Fish Physiologically Based Toxicokinetic Modeling Approach for In Vitro-In Vivo and Cross-Species Extrapolation of Endocrine-Disrupting Chemicals in Risk Assessment.

High-throughput in vitro assays combined with in vitro-in vivo extrapolation (IVIVE) leverage in vitro responses to predict the corresponding in vivo exposures and thresholds of concern. The integrated approach is also expected to offer the potential for efficient tools to provide estimates of chemical toxicity to various wildlife species instead of animal testing. However, developing fish physiologically based toxicokinetic (PBTK) models for IVIVE in ecological applications is challenging, especially for plausible estimation of an internal effective dose, such as fish equivalent concentration (FEC). Here, a fish PBTK model linked with the IVIVE approach was established, with parameter optimization of chemical unbound fraction, pH-dependent ionization and hepatic clearance, and integration of temperature effect and growth dilution. The fish PBTK-IVIVE approach provides not only a more precise estimation of tissue-specific concentrations but also a reasonable approximation of FEC targeting the estrogenic potency of endocrine-disrupting chemicals. Both predictions were compared with in vivo data and were accurate for most indissociable/dissociable chemicals. Furthermore, the model can help determine cross-species variability and sensitivity among the five fish species. Using the available IVIVE-derived FEC with target pathways is helpful to develop predicted no-effect concentration for chemicals with similar mode of action and support screening-level ecological risk assessment.

Calretinin-Expressing Synapses Show Improved Synaptic Efficacy with Reduced Asynchronous Release during High-Rate Activity.

Calretinin (CR) is a major calcium binding protein widely expressed in the CNS. However, its synaptic function remains largely elusive. At the auditory synapse of the endbulb of Held, CR is selectively expressed in different subtypes. Combining electrophysiology with immunohistochemistry, we investigated the synaptic transmission at the endbulb of Held synapses with and without endogenous CR expression in mature CBA/CAJ mice of either sex. Two synapse subtypes showed similar basal synaptic transmission, except a larger quantal size in CR-expressing synapses. During high-rate stimulus trains, CR-expressing synapses showed improved synaptic efficacy with significantly less depression and lower asynchronous release, suggesting more efficient exocytosis than non-CR-expressing synapses. Conversely, CR-expressing synapses had a smaller readily releasable pool size, which was countered by higher release probability and faster synaptic recovery to support sustained release during high-rate activity. EGTA-AM treatment did not change the synaptic transmission of CR-expressing synapses, but reduced synaptic depression and decreased asynchronous release at non-CR-expressing synapses, suggesting that CR helps to minimize calcium accumulation during high-rate activity. Both synapses express parvalbumin, another calcium-binding protein with slower kinetics and higher affinity than CR, but not calbindin. Furthermore, CR-expressing synapses only express the fast isoform of vesicular glutamate transporter 1 (VGluT1), while most non-CR-expressing synapses express both VGluT1 and the slower VGluT2, which may underlie their lagged synaptic recovery. The findings suggest that, paired with associated synaptic machinery, differential CR expression regulates synaptic efficacy among different subtypes of auditory nerve synapses to accomplish distinctive physiological functions in transmitting auditory information at high rates.SIGNIFICANCE STATEMENT CR is a major calcium-binding protein in the brain. It remains unclear how endogenous CR impacts synaptic transmission. We investigated the question at the large endbulb of Held synapses with selective CR expression and found that CR-expressing and non-CR-expressing synapses had similar release properties under basal synaptic transmission. During high-rate activity, however, CR-expressing synapses showed improved synaptic efficacy with less depression, lower asynchronous release, and faster recovery. Furthermore, CR-expressing synapses use exclusive VGluT1 to refill synaptic vesicles, while non-CR-expressing synapses use both VGluT1 and the slower isoform of VGluT2. Our findings suggest that CR may play significant roles in promoting synaptic efficacy during high-rate activity, and selective CR expression can differentially impact signal processing among different synapses.

Neural Adaptation of the Electrically Stimulated Auditory Nerve Is Not Affected by Advanced Age in Postlingually Deafened, Middle-aged, and Elderly Adult Cochlear Implant Users.

