Exposure to N,N-Diethyl-Meta-Toluamide Insect Repellent and Human Health Markers: Population Based Estimates from the National Health and Nutrition Examination Survey.

N,N-diethyl-meta-toluamide (DEET) is one of the most commonly used insect repellants in the United States, yet the existing literature regarding DEET's potential deleterious impact on humans is mixed and is based mostly on case reports. The primary aim of this study was to address this lack of population-based evidence of the effects of DEET exposure on human health in the United States. Our primary outcome measures were biomarkers related to systemic inflammation (high sensitivity C-reactive protein), immune function (lymphocyte), liver function (aspartate aminotransferase, alanine aminotransferase, and γ-glutamyl transferace), and kidney function (estimated glomerular filtration rate). We analyzed data from the population-based National Health and Nutrition Examination Survey, 2015-2016, and identified 1,205 patients (age 20+ years) who had DEET metabolite levels recorded at or above detection limits. A Pearson correlation was used to assess the relationship between DEET metabolite, and each biomarker found there was no significant correlation. Thus, there is no evidence that DEET exposure has any impact on the biomarkers identified.

The Association Between Admiration of Antisocial Peers and Past 30-Day Opioid Misuse Among Justice-Involved children.

AIM: Prevention of illicit or nonmedical opioid use, called opioid misuse (OM) is a key public health concern that requires research on the factors that influence OM initiation among high-risk populations. Justice-involved children (JIC) have more risk factors and fewer resources. Antisocial peers have been linked to adolescent substance abuse and delinquency. However, the association between the admiration of antisocial peers and OM among JIC has not yet been studied. This study hypothesizes that admiration of antisocial peers will be associated with a higher likelihood of OM among Florida JIC. METHODS: Cross-sectional data on 79,960 JIC from the Florida Department of Juvenile Justice (FLDJJ) were examined. To test the hypothesis, bivariate and multivariate logistic regression analyses were employed. The multivariate models controlled for gender, race, age in 2007, family income, history of mental health, history of depression, and optimism. RESULTS: Nearly 2.7% of the sample met the criteria for past 30-day OM, and over 75% of those current users admired or somewhat admired their antisocial peers. Compare to JIC who did not admire their antisocial peers, those who had some admiration of antisocial peers were 2.39 times more likely to misuse opioids in the past 30-days and those who admired their antisocial peers were 4.40 times more likely to meet the criteria for past 30-day OM. CONCLUSIONS: Cultivating positive peer interactions and providing positive peer role models may help to reduce illicit opioid use among JIC.

Urotensin-II induces ear flushing in rats.

BACKGROUND AND PURPOSE: While investigating the effects of systemic urotensin II (U-II), a potent vasoactive peptide acting at the UT receptor, we observed ear pinna flushing after systemic administration to conscious rats. In the present study, U-II-induced ear flushing was quantified in terms of ear pinna temperature change and potential mechanisms were explored. EXPERIMENTAL APPROACH: U-II-induced ear flushing was quantified by measuring lateral ear pinna temperature changes and compared to that of calcitonin gene-related peptide (CGRP), a known cutaneous vasodilator. Further, the effects of a variety of pharmacological agents on U-II-induced ear flushing were explored. KEY RESULTS: Subcutaneous injection of U-II (9 microg kg(-1))produced localized ear pinna flushing with an onset of approximately 15 min, a duration of approximately 30 min and a maximal temperature change of 9 degrees C. In contrast, CGRP caused cutaneous flushing within multiple cutaneous beds including the ear pinna with a shorter onset and greater duration than U-II. A potent UT receptor antagonist, urantide, blocked U-II-induced ear flushing but did not affect CGRP-induced ear flushing. Pretreatment with indomethacin or L-Nomega-nitroarginine methylester (L-NAME) abolished U-II-induced ear flushing. Mecamylamine or propranolol did not affect this response to U-II. Direct intracerebroventricular injection studies suggested that the ear flushing response to U-II was not mediated directly by the CNS. CONCLUSION AND IMPLICATIONS: Our results suggest that U-II-induced ear flushing and temperature increase is mediated by peripheral activation of the UT receptor and involves prostaglandin- and nitric oxide-mediated vasodilation of small capillary beds in the rat ear pinna.

