A Path to Real-World Evidence in Critical Care Using Open-Source Data Harmonization Tools.

COVID-19 highlighted the need for use of real-world data (RWD) in critical care as a near real-time resource for clinical, research, and policy efforts. Analysis of RWD is gaining momentum and can generate important evidence for policy makers and regulators. Extracting high quality RWD from electronic health records (EHRs) requires sophisticated infrastructure and dedicated resources. We sought to customize freely available public tools, supporting all phases of data harmonization, from data quality assessments to de-identification procedures, and generation of robust, data science ready RWD from EHRs. These data are made available to clinicians and researchers through CURE ID, a free platform which facilitates access to case reports of challenging clinical cases and repurposed treatments hosted by the National Center for Advancing Translational Sciences/National Institutes of Health in partnership with the Food and Drug Administration. This commentary describes the partnership, rationale, process, use case, impact in critical care, and future directions for this collaborative effort.

Assessment of Postsystolic Shortening and Global Longitudinal Strain Improves the Sensitivity of Dobutamine Stress Echocardiography in End-Stage Liver Disease.

BACKGROUND: Dobutamine stress echocardiography (DSE) remains a widely used method for detection of coronary artery disease (CAD) in patients with end-stage liver disease (ESLD) despite low sensitivity. Speckle-tracking assessment of strain may enhance the sensitivity of DSE in the general population, but the value of strain analysis in ESLD is unknown. METHODS: Dobutamine stress echocardiography with two-dimensional speckle-tracking and quantitative coronary angiography were performed in 146 patients with ESLD. Thirty-six patients (25%) had CAD (≥50% diameter stenosis of a major vessel). Global longitudinal strain at rest (GLSr) and at peak stress (GLSp) and an index of postsystolic (PSSi) shortening ([maximal extent of shortening - extent of shortening in systole]/[extent of shortening in systole]) were determined. A PSSi of ≥ 0.25 was considered evidence for CAD. Receiver operating characteristic analysis was used to determine the optimal thresholds of GLSr and GLSp for CAD and to assess the diagnostic performance of visual assessment of wall motion (WMA) and strain parameters. The sensitivity and specificity of WMA, GLSr, GLSp, and PSSi were compared. RESULTS: Thirty-six patients (25%) had significant CAD. The areas under the curve for WMA, GLSr, GLSp, and PSSi were 0.60, 0.72, 0.68, and 0.78, respectively. Visual assessment of wall motion had a sensitivity of 28%. The sensitivity of each of the strain parameters, GLSr (53%, P = .016), GLSp (69%, P = .004), and PSSi (78%, P < .001), exceeded the sensitivity for WMA. Visual assessment of wall motion specificity was 92%, which exceeded the specificity for each of the strain parameters (GLSr = 82%, P = .037; GLSp = 63%, P < .001; and PSSi =78%, P = .009). Of the strain parameters, PSSi had the best balance between sensitivity and specificity (both 78%). CONCLUSION: Assessment of GLS and PSSi with DSE yields better sensitivity than WMA in ELSD patients. Index of postsystolic shortening had the best diagnostic performance of all parameters in this population with a low prevalence of CAD.

Comparison of Warming Capabilities Between Buddy Lite, enFlow, and Thermal Angel for US Army Medical Personnel in Austere Conditions: A Literature Review.

US Army Forward Surgical Elements (FSEs) are highly mobile teams that provide damage control surgery (DCS) and damage control resuscitation (DCR) in austere locations that often lack standard hospital utilities (electricity, heat, food, and water). FSEs rely on portable battery-operated intravenous (IV) fluid warmers to remain light and mobile. However, their ability to warm blood in a massive resuscitation requires additional analysis. The purpose of this literature review is to examine the three most common battery-operated IV fluid warmers as determined by type and quantity listed on the Mission Table of Organization and Equipment (MTOE) of organic mobile medical units. These include the Buddy Lite, enFlow, and Thermal Angel, which are available to deployed US Army FSEs for blood resuscitation therapy. Based on limited available evidence, the enFlow produced higher outlet temperatures, effectively warmed greater volumes, reached the time to peak temperature faster, and produced greatest flow rates, with cool saline (5-10°C), compared to the Thermal Angel and Buddy Lite. However, recently the US Food and Drug Administration (FDA) issued a Class 1 recall on enFlow cartridges. Testing demonstrated aluminum elution from enFlow cartridges into IV solutions, thereby exposing patients to potentially unsafe aluminum levels. The authors recommend FSE units conduct a 100% enFlow cartridge inventory and seek an alternative IV fluid warming system prior to enFlow cartridge disposal. If an alternative does not exist, or the alternative warming system does not fit mission requirements, then medical personnel must carefully weigh the risks and benefits associated with the enFlow delivery system.

