NRF1 association with AUTS2-Polycomb mediates specific gene activation in the brain.

The heterogeneous family of complexes comprising Polycomb repressive complex 1 (PRC1) is instrumental for establishing facultative heterochromatin that is repressive to transcription. However, two PRC1 species, ncPRC1.3 and ncPRC1.5, are known to comprise novel components, AUTS2, P300, and CK2, that convert this repressive function to that of transcription activation. Here, we report that individuals harboring mutations in the HX repeat domain of AUTS2 exhibit defects in AUTS2 and P300 interaction as well as a developmental disorder reflective of Rubinstein-Taybi syndrome, which is mainly associated with a heterozygous pathogenic variant in CREBBP/EP300. Moreover, the absence of AUTS2 or mutation in its HX repeat domain gives rise to misregulation of a subset of developmental genes and curtails motor neuron differentiation of mouse embryonic stem cells. The transcription factor nuclear respiratory factor 1 (NRF1) has a novel and integral role in this neurodevelopmental process, being required for ncPRC1.3 recruitment to chromatin.

PRC2 is high maintenance.

As the process that silences gene expression ensues during development, the stage is set for the activity of Polycomb-repressive complex 2 (PRC2) to maintain these repressed gene profiles. PRC2 catalyzes a specific histone posttranslational modification (hPTM) that fosters chromatin compaction. PRC2 also facilitates the inheritance of this hPTM through its self-contained "write and read" activities, key to preserving cellular identity during cell division. As these changes in gene expression occur without changes in DNA sequence and are inherited, the process is epigenetic in scope. Mutants of mammalian PRC2 or of its histone substrate contribute to the cancer process and other diseases, and research into these aberrant pathways is yielding viable candidates for therapeutic targeting. The effectiveness of PRC2 hinges on its being recruited to the proper chromatin sites; however, resolving the determinants to this process in the mammalian case was not straightforward and thus piqued the interest of many in the field. Here, we chronicle the latest advances toward exposing mammalian PRC2 and its high maintenance.

Automethylation of PRC2 promotes H3K27 methylation and is impaired in H3K27M pediatric glioma.

The histone methyltransferase activity of PRC2 is central to the formation of H3K27me3-decorated facultative heterochromatin and gene silencing. In addition, PRC2 has been shown to automethylate its core subunits, EZH1/EZH2 and SUZ12. Here, we identify the lysine residues at which EZH1/EZH2 are automethylated with EZH2-K510 and EZH2-K514 being the major such sites in vivo. Automethylated EZH2/PRC2 exhibits a higher level of histone methyltransferase activity and is required for attaining proper cellular levels of H3K27me3. While occurring independently of PRC2 recruitment to chromatin, automethylation promotes PRC2 accessibility to the histone H3 tail. Intriguingly, EZH2 automethylation is significantly reduced in diffuse intrinsic pontine glioma (DIPG) cells that carry a lysine-to-methionine substitution in histone H3 (H3K27M), but not in cells that carry either EZH2 or EED mutants that abrogate PRC2 allosteric activation, indicating that H3K27M impairs the intrinsic activity of PRC2. Our study demonstrates a PRC2 self-regulatory mechanism through its EZH1/2-mediated automethylation activity.

Cortical profiles of numerous psychiatric disorders and normal development share a common pattern.

The neurobiological bases of the association between development and psychopathology remain poorly understood. Here, we identify a shared spatial pattern of cortical thickness (CT) in normative development and several psychiatric and neurological disorders. Principal component analysis (PCA) was applied to CT of 68 regions in the Desikan-Killiany atlas derived from three large-scale datasets comprising a total of 41,075 neurotypical participants. PCA produced a spatially broad first principal component (PC1) that was reproducible across datasets. Then PC1 derived from healthy adult participants was compared to the pattern of CT differences associated with psychiatric and neurological disorders comprising a total of 14,886 cases and 20,962 controls from seven ENIGMA disease-related working groups, normative maturation and aging comprising a total of 17,697 scans from the ABCD Study® and the IMAGEN developmental study, and 17,075 participants from the ENIGMA Lifespan working group, as well as gene expression maps from the Allen Human Brain Atlas. Results revealed substantial spatial correspondences between PC1 and widespread lower CT observed in numerous psychiatric disorders. Moreover, the PC1 pattern was also correlated with the spatial pattern of normative maturation and aging. The transcriptional analysis identified a set of genes including KCNA2, KCNS1 and KCNS2 with expression patterns closely related to the spatial pattern of PC1. The gene category enrichment analysis indicated that the transcriptional correlations of PC1 were enriched to multiple gene ontology categories and were specifically over-represented starting at late childhood, coinciding with the onset of significant cortical maturation and emergence of psychopathology during the prepubertal-to-pubertal transition. Collectively, the present study reports a reproducible latent pattern of CT that captures interregional profiles of cortical changes in both normative brain maturation and a spectrum of psychiatric disorders. The pubertal timing of the expression of PC1-related genes implicates disrupted neurodevelopment in the pathogenesis of the spectrum of psychiatric diseases emerging during adolescence.