OBJECTIVE: This study aimed to investigate the associations between advanced age and the amount and the speed of neural adaptation of the electrically stimulated auditory nerve (AN) in postlingually deafened adult cochlear implant (CI) users. DESIGN: Study participants included 26 postlingually deafened adult CI users, ranging in age between 28.7 and 84.0 years (mean: 63.8 years, SD: 14.4 years) at the time of testing. All study participants used a Cochlear Nucleus device with a full electrode array insertion in the test ear. The stimulus was a 100-ms pulse train with a pulse rate of 500, 900, 1800, or 2400 pulses per second (pps) per channel. The stimulus was presented at the maximum comfortable level measured at 2400 pps with a presentation rate of 2 Hz. Neural adaptation of the AN was evaluated using electrophysiological measures of the electrically evoked compound action potential (eCAP). The amount of neural adaptation was quantified by the adaptation index (AI) within three time windows: around 0 to 8 ms (window 1), 44 to 50 ms (window 2), and 94 to 100 ms (window 3). The speed of neural adaptation was quantified using a two-parameter power law estimation. In 23 participants, four electrodes across the electrode array were tested. In three participants, three electrodes were tested. Results measured at different electrode locations were averaged for each participant at each pulse rate to get an overall representation of neural adaptation properties of the AN across the cochlea. Linear-mixed models (LMMs) were used (1) to evaluate the effects of age at testing and pulse rate on the speed of neural adaptation and (2) to assess the effects of age at testing, pulse rate, and duration of stimulation (i.e., time window) on the amount of neural adaptation in these participants. RESULTS: There was substantial variability in both the amount and the speed of neural adaptation of the AN among study participants. The amount and the speed of neural adaptation increased at higher pulse rates. In addition, larger amounts of adaptation were observed for longer durations of stimulation. There was no significant effect of age on the speed or the amount of neural adaptation. CONCLUSIONS: The amount and the speed of neural adaptation of the AN are affected by both the pulse rate and the duration of stimulation, with higher pulse rates and longer durations of stimulation leading to faster and greater neural adaptation. Advanced age does not affect neural adaptation of the AN in postlingually deafened, middle-aged and elderly adult CI users.

Oxidative Damage in Roots of Rice (Oryza sativa L.) Seedlings Exposed to Microplastics or Combined with Cadmium.

This study aimed to investigate the effect of 10-40 mg L-1 polystyrene microplastics (PS-MPs), 0.05 mg L-1 cadmium (Cd) and their combination on the growth and related physiological and toxicological responses in Oryza sativa L. seedling roots. Results showed that the fresh weight, dry weight and root lengths of treatments by PS-MPs, Cd single and combinative were all lower than the control, and opposite phenomenon appeared in production of superoxide radical (O2-.), malondialdehyde (MDA) and carbonylated protein. Superoxide dismutase (SOD) and guaiacol peroxidase (POD) activities induced by 10-40 mg L-1 PS-MPs and combination with Cd were almost higher than those by Cd alone, expression of heat shock protein (HSP)70 and carbonylated protein slightly decreased. In compound exposure, 10-20 mg L-1 PS-MPs alleviated Cd damage and promoted root growth by increasing SOD and POD activities, but 40 mg L-1 PS-MPs accelerated the accumulation of Cd, MDA, and O2-., which was responsible for decreasing root biomass and the aggravating necrosis of root tip cells.

Biased auditory nerve central synaptopathy is associated with age-related hearing loss.