Superior canal dehiscence: mechanisms of pressure sensitivity in a chinchilla model.

BACKGROUND: Patients with superior canal dehiscence syndrome may experience vertigo and nystagmus when pressure changes occur in the external auditory canal, the middle ear, or the intracranial space. The cause is a defect in the bone of the superior canal. OBJECTIVE: To study the mechanisms of pressure sensitivity of the labyrinth in superior canal dehiscence syndrome and its surgical repair in a chinchilla model. METHODS: We investigated the changes in firing rates of vestibular nerve afferents in the chinchilla in response to changes in external auditory canal pressure before and after fenestration of the superior canal, and after repair of the fenestra. RESULTS: Before superior canal fenestration, external auditory canal pressure changes caused no responses in horizontal canal or otolith afferents, and only 1 of 9 superior canal afferents responded to pressure. After fenestration, all superior canal afferents were excited by positive pressure and inhibited by negative pressure. Half of 18 otolith and most (21 of 33) horizontal canal afferents were unaffected by pressure. The superior canal afferents had higher pressure gain than the horizontal canal afferents (P =.03). Pressure responses could be abolished only by applying a rigid seal to the fenestra. CONCLUSIONS: Fenestration of the superior canal rendered all superior canal afferents sensitive to pressure, whereas less than half of the other afferents became pressure sensitive. The direction of the superior canal afferent responses agreed with the predictions of our model of endolymph flow within the superior canal. A rigid seal applied to the fenestra abolished pressure sensitivity while maintaining physiologic rotational sensitivity.

Quantitative analysis of agonist-dependent parathyroid hormone receptor trafficking in whole cells using a functional green fluorescent protein conjugate.

Many G-protein coupled receptors (GPCRs) undergo ligand-dependent internalization upon activation. The parathyroid hormone (PTH) receptor undergoes endocytosis following prolonged exposure to ligand although the ultimate fate of the receptor following internalization is largely unknown. To investigate compartmentalization of the PTH receptor, we have established a stable cell line expressing a PTH receptor-green fluorescent protein (PTHR-GFP) conjugate and an algorithm to quantify PTH receptor internalization. HEK 293 cells expressing the PTHR-GFP were compared with cells expressing the wild-type PTH receptor in whole-cell binding and functional assays. 125I-PTH binding studies revealed similar Bmax and kD values in cells expressing either the PTHR-GFP or the wild-type PTH receptor. PTH-induced cAMP accumulation was similar in both cell lines suggesting that addition of the GFP to the cytoplasmic tail of the PTH receptor does not alter the ligand binding or G-protein coupling properties of the receptor. Using confocal fluorescence microscopy, we demonstrated that PTH treatment of cells expressing the PTHR-GFP conjugate produced a time-dependent redistribution of the receptor to the endosomal compartment which was blocked by pretreatment with PTH antagonist peptides. Treatment with hypertonic sucrose prevented PTH-induced receptor internalization, suggesting that the PTH receptor internalizes via a clathrin-dependent mechanism. Moreover, co-localization with internalized transferrin showed that PTHR-GFP trafficking utilized the endocytic recycling compartment. Experiments using cycloheximide to inhibit protein synthesis demonstrated that recycling of the PTHR-GFP back to the plasma membrane was complete within 1-2 h of ligand removal and was partially blocked by pretreatment with cytochalasin D, but not nocodazole. We also demonstrated that the PTH receptor, upon recycling to the plasma membrane, is capable of undergoing a second round of internalization, a finding consistent with a role for receptor recycling in functional resensitization.

Symptoms and signs in superior canal dehiscence syndrome.

Patients with superior canal dehiscence (SCD) syndrome experience vertigo and oscillopsia in response to loud sounds and to stimuli that result in changes in middle ear or intracranial pressure. They may also experience hyperacusis to bone-conducted sounds. The evoked eye movements in this syndrome align with the plane of the dehiscent superior canal. The symptoms and signs can be understood in terms of the effect of the dehiscence in creation of a third mobile window into the inner ear. The SCD syndrome has been diagnosed in 28 patients who were examined in the neuro-otology clinics at the Johns Hopkins Medical Institutions from May 1995 through January 2001. The diagnosis is best established based upon the symptoms that are characteristic for the syndrome, the vertical-torsional eye movements evoked by sound or pressure stimuli noted on examination performed with Frenzel goggles, the lowered thresholds for responses to vestibular-evoked myogenic potentials, and CT imaging of the temporal bones.