Expression analysis of Carassius auratus-leukocyte-immune-type receptors (CaLITRs) during goldfish kidney macrophage development and in activated kidney leukocyte cultures.

Carassius auratus leukocyte immune-type receptors (CaLITRs) were recently discovered immunoregulatory receptors in goldfish that have diverse immunoglobulin-like (Ig-like) ectodomains and intracellular signaling motifs. Genomic analysis shows that CaLITR-types are also located as distinct gene clusters across multiple goldfish chromosomes. For example, CaLITR1 (unplaced) is a functionally ambiguous receptor having two Ig-like domains, a transmembrane domain (TM), and a short cytoplasmic tail (CYT) devoid of any recognizable signaling motifs. CaLITR2 (Chr47) is a putative inhibitory receptor containing four Ig-like domains, a TM, and a long CYT with an immunoreceptor tyrosine-based inhibition motif (ITIM) and immunoreceptor tyrosine-based switch motif (ITSM). A putative activating receptor-type, CaLITR3 (Chr3), has four Ig-like domains, a TM, and a short CYT containing a positively charged histidine residue and CaLITR4 (ChrLG28B) is a receptor with putative multifunctional signaling potential as well as five Ig-like domains, a TM, and a long tyrosine-motif containing CYT region. The variable genomic locations of the CaLITRs suggest that they are likely under the influence of different cis- and/or trans-regulatory elements. To better understand the transcriptional activities of select CaLITRs from variable genomic regions, we used an RT-qPCR-based approach to examine the expression of CaLITR1, CaLITR2, CaLITR3, and CaLITR4 during goldfish primary kidney macrophage (PKM) development and in mixed leukocyte reaction cultures (MLRs) of the goldfish. Our results showed that the select CaLITRs are differentially expressed during PKM development and in goldfish MLRs exposed to T-cell mitogens/immunosuppressive drugs, supporting that the transcription of these CaLITRs is likely regulated by distinct cis- and/or trans-regulatory elements.

Abiotic and biotic constituents of oil sands process-affected waters.

The oil sands in Northern Alberta are the largest oil sands in the world, providing an important economic resource for the Canadian energy industry. The extraction of petroleum in the oil sands begins with the addition of hot water to the bituminous sediment, generating oil sands process-affected water (OSPW), which is acutely toxic to organisms. Trillions of litres of OSPW are stored on oil sands mining leased sites in man-made reservoirs called tailings ponds. As the volume of OSPW increases, concerns arise regarding the reclamation and eventual release of this water back into the environment. OSPW is composed of a complex and heterogeneous mix of components that vary based on factors such as company extraction techniques, age of the water, location, and bitumen ore quality. Therefore, the effective remediation of OSPW requires the consideration of abiotic and biotic constituents within it to understand short and long term effects of treatments used. This review summarizes selected chemicals and organisms in these waters and their interactions to provide a holistic perspective on the physiochemical and microbial dynamics underpinning OSPW .

Using immune cell-based bioactivity assays to compare the inflammatory activities of oil sands process-affected waters from a pilot scale demonstration pit lake.

In this study, we provide evidence that oil sands process-affected waters (OSPW) contain factors that activate the antimicrobial and proinflammatory responses of immune cells. Specifically, using the murine macrophage RAW 264.7 cell line, we establish the bioactivity of two different OSPW samples and their isolated fractions. Here, we directly compared the bioactivity of two pilot scale demonstration pit lake (DPL) water samples, which included expressed water from treated tailings (termed the before water capping sample; BWC) as well as an after water capping (AWC) sample consisting of a mixture of expressed water, precipitation, upland runoff, coagulated OSPW and added freshwater. Significant inflammatory (i.e. macrophage activating) bioactivity was associated with the AWC sample and its organic fraction (OF), whereas the BWC sample had reduced bioactivity that was primarily associated with its inorganic fraction (IF). Overall, these results indicate that at non-toxic exposure doses, the RAW 264.7 cell line serves as an acute, sensitive and reliable biosensor for the screening of inflammatory constituents within and among discrete OSPW samples.