KEY POINTS: Sound information is transmitted by different subtypes of spiral ganglion neurons (SGN) from the ear to the brain. Selective damage of SGN peripheral synapses (cochlear synaptopathy) is widely recognized as one of the primary mechanisms of hearing loss, whereas the mechanisms at the SGN central synapses remain unclear. We report that different subtypes of SGN central synapses converge at different ratios onto individual target cochlear nucleus neurons with distinct physiological properties, and show biased morphological and physiological changes during age-related hearing loss (ARHL). The results reveal a new dimension in cochlear nucleus neural circuitry that systematically reassembles and processes auditory information from different SGN subtypes, which is altered during ageing and probably contributes to the development of ARHL. In addition to known cochlear synaptopathy, the present study shows that SGN central synapses are also pathologically changed during ageing, which collectively helps us better understand the structure and function of SGNs during ARHL. ABSTRACT: Sound information is transmitted from the cochlea to the brain by different subtypes of spiral ganglion neurons (SGN), which show varying degrees of vulnerability under pathological conditions. Selective cochlear synaptopathy, the preferential damage of certain subtypes of SGN peripheral synapses, has been recognized as one of the main mechanisms of hearing loss. The organization and function of the auditory nerve (AN) central synapses from different subtypes of SGNs remain unclear, including how different AN synapses reassemble onto individual neurons in the cochlear nucleus, as well as how they differentially change during hearing loss. Combining immunohistochemistry with electrophysiology, we investigated the convergence pattern and subtype-specific synaptopathy of AN synapses at the endbulb of Held, as well as the response properties of their postsynaptic bushy neurons in CBA/CaJ mice of either sex under normal hearing and age-related hearing loss (ARHL). We found that calretinin-expressing (type Ia ) and non-calretinin-expressing (type Ib /Ic ) endbulbs converged along a continuum of different ratios onto individual bushy neurons with varying physiological properties. Endbulbs degenerated during ageing in parallel with ARHL. Furthermore, the degeneration was more severe in non-calretinin-expressing synapses, which correlated with a gradual decrease in bushy neuron subpopulation predominantly innervated by these inputs. These synaptic and cellular changes were profound in middle-aged mice when their hearing thresholds were still relatively normal and prior to severe ARHL. Our findings suggest that biased AN central synaptopathy and the correlated shift in cochlear nucleus neuronal composition play significant roles in weakened auditory input and altered central auditory processing during ARHL.

eDNA metabarcoding revealed differential structures of aquatic communities in a dynamic freshwater ecosystem shaped by habitat heterogeneity.

Freshwater ecosystems have been threatened by complicated disturbances from both natural and anthropogenic variables, especially in dynamic and complex river basins. The environmental DNA (eDNA)-based approach provides a broader spectrum and higher throughput way of biomonitoring for biodiversity assessment compared with traditional morphological survey. Most eDNA metabarcoding studies have been limited to a few specific taxa/groups and habitat scopes. Here we applied the eDNA metabarcoding to characterize the structures and spatial variations of zooplankton and fish communities among different habitat types in a highly dynamic and complex freshwater ecosystem of the Daqing River basin (DRB). The results showed that varied species spectra of zooplankton and fish communities were identified and unique dominant species occurred across habitats. Additionally, markedly spatial distributions of biotic community structures were observed in areas with different habitat characteristics. Natural variables, including geographic distances and gradient ratio, as well as anthropogenic factors of chemical oxygen demand (COD) and organic chemicals demonstrated significant effects but different outcomes on the structures of zooplankton and fish communities. Moreover, the relative abundances of specific aquatic taxa were associated with the gradient of particular environmental variables. This case study verified the distribution patterns and differentiation mechanisms of biotic communities under habitat heterogeneity could be captured by application of eDNA biomonitoring. And habitat-specific and even species-specific environmental stressors would be diagnosed for improving management of complex river basins.

Synaptic transmission at the endbulb of Held deteriorates during age-related hearing loss.

