Prevalence, time course, and visual impact of peripapillary hyperreflective ovoid masslike structures (PHOMS) in pediatric patients with optic nerve pathologies.

BACKGROUND: Peripapillary hyperreflective ovoid masslike structures (PHOMS) are a recently defined optical coherence tomography (OCT) finding. The purpose of this study was to characterize the presence of PHOMS and their visual significance in pediatric patients with and without optic nerve pathologies. METHODS: This retrospective study evaluated 400 patients (<18 years of age) including normal control subjects and patients with optic neuritis, papillitis, optic nerve head drusen (ONHD), and papilledema. Information on demographics, visual function, and structural parameters were obtained. RESULTS: PHOMS were found in 7 of 2,582 of normal control eyes (7%), 9 of 59 eyes with optic neuritis (15.3%), 58 of 76 eyes with ONHD (76.3%), 3 of 11 eyes with papillitis (27.3%), and 180 of 308 eyes with papilledema (58.4%). PHOMS were more prevalent in the papilledema (P < 0.001), ONHD (P < 0.001), and optic neuritis (P = 0.028) eyes than in control eyes. We identified 5 cases where PHOMS developed de novo. This occurred over an average of 2.3 years (range, 0.2-7.4 years). Sixteen cases of PHOMS resolved over an average of 1.1 years (range, 0.3-4.0 years). Cross-sectionally, PHOMS were not associated with visual acuity (P = 0.551), retinal nerve fiber layer thickness (P = 0.068), ganglion cell volume (P = 0.375), or visual field mean deviation (P = 0.795). CONCLUSIONS: PHOMS are present in a majority of children with papilledema or ONHD. PHOMS are dynamic and may form de novo over time with optic nerve pathology and may resolve either through treatment or atrophy. There was no relationship between the presence of PHOMS and poor visual function in our study cohort.

Identifying thresholds in air exposure, water temperature and fish size that determine reflex impairment in brook trout exposed to catch-and-release angling.

Understanding the factors that contribute to fish impairment and survival from angling events is essential to guide best angling practices for catch-and-release (C&R) recreational fisheries. Complex interactions often exist between angler behaviour, environmental conditions, and fish characteristics that ultimately determine biological outcomes for fish. Yet, few studies focus on identifying biologically relevant thresholds. We therefore examined the effects of water temperature, air exposure and fish size on reflex impairment and mortality in brook trout Salvelinus fontinalis exposed to experimental and simulated angling stressors (n = 337). Using conditional inference trees, we identified interactions among these factors as well as threshold values within them that determine brook trout reflex impairment as an indicator of whole animal stress. Specifically, longer air exposure times (>30 sec) and warmer temperatures (>19.5°C) had a synergistic effect leading to higher reflex impairment scores. Further, larger fish (>328 mm) were more sensitive to air exposure durations >10 sec. Of the reflex impairment measures, loss of equilibrium and time to regain equilibrium were strongly and moderately associated with brook trout mortality (18-24 h monitoring), although mortality rates were generally low (6%). These findings support previous research that has established strong links between these reflex impairment measures and fish health outcomes in other species. They also highlight the important interactions among air exposure duration, water temperature and fish size that determine impairment in brook trout, providing specific thresholds to guide best angling practices for C&R fisheries. This approach may be widely applicable to generate similar thresholds that can be encouraged by regulators and adopted by anglers for other common C&R fishes.

An overview of peripapillary hyperreflective ovoid mass-like structures.

PURPOSE OF REVIEW: The purpose of this review is to provide an overview of the ophthalmic findings associated with peripapillary hyperreflective ovoid mass-like structures (PHOMS) in both adult and pediatric patients. RECENT FINDINGS: PHOMS have recently been identified in a number of different ophthalmic disease entities ranging from nonpathologic to pathologic, including but not limited to anatomic abnormalities (tilting in myopia), optic nerve head drusen, optic disc edema from inflammation (optic neuritis, white dot syndromes), vascular insults (ischemic optic neuropathy, retinal vascular occlusion), and papilledema. The mechanism underlying the formation of PHOMS has not been fully elucidated although it has been hypothesized that PHOMS occur secondary to axoplasmic stasis from crowding at the optic nerve head. SUMMARY: Although the clinical significance of the presence of PHOMS remains unclear, PHOMS are associated with several disease processes. Understanding the mechanism behind their formation and their impact on optic nerve head structure and visual function may be relevant in patients with optic nerve head pathology. The presence of PHOMS may also correlate with disease severity and duration. Future studies to evaluate whether the formation of PHOMS may be useful as an early indicator of disease or a prognostic tool are warranted.