Developmental differences across the lifespan in the use of perceptual information to guide action-based decisions.

Perceptual information about unfolding events is important for guiding decisions about when and how to move in real-world action situations. As an exemplary case, road-crossing is a perceptual-motor task where age has been shown to be a strong predictor of risk due to errors in action-based decisions. The present study investigated age differences between three age groups (Children: 10-12 years old; Adults: 19-39 years old; Older Adults: 65 + year olds) in the use of perceptual information for selection, timing, and control of action when crossing a two-way street in an immersive, interactive virtual reality environment. Adults and children selected gaps to cross that were consistent with the use of a time-based information variable (tau), whereas older adults tuned less into the time-based variable (tau) to guide road-crossing decisions. For action initiation and control, children and adults also showed a strong ability to precisely time their entry with respect to the lead vehicle maximising the available time to cross and coordinating walking movements with the tail vehicle to ensure they were not on a collision course. In contrast, older adults delayed action initiation and showed difficulty coordinating self-movement with the approaching vehicle. This study and its results tie together age-based differences in the three components of action decision-making (selection, timing and control) within a unified framework based on perceptual information. The implications of these age-related differences in action decisions and crossing behaviours are discussed in the context of road safety.

Identification of distinct LRC- and Fc receptor complex-like chromosomal regions in fish supports that teleost leukocyte immune-type receptors are distant relatives of mammalian Fc receptor-like molecules.

Leukocyte immune-type receptors (LITRs) are a large family of immunoregulatory receptor-types originally identified in the channel catfish (Ictalurus punctatus (Ip)LITRs). Phylogenetic analyses of LITRs show that they share distant evolutionary relationships with important mammalian immunoregulatory receptors belonging to the Fc receptors family and the leukocyte receptor complex (LRC), but their syntenic relationships with these immunoglobulin superfamily members have not been investigated. To further examine the possible evolutionary connections between teleost LITRs and various mammalian immunoregulatory receptor-types, we surveyed the genomic databases of representative vertebrate taxa and our results show that teleost LITRs generally exist in large genomic clusters, which are linked to vangl2, arhgef11, and slam family genes, features that are also shared by amphibian and mammalian Fc receptor-like molecules (FCRLs). Moreover, detailed phylogenetic comparisons between the individual Ig-like domains of LITRs and mammalian FCRLs shows that these receptors share related Ig-like domains indicative of their common ancestry. However, contrary to our previous reports, no supportive evidence for phylogenetic relationships between the Ig-like domains of LITRs with the Ig-like domains of LRC-encoded mammalian immunoregulatory receptors was found. We also identified an LRC-like region in the zebrafish genome, but no expanded litr-related genes were located in this region. Similarly, no lilr-related genes were found in spotted gar, a representative basal ray-finned fish. Finally, two distantly related fcrls and an LRC-like gene were identified in the elephant shark genome, suggesting that the loss of an immunoregulatory receptor-containing LRC region may be unique to ray-finned fish.

Effect of amino acid composition of elastin-like polypeptide nanoparticles on nonspecific protein adsorption, macrophage cell viability and phagocytosis.

Development of elastin-like polypeptide (ELP) biomaterials is widespread, but information critical for clinical deployment is limited, with biocompatibility studies focused on a narrow cross-section of ELP sequences. Macrophages can impair biomaterial systems by degrading or isolating the biomaterial and by activating additional immune functions. Their phagocytic response will reveal early immune biocompatibility of ELP nanoparticles (NPs). This study examines that response, induced by the adsorbed protein corona, as a function of ELP guest amino acid, chain length and NP diameter. The breadth of proteins adsorbed to ELP NPs varied, with valine-containing ELP NPs adsorbing fewer types of proteins than leucine-containing constructs. Particle diameter was also a factor, with smaller leucine-containing ELP NPs adsorbing the broadest range of proteins. Macrophage viability was unaffected by the ELP NPs, and their phagocytic capabilities were unimpeded except when incubated with a 500 nm valine-containing 40-mer. This NP significantly decreased the phagocytic capacity of macrophages relative to the control and to a corresponding 500 nm leucine-containing 40-mer. NP size and the proportion of opsonin to dysopsonin proteins likely influenced this outcome. These results suggest that certain combinations of ELP sequence and particle size can result in an adsorbed protein corona, which may hinder macrophage function.

