Stephen G. Weber

Information about the rates of hydrolysis of neuropeptidesby extracellularpeptidases can lead to a quantitative understanding of how the steady-stateand transient concentrations of neuropeptides are controlled. We havecreated a small microfluidic device that electroosmotically infusespeptides into, through, and out of the tissue to a microdialysis probeoutside the head. The device is created by two-photon polymerization(Nanoscribe). Inferring quantitative estimates of a rate process fromthe change in concentration of a substrate that has passed through tissue is challenging for two reasons. One is that diffusion is significant,so there is a distribution of peptide substrate residence times inthe tissue. This affects the product yield. The other is that thereare multiple paths taken by the substrate as it passes through tissue,so there is a distribution of residence times and thus reaction times.Simulation of the process is essential. The simulations presentedhere imply that a range of first order rate constants of more than3 orders of magnitude is measurable and that 5-10 min is requiredto reach a steady state value of product concentration following initiationof substrate infusion. Experiments using a peptidase-resistant d-amino acid pentapeptide, yaGfl, agree with simulations.

Due to significant advances in instrumentation, many previously specialized techniques have become “routine” in user facilities. However, detailed knowledge held by experts has often not been relayed to general users, so they often rely on entry-level information, basic principles, and comparison with literature results for data analysis. As a result, major errors in the data analysis of multiple surface and material analysis techniques, including in x-ray photoelectron spectroscopy (XPS), have been appearing in the scientific literature. Representative examples of serious errors in XPS data analysis are shown in this work. We suggest that surface and material analysis, and perhaps even science in general, are in a state of “pre-crisis.” We use two (logistic) models from population biology to suggest that bad analyses self-correct if they remain below a critical number. However, beyond a threshold, the literature can become useless because of the perpetuation of faulty analyses and concomitant loss of its self-correcting ability. XPS is used by scientists in many communities because of the power of the technique and high-quality instrumentation that is commercially available. Those who make new surfaces and materials face unique challenges because of the large number of surface and material analytical techniques that are often needed to characterize their materials. Graduate students and post-docs are often provided with only minimal instruction on using surface and material characterization methods. High fees for instruments may affect both the quality and the quantity of the data people collect. The Prisoner's Dilemma is a model from game theory that describes situations with reward structures that encourage uncooperative behavior and lead to suboptimal outcomes. However, the outcomes of Prisoner's Dilemma are not inevitable—their consequences change if their reward structures change. The current system does not appear to incentivize detailed learning of surface and material characterization techniques and careful material characterization. Prisoner's dilemmas appear to lead to other undesirable consequences in science. The concerns raised in this work suggest that many manuscripts are incompletely reviewed at present. The different stakeholders in this problem, including authors, research advisers, subject matter experts, reviewers, scientists who notice examples of faulty data analysis, editors, journals and publishers, funding agencies, scientific societies, leaders at universities and research centers, and instrument vendors, can improve the current situation. This work provides specific recommendations for each of these stakeholders. For example, we believe that authors are primarily responsible for the correctness of their work, not reviewers or editors; we question the wisdom of listing the names of the editor and reviewers on a paper; we are grateful for the significant contributions that have been made by subject matter experts to produce standards and tutorial information; the high cost of instrument time at some institutions may limit student access and result in suboptimal analyses; staff scientists often need to be better recognized for their intellectual contributions to studies; publishers may wish to allow selective reviewing of specific sections of papers related to material characterization; the reviewing at some open access journals may be inadequate; while it had its shortcomings, the pre-open access model of publishing incentivized the production and publication of high-quality work; audits of the products (scientific papers) of funding agencies may be necessary; collaboration needs to be encouraged to a greater extent at some institutions; and instrument vendors should not suggest to potential customers that surface characterization, e.g., by XPS, is trivial or simple.

In the increasingly complex health care system, physicians require skills and knowledge to participate with multidisciplinary team members in quality improvement (QI) that adds value to health care organizations. The Educational and Clinical Leaders Improving Performance with Structured E3L training (ECLIPSE) program was developed to address this challenge. Clinically relevant components of lean management were leveraged to create an online, flipped-classroom curriculum, and this was paired with Kaizen adapted specifically for physicians and multidisciplinary clinicians to promote experiential skills utilization. The focus of each adapted Kaizen was a topic of institutional QI priority, such as improving patient throughput or reducing readmission rates. Participants were awarded certification in the E3 Leadership management system—a patient-centered, equity-focused system based on lean principles. After 4 years, 50 E3 Leadership certificates were awarded to multidisciplinary clinicians, including 30 to physicians; participants scored an average 85% on module quizzes. The ECLIPSE program has improved physician participation in multidisciplinary QI projects with institutional alignment.