Host-pathogen-environment interactions predict survival outcomes of adult sockeye salmon (Oncorhynchus nerka) released from fisheries.

Incorporating host-pathogen(s)-environment axes into management and conservation planning is critical to preserving species in a warming climate. However, the role pathogens play in host stress resilience remains largely unexplored in wild animal populations. We experimentally characterized how independent and cumulative stressors (fisheries handling, high water temperature) and natural infections affected the health and longevity of released wild adult sockeye salmon (Oncorhynchus nerka) in British Columbia, Canada. Returning adults were collected before and after entering the Fraser River, yielding marine- and river-collected groups, respectively (N = 185). Fish were exposed to a mild (seine) or severe (gill net) fishery treatment at collection, and then held in flow-through freshwater tanks for up to four weeks at historical (14°C) or projected migration temperatures (18°C). Using weekly nonlethal gill biopsies and high-throughput qPCR, we quantified loads of up to 46 pathogens with host stress and immune gene expression. Marine-collected fish had less severe infections than river-collected fish, a short migration distance (100 km, 5-7 days) that produced profound infection differences. At 14°C, river-collected fish survived 1-2 weeks less than marine-collected fish. All fish held at 18°C died within 4 weeks unless they experienced minimal handling. Gene expression correlated with infections in river-collected fish, while marine-collected fish were more stressor-responsive. Cumulative stressors were detrimental regardless of infections or collection location, probably due to extreme physiological disturbance. Because river-derived infections correlated with single stressor responses, river entry probably decreases stressor resilience of adult salmon by altering both physiology and pathogen burdens, which redirect host responses toward disease resistance.

Conservation physiology of animal migration.

Migration is a widespread phenomenon among many taxa. This complex behaviour enables animals to exploit many temporally productive and spatially discrete habitats to accrue various fitness benefits (e.g. growth, reproduction, predator avoidance). Human activities and global environmental change represent potential threats to migrating animals (from individuals to species), and research is underway to understand mechanisms that control migration and how migration responds to modern challenges. Focusing on behavioural and physiological aspects of migration can help to provide better understanding, management and conservation of migratory populations. Here, we highlight different physiological, behavioural and biomechanical aspects of animal migration that will help us to understand how migratory animals interact with current and future anthropogenic threats. We are in the early stages of a changing planet, and our understanding of how physiology is linked to the persistence of migratory animals is still developing; therefore, we regard the following questions as being central to the conservation physiology of animal migrations. Will climate change influence the energetic costs of migration? Will shifting temperatures change the annual clocks of migrating animals? Will anthropogenic influences have an effect on orientation during migration? Will increased anthropogenic alteration of migration stopover sites/migration corridors affect the stress physiology of migrating animals? Can physiological knowledge be used to identify strategies for facilitating the movement of animals? Our synthesis reveals that given the inherent challenges of migration, additional stressors derived from altered environments (e.g. climate change, physical habitat alteration, light pollution) or interaction with human infrastructure (e.g. wind or hydrokinetic turbines, dams) or activities (e.g. fisheries) could lead to long-term changes to migratory phenotypes. However, uncertainty remains because of the complexity of biological systems, the inherently dynamic nature of the environment and the scale at which many migrations occur and associated threats operate, necessitating improved integration of physiological approaches to the conservation of migratory animals.

Watershed-Scale Land Use Activities Influence the Physiological Condition of Stream Fish.