Coagulation-flocculation followed by catalytic ozonation processes for enhanced primary treatment during wet weather conditions.

Combined sewer overflows (CSO), generated during the wet weather flow from the combination of the inflow and stormwater runoff in sewer system, result in an overflow of untreated wastewater from sewer system, which might ultimately contain different micropollutants (MPs). In this study, a coagulation-flocculation-sedimentation (CFS) pretreated CSO spiked with MPs was treated by catalytic ozonation using carbon, iron, and peroxide-based catalysts. The catalysts were characterized and their activity on MPs removal was studied at two different ozone (O3) doses (5 and 10 mg L-1). The effect of the treatment on the spiked CSO effluent was also assessed from the acute toxicity of the effluent using Microtox®, Yeast, and Macrophage cell-line toxicity assay tests. All the carbon-based catalysts showed large surface area, which was strongly influenced by the activation technique in the preparation of the catalysts. The CFS treatment strongly reduced the turbidity (≥60%) but had marginal effect on the UV254, dissolved organic carbon (DOC), and pH. Sludge Based Carbon (SBC) showed strong adsorption capacity (≥60% removal efficiency) for all MPs studied compared to other carbon and iron-based catalysts. Ozonation alone was effective for the degradation of easily oxidizable MPs (sulfamethoxazole, mecoprop, and 2,4-dichlorophenoxyl acetic acid), achieving more than 80% degradation efficiency at 10 mg L-1 of ozone, but not effective for atrazine (≤60% degradation efficiency) at similar O3 dose. Catalytic ozonation (at 10 mg L-1 O3 dose) improved the degradation of the MPs at low catalyst dosage but higher dosage strongly inhibited their degradation. In all cases, the effluents showed negligible acute toxicity, indicating the suitability of the process for the treatment of CSO.

An AUTS2-Polycomb complex activates gene expression in the CNS.

Naturally occurring variations of Polycomb repressive complex 1 (PRC1) comprise a core assembly of Polycomb group proteins and additional factors that include, surprisingly, autism susceptibility candidate 2 (AUTS2). Although AUTS2 is often disrupted in patients with neuronal disorders, the mechanism underlying the pathogenesis is unclear. We investigated the role of AUTS2 as part of a previously identified PRC1 complex (PRC1-AUTS2), and in the context of neurodevelopment. In contrast to the canonical role of PRC1 in gene repression, PRC1-AUTS2 activates transcription. Biochemical studies demonstrate that the CK2 component of PRC1-AUTS2 neutralizes PRC1 repressive activity, whereas AUTS2-mediated recruitment of P300 leads to gene activation. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) demonstrated that AUTS2 regulates neuronal gene expression through promoter association. Conditional targeting of Auts2 in the mouse central nervous system (CNS) leads to various developmental defects. These findings reveal a natural means of subverting PRC1 activity, linking key epigenetic modulators with neuronal functions and diseases.

A Fish Leukocyte Immune-Type Receptor Uses a Novel Intracytoplasmic Tail Networking Mechanism to Cross-Inhibit the Phagocytic Response.

Channel catfish (Ictalurus punctatus) leukocyte immune-type receptors (IpLITRs) are a family of immunoregulatory proteins shown to regulate several innate immune cell effector responses, including phagocytosis. The precise mechanisms of IpLITR-mediated regulation of the phagocytic process are not entirely understood, but we have previously shown that different IpLITR-types use classical as well as novel pathways for controlling immune cell-mediated target engulfment. To date, all functional assessments of IpLITR-mediated regulatory actions have focused on the independent characterization of select IpLITR-types in transfected cells. As members of the immunoglobulin superfamily, many IpLITRs share similar extracellular Ig-like domains, thus it is possible that various IpLITR actions are influenced by cross-talk mechanisms between different IpLITR-types; analogous to the paired innate receptor paradigm in mammals. Here, we describe in detail the co-expression of different IpLITR-types in the human embryonic AD293 cell line and examination of their receptor cross-talk mechanisms during the regulation of the phagocytic response using imaging flow cytometry, confocal microscopy, and immunoprecipitation protocols. Overall, our data provides interesting new insights into the integrated control of phagocytosis via the antagonistic networking of independent IpLITR-types that requires the selective recruitment of inhibitory signaling molecules for the initiation and sustained cross-inhibition of phagocytosis.