Asystole in a COVID-19 patient without systemic illness: a case report.

There is growing evidence that patients with severe systemic illness from coronavirus disease 2019 (COVID-19) are at risk for developing a variety of cardiac arrhythmias. Less is known about patients with milder symptoms. Here, we report on the case of a 62-year-old male, admitted to the hospital following an episode of syncope, who experienced multiple episodes of cardiac arrest due to asystole lasting up to 30 seconds. History revealed a recent asymptomatic COVID-19 infection, and recurrent episodes of prolonged asystole necessitated permanent pacemaker placement. To our knowledge, this is the first report of an asymptomatic COVID-19 patient experiencing prolonged asystole. Cardiac arrhythmias in asymptomatic or oligosymptomatic COVID-19 patients may be underestimated.

Lymphocytic myocarditis with suspected granulomatosis with polyangiitis presenting as cardiogenic shock, restrictive cardiomyopathy and complete heart block.

We report a case of restrictive cardiomyopathy from lymphocytic myocarditis in a patient with suspected granulomatosis with polyangiitis (GPA). The case was complicated by complete heart block and renal failure. The diagnosis was supported by upper airway involvement, elevated serum serine proteinase 3 antibodies, and endomyocardial biopsy with lymphocytic infiltration. The patient responded appropriately to aggressive immunosuppressive therapy. .

Morphine-sensitive synaptic transmission emerges in embryonic rat microphysiological model of lower afferent nociceptive signaling.

Debilitating chronic pain resulting from genetic predisposition, injury, or acquired neuropathy is becoming increasingly pervasive. Opioid analgesics remain the gold standard for intractable pain, but overprescription of increasingly powerful and addictive opioids has contributed to the current prescription drug abuse epidemic. There is a pressing need to screen experimental compounds more efficiently for analgesic potential that remains unmet by conventional research models. The spinal cord dorsal horn is a common target for analgesic intervention, where peripheral nociceptive signals are relayed to the central nervous system through synaptic transmission. Here, we demonstrate that coculturing peripheral and dorsal spinal cord nerve cells in a novel bioengineered microphysiological system facilitates self-directed emergence of native nerve tissue macrostructure and concerted synaptic function. The mechanistically distinct analgesics-morphine, lidocaine, and clonidine-differentially and predictably modulate this microphysiological synaptic transmission. Screening drug candidates for similar microphysiological profiles will efficiently identify therapeutics with analgesic potential.

Spatial variation in direct and indirect contact rates at the wildlife-livestock interface for informing disease management.

Little is known about disease transmission relevant contact rates at the wildlife-livestock interface and the factors shaping them. Indirect contact via shared resources is thought to be important but remains unquantified in most systems, making it challenging to evaluate the impact of livestock management practices on contact networks. Free-ranging wild pigs (Sus scrofa) in North America are an invasive, socially-structured species with an expanding distribution that pose a threat to livestock health given their potential to transmit numerous livestock diseases, such as pseudorabies, brucellosis, trichinellosis, and echinococcosis, among many others. Our objective in this study was to quantify the spatial variations in direct and indirect contact rates among wild pigs and cattle on a commercial cow-calf operation in Florida, USA. Using GPS data from 20 wild pigs and 11 cattle and a continuous-time movement model, we extracted three types of spatial contacts between wild pigs and cattle, including direct contact, indirect contact in the pastoral environment (unknown naturally occurring resources), and indirect contact via anthropogenic cattle resources (feed supplements and water supply troughs). We examined the effects of sex, spatial proximity, and cattle supplement availability on contact rates at the species level and characterized wild pig usage of cattle supplements. Our results suggested daily pig-cattle direct contacts occurred only occasionally, while a significant number of pig-cattle indirect contacts occurred via natural resources distributed heterogeneously across the landscape. At cattle supplements, more indirect contacts occurred at liquid molasses than water troughs or molasses-mineral block tubs due to higher visitation rates by wild pigs. Our results can be directly used for parameterizing epidemiological models to inform risk assessment and optimal control strategies for controlling transmission of shared diseases.