Translational vestibulo-ocular reflex evoked by a "head heave" stimulus.

The gain and symmetry of vestibulo-ocular reflexes for high-frequency, high-acceleration movements of the head are altered following unilateral vestibular lesions. These changes have been well characterized for rotational head movements (thrusts), and provide reliable markers of dysfunction in individual semicircular canals. Alterations in the vestibulo-ocular reflex (VOR) evoked by lateral, whole-body translations have also been observed. In an approach directed at the development of a bedside test of otolith function, we have recorded (scleral search coil) the VOR evoked by brief, high-acceleration lateral translations (heaves). We delivered these stimuli manually and also developed a "head sled" device that minimizes any rotational contaminating component of the stimulus. Our geometrical analysis of the stimuli enables us to take into account the translational and rotational components of the movement, and to calculate an ideal response required for stabilization of images on the fovea at different fixation distances. We observed a tracking response (visually assisted VOR) that was close to ideal for image stabilization when these methods were used to analyze responses to slow, low-amplitude lateral translations of the head. When applied to rapid, high-acceleration (0.5 g) translations, the VOR was found to be less than compensatory in subjects with normal vestibular function. In a patient with unilateral vestibular hypofunction following intratympanic gentamicin injections, both the rotational and the translational VOR were asymmetric. Responses for translations toward the treated side had lower gain than those for translations toward the normal side. These findings provide a basis for further development of this technique as a clinical test and as a method for quantitative evaluation of otolith function.

Horizontal vestibuloocular reflex evoked by high-acceleration rotations in the squirrel monkey. IV. Responses after spectacle-induced adaptation.

The horizontal angular vestibuloocular reflex (VOR) evoked by sinusoidal rotations from 0.5 to 15 Hz and acceleration steps up to 3,000 degrees /s(2) to 150 degrees /s was studied in six squirrel monkeys following adaptation with x2.2 magnifying and x0.45 minimizing spectacles. For sinusoidal rotations with peak velocities of 20 degrees /s, there were significant changes in gain at all frequencies; however, the greatest gain changes occurred at the lower frequencies. The frequency- and velocity-dependent gain enhancement seen in normal monkeys was accentuated following adaptation to magnifying spectacles and diminished with adaptation to minimizing spectacles. A differential increase in gain for the steps of acceleration was noted after adaptation to the magnifying spectacles. The gain during the acceleration portion, G(A), of a step of acceleration (3,000 degrees /s(2) to 150 degrees /s) increased from preadaptation values of 1.05 +/- 0.08 to 1.96 +/- 0.16, while the gain during the velocity plateau, G(V), only increased from 0.93 +/- 0.04 to 1.36 +/- 0.08. Polynomial fits to the trajectory of the response during the acceleration step revealed a greater increase in the cubic than the linear term following adaptation with the magnifying lenses. Following adaptation to the minimizing lenses, the value of G(A) decreased to 0.61 +/- 0.08, and the value of G(V) decreased to 0.59 +/- 0.09 for the 3,000 degrees /s(2) steps of acceleration. Polynomial fits to the trajectory of the response during the acceleration step revealed that there was a significantly greater reduction in the cubic term than in the linear term following adaptation with the minimizing lenses. These findings indicate that there is greater modification of the nonlinear as compared with the linear component of the VOR with spectacle-induced adaptation. In addition, the latency to the onset of the adapted response varied with the dynamics of the stimulus. The findings were modeled with a bilateral model of the VOR containing linear and nonlinear pathways that describe the normal behavior and adaptive processes. Adaptation for the linear pathway is described by a transfer function that shows the dependence of adaptation on the frequency of the head movement. The adaptive process for the nonlinear pathway is a gain enhancement element that provides for the accentuated gain with rising head velocity and the increased cubic component of the responses to steps of acceleration. While this model is substantially different from earlier models of VOR adaptation, it accounts for the data in the present experiments and also predicts the findings observed in the earlier studies.

Vestibular-evoked myogenic potentials in the diagnosis of superior canal dehiscence syndrome.