Exposure to Organic Fraction Extracted from Oil Sands Process-Affected Water Has Negligible Impact on Pregnancy and Lactation of Mice.

Dissolved organic compounds are major contaminants in oil sands process-affected water (OSPW), of which naphthenic acids (NAs) are one of the main persistent toxicants. In the present study, we explore the toxic effects of the organic fraction extracted from OSPW (OSPW-OF) in mice during pregnancy and lactation. Here, we report that acute oral exposure of female Balb/c mice during gestation, and subchronic exposure throughout gestation and lactation to OSPW-OF (containing naturally occurring levels of NAs found in tailings ponds), had negligible effects on their reproductive performance. Specifically, mating behavior, pregnancy success, embryonic implantation, gestation period, litter size, and offspring viability were not affected by OSPW-OF containing up to 55 mg/L NAs. OSPW-OF exposure also did not affect plasma concentrations of pregnancy-associated hormones or pro- and anti-inflammatory cytokines, and it had minimal effects on liver stress gene expression. This study presents the first comprehensive in vivo analysis of mammalian toxicity associated with OSPW-OF exposure. Overall, our results suggest that the risk of acute and subchronic toxicity to mice exposed to OSPW-OF at environmentally relevant concentrations of NAs in contaminated drinking water is likely negligible.

Connected component masking accurately identifies the ratio of phagocytosed and cell-bound particles in individual cells by imaging flow cytometry.

Innate immune cell-mediated recognition, capture, and engulfment of large particulate targets such as bacteria is known as phagocytosis. This highly dynamic cellular process involves a series of steps including receptor-mediated target binding, phagocytic cup formation, pseudopod extension, and phagosome closure, which depend on distinct actin polymerization events. Using flow cytometry, precise determination of target locations relative to cell membranes (i.e., surface-bound vs. fully engulfed/internalized) during the phagocytic process is difficult to quantify. Here, we describe the application of new analysis features within the IDEAS® software to distinguish internalized and surface-bound particles on individual cells with a high degree of accuracy and reproducibility. Through the use of connected component masks, the accurate discrimination of surface-bound beads versus those internalized is clearly demonstrated. In addition, we were able to further analyze the ratio of beads that had been surface-bound or internalized within individual cells. This novel method of analyzing the phagocytic process provides more accurate determination of target-cell interactions that will assist in examination of the signalling events that occur during the various stages of phagocytosis. © 2017 International Society for Advancement of Cytometry.

Comparison of the Acute Immunotoxicity of Nonfractionated and Fractionated Oil Sands Process-Affected Water Using Mammalian Macrophages.

OSPW is a complex mixture of inorganic and organic substances and its principal toxic components have yet to be fully characterized. Previously, we showed in vitro that the oil sands process-affected water (OSPW) organic fraction (OF) caused a concentration-dependent immunotoxicity in mammals. In the present study we further explore the immunotoxicological properties of OSPW in mammals using a series of in vitro bioassays. Specifically, using the RAW 264.7 mouse macrophage cell line we show that whole OSPW containing naphthenic acid (NA) concentrations ranging from 12 to 18 mg/L, significantly inhibited cell proliferation, reduced cell viability, and was directly cytotoxic, whereas the exposure of cells to equivalent doses of the OSPW-OF had no measurable effects. Whole OSPW exposures also caused morphological changes in RAW 264.7 cells, and at sublethal doses (i.e., 10 mg/L) it induced the early expression of the stress genes hmox1 and gadd45. In addition, at NA concentrations of 10 mg/L, whole OSPW but not the OSPW-OF had significant effects on pro-inflammatory cytokine mRNA levels and cytokine protein secretion activities. Finally, whole OSPW also impaired the ability of RAW 264.7 cells to perform phagocytosis. Overall, we demonstrate that exposure to whole OSPW (at NA doses ranging from 10 to 20 mg/L), but not the OSPW-OF caused both cytotoxic and immunomodulatory changes in mouse macrophages. This suggests that the complex mixture of inorganic and organic components found in whole OSPW are acutely toxic at much lower doses than we previously reported for the OSPW-OF (i.e., 50 mg/L) due to unknown additive and/or synergistic interactions that likely occur between the various components present in whole OSPW.

Large-scale topology and the default mode network in the mouse connectome.