Land use changes within watersheds can have large effects on stream ecosystems, but the mechanistic basis of those effects remains poorly understood. While changes to population size presumably reflect underlying variation in organismal health and condition, such individual-level metrics are rarely evaluated in the context of ecosystem disturbance. To address this deficiency, we combined physiological sampling with geographic information systems to quantify the effects of land use on physiological indicators of health in largemouth bass. More specifically, we first quantified blood metrics relating to nutrition, oxidative stress, and the glucocorticoid stress response from largemouth bass residing in eight watersheds. We then used Akaike's information criterion to define relationships between these blood metrics and land cover, including forests, agricultural areas, urban areas, and wetlands. The proportion of forest cover in a watershed was the best predictor of blood metrics representing recent feeding and resistance to oxidative stress, whereas the proportion of wetlands was the best predictor of glucocorticoid function; however, further investigation is needed, as the explanatory power of the models was relatively low. Patterns in energy reserves were not influenced by any land use practices. Interestingly, anthropogenic land use categories, such as urban and agricultural areas, were not the best predictor for any blood metrics. Together, our results indicate that fish health is most related to natural features of a landscape rather than anthropogenic land uses. Furthermore, these findings suggest that physiological methods could supplement traditional population and community assessments to develop a more comprehensive understanding of ecosystem interactions and improve stream management.

Stress in the neighborhood: Tissue glucocorticoids relative to stream quality for five species of fish.

Anthropogenic alterations to terrestrial habitat (e.g., urbanization, deforestation, agriculture) can have a variety of negative effects on watercourses that flow through disturbed landscapes. Currently, the relationship between stream habitat quality and fish condition remains poorly understood. The use of physiological metrics such as glucocorticoids (GCs) provides a useful tool for quantifying these effects by relating the health of resident fishes to stream quality. To date, however, most studies that measure GC levels tend to focus on a single, large-bodied species, rather than evaluating how GCs may be influenced differently between species in a community. In this study, we measured cortisol, the glucocorticoid found in fishes, from fish tissues to quantify effects of habitat degradation on the glucocorticoid function of five species of juvenile and small-bodied stream fish which differ ecologically and phylogenetically. Largemouth bass Micropterus salmoides, brown bullhead Ameiurus nebulosus, white sucker Catostomus commersonii, pumpkinseed Lepomis gibbosus, and logperch Percina caprodes were sampled from a reference and a degraded stream. Upon capture, fish were either euthanized immediately, to quantify baseline stress parameters, or following a standardized stressor, to quantify GC responsiveness. As a result of stream degradation largemouth bass possessed altered baseline GC concentrations and brown bullhead and logperch had altered GC responses to a stressor. White sucker and pumpkinseed did not demonstrate any alteration in baseline or post-stress GC concentrations. Together, our results show that different species residing in identical habitats can demonstrate a variety of responses to environmental stress, highlighting the variation in physiological ability to cope under poor environmental conditions, as well as the difficulty of predicting GC dynamics in wild animals. Understanding the relationships between GC function, habitat quality, and population-level processes will increase the ability of researchers and managers to predict how fish communities and aquatic ecosystems will be shaped by anthropogenic environmental change.

Linking Landscape-Scale Disturbances to Stress and Condition of Fish: Implications for Restoration and Conservation.

Humans have dramatically altered landscapes as a result of urban and agricultural development, which has led to decreases in the quality and quantity of habitats for animals. This is particularly the case for freshwater fish that reside in fluvial systems, given that changes to adjacent lands have direct impacts on the structure and function of watersheds. Because choices of habitat have physiological consequences for organisms, animals that occupy sub-optimal habitats may experience increased expenditure of energy or homeostatic overload that can cause negative outcomes for individuals and populations. With the imperiled and threatened status of many freshwater fish, there is a critical need to define relationships between land use, quality of the habitat, and physiological performance for resident fish as an aid to restoration and management. Here, we synthesize existing literature to relate variation in land use at the scale of watersheds to the physiological status of resident fish. This examination revealed that landscape-level disturbances can influence a host of physiological properties of resident fishes, ranging from cellular and genomic levels to the hormonal and whole-animal levels. More importantly, these physiological responses have been integrated into traditional field-based monitoring protocols to provide a mechanistic understanding of how organisms interact with their environment, and to enhance restoration. We also generated a conceptual model that provides a basis for relating landscape-level changes to physiological responses in fish. We conclude that physiological sampling of resident fish has the potential to assess the effects of landscape-scale disturbances on freshwater fish and to enhance restoration and conservation.

Clear as mud: a meta-analysis on the effects of sedimentation on freshwater fish and the effectiveness of sediment-control measures.

