Automatic quantitative analysis of experimental primary and secondary retinal neurodegeneration: implications for optic neuropathies.

Secondary neurodegeneration is thought to play an important role in the pathology of neurodegenerative disease, which potential therapies may target. However, the quantitative assessment of the degree of secondary neurodegeneration is difficult. The present study describes a novel algorithm from which estimates of primary and secondary degeneration are computed using well-established rodent models of partial optic nerve transection (pONT) and ocular hypertension (OHT). Brn3-labelled retinal ganglion cells (RGCs) were identified in whole-retinal mounts from which RGC density, nearest neighbour distances and regularity indices were determined. The spatial distribution and rate of RGC loss were assessed and the percentage of primary and secondary degeneration in each non-overlapping segment was calculated. Mean RGC number (82 592±681) and RGC density (1695±23.3 RGC/mm(2)) in naïve eyes were comparable with previous studies, with an average decline in RGC density of 71±17 and 23±5% over the time course of pONT and OHT models, respectively. Spatial analysis revealed greatest RGC loss in the superior and central retina in pONT, but significant RGC loss in the inferior retina from 3 days post model induction. In comparison, there was no significant difference between superior and inferior retina after OHT induction, and RGC loss occurred mainly along the superior/inferior axis (~30%) versus the nasal-temporal axis (~15%). Intriguingly, a significant loss of RGCs was also observed in contralateral eyes in experimental OHT. In conclusion, a novel algorithm to automatically segment Brn3a-labelled retinal whole-mounts into non-overlapping segments is described, which enables automated spatial and temporal segmentation of RGCs, revealing heterogeneity in the spatial distribution of primary and secondary degenerative processes. This method provides an attractive means to rapidly determine the efficacy of neuroprotective therapies with implications for any neurodegenerative disorder affecting the retina.

Multicenter Evaluation of the Xpert Norovirus Assay for Detection of Norovirus Genogroups I and II in Fecal Specimens.

Norovirus is the most common cause of sporadic gastroenteritis and outbreaks worldwide. The rapid identification of norovirus has important implications for infection prevention measures and may reduce the need for additional diagnostic testing. The Xpert Norovirus assay recently received FDA clearance for the detection and differentiation of norovirus genogroups I and II (GI and GII), which account for the vast majority of infections. In this study, we evaluated the performance of the Xpert Norovirus assay with both fresh, prospectively collected (n = 914) and frozen, archived (n = 489) fecal specimens. A Centers for Disease Control and Prevention (CDC) composite reference method was used as the gold standard for comparison. For both prospective and frozen specimens, the Xpert Norovirus assay showed positive percent agreement (PPA) and negative percent agreement (NPA) values of 98.3% and 98.1% for GI and of 99.4% and 98.2% for GII, respectively. Norovirus prevalence in the prospective specimens (collected from March to May of 2014) was 9.9% (n = 90), with the majority of positives caused by genogroup II (82%, n = 74). The positive predictive value (PPV) of the Xpert Norovirus assay was 75% for GI-positive specimens, whereas it was 86.5% for GII-positive specimens. The negative predictive values (NPV) for GI and GII were 100% and 99.9%, respectively.

Inhibition of return in aging and Alzheimer's disease: performance as a function of task demands and stimulus timing.

Inhibition of return (IOR) is a phenomenon of spatial attention that biases attention toward novel events in the environment. Recent evidence suggests that the magnitude and timing of IOR varies as a function of task conditions (e.g., detection vs. discrimination tasks, short vs. long cue-target intervals, intrinsic vs. extrinsic cues). Although IOR appears relatively preserved with both normal aging and Alzheimer's disease (AD), it has been tested under relatively simple task conditions. To test whether IOR is resistant to age and / or AD when cognitive demands are increased, we employed a double-cue IOR paradigm that required categorization as well as detection responses. The stimulus onset asynchrony (SOA) between the cue and target events was varied to determine whether group differences existed in IOR effects over time. Younger normal adults and older normal adults exhibited significant IOR effects on both the detection task and the categorization task at a short cue-target SOA (950 ms). In contrast, AD patients exhibited significant IOR effects at the short SOA on the detection task but not on the categorization task. From the short to the long SOA (3500 ms), IOR effects exhibited by younger normal adults declined significantly during both the detection and the categorization tasks, suggesting that inhibition resolved over time. In contrast, neither older normal adults nor AD patients exhibited SOA-related IOR reductions on the detection task. These results suggest that IOR may show differential age- and AD-related vulnerabilities depending on task conditions and timing characteristics.

Functional neuroimaging of memory: implications for cognitive aging.