Advances in 3D neuronal microphysiological systems: towards a functional nervous system on a chip.

Microphysiological systems (MPS) designed to study the complexities of the peripheral and central nervous systems have made marked improvements over the years and have allowed researchers to assess in two and three dimensions the functional interconnectivity of neuronal tissues. The recent generation of brain organoids has further propelled the field into the nascent recapitulation of structural, functional, and effective connectivities which are found within the native human nervous system. Herein, we will review advances in culture methodologies, focused especially on those of human tissues, which seek to bridge the gap from 2D cultures to hierarchical and defined 3D MPS with the end goal of developing a robust nervous system-on-a-chip platform. These advances have far-reaching implications within basic science, pharmaceutical development, and translational medicine disciplines.

Effects of social structure and management on risk of disease establishment in wild pigs.

Contact heterogeneity among hosts determines invasion and spreading dynamics of infectious disease, thus its characterization is essential for identifying effective disease control strategies. Yet, little is known about the factors shaping contact networks in many wildlife species and how wildlife management actions might affect contact networks. Wild pigs in North America are an invasive, socially structured species that pose a health concern for domestic swine given their ability to transmit numerous devastating diseases such as African swine fever (ASF). Using proximity loggers and GPS data from 48 wild pigs in Florida and South Carolina, USA, we employed a probabilistic framework to estimate weighted contact networks. We determined the effects of sex, social group and spatial distribution (monthly home-range overlap and distance) on wild pig contact. We also estimated the impacts of management-induced perturbations on contact and inferred their effects on ASF establishment in wild pigs with simulation. Social group membership was the primary factor influencing contacts. Between-group contacts depended primarily on space use characteristics, with fewer contacts among groups separated by >2 km and no contacts among groups >4 km apart within a month. Modelling ASF dynamics on the contact network demonstrated that indirect contacts resulting from baiting (a typical method of attracting wild pigs or game species to a site to enhance recreational hunting) increased the risk of disease establishment by ~33% relative to direct contact. Low-intensity population reduction (<5.9% of the population) had no detectable impact on contact structure but reduced predicted ASF establishment risk relative to no population reduction. We demonstrate an approach for understanding the relative role of spatial, social and individual-level characteristics in shaping contact networks and predicting their effects on disease establishment risk, thus providing insight for optimizing disease control in spatially and socially structured wildlife species.

Bundling of axons through a capillary alginate gel enhances the detection of axonal action potentials using microelectrode arrays.

Microelectrode arrays (MEAs) have become important tools in high throughput assessment of neuronal activity. However, geometric and electrical constraints largely limit their ability to detect action potentials to the neuronal soma. Enhancing the resolution of these systems to detect axonal action potentials has proved both challenging and complex. In this study, we have bundled sensory axons from dorsal root ganglia through a capillary alginate gel (Capgel™) interfaced with an MEA and observed an enhanced ability to detect spontaneous axonal activity compared with two-dimensional cultures. Moreover, this arrangement facilitated the long-term monitoring of spontaneous activity from the same bundle of axons at a single electrode. Finally, using waveform analysis for cultures treated with the nociceptor agonist capsaicin, we were able to dissect action potentials from multiple axons on an individual electrode, suggesting that this model can reproduce the functional complexity associated with sensory fascicles in vivo. This novel three-dimensional functional model of the peripheral nerve can be used to study the functional complexities of peripheral neuropathies and nerve regeneration as well as being utilized in the development of novel therapeutics.

Wild pigs as sentinels for hard ticks: A case study from south-central Florida.