Increase in fine sediments in freshwater resulting from anthropogenic development is a potential stressor for fish and thus may cause population declines. Though a large body of literature exists on the topic, there have been few attempts to synthesize this information in a quantitative manner. Through meta-analysis we investigated the effects of sediment in lotic environments on resident ichthyofauna using ecologically-relevant endpoints for tolerant (e.g., northern pike Esox lucius) and intolerant (e.g., brook trout Salvelinus fontinalis) species. Further, the efficiency of sediment-control devices was explored to inform mitigation measures. An increase in suspended and deposited sediments was demonstrated to have a negative effect on all parameters and tolerances tested (feeding behavior [feeding rate, reaction distance to food item]; spawning success [survival of fry to eyed stage, fry emergence]; species richness; P < 0.001) except fish abundance (P = 0.058). Heterogeneity between studies was a factor in all analyses. Although there were insufficient studies to conduct meta-analysis on sediment-control devices, weighted percent efficiency estimates revealed that properly installed sediment-control fences tended to have a higher percent efficiency (73-80%) than sediment traps and basins (40-52%). These results highlight the negative impact that increases in suspended and deposited sediments can have on resident fishes from the individual to the population, and the need for more transparent and thorough statistical reporting. The analysis also identifies a clear need for rigorous experimental studies contrasting different sediment-control devices and strategies given that little such work has been published. That alone is remarkable given that sediment-control devices are often a requirement of regulators for riparian development activities, yet the evidence to support the effectiveness of the primary mitigative strategies is weak.

P-glycoprotein antagonists confer synergistic sensitivity to short-chain ceramide in human multidrug-resistant cancer cells.

P-glycoprotein (P-gp) antagonists inhibit ceramide metabolism at the juncture of glycosylation. The purpose of this study was to test whether targeting P-gp would be a viable alternative to targeting glucosylceramide synthase (GCS) for enhancing ceramide cytotoxicity. A2780 wild-type, and multidrug-resistant 2780AD and NCI/ADR-RES human ovarian cancer cell lines and the cell-permeable ceramide analog, C6-ceramide (C6-cer), were employed. Compared to P-gp-poor A2780 cells, P-gp-rich 2780AD cells converted 3.7-fold more C6-cer to nontoxic C6-glucosylceramide (C6-GC), whereas cell-free GCS activities were equal. 2780AD cells displayed resistance to C6-cer (10 µM) that was reversed by inclusion of the P-gp antagonist tamoxifen (5 µM) but not by inclusion of a GCS inhibitor. Co-administration of C6-cer and P-gp antagonists was also effective in NCI/ADR-RES cells. For example, C6-cer, VX-710 (Biricodar), and cyclosporin A (cyc A) exposure resulted in viabilities of ~90% of control; however, C6-cer/VX-710 and C6-cer/cyc A additions were synergistic and resulted in viabilities of 22% and 17%, respectively. Further, whereas C6-ceramide and cyc A imparted 1.5- and 0-fold increases in caspase 3/7 activity, the combination produced a 3.5-fold increase. Although the upstream elements of cell death have not been elucidated, the novel C6-ceramide/P-gp antagonist combination merits further study and assessment of clinical translational potential.

Expression of P-glycoprotein in HeLa cells confers resistance to ceramide cytotoxicity.