KEY POINTS: Synaptic transmission at the endbulb of Held was assessed by whole-cell patch clamp recordings from auditory neurons in mature (2-4 months) and aged (20-26 months) mice. Synaptic transmission is degraded in aged mice, which may contribute to the decline in neural processing of the central auditory system during age-related hearing loss. The changes in synaptic transmission in aged mice can be partially rescued by improving calcium buffering, or decreasing action potential-evoked calcium influx. These experiments suggest potential mechanisms, such as regulating intraterminal calcium, that could be manipulated to improve the fidelity of transmission at the aged endbulb of Held. ABSTRACT: Age-related hearing loss (ARHL) is associated with changes to the auditory periphery that raise sensory thresholds and alter coding, and is accompanied by alterations in excitatory and inhibitory synaptic transmission, and intrinsic excitability in the circuits of the central auditory system. However, it remains unclear how synaptic transmission changes at the first central auditory synapses during ARHL. Using mature (2-4 months) and old (20-26 months) CBA/CaJ mice, we studied synaptic transmission at the endbulb of Held. Mature and old mice showed no difference in either spontaneous quantal synaptic transmission or low frequency evoked synaptic transmission at the endbulb of Held. However, when challenged with sustained high frequency stimulation, synapses in old mice exhibited increased asynchronous transmitter release and reduced synchronous release. This suggests that the transmission of temporally precise information is degraded at the endbulb during ARHL. Increasing intraterminal calcium buffering with EGTA-AM or decreasing calcium influx with ω-agatoxin IVA decreased the amount of asynchronous release and restored synchronous release in old mice. In addition, recovery from depression following high frequency trains was faster in old mice, but was restored to a normal time course by EGTA-AM treatment. These results suggest that intraterminal calcium in old endbulbs may rise to abnormally high levels during high rates of auditory nerve firing, or that calcium-dependent processes involved in release are altered with age. These observations suggest that ARHL is associated with a decrease in temporal precision of synaptic release at the first central auditory synapse, which may contribute to perceptual deficits in hearing.

A High Performance Piezoelectric Sensor for Dynamic Force Monitoring of Landslide.

Due to the increasing influence of human engineering activities, it is important to monitor the transient disturbance during the evolution process of landslide. For this purpose, a high-performance piezoelectric sensor is presented in this paper. To adapt the high static and dynamic stress environment in slope engineering, two key techniques, namely, the self-structure pressure distribution method (SSPDM) and the capacitive circuit voltage distribution method (CCVDM) are employed in the design of the sensor. The SSPDM can greatly improve the compressive capacity and the CCVDM can quantitatively decrease the high direct response voltage. Then, the calibration experiments are conducted via the independently invented static and transient mechanism since the conventional testing machines cannot match the calibration requirements. The sensitivity coefficient is obtained and the results reveal that the sensor has the characteristics of high compressive capacity, stable sensitivities under different static preload levels and wide-range dynamic measuring linearity. Finally, to reduce the measuring error caused by charge leakage of the piezoelectric element, a low-frequency correction method is proposed and experimental verified. Therefore, with the satisfactory static and dynamic properties and the improving low-frequency measuring reliability, the sensor can complement dynamic monitoring capability of the existing landslide monitoring and forecasting system.

Transmission of auditory sensory information decreases in rate and temporal precision at the endbulb of Held synapse during age-related hearing loss.

Age-related hearing loss (ARHL) is largely attributed to structural changes and functional declines in the peripheral auditory system, which include synaptopathy at the inner hair cell/spiral ganglion cell (SGC) connection and the loss of SGCs. However, functional changes at the central terminals of SGCs, namely the auditory nerve synapses in the cochlear nucleus, are not yet fully understood during ARHL. With the use of young (1-3 mo) and old (25-30 mo) CBA/CaJ mice, this study evaluated the intrinsic properties of the bushy neurons postsynaptic to the endbulb of Held synapses, and the firing properties of these neurons to direct current injections as well as to synaptic inputs from the auditory nerve. Results showed that bushy neurons in old mice are more excitable and are able to fire spikes at similar rate and timing to direct current injections as those in young mice. In response to synaptic inputs, however, bushy neurons from old mice fired spikes with significantly decreased rate and reduced temporal precision to stimulus trains at 100 and 400 Hz, with the drop in firing probability more profound at 400 Hz. It suggests that transmission of auditory information at the endbulb is declined in both rate and timing during aging, which signifies the loss of sensory inputs to the central auditory system under ARHL. The study proposes that, in addition to damages at the peripheral terminals of SGCs as well as the loss of SGCs, functional decline at the central terminals of surviving SGCs is also an essential component of ARHL.