As a result of shifts in the habitable range of ticks due to climate change and the ongoing threat of exotic tick species introductions, efficient surveillance tools for these pests and disease vectors are needed. Wild pigs are habitat generalists, distributed throughout most of the United States, and often hunted recreationally or removed as part of management programs, making them potentially useful sentinel hosts for ticks. We compared ticks collected from captured wild pigs and standard tick dragging methods on a south-central Florida cattle ranch from May 2015-August 2017. Three hundred and sixteen wild pigs were surveyed, and 84 km spanning three habitat types (seminative pasture, improved pasture, and hammock) were dragged. In total, 1023 adults of four species (Amblyomma auricularium, Amblyomma maculatum, Dermacentor variabilis, and Ixodes scapularis) were collected from wild pigs, while 39 adults of three species (A. auricularium, A. maculatum, and I. scapularis) were collected from drags. Only one immature specimen, a nymph, was collected from a pig, while dragging collected 2808 larvae and 150 nymphs. Amblyomma maculatum comprised 96% of adults collected from pigs, while A. maculatum, I. scapularis, and A. auricularium comprised 38%, 33%, and 28% of adults collected from drags, respectively. Adults of all tick species found on drags were found on pigs, and wild pig surveillance detected adults of an additional species not found on drags. Dragging was far superior for collection of immatures but not for adults of most species found in this study. These findings suggest wild pigs could be used as a sentinel for the detection of tick species. When combined with ongoing wild pig research, hunting, or management, wild pig surveillance can provide an effective method to survey for adult tick presence of some species of interest and may assist in tracking the range expansion of some tick species.

Use of a capillary alginate gel (Capgel™) to study the three-dimensional development of sensory nerves reveals the formation of a rudimentary perineurium.

BACKGROUND: Peripheral neuropathies affect approximately 20 million people in the United States and often stem from other chronic conditions, such as diabetes. In vitro methodologies to facilitate the understanding and treatment of these disorders often lack the cellular and functional complexity required to accurately model peripheral neuropathies. In particular, they are often 2D and fail to faithfully reproduce the 3D in vivo microenvironment. NEW METHOD: Embryonic dorsal root ganglion (DRG) explants were inserted into laminin derivatized capillary alginate gel (Capgel™), a bioabsorbable, self-assembling biomaterial, possessing parallel microchannel architecture, and cultured to mimic normal nerve development, including Schwann cell myelination. RESULTS: Laminin derivatization of the microchannels improved nerve growth through the gel. Axon bundles containing myelinating Schwann cells migrated through the gel and were ensheathed by rudimentary perineurium up to 1 mm from the DRG explant site. COMPARISON WITH EXISTING METHODS: Other nerve models are two-dimensional in nature and/or fail to conserve the complicated architecture and cellular milieu observed in vivo. Our nerve model shows the simple culture technique of cells grown in 3D, which allows for a more advanced structural organization that more accurately mimics the in vivo nerve fascicle. CONCLUSIONS: When embryonic DRG explants are cultured in this system, they show a striking resemblance to in vivo peripheral nerve fascicles, including myelinated axons and the formation of a rudimentary perineurium, suggesting that both neuronal and non-neuronal cells within the DRG explant are capable of recreating the 3D structure of a developing sensory fascicle within the microchannel architecture.

Alzheimer disease brain atrophy subtypes are associated with cognition and rate of decline.

OBJECTIVE: To test the hypothesis that cortical and hippocampal volumes, measured in vivo from volumetric MRI (vMRI) scans, could be used to identify variant subtypes of Alzheimer disease (AD) and to prospectively predict the rate of clinical decline. METHODS: Amyloid-positive participants with AD from the Alzheimer's Disease Neuroimaging Initiative (ADNI) 1 and ADNI2 with baseline MRI scans (n = 229) and 2-year clinical follow-up (n = 100) were included. AD subtypes (hippocampal sparing [HpSpMRI], limbic predominant [LPMRI], typical AD [tADMRI]) were defined according to an algorithm analogous to one recently proposed for tau neuropathology. Relationships between baseline hippocampal volume to cortical volume ratio (HV:CTV) and clinical variables were examined by both continuous regression and categorical models. RESULTS: When participants were divided categorically, the HpSpMRI group showed significantly more AD-like hypometabolism on 18F-fluorodeoxyglucose-PET (p < 0.05) and poorer baseline executive function (p < 0.001). Other baseline clinical measures did not differ across the 3 groups. Participants with HpSpMRI also showed faster subsequent clinical decline than participants with LPMRI on the Alzheimer's Disease Assessment Scale, 13-Item Subscale (ADAS-Cog13), Mini-Mental State Examination (MMSE), and Functional Assessment Questionnaire (all p < 0.05) and tADMRI on the MMSE and Clinical Dementia Rating Sum of Boxes (CDR-SB) (both p < 0.05). Finally, a larger HV:CTV was associated with poorer baseline executive function and a faster slope of decline in CDR-SB, MMSE, and ADAS-Cog13 score (p < 0.05). These associations were driven mostly by the amount of cortical rather than hippocampal atrophy. CONCLUSIONS: AD subtypes with phenotypes consistent with those observed with tau neuropathology can be identified in vivo with vMRI. An increased HV:CTV ratio was predictive of faster clinical decline in participants with AD who were clinically indistinguishable at baseline except for a greater dysexecutive presentation.