Our understanding of the ways in which changes in specific neural systems mediate adult age differences in memory is rapidly increasing, due in no small part to the advent of functional neuroimaging techniques. This article reviews age-related changes in memory performance obtained with behavioral measures, describes models of the neural mechanisms of memory, and derives predictions from these models regarding age-related changes in brain activation patterns. The neuroimaging findings obtained to date support models emphasizing the role of prefrontal cortex in age-related changes in memory functioning, especially for episodic memory retrieval. In general, neural activation associated with episodic memory encoding is regionally similar for younger and older adults but relatively lower in magnitude for older adults. During retrieval, activation that is restricted to the right prefrontal cortex for younger adults is more likely to be bilateral for older adults. Prefrontal activation exhibits an age-related increase when working memory tasks require simple storage and an age-related decrease when working memory requires higher-level executive processes. Although the evidence is limited, behavioral performance and activation patterns appear to be similar among younger and older adults on tests of semantic (context-independent) and implicit memory. We conclude that several methodological issues, such as defining the relation between brain structure and function, and determining the relationship between performance and activation, are particularly important for understanding age-related changes. Future directions for aging research include further investigation of the relation between encoding and retrieval and the identification of both spared and impaired neural systems.

Nongenomic action of progesterone: activation of Xenopus oocyte phospholipase C through a plasma membrane-associated tyrosine kinase.

Using a plasma membrane-cortex preparation (wherein the nucleus and >90% of the total cell protein are removed), progesterone stimulated tyrosine kinase activity that stimulated phospholipase C. Although it has been known for over 20 yr that progesterone acts at the plasma membrane of Xenopus oocytes to induce oocyte maturation, this is the first report that progesterone stimulates this tyrosine kinase activity that is associated with the oocyte plasma membrane and cortex. A tyrosine kinase inhibitor (tyrphostin B46) inhibited steroid stimulation of tyrosine kinase and phospholipase C (PLC) activities, but did not block lipase C stimulation by G protein activators. A fusion protein that contains tandem N- and C-terminal SH2 domains of PLCgamma also blocked progesterone stimulation of PLC (a fusion protein with the SH2 domain from Shc was ineffective). Lowering the Ca2+ concentration in the medium inhibited progesterone, but not guanosine 5'-O-(3-thiotriphosphate), stimulation of PLC, and the effects of progesterone and a G protein agonist were additive. However, neither progesterone nor insulin increased phosphotyrosine on PLCgamma. To evaluate another tyrosine kinase path involving phosphatidylinositol 3-kinase, we added wortmannin to our membrane preparation, but wortmannin did not inhibit progesterone's ability to activate PLC.

Inhibition and habituation: preserved mechanisms of attentional selection in aging and Alzheimer's disease.

Are inhibition and habituation, processes that contribute to selective attention, impaired by aging or Alzheimer's disease (AD)? Younger adults, older adults, and adults with AD read lists of letters presented either alone or paired with distractor letters. Slower reading times for lists containing distractors relative to lists without distractors indexed concurrent interference (distraction). Slower reading times for lists in which distractors subsequently became targets relative to lists in which distractors and targets were unrelated indexed negative priming (inhibition). Faster reading times when distractors were constant in identity or location rather than random indexed repeated distractor effects (habituation). Distraction increased with aging and AD, whereas inhibition and habituation showed no age- or AD-related decline, suggesting that inhibition and habituation still function to aid attentional selection in older adults and adults with AD.

Comparison of UW solution and St. Thomas' solution in the rat: importance of potassium concentration.

BACKGROUND: University of Wisconsin solution (UW) is in limited clinical use for heart transplantation, but there are doubts about its efficacy and concerns about the effect of its high K+ concentration on endothelium. St. Thomas' solution with or without aspartate is widely used and is of proven efficacy. METHODS: Using a modified (starch-free) variant of UW (MUW) we studied: (1) recovery of function with UW compared with aspartate-containing St. Thomas' solution; (2) effect of elevation of K+ in St. Thomas' solution to the level in UW; and (3) effect of reduction of K+ in UW and addition of Ca2+ or aspartate. Isolated rat hearts underwent 7 hours of arrest at 1 degrees C using MUW with or without 20 mmol/L aspartate or using aspartate-containing St. Thomas' solution. RESULTS: Functional recovery with MUW (51.8% +/- 2.5%) was superior to that with aspartate-containing St. Thomas' solution (37.1% +/- 4.3%; p < 0.01). Addition of aspartate to MUW had no effect. During 6 hours of arrest, lowering the K+ in MUW from 125 mmol/L to 20 mmol/L reduced functional recovery from 59.9% +/- 4.2% to 42.3% +/- 4.3% (p < 0.01). The addition of 1 mmol/L Ca2+ had no effect. Elevation of K+ in St. Thomas' solution produced more rapid arrest but no improvement in recovery. CONCLUSIONS: The protective effect of starch-free UW is greater (+13%) than that of aspartate-enriched St. Thomas' solution. Reduction of K+ in UW to lessen possible deleterious effects would decrease its protective effect by about 30% to a level comparable with that of St. Thomas' solution.