Patients with superior canal dehiscence (SCD) syndrome have vertigo and oscillopsia induced by loud noises and by stimuli that result in changes in middle ear or intracranial pressure. We recorded vestibular-evoked myogenic potentials (VEMP responses) in 10 patients with SCD syndrome. The diagnosis had been confirmed in each case by evoked eye movements and by high-resolution CT scans of the temporal bones that showed a dehiscence overlying the affected superior canal. For the 8 patients without prior middle ear disease, the VEMP threshold from the dehiscent ears measured 72 +/- 8 dB NHL (normal hearing level) whereas the threshold from normal control subjects was 96 +/- 5 dB NHL (p < 0.0001). The VEMP threshold measured from the contralateral ear in patients with unilateral dehiscence was 98 +/- 4 dB NHL (p > 0.9 with respect to normal controls). Two patients with apparent conductive hearing loss from middle ear disease, and SCD, had VEMP responses from the affected ears. In the absence of dehiscence, VEMP responses would not have been expected in the setting of conductive hearing loss. These findings confirm earlier studies demonstrating that patients with SCD syndrome have lowered VEMP thresholds. Conditions other than SCD syndrome may also lead to lowered VEMP thresholds. Rather than being based upon a single test, the diagnosis of SCD syndrome is best established when the characteristic symptoms, signs, VEMP response, and CT imaging all indicate SCD.

5-hydroxytryptamine3 (5-HT3) receptors mediate spinal 5-HT antinociception: an antisense approach.

To examine the role of the 5-hydroxytryptamine(1B) (5-HT1B) and 5-HT3 receptor subtypes in the analgesia produced by 5-HT (serotonin) agonists, we assessed the effect of antisense oligodeoxynucleotides (AODNs) designed to "knock down" the number of these receptor subtypes on analgesia produced by intrathecal (i.t.) 5-HT, the 5-HT1B receptor agonist, 7-trifluoromethyl-4-(4-methyl-1-piperazinyl)-pyrrolo[1,2-a]quinoxaline maleate (CGS-12066A), and the 5-HT3 receptor agonist, 2-methyl-5-HT. Groups of mice (n = 17-20) were injected i.t. on days 1, 3, and 5 with one of the AODNs, a mismatch oligo, or saline. On day 6, all mice were injected i.t. with 70.5 nmol of 5-HT, 44.4 nmol of CGS-12066A, or 49 nmol of 2-methyl-5-HT by lumbar puncture. Following testing, spinal cords were rapidly removed and prepared for receptor binding assays. Treatment with AODN for 5-HT1B receptors produced a 70% reduction in ligand binding to this receptor subtype. After treatment with AODN for 5-HT3 receptors, ligand binding to this receptor subtype was undetectable. In mice tested with i.t. 5-HT, tail-flick analgesia was attenuated only in mice treated with the 5-HT3 receptor AODN. Mice treated with the AODN designed to knock down 5-HT(1B) receptors or with its mismatch oligo were not significantly different from controls. In mice tested with i.t. administration of CGS-12066A, none of the oligo treatments produced a significant attenuation of analgesia. In mice tested with i.t. administration of 2-methyl-5-HT, only 5-HT3 receptor AODN attenuated analgesia. Thus, 5-HT and 2-methyl-5-HT analgesia are mediated by the 5-HT3 receptor subtype. However, spinal CGS-12066A analgesia appears not to be mediated by either the 5-HT1B or the 5-HT3 receptor subtypes.

Relationship between time- and frequency-domain analyses of angular head movements in the squirrel monkey.

We used the three-dimensional magnetic search-coil recording technique to study the range of active angular head movements made by squirrel monkeys. There were two goals in this study: (1) to determine the range of angular velocities and accelerations as well as the bandwidth and other frequency characteristics of active head movements and (2) to compare analyses of transients of velocity and acceleration that are studied by residual analysis, Fourier transform, and wavelet transform of the head velocity signal. The residual analysis showed that the shape and duration of the transients affected the bandwidth. During the time after the head had begun to accelerate, the frequency content of the head movement extended into the range of 6 to 12 Hz. When considering all three planes of rotation, approximately 75% of the transients had peak acceleration between 2,000 and 10,000 deg/s(2) and a peak velocity of 50 to 400 deg/s. A peak acceleration of >10,000 deg/s(2) was recorded in 10% of the transients. These findings indicate that active head movements in squirrel monkeys cover a higher range of frequencies, accelerations, and velocities than have typically been used in previous eye-movement and neuronal studies of the reflexes that control gaze. We further conclude that the choice of a method for analyzing transient, time-varying biological signals is dependent on the desired information. Residual analysis provides detailed resolution in the time domain, but estimation of the frequency content of the signal is dependent on the portions selected for analysis and the choice of filters. Fourier transform provides a representation of the power spectrum in the frequency domain but without any inherent temporal resolution. We show that the wavelet transform, a novel method as applied to the signal analysis goals of this study, is the most useful technique for relating time- and frequency-domain information during a continuous signal.