Target-specific IPSC kinetics promote temporal processing in auditory parallel pathways.

The acoustic environment contains biologically relevant information on timescales from microseconds to tens of seconds. The auditory brainstem nuclei process this temporal information through parallel pathways that originate in the cochlear nucleus from different classes of cells. Although the roles of ion channels and excitatory synapses in temporal processing have been well studied, the contribution of inhibition is less well understood. Here, we show in CBA/CaJ mice that the two major projection neurons of the ventral cochlear nucleus, the bushy and T-stellate cells, receive glycinergic inhibition with different synaptic conductance time courses. Bushy cells, which provide precisely timed spike trains used in sound localization and pitch identification, receive slow inhibitory inputs. In contrast, T-stellate cells, which encode slower envelope information, receive inhibition that is eightfold faster. Both types of inhibition improved the precision of spike timing but engage different cellular mechanisms and operate on different timescales. Computer models reveal that slow IPSCs in bushy cells can improve spike timing on the scale of tens of microseconds. Although fast and slow IPSCs in T-stellate cells improve spike timing on the scale of milliseconds, only fast IPSCs can enhance the detection of narrowband acoustic signals in a complex background. Our results suggest that target-specific IPSC kinetics are critical for the segregated parallel processing of temporal information from the sensory environment.

Mechanisms of age-related hearing loss at the auditory nerve central synapses and postsynaptic neurons in the cochlear nucleus.

Sound information is transduced from mechanical vibration to electrical signals in the cochlea, conveyed to and further processed in the brain to form auditory perception. During the process, spiral ganglion neurons (SGNs) are the key cells that connect the peripheral and central auditory systems by receiving information from hair cells in the cochlea and transmitting it to neurons of the cochlear nucleus (CN). Decades of research in the cochlea greatly improved our understanding of SGN function under normal and pathological conditions, especially about the roles of different subtypes of SGNs and their peripheral synapses. However, it remains less clear how SGN central terminals or auditory nerve (AN) synapses connect to CN neurons, and ultimately how peripheral pathology links to structural alterations and functional deficits in the central auditory nervous system. This review discusses recent progress about the morphological and physiological properties of different subtypes of AN synapses and associated postsynaptic CN neurons, their changes during aging, and the potential mechanisms underlying age-related hearing loss.

Glycinergic synaptic transmission in the cochlear nucleus of mice with normal hearing and age-related hearing loss.

The principal inhibitory neurotransmitter in the mammalian cochlear nucleus (CN) is glycine. During age-related hearing loss (AHL), glycinergic inhibition becomes weaker in CN. However, it is unclear what aspects of glycinergic transmission are responsible for weaker inhibition with AHL. We examined glycinergic transmission onto bushy cells of the anteroventral CN in normal-hearing CBA/CaJ mice and in DBA/2J mice, a strain that exhibits an early onset AHL. Glycinergic synaptic transmission was examined in brain slices of mice at 10-15 postnatal days old, 20-35 days old, and at 6-7 mo old. Spontaneous inhibitory postsynaptic current (sIPSC) event frequency and amplitude were the same among all three ages in both strains of mice. However, the amplitudes of IPSCs evoked (eIPSC) from stimulating the dorsal CN were smaller, and the failure rate was higher, with increasing age due to decreased quantal content in both mouse strains, independent of hearing status. The coefficient of variation of the eIPSC amplitude also increased with age. The decay time constant (τ) of sIPSCs and eIPSCs were constant in CBA/CaJ mice at all ages, but were significantly slower in DBA/2J mice at postnatal days 20-35, following the onset of AHL, and not at earlier or later ages. Our results suggest that glycinergic inhibition at the synapses onto bushy cells becomes weaker and less reliable with age through changes in release. However, the hearing loss in DBA/2J mice is accompanied by a transiently enhanced inhibition, which could disrupt the balance of excitation and inhibition.

Apical-basal distribution of different subtypes of spiral ganglion neurons in the cochlea and the changes during aging.