Further Evaluation of Mechanisms Associated with the Antidepressantlike Signature of Scopolamine in Mice.

BACKGROUND: Conventional antidepressants lack efficacy for many patients (treatmentresistant depression or TRD) and generally take weeks to produce full therapeutic response in others. Emerging data has identified certain drugs such as ketamine as rapidly-acting antidepressants for major depressive disorder and TRD. Scopolamine, a drug used to treat motion sickness and nausea, has also been demonstrated to function as a rapidly-acting antidepressant. The mechanisms associated with efficacy in TRD patients and rapid onset of action have been suggested to involve a-Amino-3-hydroxy- 5-methyl-4-isoxazolepropionic acid (AMPA) receptor and mammalian target of rapamycin (mTOR) signaling. Since the work on these mechanisms with scopolamine has been limited, the present set of experiments was designed to further explore these mechanisms of action. METHOD: Male, NIH Swiss mice demonstrated a robust and immediate antidepressant signature with ketamine or scopolamine when studied under the forced-swim test. RESULTS: The AMPA receptor antagonist NBQX prevented this antidepressant-like effect of scopolamine and ketamine. An orally-bioavilable mTOR inhibitor (AZD8055) also attenuated the antidepressant- like effects of scopolamine and ketamine. Scopolamine was also shown to augment the antidepressant- like effect of the selective serotonin reuptake inhibitor citalopram. When given in combination, scopolamine and ketamine acted synergistically to produce antidepressant-like effects. Although drug interaction data suggested that additional mechanisms might be at play, metabolomic analysis of frontal cortex and plasma from muscarinic M1+/+ and M1 -/- mice given scopolamine or vehicle did not reveal any hints as to the nature of these additional mechanisms of action. CONCLUSION: Overall, the data substantiate and extend the idea that AMPA and mTOR signaling pathways are necessary for the antidepressant-like effects of scopolamine and ketamine, mechanisms that appear to be of general significance for TRD therapeutic agents.

In vitro pharmacological and rat pharmacokinetic characterization of LY3020371, a potent and selective mGlu2/3 receptor antagonist.