The role of glucosylceramide synthase (GCS) in regulating ceramide-induced apoptosis has been widely studied. The purpose of this investigation was to evaluate the role of P-glycoprotein (P-gp) in regulating ceramide cytotoxicity by using C6-ceramide. To accomplish this, we employed HeLa cells with conditional expression of the multidrug resistance gene 1/P-gp. HeLa cells expressing P-gp (P-gp/on cells) challenged with [14C]C6-ceramide (6 µM), synthesized 4.5-fold the amount of C6-glucosylceramide (GC) compared to HeLa cells with suppressed expression of P-gp (P-gp/off cells), whereas the generated levels of C6-sphingomyelin were almost equal (33 and 29% of intracellular 14C, respectively). Tamoxifen, a P-gp antagonist, decreased the C6-GC levels from 3.5-1.0% in the P-gp/off and from 17-2.8% of the total lipid 14C levels in the P-gp/on cells. Tamoxifen did not inhibit cell-free C6-GC synthesis in the P-gp/off or P-gp/on homogenates. However, a specific GCS inhibitor, ethylenedioxy-1-phenyl-2-hexadecanoylamino-3-pyrrolidino-1-propanol (ethylenedioxy-P4), blocked synthesis by 90%. In the cytotoxicity assays, the P-gp/off cells were sensitive to C6-ceramide and the P-gp/on cells were resistant. Resistance to C6-ceramide in the P-gp/on cells was reversed by tamoxifen but not by ethylenedioxy-P4. Experiments in another cervical cancer model showed that multidrug-resistant P-gp-rich KB-V1 cells synthesized 3-fold more C6-GC from C6-ceramide than the parental, P-gp-poor KB-3-1 cells, and whereas tamoxifen had no effect on the C6-GC synthesis in the KB-3-1 cells, it inhibited synthesis by 70% in the KB-V1 cells. This study demonstrates that P-gp potentiates C6-ceramide glycosylation and if antagonized augments C6-ceramide sensitivity, both features previously ascribed to GCS. We propose that P-gp can be an effective target for enhancing short-chain ceramide cytotoxicity in the treatment of drug-resistant cancer.

Metabolism of short-chain ceramide by human cancer cells--implications for therapeutic approaches.

Due to recent use of short-chain ceramides in preclinical studies, we characterized C6-ceramide metabolism in cancer cell lines and assessed metabolic junctures for enhancing efficacy. MDA-MB-231 breast cancer cells decreased the amount of C6-ceramide metabolized to C6-sphingomyelin (C6-SM) and increased the amount metabolized to C6-glucosylceramide (C6-GC) in response to increasing concentrations. A similar trend was seen in DU-145 (prostate cancer), PANC-1 (pancreatic cancer), and LoVo (colorectal cancer) cells. KG-1 leukemia cells favored C6-SM synthesis at low (0.6muM) and high-dose (12muM) C6-ceramide. Partnering C6-ceramide with tamoxifen, a P-glycoprotein antagonist that impedes ceramide glycosylation, was an effective regimen for enhancing cytotoxicity in cells. Experiments to assess the mechanism of cell death using KG-1 cells showed that tamoxifen inhibited synthesis of C6-GC and C6-SM from C6-ceramide by 80% and 50%, respectively, which was accompanied by enhanced apoptosis. Radiolabeling of KG-1 cells with [(3)H]palmitic acid produced a 2-fold increase in (3)H-long-chain ceramides when unlabeled C6-ceramide was added and a 9-fold increase when C6-ceramide and tamoxifen were added. The increase in (3)H-palmitate radiolabeling of long-chain ceramides was blocked by inclusion of a ceramide synthase inhibitor; however, inhibiting synthesis of long-chain ceramide did not rescue cells. These studies show that tamoxifen enhances the apoptotic effects of C6-ceramide. The proposed mechanism involves blocking short-chain ceramide anabolism to favor hydrolysis and generation of sphingosine. We propose that use of tamoxifen and other P-glycoprotein antagonists can be an effective means for enhancing cytotoxic potential of short-chain ceramides in the treatment of cancer.

Full pharmacological efficacy of a novel S1P1 agonist that does not require S1P-like headgroup interactions.