Noninvasive functional imaging holds great promise for serving as a translational bridge between human and animal models of various neurological and psychiatric disorders. However, despite a depth of knowledge of the cellular and molecular underpinnings of atypical processes in mouse models, little is known about the large-scale functional architecture measured by functional brain imaging, limiting translation to human conditions. Here, we provide a robust processing pipeline to generate high-resolution, whole-brain resting-state functional connectivity MRI (rs-fcMRI) images in the mouse. Using a mesoscale structural connectome (i.e., an anterograde tracer mapping of axonal projections across the mouse CNS), we show that rs-fcMRI in the mouse has strong structural underpinnings, validating our procedures. We next directly show that large-scale network properties previously identified in primates are present in rodents, although they differ in several ways. Last, we examine the existence of the so-called default mode network (DMN)--a distributed functional brain system identified in primates as being highly important for social cognition and overall brain function and atypically functionally connected across a multitude of disorders. We show the presence of a potential DMN in the mouse brain both structurally and functionally. Together, these studies confirm the presence of basic network properties and functional networks of high translational importance in structural and functional systems in the mouse brain. This work clears the way for an important bridge measurement between human and rodent models, enabling us to make stronger conclusions about how regionally specific cellular and molecular manipulations in mice relate back to humans.

Channel catfish (Ictalurus punctatus) leukocytes express estrogen receptor isoforms ERα and ERß2 and are functionally modulated by estrogens.

Estrogens are recognized as modulators of immune responses in mammals and teleosts. While it is known that the effects of estrogens are mediated via leukocyte-specific estrogen receptors (ERs) in humans and mice, leucocyte-specific estrogen receptor expression and the effects of estrogens on this cell population is less explored and poorly understood in teleosts. Here in, we verify that channel catfish (Ictalurus punctaus) leukocytes express ERα and ERß2. Transcripts of these isoforms were detected in tissue-associated leukocyte populations by PCR, but ERß2 was rarely detected in PBLs. Expression of these receptors was temporally regulated in PBLs following polyclonal activation by concanavalin A, lipopolysaccharide or alloantigen based on evaluation by quantitative and end-point PCR. Examination of long-term leukocyte cell lines demonstrated that these receptors are differentially expressed depending on leukocyte lineage and phenotype. Expression of ERs was also temporally dynamic in some leukocyte lineages and may reflect stage of cell maturity. Estrogens affect the responsiveness of channel catfish peripheral blood leukocytes (PBLs) to mitogens in vitro. Similarly, bactericidal activity and phorbol 12-myristate 13-acetate induced respiratory burst was modulated by 17ß-estradiol. These actions were blocked by the pure ER antagonist ICI 182780 indicating that response is, in part, mediated via ERα. In summary, estrogen receptors are expressed in channel catfish leukocytes and participate in the regulation of the immune response. This is the first time leukocyte lineage expression has been reported in teleost cell lines.

Vocoder simulations of highly focused cochlear stimulation with limited dynamic range and discriminable steps.

OBJECTIVES: There is recent interest in focused stimulation of the cochlea via modalities such as tripolar electrical and infrared neural stimulation to improve speech in noise comprehension and music perception. The purpose of this work was to use vocoder-based simulations to investigate speech recognition for broad stimulation (standard monopolar paradigm) versus more focused stimulation under a variety of signal-to-noise ratios, dynamic ranges, and numbers of discriminable loudness steps. DESIGN: Vocoder simulations were used to assess the intelligibility of sentences, consonants, and vowels that were noise vocoded and presented to 7 normal-hearing listeners for identification. A novel aspect of the simulations presented here was the use of nonuniform quantization steps within the dynamic range to more closely simulate the Weber functions observed in cochlear implant users. Intelligibility was assessed for the different filter slopes under a variety of signal-to-noise ratio levels, dynamic ranges, and numbers of discriminable steps. RESULTS: Speech processed via vocoder simulations representing focused stimulation was found to be substantially more intelligible than speech processed via a monopolar electric vocoder simulation, with differences of up to 60 percentage points. There were no significant differences, however, seen between the two focused approaches (signal attenuations of 10 and 17 dB/mm) for the conditions investigated. Speech processed via the highly focused vocoder (17 dB/mm) was robust to constraints on small envelope dynamic range and small number of discriminable steps within the dynamic range, as high performance was maintained with at least a 5 dB dynamic range and eight or more discriminable steps. Significant drops in intelligibility were noted when the number of steps fell below eight. CONCLUSIONS: Highly focused stimulation-tripolar electrical and infrared neural stimulation-has potential for increased performance in noise compared with monopolar stimulation, but much work remains to bear this potential out and to take full advantage of each modality's strengths.