Metabotropic glutamate 2/3 (mGlu2/3) receptors are of considerable interest owing to their role in modulating glutamate transmission via presynaptic, postsynaptic and glial mechanisms. As part of our ongoing efforts to identify novel ligands for these receptors, we have discovered (1S,2R,3S,4S,5R,6R)-2-amino-3-[(3,4-difluorophenyl)sulfanylmethyl]-4-hydroxy-bicyclo[3.1.0]hexane-2,6-dicarboxylic acid; (LY3020371), a potent and selective orthosteric mGlu2/3 receptor antagonist. In this account, we characterize the effects of LY3020371 in membranes and cells expressing human recombinant mGlu receptor subtypes as well as in native rodent and human brain tissue preparations, providing important translational information for this molecule. In membranes from cells expressing recombinant human mGlu2 and mGlu3 receptor subtypes, LY3020371.HCl competitively displaced binding of the mGlu2/3 agonist ligand [3H]-459477 with high affinity (hmGlu2 Ki = 5.26 nM; hmGlu3 Ki = 2.50 nM). In cells expressing hmGlu2 receptors, LY3020371.HCl potently blocked mGlu2/3 agonist (DCG-IV)-inhibited, forskolin-stimulated cAMP formation (IC50 = 16.2 nM), an effect that was similarly observed in hmGlu3-expressing cells (IC50 = 6.21 nM). Evaluation of LY3020371 in cells expressing the other human mGlu receptor subtypes revealed high mGlu2/3 receptor selectivity. In rat native tissue assays, LY3020371 demonstrated effective displacement of [3H]-459477 from frontal cortical membranes (Ki = 33 nM), and functional antagonist activity in cortical synaptosomes measuring both the reversal of agonist-suppressed second messenger production (IC50 = 29 nM) and agonist-inhibited, K+-evoked glutamate release (IC50 = 86 nM). Antagonism was fully recapitulated in both primary cultured cortical neurons where LY3020371 blocked agonist-suppressed spontaneous Ca2+ oscillations (IC50 = 34 nM) and in an intact hippocampal slice preparation (IC50 = 46 nM). Functional antagonist activity was similarly demonstrated in synaptosomes prepared from epileptic human cortical or hippocampal tissues, suggesting a translation of the mGlu2/3 antagonist pharmacology from rat to human. Intravenous dosing of LY3020371 in rats led to cerebrospinal fluid drug levels that are expected to effectively block mGlu2/3 receptors in vivo. Taken together, these results establish LY3020371 as an important new pharmacological tool for studying mGlu2/3 receptors in vitro and in vivo. This article is part of the Special Issue entitled 'Metabotropic Glutamate Receptors, 5 years on'.

An Allosteric Potentiator of the Dopamine D1 Receptor Increases Locomotor Activity in Human D1 Knock-In Mice without Causing Stereotypy or Tachyphylaxis.

Allosteric potentiators amplify the sensitivity of physiologic control circuits, a mode of action that could provide therapeutic advantages. This hypothesis was tested with the dopamine D1 receptor potentiator DETQ [2-(2,6-dichlorophenyl)-1-((1S,3R)-3-(hydroxymethyl)-5-(2-hydroxypropan-2-yl)-1-methyl-3,4-dihydroisoquinolin-2(1H)-yl)ethan-1-one]. In human embryonic kidney 293 (HEK293) cells expressing the human D1 receptor, DETQ induced a 21-fold leftward shift in the cAMP response to dopamine, with a Kb of 26 nM. The maximum response to DETQ alone was ∼12% of the maximum response to dopamine, suggesting weak allosteric agonist activity. DETQ was ∼30-fold less potent at rat and mouse D1 receptors and was inactive at the human D5 receptor. To enable studies in rodents, an hD1 knock-in mouse was generated. DETQ (3-20 mg/kg orally) caused a robust (∼10-fold) increase in locomotor activity (LMA) in habituated hD1 mice but was inactive in wild-type mice. The LMA response to DETQ was blocked by the D1 antagonist SCH39166 and was dependent on endogenous dopamine. LMA reached a plateau at higher doses (30-240 mg/kg) even though free brain levels of DETQ continued to increase over the entire dose range. In contrast, the D1 agonists SKF 82958, A-77636, and dihydrexidine showed bell-shaped dose-response curves with a profound reduction in LMA at higher doses; video-tracking confirmed that the reduction in LMA caused by SKF 82958 was due to competing stereotyped behaviors. When dosed daily for 4 days, DETQ continued to elicit an increase in LMA, whereas the D1 agonist A-77636 showed complete tachyphylaxis by day 2. These results confirm that allosteric potentiators may have advantages compared with direct-acting agonists.

Service Block Time Allocation in the US Army Medical Command.

Service block time allocation is a critical requirement for the optimization of patient throughput and access to care in the Surgical Services Service Line of the US Army Medical Command. The procedure complexity, volume, and diversity across 25 facilities create significant variation in service block time. This variation requires the involvement of both the informatics and leadership teams for block time allocation to be effective. This article describes our use of the Army's Surgery Scheduling System, which includes service block time as an embedded function, to develop a standardized process that helps ensure service block time is optimized. We also present guidelines for block time allocation and offer case studies that demonstrate the application of these guidelines.

Dogs Can Be Successfully Trained to Alert to Hypoglycemia Samples from Patients with Type 1 Diabetes.

