray free electron lasers.

Echo-enabled harmonic generation free electron lasers hold great promise for the generation of fully coherent radiation in x-ray wavelengths. Here we report the first evidence of high harmonics from the echo-enabled harmonic generation technique in the realistic scenario where the laser energy modulation is comparable to the beam slice energy spread. In this experiment, coherent radiation at the seventh harmonic of the second seed laser is generated when the energy modulation amplitude is about 2-3 times the slice energy spread. The experiment confirms the underlying physics of echo-enabled harmonic generation and may have a strong impact on emerging seeded x-ray free electron lasers that are capable of generating laserlike x rays which will advance many areas of science.

Comparison of portable and benchtop electrochemical instruments for detection of inorganic and organic gunshot residues in authentic shooter samples.

Analysis of gunshot residue currently lacks effective screening methods that can be implemented in real time at the crime scene. Historically, SEM-EDS has been the standard for analysis; however, advances in technology have brought portable instrumentation to the forefront of forensic science disciplines, including the screening of GSR. This study proposes electrochemical methods with disposable screen-printed carbon electrodes for GSR screening at the laboratory and points of care due to their rapid, cost-efficient, and compact platform. GSR residues were extracted from typical aluminum/carbon adhesive collection stubs and analyzed via square-wave anodic stripping voltammetry. Benchtop and portable electrochemical instruments were compared for the assessment and classification of authentic shooter samples by monitoring a panel of inorganic and organic GSR elements and compounds including lead, antimony, copper, 2,4-dinitrotoluene, diphenylamine, nitroglycerin, and ethyl centralite. The evaluation included the assessment of figures of merit and performance measures from quality controls, nonshooter, and shooter data sets. Samples collected from the hands of 200 background individuals (nonshooters), and shooters who fired leaded ammunition (100) and lead-free ammunition (50) were analyzed by the benchtop and portable systems with accuracies of 95.7% and 96.5%, respectively. The findings indicate that electrochemical methods are fast, sensitive, and specific for the identification of inorganic and organic gunshot residues. The portable potentiostat provided results comparable with the benchtop system, serving as a proof-of-concept to transition this methodology to crime scenes for a practical and inexpensive GSR screening that could reduce backlogs, improve investigative leads, and increase the impact of gunshot residues in forensic science.

Low-Frequency Oscillations of In Vivo Ambient Extracellular Brain Serotonin.

Serotonin is an important neurotransmitter that plays a major role in many aspects of neuroscience. Fast-scan cyclic voltammetry measures fast in vivo serotonin dynamics using carbon fiber microelectrodes. More recently, fast-scan controlled-adsorption voltammetry (FSCAV) has been developed to measure slower, minute-to-minute changes in ambient extracellular serotonin. We have previously demonstrated that FSCAV measurements of basal serotonin levels give critical information regarding brain physiology and disease. In this work, we revealed the presence of low-periodicity fluctuations in serotonin levels in mouse hippocampi, measured in vivo with FSCAV. Using correlation analyses, we found robust evidence of oscillations in the basal serotonin levels, which had a period of 10 min and were not present in vitro. Under control conditions, the oscillations did not differ between male and female mice, nor do they differ between mice that underwent a chronic stress paradigm and those in the control group. After the acute administration of a selective serotonin reuptake inhibitor, we observed a shift in the frequency of the oscillations, leading us to hypothesize that the newly observed fluctuations were transporter regulated. Finally, we optimized the experimental parameters of the FSCAV to measure at a higher temporal resolution and found more pronounced shifts in the oscillation frequency, along with a decreased oscillation amplitude. We postulate that this work may serve as a potential bridge for studying serotonin/endocrine interactions that occur on the same time scale.

Novel, User-Friendly Experimental and Analysis Strategies for Fast Voltammetry: Next Generation FSCAV with Artificial Neural Networks.

Fast-scan adsorption-controlled voltammetry (FSCAV) was recently derived from fast-scan cyclic voltammetry to estimate the absolute concentrations of neurotransmitters by using the innate adsorption properties of carbon fiber microelectrodes. This technique has improved our knowledge of serotonin dynamics in vivo. However, the analysis of FSCAV data is laborious and technically challenging. First, each electrode requires post-experimental in vitro calibration. Second, current analysis methods are semi-manual and time-consuming and require a steep learning curve. Finally, the calibration methods used do not adapt to nonlinear electrode responses. In this work, we provide freely accessible computational solutions to these issues. First, we design an artificial neural network (ANN) and train it with a large data set (calibrations from 140 electrodes by six different researchers) to achieve calibration-free estimations and improve predictive error. We discuss the power of the ANN to obtain a low predictive error without electrode-specific calibrations as a function of being able to predict the sensitivity of the electrode. We use the ANN to successfully predict the absolute serotonin concentrations of real in vivo data. Finally, we create a fast and user-friendly, fully automated analysis web platform to simplify and reduce the expertise required for the postanalysis of FSCAV signals.