Differing protection with aspartate and glutamate cardioplegia in the isolated rat heart.

Aspartate and glutamate each have been shown to improve cardiac recovery after hypoxia or ischemia under normothermic conditions, but whether their effects are additive and to what extent they are modified by hypothermia has not been studied systematically. We set out to compare the individual and combined protective effects of aspartate and glutamate during cardioplegic arrest under normothermic and hypothermic conditions in the rat. Using isolated working rat hearts, functional and metabolic recovery was assessed after 0.5 hours of potassium arrest at 37 degrees C or 5 hours at 2 degrees C in control hearts (C) and in hearts in which 20 mmol/L glutamate (G), 20 mmol/L aspartate (A), or both (A + G) was added to the cardioplegic solution. Under normothermic conditions, percentage recovery of prearrest work (mean +/- standard error of the mean) was as follows: C = 31.7 +/- 2.8, G = 34.8 +/- 0.2, A = 49.6 +/- 2.8*, A + G = 53.7 +/- 2.3*. Under hypothermic conditions, the values were as follows: C = 40.4 +/- 4.0, G = 45.2 +/- 2.3, A = 59.4 +/- 1.8*, A + G = 54.1 +/- 1.2* (*p < 0.01 versus C and G). Recovery of postischemic high-energy phosphate content followed the same pattern: A = A + G > G or C. Measurement of postischemic myocardial content of amino acids showed that recovery of function and energy status correlated with maintenance of myocardial levels of aspartate (r = 0.9; p < 0.01) but not glutamate.(ABSTRACT TRUNCATED AT 250 WORDS)

Nicotine patches in Alzheimer's disease: pilot study on learning, memory, and safety.

In view of the cholinergic deficits present in patients with Alzheimer's disease (AD), a widely investigated treatment strategy for the cognitive deficits in AD is cholinergic stimulation. Although nicotinic cholinergic receptor binding has been demonstrated to be deficient in the AD brain, the predominant theoretical and therapeutic focus to date has been on muscarinic cholinergic receptors and systems. The purpose of the present study was to evaluate the effects of sustained nicotine administration on behavior, cognition, and physiology. A double-blind placebo-controlled trial was conducted in which six patients with probable AD were exposed to 7, 8, and 7 days of placebo, nicotine, and washout, respectively. Daily sessions evaluating learning, memory, and behavior were conducted. Global cognitive functioning, rest and activity levels, cardiac activity, and blood levels were also measured. Findings included improved learning during the nicotine condition, which persisted throughout washout. Memory, behavior, and global cognition were not significantly affected. Sustained administration of nicotine appeared to be safe, although sleep showed a significant decrease.

Targeting periodic oscillations of external cavity laser diodes.

A simple method of targeting periodic dynamics appearing in semiconductor lasers subject to optical feedback is demonstrated. Possible applications of the technique in chaotic communications are identified, and its advantages for chaos control are discussed.

Dynamical and noise properties of laser diodes subject to strong optical feedback.

Dynamical and noise properties of laser diodes subject to strong optical feedback have been investigated. Under strong optical feedback a conventional rate-equation approach is not valid, and thus an iterative traveling-wave model has been developed to describe this operating regime. Attention has been given to identifying the transition from stable strong-feedback operation to the coherence collapse regime obtained with moderate optical feedback.

Monitoring universal precautions: a new assessment tool.

OBJECTIVES: Two pilot studies were conducted to produce efficacy data on an observational tool designed to assess the use of Universal Precautions (UP) in patient care settings. The instrument addresses barrier precautions, hand-washing, handling of sharps, and avoidance of unprotected mouth to mouth resuscitation. DESIGN: The Universal Precautions Assessment Tool was submitted to a panel of 3 experts to establish consensual validity. It was pilot tested by 2 simultaneous observers to establish interrater reliability. SETTING: Pilot Study I was conducted in 3 different units within a 100-bed U.S. Army hospital. Pilot Study II was conducted in the emergency department of a large university-based hospital. PARTICIPANTS: Subjects observed were registered nurses providing acute patient care. RESULTS: Two simultaneous raters calculated UP compliance rates of 76.4% and 78.6%, respectively, for 9 nurses in Pilot Study I, and 62% and 65%, respectively, for 5 nurses in Pilot Study II. The intraclass correlation coefficient for the raters' scores in Pilot Study I was 0.992 with a 95% confidence interval (0.979, 0.997). Consensual validity was established. CONCLUSIONS: The instrument has acceptable interrater reliability under the conditions used. Limitations to use include the possibility of a Hawthorne effect and the fact that assessing proper implementation of UP occasionally relies on a "judgment call" by the observer. With test conditions adjusted to minimize these limitations and with proper consideration of sample size, the tool can be used by researchers and by monitors of hospital quality control to measure UP compliance of caregivers individually or collectively.