INTRODUCTION: Hypoglycemia (Hypo) is the most common side effect of insulin therapy in people with type 1 diabetes (T1D). Over time, patients with T1D become unaware of signs and symptoms of Hypo. Hypo unawareness leads to morbidity and mortality. Diabetes alert dogs (DADs) represent a unique way to help patients with Hypo unawareness. Our group has previously presented data in abstract form which demonstrates the sensitivity and specificity of DADS. The purpose of our current study is to expand evaluation of DAD sensitivity and specificity using a method that reduces the possibility of trainer bias. METHODS: We evaluated 6 dogs aging 1-10 years old who had received an average of 6 months of training for Hypo alert using positive training methods. Perspiration samples were collected from patients during Hypo (BG 46-65 mg/dL) and normoglycemia (BG 85-136 mg/dl) and were used in training. These samples were placed in glass vials which were then placed into 7 steel cans (1 Hypo, 2 normal, 4 blank) randomly placed by roll of a dice. The dogs alerted by either sitting in front of, or pushing, the can containing the Hypo sample. Dogs were rewarded for appropriate recognition of the Hypo samples using a food treat via a remote control dispenser. The results were videotaped and statistically evaluated for sensitivity (proportion of lows correctly alerted, "true positive rate") and specificity (proportion of blanks + normal samples not alerted, "true negative rate") calculated after pooling data across all trials for all dogs. RESULTS: All DADs displayed statistically significant (p value <0.05) greater sensitivity (min 50.0%-max 87.5%) to detect the Hypo sample than the expected random correct alert of 14%. Specificity ranged from a min of 89.6% to a max of 97.9% (expected rate is not defined in this scenario). CONCLUSIONS: Our results suggest that properly trained DADs can successfully recognize and alert to Hypo in an in vitro setting using smell alone.

Inquiries into the Biological Significance of Transmembrane AMPA Receptor Regulatory Protein (TARP) γ-8 Through Investigations of TARP γ-8 Null Mice§.

Transmembrane AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor regulatory protein (TARP) γ-8 is an auxiliary protein associated with some AMPA receptors. Most strikingly, AMPA receptors associated with this TARP have a relatively high localization in the hippocampus. TARP γ-8 also modifies the pharmacology and trafficking of AMPA receptors. However, to date there is little understanding of the biological significance of this auxiliary protein. In the present set of studies we provide a characterization of the differential pharmacology and behavioral consequences of deletion of TARP γ-8 by comparing the wild type (WT) and γ-8 -/- (knock-out, KO) mouse. KO mice were mildly hyperactive in a locomotor arena but not in other environments compared to WT mice. Additionally, the KO mice demonstrated enhanced locomotor stimulatory effects of both d-amphetamine and phencyclidine. Marble-burying and digging behaviors were dramatically reduced in KO mice. In another assay that can detect anxiety-like phenotypes, the elevated plus maze, no differences were observed in overall movement or open arm entries. In the forced-swim assay, KO mice displayed decreases in immobility time like the antidepressant imipramine and the AMPA receptor potentiator, LY392098. In KO mice, the antidepressant-like effects of LY392098 were prevented whereas the effects of imipramine were unaffected. Convulsions were induced by pentylenetetrazole, N-methyl-D-aspartate, and by kainic acid. However, in KO mice, kainic acid produced less tonic convulsions and lethality. KO mice had reduced levels of norepinephrine in hippocampus and cerebellum but not in hypothalamus or prefrontal cortex, decreased levels of cAMP in hippocampus, and increased levels of acetylcholine in the hypothalamus and prefrontal cortex. KO mice displayed decreased turnover of dopamine and increased histamine turnover in multiple brain areas In contrast, serotonin and its metabolites were not significantly affected by deletion of the γ-8 protein. Of a large panel of plasma lipids, only two monoacylglycerols (1OG and 2OG) were marginally but nonsignificantly altered in WT vs KO mice. Overall, the data suggest genetic inactivation of this specific population of AMPA receptors results in modest changes in behavior characterized by a mild hyperactivity which is condition dependent and a marked reduction in digging and burying behaviors. Despite deletion of TARP γ-8, chemoconvulsants were still active. Consistent with their predicted pharmacological actions, the convulsant effects of kainate and the antidepressant-like effects of an AMPA receptor potentiator (both acting upon AMPA receptors) were reduced or absent in KO mice.