Strong evidence exists for interactions of zwitterionic phosphate and amine groups in sphingosine-1 phosphate (S1P) to conserved Arg and Glu residues present at the extracellular face of the third transmembrane domain of S1P receptors. The contribution of Arg(120) and Glu(121) for high-affinity ligand-receptor interactions is essential, because single-point R(120)A or E(121)A S1P(1) mutants neither bind S1P nor transduce S1P function. Because S1P receptors are therapeutically interesting, identifying potent selective agonists with different binding modes and in vivo efficacy is of pharmacological importance. Here we describe a modestly water-soluble highly selective S1P(1) agonist [2-(4-(5-(3,4-diethoxyphenyl)-1,2,4-oxadiazol-3-yl)-2,3-dihydro-1H-inden-1-yl amino) ethanol (CYM-5442)] that does not require Arg(120) or Glu(121) residues for activating S1P(1)-dependent p42/p44 mitogen-activated protein kinase phosphorylation, which defines a new hydrophobic pocket in S1P(1). CYM-5442 is a full agonist in vitro for S1P(1) internalization, phosphorylation, and ubiquitination. It is noteworthy that CYM-5442 was a full agonist for induction and maintenance of S1P(1)-dependent blood lymphopenia, decreasing B lymphocytes by 65% and T lymphocytes by 85% of vehicle. Induction of CYM-5442 lymphopenia was dose- and time-dependent, requiring serum concentrations in the 50 nM range. In vitro measures of S1P(1) activation by CYM-5442 were noncompetitively inhibited by a specific S1P(1) antagonist [(R)-3-amino-(3-hexylphenylamino)-4-oxobutylphosphonic acid (W146)], competitive for S1P, 2-amino-2-(4-octylphenethyl)propane-1,3-diol (FTY720-P), and 5-[4-phenyl-5-(trifluoromethyl)-2-thienyl]-3-[3-(trifluoromethyl)phenyl]-1,2, 4-oxadiazole (SEW2871). In addition, lymphopenia induced by CYM-5442 was reversed by W146 administration or upon pharmacokinetic agonist clearance. Pharmacokinetics in mice also indicated that CYM-5442 partitions significantly in central nervous tissue. These data show that CYM-5442 activates S1P(1)-dependent pathways in vitro and to levels of full efficacy in vivo through a hydrophobic pocket separate from the orthosteric site of S1P binding that is headgroup-dependent.

Ligand-binding pocket shape differences between sphingosine 1-phosphate (S1P) receptors S1P1 and S1P3 determine efficiency of chemical probe identification by ultrahigh-throughput screening.

We have studied the sphingosine 1-phosphate (S1P) receptor system to better understand why certain molecular targets within a closely related family are much more tractable when identifying compelling chemical leads. Five medically important G-protein-coupled receptors for S1P regulate heart rate, coronary artery caliber, endothelial barrier integrity, and lymphocyte trafficking. Selective S1P receptor agonist probes would be of great utility to study receptor subtype-specific function. Through systematic screening of the same libraries, we identified novel selective agonist chemotypes for each of the S1P1 and S1P3 receptors. Ultrahigh-throughput screening (uHTS) for S1P1 was more effective than that for S1P3, with many selective, low nanomolar hits of proven mechanism emerging. Receptor structure modeling and ligand docking reveal differences between the receptor binding pockets, which are the basis for subtype selectivity. Novel selective agonists interact primarily in the hydrophobic pocket of the receptor in the absence of headgroup interactions. Chemistry-space and shape-based analysis of the screening libraries in combination with the binding models explain the observed differential hit rates and enhanced efficiency for lead discovery for S1P1 versus S1P3 in this closely related receptor family.

Turnover rate of the gamma-aminobutyric acid transporter GAT1.

We combined electrophysiological and freeze-fracture methods to estimate the unitary turnover rate of the gamma-aminobutyric acid (GABA) transporter GAT1. Human GAT1 was expressed in Xenopus laevis oocytes, and individual cells were used to measure and correlate the macroscopic rate of GABA transport and the total number of transporters in the plasma membrane. The two-electrode voltage-clamp method was used to measure the transporter-mediated macroscopic current evoked by GABA (I(NaCl)(GABA)), macroscopic charge movements (Q (NaCl)) evoked by voltage pulses and whole-cell capacitance. The same cells were then examined by freeze-fracture and electron microscopy in order to estimate the total number of GAT1 copies in the plasma membrane. GAT1 expression in the plasma membrane led to the appearance of a distinct population of 9-nm freeze-fracture particles which represented GAT1 dimers. There was a direct correlation between Q (NaCl) and the total number of transporters in the plasma membrane. This relationship yielded an apparent valence of 8 +/- 1 elementary charges per GAT1 particle. Assuming that the monomer is the functional unit, we obtained 4 +/- 1 elementary charges per GAT1 monomer. This information and the relationship between I(NaCl)(GABA) and Q (NaCl) were used to estimate a GAT1 unitary turnover rate of 15 +/- 2 s(-1) (21 degrees C, -50 mV). The temperature and voltage dependence of GAT1 were used to estimate the physiological turnover rate to be 79-93 s(-1) (37 degrees C, -50 to -90 mV).