The effect of orotic acid on the response of the recently infarcted rat heart to hypothermic cardioplegia.

Patients with a recent myocardial infarction have a higher morbidity and mortality than comparable patients with chronic myocardial ischaemia. We postulated that this might be due to a reduced overall tolerance of the heart to cardioplegic arrest in the presence of a recent infarct. We postulated that orotic acid, a pyrimidine precursor which augments the rate of protein synthesis, might improve the response of the recently infarcted heart to cardioplegic arrest. Myocardial infarction was produced in rats by coronary ligation. The rats were then divided into two groups according to whether they were treated with oral orotic acid (10 mg/kg per day) or untreated. A sham-operated (non-infarcted) group served as normal controls. After 2 days, the hearts (n = 12 per group) underwent 1 h of cardioplegic arrest at 23 degrees C on the isolated working heart apparatus. Before arrest, maximum cardiac function in the untreated infarct group was lower than in the normal group (P less than 0.05), whereas in the treated group, function was similar to the normal group. After arrest there was severe depression of cardiac function in the untreated infarct group: only 57% recovery of the pre-arrest value compared with 86% in the normal group (P less than 0.001). In the orotic acid treated group, recovery (90%) was significantly greater than in the untreated group (P less than 0.001) and equivalent to the normal group. Oxygen utilisation, when corrected for external work, was higher in both infarct groups than in the normal group before and after arrest (P less than 0.05 in both cases). Total uridine nucleotide content of the infarcted and non-infarcted zones of the heart was increased. Treatment with orotic acid produced a further upward trend in uridine nucleotide levels. We conclude that an established, recent infarct reduces the overall tolerance of the heart to hypothermic cardioplegia. Treatment with orotic acid improves the function of the infarcted heart following cardioplegic arrest, and may therefore improve the results of urgent cardiac surgery in patients with myocardial infarction.

Development of an in vitro model of primate cervical goblet cells.

Mucins, are densely packed in secretory granules of goblet cells. Upon exocytosis they undergo massive hydration that results in the formation of the mucus gel. Because the mucin polymer network is held together by tangles and low energy bonds, the rheological properties of this gel are mainly determined by the degree of postexocytotic hydration. Hydration in mucus is governed by a Donnan equilibrium as it is driven by electrostatic interaction among the polyionic charges of the mucins and other fixed polyions. Although, variations of charge density of the mucin polymer could be an efficient physiologic mechanism to control the rate of mucus hydration and rheology, this subject has not been investigated. In here we describe a primary tissue culture system of cervical goblet cells of the monkey uterus. This preparation allows to measure directly the kinetic of hydration of exocytosed mucins. Because the physicochemical parameters of the bathing medium can be effectively controlled, variations in the kinetic of mucins swelling upon exocytosis, can be used as a convenient indicator of fluctuations of charge density in secretory products. Since the cervical mucosa readily respond to endocrine influences, this preparation can provide a useful model to study the effect of hormones or other transmitters on polyionic charge density of secretory product.

In vitro growth of Chlamydia trachomatis in conjunctival and corneal epithelium.

This study used primary cultures of conjunctival and corneal epithelial cells from rabbit, monkey and humans to investigate the infection process and host interactions of chlamydia. The epithelial cells were isolated from bulbar and palpebral conjunctivae and the cornea following incubation with EDTA or dispase and microdissection. The cells were trypsinized and grown in microtest wells or Rose Chambers (gelatin-coated glass substratum). The cell origin of the cultured epithelial cells was verified by immunofluorescence staining with anticytokeratins. The cells were nonreactive when stained with antivimentin, a fibroblast marker. The growth kinetics of the cultured epithelial cells were determined by morphological criteria. Cells reached confluency by 5-7 days, and remained as such for 18 days. The cells were maintained as long as 32 days in primary culture. After the cells had attained confluency (5-7 days), they were infected with ocular strains B/TW-5/OT or C/TW-3/OT C. trachomatis. Chlamydial inclusions were identified by light and electron microscopy. Direct FITC C. trachomatis staining (species-specific monoclonal antibody) of the inclusions was used to determine the infectivity of the various types of epithelial cells. The dynamics of the infection process was documented by time lapse photomicroscopy. Small inclusions were identified by 18 hr postinoculation (pi). By 48 hr pi, the inclusions had dramatically increased in size, occupying much of the host perikarya. The reticulate bodies were very active at this time. By 70 to 100 hr pi, the inclusions contained the highly infectious elementary bodies. At this time, rupture of the inclusion and lysis of the infected cells usually occurred.(ABSTRACT TRUNCATED AT 250 WORDS)