Sound information is transmitted from the cochlea to the brain mainly by type I spiral ganglion neurons (SGNs), which consist of different subtypes with distinct physiological properties and selective expression of molecular markers. It remains unclear how these SGN subtypes distribute along the tonotopic axis, and whether the distribution pattern changes during aging that might underlie age-related hearing loss (ARHL). We investigated these questions using immunohistochemistry in three age groups of CBA/CaJ mice of either sex, including 2-5 months (young), 17-19 months (middle-age), and 28-32 months (old). Mouse cochleae were cryo-sectioned and triple-stained using antibodies against Tuj1, calretinin (CR) and calbindin (CB), which are reportedly expressed in all type I, subtype Ia, and subtype Ib SGNs, respectively. Labeled SGNs were classified into four groups based on the expression pattern of stained markers, including CR+ (subtype Ia), CB+ (subtype Ib), CR+CB+ (dual-labeled Ia/Ib), and CR-CB- (subtype Ic) neurons. The distribution of these SGN groups was analyzed in the apex, middle, and base regions of the cochleae. It showed that the prevalence of subtype Ia, Ib and dual-labeled Ia/Ib SGNs are high in the apex and low in the base. In contrast, the distribution pattern is reversed in Ic SGNs. Such frequency-dependent distribution is largely maintained during aging except for a preferential reduction of Ic SGNs, especially in the base. These findings corroborate the prior study based on RNAscope that SGN subtypes show differential vulnerability during aging. It suggests that sound processing of different frequencies involves distinct combinations of SGN subtypes, and the age-dependent loss of Ic SGNs in the base may especially impact high-frequency hearing during ARHL.

Dendritic Degeneration and Altered Synaptic Innervation of a Central Auditory Neuron During Age-related Hearing Loss.

Cellular morphology and synaptic configuration are key determinants of neuronal function and are often modified under pathological conditions. In the first nucleus of the central auditory system, the cochlear nucleus (CN), principal bushy neurons specialize in processing temporal information of sound critical for hearing. These neurons alter their physiological properties during aging that contribute to age-related hearing loss (ARHL). The structural basis of such changes remains unclear, especially age-related modifications in their dendritic morphology and the innervating auditory nerve (AN) synapses. Using young (2-5 months) and aged (28-33 months) CBA/CaJ mice of either sex, we filled individual bushy neurons with fluorescent dye in acute brain slices to characterize their dendritic morphology, followed by immunostaining against vesicular glutamate transporter 1 (VGluT1) and calretinin (CR) to identify innervating AN synapses. We found that dendritic morphology of aged bushy neurons had significantly reduced complexity, suggesting age-dependent dendritic degeneration, especially in neurons with predominantly non-CR-expressing synapses on the soma. These dendrites were innervated by AN bouton synapses, which were predominantly non-CR-expressing in young mice but had increased proportion of CR-expressing synapses in old mice. While somatic AN synapses degenerated substantially with age, as quantified by VGluT1-labeled puncta volume, no significant difference was observed in the total volume of dendritic synapses between young and old mice. Consequently, synaptic density on dendrites was significantly higher in old mice. The findings suggest that dendritic degeneration and altered synaptic innervation in bushy neurons during aging may underlie their changed physiological activity and contribute to the development of ARHL.

Potential uses of auditory nerve stimulation to modulate immune responses in the inner ear and auditory brainstem.

Bioelectronic medicine uses electrical stimulation of the nervous system to improve health outcomes throughout the body primarily by regulating immune responses. This concept, however, has yet to be applied systematically to the auditory system. There is growing interest in how cochlear damage and associated neuroinflammation may contribute to hearing loss. In conjunction with recent findings, we propose here a new perspective, which could be applied alongside advancing technologies, to use auditory nerve (AN) stimulation to modulate immune responses in hearing health disorders and following surgeries for auditory implants. In this article we will: (1) review the mechanisms of inflammation in the auditory system in relation to various forms of hearing loss, (2) explore nerve stimulation to reduce inflammation throughout the body and how similar neural-immune circuits likely exist in the auditory system (3) summarize current methods for stimulating the auditory system, particularly the AN, and (4) propose future directions to use bioelectronic medicine to ameliorate harmful immune responses in the inner ear and auditory brainstem to treat refractory conditions. We will illustrate how current knowledge from bioelectronic medicine can be applied to AN stimulation to resolve inflammation associated with implantation and disease. Further, we suggest the necessary steps to get discoveries in this emerging field from bench to bedside. Our vision is a future for AN stimulation that includes additional protocols as well as advances in devices to target and engage neural-immune circuitry for therapeutic benefits.