Central versus peripheral origin of vestibuloocular reflex recovery following semicircular canal plugging in rhesus monkeys.

We have previously shown that there is a slowly progressing, frequency-specific recovery of the gain and phase of the horizontal vestibuloocular reflex (VOR) in rhesus monkeys following plugging of the lateral semicircular canals. The adapted VOR response exhibited both dynamic and spatial characteristics that were distinctly different from responses in intact animals. To discriminate between adaptation or recovery of central versus peripheral origin, we have tested the recovered vestibuloocular responses in three rhesus monkeys in which either one or both coplanar pairs of vertical semicircular canals had been plugged previously by occluding the remaining semicircular canals in a second plugging operation. We measured the spatial tuning of the VOR in two or three different mutually orthogonal planes in response to sinusoidal oscillations (1.1 Hz, +/-5 degrees, +/-35 degrees /s) over a period of 2-3 and 12-14 mo after each operation. Apart from a significant recovery of the torsional/vertical VOR following the first operation we found that these recovered responses were preserved following the second operation, whereas the responses from the newly operated semicircular canals disappeared acutely as expected. In the follow-up period of up to 3 mo after the second operation, responses from the last operated canals showed recovery in two of three animals, whereas the previously recovered responses persisted. The results suggest that VOR recovery following plugging may depend on a regained residual sensitivity of the plugged semicircular canals to angular head acceleration.

Cell-based, high-content screen for receptor internalization, recycling and intracellular trafficking.

A variety of physiologically important receptors are internalized and then recycled back to the plasma membrane by the endocytic recycling compartment. These include the transferrin receptor and many G-protein coupled receptors (GPCRs). The internalization of GPCRs is a result of agonist stimulation. A cell-based fluorescent imaging assay is described that detects and quantifies the presence of fluorescently labeled receptors and macromolecules in the recycling compartment. This High Content Screening application is conducted on the ArrayScan II System that includes fluorescent reagents, imaging instrumentation and the informatics tools necessary to screen for compounds that affect receptor internalization, recycling and GPCR activation. We demonstrate the Receptor Internalization and Trafficking application by quantifying (i) the internalization and recycling of the transferrin receptor using a fluorescently labeled ligand and (ii) the internalization of a physiologically functional model GPCR, a GFP-parathyroid hormone receptor chimera. These assays give high signal-to-noise ratios, broad dynamic ranges between stimulated and unstimulated conditions and low variability across different screening runs. Thus, the Receptor Internalization and Trafficking application, in conjunction with the ArrayScan II System, forms the basis of a robust, information-rich and automated screen for GPCR activation.

A branched DNA signal amplification assay to quantitate messenger RNA of human uncoupling proteins 1, 2, and 3.

Uncoupling proteins (UCP) are inner mitochondrial membrane transporters which dissipate the proton gradient, releasing stored energy as heat. Three subtypes of UCP have been identified so far. The regulation of UCP expression is mainly controlled at the transcriptional level, thus making the measurement of UCP mRNA beneficial for both diagnosis and research of weight disorders and diabetes. We have developed an assay using the branched DNA signal amplification assay (bDNA assay) to quantitatively measure the mRNA levels for human UCP1, 2, and 3. UCP-subtype-specific primers were designed for the assay. RNA transcripts of each UCP generated by in vitro transcription were used to validate the specificity and sensitivity of the assay. The quantitative measurement of UCP mRNA was further demonstrated with cultured cells and human tissue. A comprehensive survey of UCP expression from 17 human tissues measured by the newly developed assay is provided. The method described here offers a rapid, sensitive, specific, and quantitative assay for measurement of human UCP mRNA.