There has not been a comprehensive investigation of the effect of voltammetric parameters: amplitude, dc bias potential, and excitation frequency of the sine waveform in Fourier Transform Sine‐Wave Voltammtery (FT‐SWV) on background, signal, and signal‐to‐background ratio as well as linearity with respect to the concentration of an analyte. FT‐SWV of ruthenium hexamine (III) chloride (RuHex) demonstrates that the dc bias potential can be used to adjust the phase angle difference between the signal and background to obtain out‐of‐phase background. Several conditions show a high signal‐to‐noise ratio and good concentration linearity. The lessons from RuHex guided us to conditions for good signal‐to‐background ratio and concentration linearity for dopamine (DA).

Traumatic brain injury (TBI) induces a pathophysiologic state that can be worsened by secondary injury. Monitoring brain metabolism with intracranial microdialysis can provide clinical insights to limit secondary injury in the days following TBI. Recent enhancements to microdialysis include the implementation of continuously operating electrochemical biosensors for monitoring the dialysate sample stream in real time and dexamethasone retrodialysis to mitigate the tissue response to probe insertion. Dexamethasone-enhanced continuous-online microdialysis (Dex-enhanced coMD) records long-lasting declines of glucose after controlled cortical impact in rats and TBI in patients. The present study employed retrodialysis and fluorescence microscopy to investigate the mechanism responsible for the decline of dialysate glucose after injury of the rat cortex. Findings confirm the long-term functionality of Dex-enhanced coMD for monitoring brain glucose after injury, demonstrate that intracranial glucose microdialysis is coupled to glucose utilization in the tissues surrounding the probes, and validate the conclusion that aberrant glucose utilization drives the postinjury glucose decline.

There are many processes that actively alter the concentrations of solutes in the extracellular space. Enzymatic reactions, either by soluble enzymes or membrane-bound ectoenzymes, and uptake or clearance are two such processes. Investigations of ectoenzymatic reactions in vivo is challenging, particularly in the brain. Studies using microdialysis have revealed some qualitative information about what enzymes may be present, but microdialysis is a sampling technique so it is not designed to control conditions such as a substrate concentration outside the probe. Micropush–pull perfusion has been used to determine which nitric oxide synthase enzymes are active in discrete regions of the rat retina. Ectopeptidases are a particularly important class of ectoenzymes. As far as it is known, the extracellular activity of active peptides in the brain is controlled by ectopeptidases. To understand ectopeptidase activity, we developed a physical probe and an accompanying method. The probe has a two-channel source that supplies substrate or substrate plus inhibitor using electroosmotic perfusion (EOP). It also has a microdialysis probe to collect products and unreacted substrate. The method provides quantitative estimates of substrate-to-product conversion and the influence of inhibitors on this process. The quantitative estimates are made possible by including a d-amino acid-containing peptide analog of the substrate in the substrate-containing solution infused. Quantitative analysis of substrate, substrate analog, and products is carried out by quantitative, online capillary liquid chromatography-tandem mass spectrometry. The electroosmotic perfusion-microdialysis probe and associated method were used to determine the effect of the selective inhibitor HFI-419 on insulin-regulated aminopeptidase (EC 3.4.11.3) in the rat neocortex.

Pressure-induced infusion of solutions into brain tissue is used both in research and in medicine. In medicine, convection enhanced delivery (CED) may be used to deliver agents to localized areas of the brain, such as with gene therapy to functional targets or with deep tumors not readily amenable to resection. However, clinical trials have demonstrated mixed results from CED. CED is limited by a lack of control of the infusion flow path and may cause damage or even neurological deficits due to neuronal distortion. In laboratory research, infusions may be achieved using pressure or using brief bursts of electrical current in iontophoresis. Electrokinetic convection enhanced delivery (ECED) has the potential to deliver drugs and other bioactive substances to local regions in the brain with improved control and lower applied pressures than pressure-based CED. ECED improves control over the infusion profile because the fluid follows the electrical current path and thus can be directed. Both small molecules and macromolecules can be delivered. Here we demonstrate proof-of-principal that electrokinetic (electroosmosis and electrophoresis) convection-enhanced delivery is a viable means for delivering solutes to the brain. We assessed the volume of tissue exposed to the infusates tris(2,2′-bipyridine)ruthenium(II) and fluorescent dextrans. Control of the direction of the transport was also achieved over distances ranging from several hundred micrometers to more than 4 mm. Electrokinetic delivery has the potential to improve control over infusions.