First metabolic profiling of 4-n-nonylphenol in human liver microsomes by integrated approaches to testing and assessment: Metabolites, pathways, and biological effects.

4-n-nonylphenol (4-n-NP), a typical endocrine disrupting chemical, has been so far frequently detected in various environmental mediums and editable food. However, the specific metabolic pathways in human and potential adverse effects of metabolites have not been elucidated yet. Here, metabolic profiling of 4-n-NP in human liver microsome (HLM) was comprehensively characterized by integrated approaches of testing and assessment. A total of 21 metabolites were identified using nontarget analysis with high-resolution mass spectrum, including three groups of unique phase I metabolites first determined in HLM. Seven various metabolic pathways of 4-n-NP were identified by both in silico and in vitro, and CYP1A2, 2C19, and 2D6 were the mainly participating enzymes. Two secondary metabolites with carbonyl groups on side chains (M4, M7) presented most abundant in HLM, which were also predicted to have high binding affinities towards HPG-axis-related receptors (AR, ER, and PR). ESRs (estrogen receptors) were shared core protein targets for all metabolites revealed by protein-protein interaction networks. Biological functions enrichment analysis indicated that 4-n-NP metabolites might primarily involve in ESR-mediated signaling, GPCR ligand binding, Class A/1 (Rhodopsin-like receptors) and metabolism-related pathways. These findings of 4-n-NP metabolites, pathways, and biological effects provide insightful information for its environmental exposure and risk assessment.

In vitro to in vivo extrapolation for predicting human equivalent dose of phenolic endocrine disrupting chemicals: PBTK model development, biological pathways, outcomes and performance.

In vitro to in vivo (IVIVE) leverages in vitro high-throughput biological responses to predict the corresponding in vivo exposures and further estimate the human safe dose. However, for phenolic endocrine disrupting chemicals (EDCs) linked with complicated biological pathways and adverse outcomes (AO), such as bisphenol A (BPA) and 4-nonylphenol (4-NP), plausible estimation of human equivalent doses (HED) by IVIVE approaches considering various biological pathways and endpoints is still challenging. To explore the capabilities and limitations of IVIVE, this study conducted physiologically based toxicokinetic (PBTK)-IVIVE approaches to derive pathway-specific HEDs using BPA and 4-NP as examples. In vitro HEDs of BPA and 4-NP varied in different adverse outcomes, pathways, and testing endpoints and ranged from 0.0013 to 1.0986 mg/kg bw/day and 0.0551 to 1.7483 mg/kg bw/day, respectively. In vitro HEDs associated with reproductive AOs initiated by PPARα activation and ER agonism were the most sensitive. Model verification suggested the potential of using effective in vitro data to determine reasonable approximation of in vivo HEDs for the same AO (fold differences of most AOs ranged in 0.14-2.74 and better predictions for apical endpoints). Furthermore, system-specific parameters of cardiac output and its fraction, body weight, as well as chemical-specific parameters of partition coefficient and liver metabolic were most sensitive for the PBTK simulations. The results indicated that the application of fit for-purpose PBTK-IVIVE approach could provide credible pathway-specific HEDs and contribute to high throughput prioritization of chemicals in a more realistic scenario.

Repression of ferroptotic cell death by mitochondrial calcium signaling.

The uptake of Ca2+ into and extrusion of calcium from the mitochondrial matrix, regulated by the mitochondrial Ca2+ uniporter (MCU), is a fundamental biological process that has crucial impacts on cellular metabolism, signaling, growth and survival. Herein, we report that the embryonic lethality of Mcu-deficient mice is fully rescued by orally supplementing ferroptosis inhibitor lipophilic antioxidant vitamin E and ubiquinol. Mechanistically, we found MCU promotes acetyl-CoA-mediated GPX4 acetylation at K90 residue, and K90R mutation impaired the GPX4 enzymatic activity, a step that is crucial for ferroptosis. Structural analysis supports the possibility that GPX4 K90R mutation alters the conformational state of the molecule, resulting in disruption of a salt bridge formation with D23, which was confirmed by mutagenesis studies. Finally, we report that deletion of MCU in cancer cells caused a marked reduction in tumor growth in multiple cancer models. In summary, our study provides a first direct link between mitochondrial calcium level and sustained GPX4 enzymatic activity to regulate ferroptosis, which consequently protects cancer cells from ferroptosis.

An integrated screening strategy for novel AhR agonist candidate identification and toxicity confirmation in sediments.

Organic contaminants showing aryl hydrocarbon receptor (AhR) agonist activity are commonly detected in areas disturbed by intense human activities and they can initiate a variety of biochemical, physiological, and toxicological effects. A new integrated screening strategy for AhR agonist candidate identification and toxicity confirmation was developed to characterize the AhR-active pollutants in sediments of the contaminated Daqing River basin (DRB) in North China. The specific objectives were to (i) determine the concentrations of known AhR agonists, (ii) identify the novel AhR agonist candidates from nontarget screening (NTS) with structure alerts, computational toxicology (CompTox) Dashboard bioassays, and in silico predictions, and (iii) evaluate contributions of AhR agonists to the overall potencies and characterize the distribution and source of these pollutants. Significant AhR-mediated potencies were observed in all sediment extracts by in vitro bioassays. Concentrations of polar target chemicals in sediment extracts were much lower than nonpolar target chemicals. A total of 19 known AhR agonists explained 11.3 % to 49.1 % of bioassay-derived AhR-mediated potencies and polychlorinated biphenyls (PCB) 126 and PCB169 were found to contribute significantly to the total effects. 21 compounds screened from NTS by AhR-related structure alerts and further confirmed toxicity by high-throughput bioassays and in silico predictions were selected as AhR agonist candidates. Most of them were substituted PAHs, biphenyls, quinones, substituted phenols and heterocyclic compounds, and they primarily originated from nearby manufacturing industries. Of these compounds, 1-methy-pyrene exhibited significant AhR-mediated potency. Follow up studies should focus on toxicological mechanism, source, and fate of these novel AhR agonists in water environment.

Age-Related Hearing Loss Is Accompanied by Chronic Inflammation in the Cochlea and the Cochlear Nucleus.

Age-related hearing loss (ARHL) is a major hearing impairment characterized by pathological changes in both the peripheral and central auditory systems. Low-grade inflammation was observed in the cochlea of deceased human subjects with ARHL and animal models of early onset ARHL, which suggests that inflammation contributes to the development of ARHL. However, it remains elusive how chronic inflammation progresses during normal aging in the cochlea, and especially the accompanying changes of neuroinflammation in the central auditory system. To address this, we investigated chronic inflammation in both the cochlea and the cochlear nucleus (CN) of CBA/CaJ mice, an inbred mouse strain that undergoes normal aging and develops human, like-late-onset ARHL. Using immunohistochemistry, confocal microscopy, and quantitative image processing, we measured the accumulation and activation of macrophages in the cochlea and microglia in the CN using their shared markers: ionized calcium binding adaptor molecule 1 (Iba1) and CD68-a marker of phagocytic activity. We found progressive increases in the area covered by Iba1-labeled macrophages and enhanced CD68 staining in the osseous spiral lamina of the cochlea that correlated with elevated ABR threshold across the lifespan. During the process, we further identified significant increases in microglial activation and C1q deposition in the CN, indicating increased neuroinflammation and complement activation in the central auditory system. Our study suggests that during normal aging, chronic inflammation occurs in both the peripheral and the central auditory system, which may contribute in coordination to the development of ARHL.